Sucrose 2D NMR Spectra

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Apr 062014

Sucrose 2D NMR Spectra

The sugar sucrose can be used to illustrate homonuclear 2D NMR experiment: the TOCSY.

Sucrose

Table sugar

Sucrose (“table sugar”) is a disaccharide derived from glucose and fructose.

The interesting element from an NMR spectroscopy viewpoint is that the two monomer units are completely separate spin systems and this can be visualised in the TOCSY spectrum.

HH COSY

The HH COSY shows the coupling network within the molecule.

HH TOCSY

The HH TOCSY spectrum shows correlations that belong together in contiguous spin systems: in the sucrose example, this means that the protons in the respective glucose and fructose units can be assigned.

HMQC

In the HMQC spectrum the one-bond direct HC couplings can be viewed as cross-peaks between the proton and carbon projections.

HMBC

2D NMR spectroscopy for the structural elucidation of 4.

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Apr 062014

A multistep single-crystal-to-single-crystal bromodiacetylene dimerization

http://www.nature.com/nchem/journal/v5/n4/fig_tab/nchem.1575_F6.html

Nature Chemistry5,327–334(2013)

doi:10.1038/nchem.1575

2D NMR spectroscopy for the structural elucidation of 4.

SEE

http://www.nature.com/nchem/journal/v5/n4/extref/nchem.1575-s1.pdf

1H-1H COSY NMR

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Apr 062014

¹H-¹H COSY (COrrelated SpectroscopY) is a useful method for determining which signals arise from neighboring protons (usually up to four bonds). Correlations appear when there is spin-spin coupling between protons, but where there is no coupling, no correlation is expected to appear.

This method is very useful when the multiplets overlap or when is extensive second order coupling complicates the 1D spectrum.

There are many variants on the COSY pulse sequence. The most popular one in our laboratory is the gradient enhanced double quantum coherence (DQF-COSY) version. The ratio of gradient strengths is usually set to two to yield all COSY signals but may be set to three to yield only those correlations involving three protons, e.g., CH-CH₂. We use the gradient enhanced DQF-COSY pulse sequence shown in fig. 1.

Fig. 1. Pulse sequence for gradient DQF-COSY

cholesteryl acetate

The COSY spectrum as shown in fig. 2 for ethylbenzene (fig. 3) contains a diagonal and cross peaks (signals that are not on the diagonal and correspond to other signals on the same horizontal and vertical projections). The cross peaks indicate couplings between two mutliplets up to three, or occasionally four, bonds away. The diagonal consists of the 1D spectrum with single peaks suppressed.

The most apparent cross-peak in the spectrum is between H1′ and H2′ at 2.65 and 1.24 ppm. A much weaker four-bond correlation (see the figure below) appears between H1′ and H2 at 2.65 and 7.20 ppm. All the desired signals are antiphase. Half the multiplet is positive and half negative. In addition, artifacts (undesired signals) appear in the spectrum as vertical streaks (interference and f₁ noise) and along the inverted ‘V’ (fig. 4) whose tip is on the top axis of the sepctrum. These artifacts are rarely in phase with the desired signals and appear in specific locations.

Fig. 2. 2D COSY spectrum of ethylbenzene

COSY of ethylbenzene

Fig. 3. Structure of ethylbenzene

Ethylbenzene

Fig. 4. Artifacts in the COSY spectrum of ethylbenzene

COSY artifacts

For example, in 12,14-di^tbutylbenzo[g]chrysene (fig. 5), only a partial analysis of the regular ¹H-NMR spectrum is possible. COSY (fig. 6) provides extra information about the connectivity. No correlations (cross-peaks) are seen to the tbutyls because they are too many bonds away from the ring system.

Fig. 5. Structure of 12,14-di^tbutylbenzo[g]chrysene

12,14-ditbutylbenzo[g]chrysene

Fig. 6. Artifacts in the COSY spectrum of ethylbenzene

COSY of 12,14-ditbutylbenzo[g]chrysene

The aromatic region of the spectrum (fig. 7) shows three bond correlations strongest. These can be used to determine which protons are neighbors. For example the proton at 8.17 ppm is next to the proton at 7.34 ppm, a fact that could not be easily determined from the 1D spectrum.

Fig. 7. Aromaric region of the 2D COSY spectrum of 12,14-di^tbutylbenzo[g]chrysene showing mostly three-bond correlations (a four-bond correlation between H10 and H11 is also visible)

Aromatic region of COSY of 12,14-ditbutylbenzo[g]chrysene

Four-bond and five-bond correlations are apparent when plotted to lower contours (fig. 8). These separate the spectrum into four groups of protons in a manner that is much clearer than the 1D spectrum.

Fig. 8. Aromaric region of the 2D COSY spectrum of 12,14-di^tbutylbenzo[g]chrysene showing three, four and five-bond correlations

Aromatic region of COSY of 12,14-ditbutylbenzo[g]chrysene

Using horizontal and vertical lines, it is possible to separate each group and follow its connectivity (fig. 9). The blue group of four protons is connected in the order 8.62 ppm to 7.55 to 7.59 to 8.56, the green group of four protons in the order 8.54 to 7.34 to 7.44 to 8.17 and the red group or two protons, that correspond to H9 and 10 because they are the only group of two protons expected to have a three-bond coupling constant (8.9 Hz), are at 7.76 and 8.32 ppm. The yellow group of two protons correspond to H11 and 13 because the coupling constant is small (1.9 Hz) and consistent with a four bond correlation.

Fig. 9. Aromaric region of the 2D COSY spectrum of 12,14-di^tbutylbenzo[g]chrysene showing connectivity and separation into four color-coded proton groups

Aromatic region of COSY of 12,14-ditbutylbenzo[g]chrysene in color

The multiplet structures in COSY are anti-phase for active couplings which leads to different patterns that for pure phase. A pure singlet (A) will not appear in a DQF-COSY spectrum because it is purely single quantum and a double-quantum filter is applied. This is true for deuterated solvent signals and for some protiated solvents such as water. Advantage of this is often taken for solvent suppression. A simple doublet appears in anti-phase. Many other combinations exist. The more common ones are listed in the table below and a few examples are shown.

Table 1. multiplicities often seen in COSY spectra compared with their pure phase counterparts.^a

Multiplicity	Pure phase	Anti-phase
A	1	0
AX	1 1	1 -1
AX₂	1 2 1	1 0 -1
AXY	1 1 1 1	1 1 -1 -1
AYX	1 1 1 1	1 -1 1 -1
AX₃	1 3 3 1	1 1 -1 -1
AX₂Y	1 2 1 1 2 1	1 0 -1 1 0 -1
AY₂X	1 2 1 1 2 1	1 2 1 -1 -2 -1
AXY₂	1 1 2 2 1 1	1 -1 2 -2 1 -1
AYX₂	1 1 2 2 1 1	1 -1 0 0 1 -1
AYX₂, J_AY=2J_AX	1 2 2 2 1	1 0 0 0 -1
AXYZ	1 1 1 1 1 1 1 1	1 1 1 1 -1 -1 -1 -1
AYXZ	1 1 1 1 1 1 1 1	1 1 -1 -1 1 1 -1 -1
AYZX	1 1 1 1 1 1 1 1	1 -1 1 -1 1 -1 1 -1
AXYZ, J_AX=J_AY+J_AZ	1 1 1 2 1 1 1	1 1 1 0 -1 -1 -1
AXYZ, J_AY=J_AX+J_AZ	1 1 1 2 1 1 1	1 1 -1 0 1 -1 -1
AXYZ, J_AZ=J_AX+J_AY	1 1 1 2 1 1 1	1 -1 1 0 -1 1 -1
AX₆	1 6 15 20 15 6 1	1 2 1 0 -1 -2 -1

^aThe active coupling is from A to X. The coupling constant decreases from left to right, e.g., for AXYZ, J_AX > J_AY > J_AZ.

Figs. 10-13 show expansions of COSY multiplets. The red contours are negative and the black ones are positive. The 1D projections are only representations (in practice the sum of the projection is zero).

Fig. 10. Anti-phase AX correlation between doublets H9 and 10 of 12,14-di^tbutylbenzo[g]chrysene

Antiphase AX signal

Higher multiplicities in which all the couplings are active yield the patterns shown in table 1. For example the correlation between the CH₃ and CH₂ protons in ethylbenzene yield a 1 0 1 pattern in one direction and a 1 1 -1 – 1 pattern in the other direction (fig. 11).

Fig. 11. Anti-phase A₂X₃ correlation between the ethyl protons of ethylbenzene

Antiphase AX signal

When there are more than two multiplets coupled, then only one coupling is active in each cross-peak. This can be used to determine which coupling constant relates to which correlation, something that may not be obvious from the 1D spectrum. In figs. 12 and 13 for di^tbutylbenzo[g]chrysene, the active couplings are labeled. The top cross-peak between the protons at 7.34 and 8.17 ppm shows the largest active coupling of 8.1 Hz. The coupling pattern in the vertical (f₁) direction is 1 1 1 0 -1 -1 -1 and in the horizontal (f₂) direction it is 1 1 1 1 -1 -1 -1 -1. The multiplet below it has the smallest active coupling of 1.4 Hz. The f₁ coupling pattern is 1 -1 1 0 -1 1 -1 and the f₂ coupling pattern is 1 1 -1 -1 1 1 -1 -1. The bottom multiplet displays an active coupling of 7.0 Hz and the coupling pattern in both directions is 1 1 -1 0 1 -1 -1.

Figs. 12, 13. Comparison of three AXYZ correlations showing different active couplings (from the COSY of 12,14-di^tbutylbenzo[g]chrysene)

Antiphase AXYZ signals Antiphase AYXZ signal

EXAMPLE

COSY spectra

The information on the H that are coupling with each other is obtained by looking at the peaks inside the grid. These peaks are usually shown in a contour type format, like height intervals on a map.
In order to see where this information comes from, let’s consider an example shown below, the COSY of ethyl 2-butenoate
First look at the peak marked A in the top left corner. This peak indicates a coupling interaction between the H at 6.9 ppm and the H at 1.8 ppm. This corresponds to the coupling of the CH₃ group and the adjacent H on the alkene.
Similarly, the peak marked B indicates a coupling interaction between the H at 4.15 ppm and the H at 1.25 ppm. This corresponds to the coupling of the CH₂ and the CH₃ in the ethyl group.
Notice that there are a second set of equivalent peaks, also marked A and Bon the other side of the diagonal.

COSY spectra of ethyl 2-butenoate

The (H,H) COSY experiment establishes the connectivity of a molecule by giving cross peaks (these are the off diagnonal peaks) for pairs of protons that are in close proximity. For the example of Glutamic acid below, we obtain cross peaks for the proton pairs (2,3) and (3,4). We do not observe a crosspeak for the pair (2,4), because these protons are not directly adjacent.

A relayed COSY experiment goes one step beyond a COSY experiment by showing cross peaks not just for pairs of adjacent protons, but for triples as well. As a result, we observe additional cross peaks like the one for the pair 2,4 in Glutamic acid below. Relayed COSY experiments can give cross peaks for protons that are too distant to show coupling in the 1D NMR spectrum.500 MHz H-relayed (H,H) COSY Spectrum of Glutamic acid. 1-D spectra left and top. 10 mg of compound in 0.5 mL of D₂O, 5 mm sample tube, 256 spectra, digital resolution of 2.639 Hz/data point. Total measurement time ca. 3h.

500 MHz H-relayed (H,H) COSY Spectrum of Glutamic acid. 1-D spectra left and top. 10 mg of compound in 0.5 mL of D₂O, 5 mm sample tube, 256 spectra, digital resolution of 2.639 Hz/data point. Total measurement time ca. 3h.

With present hardware and pulse sequences, it is possible to repeat the relay step up to three times. This allows the correlation of of protons that are separated by up to six bonds (d-protons). The relaying nucleus is typically ¹H, but high abundance I =1/2 hetero-elements like ³¹P or ¹⁹F can be used as well.

PRULIFLOXACIN by Nippon Shinyaku Co.

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PRULIFLOXACIN

(RS)-6-Fluoro-1-methyl-7-[4-(5-methyl-2-oxo-1,3-dioxolen-4-yl)methyl-1-piperazinyl]-4-oxo-4H-[1,3]thiazeto[3,2-a]quinoline-3-carboxylic acid

6-Fluoro-1-methyl-7-(4-(5-methyl-2-oxo-1,3-dioxelen-4-yl)methyl-1-piperazinyl)-4-oxo-4H-(1,3)thiazeto(3,2-a)quinoline-3-carboxylic acid

123447-62-1 CAS NO

NM 441, Quisnon, Pruvel, Sword, Prulifloxacin [INN], 123447-62-1, NM-441, CCRIS 7686, NCGC00164615-01NAD-441A
OPT-99

Molecular Formula: C₂₁H₂₀FN₃O₆S

Molecular Weight: 461.463403

Launched – 2002 BY NIPPON SHINYAKU

SYNTHESIS…….http://www.drugfuture.com/synth/syndata.aspx?ID=151640

Prulifloxacin is an older synthetic chemotherapeutic antibiotic of the fluoroquinolone drug class^[1]^[2] undergoing clinical trials prior to a possible NDA (New Drug Application) submission to the U.S. Food and Drug Administration (FDA). It is a prodrug which is metabolized in the body to the active compound ulifloxacin.^[3]^[4] It was developed over two decades ago by Nippon Shinyaku Co. and was patented in Japan in 1987 and in the United States in 1989.^[5]^[6]

It has been approved for the treatment of uncomplicated and complicated urinary tract infections, community-acquired respiratory tract infections in Italy and gastroenteritis, including infectious diarrheas, in Japan.^[3]^[7] Prulifloxacin has not been approved for use in the United States.

Prulifloxacin is a novel fluoroquinolone antibiotic that was launched pursuant to a collaboration between Meiji Seika and Nippon Shinyaku in 2002 for the oral treatment of systemic bacterial infections, including acute upper respiratory tract infection, bacterial pneumonia, prostatitis, cholecystitis, bacterial enteritis, internal genital infections, otitis media, sinusitis and others. It is currently marketed in a tablet formulation. A once-daily formulation to be taken over a three-day period is in phase III clinical trials at Optimer Pharmaceuticals to be used in the treatment of bacterial gastroenteritis, including traveler’s diarrhea. The formulation had been in phase II trials at the company for the treatment of urinary tract infections, however, no recent development for this indication have been reported. The drug has also been studied at Optimer for the treatment of community-acquired respiratory tract infections, but recent progress reports for this indication have not been made available.

Prulifloxacin has in vitro activity against a wide range of gram-negative and gram-positive microorganisms. Its antibacterial action results from inhibition of DNA gyrase and topoisomerase IV, both Type II isomerases. DNA gyrase is an essential enzyme that is involved in the replication, transcription, and repair of bacterial DNA. Topoisomerase IV is an enzyme known to play a key role in the partitioning of the chromosomal DNA during bacterial cell division. Together, the Type II topoisomerases remove the positive supercoils that accumulate ahead of a translocating DNA polymerase, allowing DNA replication to continue unhindered by topological strain. Fluoroquinolones may be active against pathogens that are resistant to penicillins, cephalosporins, aminoglycosides, macrolides and tetracyclines, as they possess a distinct mechanism of action from these antibiotics.

Prulifloxacin was discovered by Nippon Shinyaku and codeveloped with Meiji Seika in Japan. Nippon Shinyaku granted Angelini a manufacturing and marketing license for Italy in 1993. Exclusive Korean manufacturing and commercialization rights were acquired by Yuhan from Nippon Shinyaku in March 2003. In June 2004, Optimer was granted exclusive development and commercialization rights to prulifloxacin in the U.S. from Nippon Shinyaku. Finally, Recordati signed a nonexclusive licensing agreement with Angelini for the marketing and sale of prulifloxacin in Spain in October 2004. In March 2009, the product was licensed to Lee’s Pharmaceuticals by Nippon Shinyaku for marketing in China as an oral treatment of bacterial infection. In 2010, prulifloxacin was licensed to Algorithm by Nippon Shinyaku in North Africa and the Middle East for the development and marketing for the treatment of bacterial infections.

History

In 1987 a European Patent (EP 315828) for prulifloxacin (Quisnon ) was issued to the Japanese based pharmaceutical company, Nippon Shinyaku Co., Ltd (Nippon). Ten years after the issuance of the European patent, marketing approval was applied for and granted in Japan (March 1997). Subsequent to being approved by the Japanese authorities in 1997 prulifloxacin (Quisnon) was co-marketed and jointly developed in Japan with Meiji Seika as licensee (Sword).^[6]

In more recent times, Angelini ACRAF SpA, under license from Nippon Shinyaku, has fully developed prulifloxacin, for the European market.^[8] Angelini is the licensee for the product in Italy. Following its launch in Italy, Angelini launched prulifloxacin in Portugal (January 2007) and it has been stated that further approvals will be sought in other European countries.^[9]^[10]

Prulifloxacin is marketed in Japan and Italy as Quisnon (Nippon Shinyaku); Sword (Meiji); Unidrox (Angelini) and generic as Pruquin.

In 1989 and 1992 United States patents (US 5086049) were issued to Nippon Shinyaku for prulifloxacin. It was not until June 2004, when Optimer Pharmaceuticals acquired exclusive rights to discover, develop and commercialize prulifloxacin (Pruvel) in the U.S. from Nippon Shinyaku Co., Ltd., that there were any attempts to seek FDA approval to market the drug in the United States. Optimer Pharmaceuticals expects to file an NDA (new drug application) for prulifloxacin some time in 2010. As the patent for prulifloxacin has already expired, Optimer Pharmaceuticals has stated that this may have an effect on the commercial prospects of prulifloxacin within the United States market.^[11]

Licensed uses

Prulifloxacin has been approved in Italy ,Japan,China,India and Greece (as indicated), for treatment of infections caused by susceptible bacteria, in the following conditions:

Italy

Acute uncomplicated lower urinary tract infections (simple cystitis)
Complicated lower urinary tract infections
Acute exacerbation of chronic bronchitis

Japan

Gastroenteritis, including infectious diarrheas

Other countries

Prulifloxacin has not been approved for use in the United States, but may have been approved in other Countries, other than that which is indicated above.

Availability

Prulifloxacin is available as:

Tablets (250 mg, 450 mg or 600 mg)

In most countries, all formulations require a prescription.

Prulifloxacin is chemically known as 6-fluoro-1-methyl-7-{4-[(5-methyl-2-oxo-1 ,3-dioxol- 4-yl)methyl]piperazin-1-yl}-4-oxo-4H-[1 ,3]-thiazeto-[3,2-a]-quinoline-3-carboxylic acid, and it has the structure as shown below as formula I:

FORMULA I

Prulifloxacin has significant antibacterial activity and has been marketed as a synthetic antibacterial agent.

Prulifloxacin was first disclosed in US 5,086,049. The patent discloses a process for the preparation of prulifloxacin by the condensation of ulifloxacin with a 4-halomethyl-5- methyl-1 ,3-dioxolen-2-one of formula III

wherein X is halo selected form chloro, bromo or iodo, in the presence or absence of an aprotic solvent and a base to obtain prulifloxacin free base which is recrytallised with chloroform-methanol. In an exemplified process, ethyl 6,7-difluoro-1-methyl-4-oxo-4H- (1 ,3)-thiazeto-(3,2-a)-quinoline-3-carboxylate is condensed with piperazine in the presence of dimethyl formamide and purified by column chromatography followed by basic hydrolysis to give ulifloxacin, which is then converted to prulifloxacin.

The above process involves column chromatography. Prulifloxacin prepared by this method has a purity of 60-65% containing impurities in unacceptable levels. Removal of these impurities by usual purification procedures, such as recrystallisation, distillation and washing, is difficult and requires extensive and expensive multiple purification processes. This further decreases the overall yield. A method involving column chromatographic purifications and multiple purifications cannot be used for large-scale operations, thereby making the process commercially non-viable.

European Patent No. 315828 disclosed a variety of quinoline carboxylic acid derivatives and pharmaceutically acceptable salts thereof. These compounds are exhibiting antibacterial activity and useful as remedies for various infectious diseases. Among them prulifloxacin, chemically (+)-6-Fluoro- 1 -methyl-7-[4-(5-methyl-2-oxo-1 ,3-dioxolen-4-ylmethyl)-1 -piperazinyl]-4-oxo-4H- [1 ,3]thiazeto[3,2-a]quinoline-3-carboxylic acid is a fluoroquinolone antibacterial prodrug which shows potent and broad-spectrum antibacterial activity both in vitro and in vivo. Prulifloxacin also showed superior activity against strains of Enterobacteriaceae and Pseudomonas aeruginosa. Prulifloxacin is represented by the following structure:

Processes for the preparation of prulifloxacin and related compounds were disclosed in European Patent No. 315828 and UK Patent Application No. GB 2190376.

In – the preparation of prulifloxacin, 6-fluoro-1-methyl-4-oxo-7-(1- piperazinyl)-4H-[1 ,3]thiazeto[3,2-a]quinoline-3-carboxylic acid of formula I:

is a key intermediate. According to the UK Patent Application No. GB 2190376, the compound of the formula I was prepared by the reaction of 3,4-difluroaniline with carbon disulfide and triethylamine to give triethylammonium N-(3,4- difluorophenyl)dithio carbamate, which by reaction with ethyl chloroformate and triethylamine in chloroform is converted into 3,4-difluorophenyl isothiocyanate, followed by reaction with diethyl malonate and KOH in dioxane affords the potassium salt, which is then treated with methoxymethyl chloride in dimethylformamide to give diethyl 1-(3,4-difluorophenylamino)-1- (methoxymethylthio)-rnethylene-rnalonate. The cyclization of the thio compound at 240⁰C in diphenyl ether affords ethyl 6,7-difluoro-4-hydroxy-2- methoxymethylthioquinoline-3-carboxylate, which by treatment with HCI in ethanol gives ethyl δy-difluoro^-hydroxy^-mercaptoquinoline-S-carboxylate. The cyclization of the mercapto compound with 1,1-dibromoethane by means of potassium carbonate and potassium iodide in hot dimethylformamide yields ethyl 6,7-difluoro-1-methyl-4-oxo-4H-[1 ,3]thiazeto[3,2-a]quinoline-3-carboxylate, which is condensed with piperazine in dimethylformamide to afford ethyl 6- fluoro-1-methyl-4-oxo-7-(1-piperazinyl)-4H-[1 ,3]thiazeto[3,2-a]quinoline-3- carboxylate, which is then subjected to hydrolysis with potassium hydroxide in hot tert-butanol to give the compound of formula I.

The compound of formula I obtained by the process described in the UK Patent Application No. GB 2190376 is not satisfactory from purity point of view, the reaction between ethyl 6,7-difluoro-1-methyl-4-oxo-4H-[1 ,3]thiazeto[3,2- a]quinoline-3-carboxylate and piperazine requires longer time about 48 hours to complete, the yield obtained is not satisfactory, and the process also involves column chromatographic purifications. Methods involving column chromatographic purifications cannot be used for large-scale operations, thereby making the process commercially not viable. According to the European Patent No. 315828, prulifloxacin is prepared by reacting 6-fluoro-1-methyl-4-oxo-7-(1-piperazinyl)-4H-[1 ,3]thiazeto[3,2-a] quinoline-3-carboxylic acid with 4-bromomethyl-5-methyl-1 ,3-dioxolen-2-one in presence of potassium bicarbonate in dimethylformamide. However, a need still remains for an improved and commercially viable process of preparing pure prulifloxacin that will solve the aforesaid problems associated with process described in the prior art and will be suitable for large- scale preparation, in terms of simplicity, purity and yield of the product.

Prulifloxacin is chemically 6-fluoro-l-methyl-7-{4-[(5-methyl-2-oxo-l,3-dioxol-4- yl)methyl]piperazin-l-yl}-4-oxo-4H-[l,3]thiazeto[3,2-α]quinoline-3-carboxylic acid of Formula I having the structure as depicted below:

FORMULA I

Prulifloxacin has significant antibacterial activity and has been marketed as a synthetic antibacterial agent. U.S. Patent No. 5,086,049 provides a process for the preparation of prulifloxacin by reacting 6-fluoro-l-methyl-4-oxo-7-piperazin-l-yl-4H- [l,3]thiazeto[3,2-α]quinoline-3-carboxylic acid of Formula II,

FORMULA II and 4-(bromomethyl)-5-methyl-l,3-dioxol-2-one of Formula III,

FORMULA III using N,N-dimethylformamide as a solvent. 4-(Bromomethyl)-5-methyl-l,3-dioxol-2-one of Formula III is used in excess to one mole of the compound of Formula II. The process provided in U.S. Patent No. 5,086,049 further involves concentrating the reaction mixture, pouring the residue into water and isolating prulifloxacin by filtration. The resulting prulifloxacin is recrystallized from chloroform-methanol.

However, U.S. Patent No. 5,086,049 does not provide any method to remove the unreacted or the excess of 4-(bromomethyl)-5-methyl-l,3-dioxol-2-one of Formula III used as a starting material. The present inventors have observed that it is difficult to obtain prulifloxacin with pharmaceutically acceptable purity by following the process provided in U.S. Patent No. 5,086,049, which is typically contaminated by process related impurities including 4-(bromomethyl)-5-methyl-l,3-dioxol-2-one

A need still remains for an improved and commercially-viable process for preparing pure prulifloxacin that will solve the aforesaid problems associated with the process described in the prior art and that will be suitable for large-scale preparation, in terms of simplicity, purity and yield of the product.

EP1626051 A1 mentions that Type I, Type II and Type III crystals of prulifloxacin are obtained by crystallization from acetonitrile as reported in lyakuhin Kenkyu, Vol. 28 (1), (1997), 1-11. However, the conditions of crystallization from acetonitrile for preparing Type I, Type II and Type III crystals are not disclosed in lyakuhin Kenkyu, Vol. 28 (1), (1997), 1-11. EP1626051A1 further mentions that Type III crystals have been marketed by considering the solubility, absorbability, therapeutic effect and the like of the respective crystal forms.

US 2007/0149540 discloses a crystal of prulifloxacin acetonitrile solvate (Compound B) which is an intermediate for producing preferentially the type III crystal of prulifloxacin. A crystal of Compound B can be preferentially precipitated by controlling the supersaturation concentration in crystallization using acetonitrile as a solvent, subsequently; the type III crystal of Compound A can be produced by performing desolvation of the crystal.

WO 2008/111018 discloses processes for the preparation of Type I, Type II and Type III crystals of prulifloxacin. There is disclosed a process for preparing Type I crystals by controlled cooling over a period of 7 to 9 hours and prolonged drying over 24 hours. The inventors of the present invention have found that Type I and Type III crystals prepared according to the WO 2008/111018 process are unstable and the process is non-reproducible.

WO 2010/0084508 discloses processes for the preparation of Type I, Type II and Type III crystals of prulifloxacin.

WO 2008/059512 discloses a process for the preparation of prulifloxacin using novel intermediates.

WO 2008/111016 discloses a process for the preparation of prulifloxacin having purity of about 99% or above. It would be a significant contribution to the art to provide a crystalline form of prulifloxacin, which is consistent and to provide industrially viable methods of preparation, pharmaceutical formulations, and methods of use thereof.

…………………

SYNTHESIS

http://www.google.com/patents/WO2012001357A1?cl=en

Scheme 1.

Formula I

[PRULIFLOXACIN]

Example 1

Preparation of ethyl-6-fluoro-1-methyl-4-oxo-7-(1-piperazinyl)-4H-(1 ,3)-thiazeto-[3,2-a]- quinoline-3-carboxylate (formula III)

5,6-difluoro-1-methyl-4-oxo-4H-[1 ,3]-thiazeto-[3,2-a]-quinoline-3-carboxylic acid ethyl ester of formula (II) (100 gms, 0.321 moles) was stirred in 500 ml of DMF at room temperature. Piperazine (76 gms, 0.882 moles) was added at room temperature and stirred for 10 minutes. The temperature was slowly raised to 50-55°C and the reaction mass was stirred at 50-55°C for 5 hours. After completion of the reaction, the reaction mass was cooled to 25-30°C and stirred for 2 hours. The reaction mass was further chilled to 10-15°C and stirred for 2 hours. The precipitated solid was filtered, washed of chilled DMF (2 x 50 ml). The solid was slurry washed with water (300 ml), filtered, washed with water ( 2 x 100 ml) and dried under vacuum at 70-75°C to yield the title compound [90 gms, 74 % yield, 95% HPLC purity].

Example 2

Preparation of 6-fluoro-1-methyl-4-oxo-7-(1-piperazinyl)-4H-[1 ,3]-thiazeto-[3,2-a]- quinoline-3-carboxylic acid (formula IV)

Ethyl-6-fluoro-1 -methyl-4-oxo-7-(1 -piperazinyl)-4H-(1 ,3)-thiazeto-[3,2-a]-quinoline-3- carboxylate (100 gms, 0.265 moles) was stirred in water (600 ml) at 25-30°C. To this potassium hydroxide solution (50 gms of potassium hydroxide flakes is dissolved in 200 ml of water) was added and the reaction mass was heated to 80-85°C. The contents were stirred for 1 hour and after completion of reaction, the reaction mass was cooled to 25-30°C. The pH of the reaction mass was adjusted to 6.5-7.0 using 1:1 aqueous acetic acid solution. The contents were stirred at room temperature for 1 hour. The precipitated solid was filtered, washed with water (2 x 100 ml). The solid was slurried in methanol (300 ml) for 1 hour at 25-30°C, filtered, washed with methanol (2 x 50 ml) and dried under vacuum at 70-75°C to yield the title compound [90 gms, 97% yield, 96% HPLC purity]. Example 3

Preparation of prulifloxacin

To a solution of 4-(chloromethyl)-5-methyl-1,3-dioxol-2-one (55 gms, 0.371 moles) in 50 ml of DMF at 25-30°C, sodium bromide (77 gms, 0.748 moles) was added and the reaction mass was slowly heated to 40-45°C. The contents were stirred at 40-45°C for 2 hours, acetone ( 500 ml) was added at 40-45°C and stirred for 3 hours. The reaction mass was filtered over hyflo, and the bed washed with acetone (100 ml). The solvent was completely distilled off under vacuum below 45°C to yield 4-(bromomethyl)-5- methyl-1 ,3-dioxol-2-one (formula V).

To a solution of 6-fluoro-1-methyl-4-oxo-7-(1-piperazinyl)-4H-[1 ,3]-thiazeto-[3,2-a]- quinoline-3-carboxylic acid of formula IV (100 gms, 0.286 moles) in 4.0 It of acetonitrile, DIPEA (70 ml , 0.402 moles)) was added at room temperature, stirred for 10 minutes. The reaction mass was cooled to 10-15°C and a solution of 4-(bromomethyl)-5-methyl- 1 ,3-dioxol-2-one (formula V) in 500 ml of acetonitrile was slowly added at 10-15°C over a period of 1 hour. The contents were stirred at 25-30°C for 20 hour, filtered over hyflo, and the bed washed with 200 ml of acetonitrile. The solvent was distilled off completely under vacuum below 50°C. Acetonitrile (100 ml) was added at 50°C and the contents were stirred for 30-60 minutes. The reaction mass was slowly chilled to 0-5°C and the precipitated solid was filtered, washed with acetonitrile (25 ml) and dried to yield 65 gms of prulifloxacin. Example 4

Preparation of Type I crystals of prulifloxacin

Prulifloxacin (65 gms) was added to 200 ml of DMF at 25-30°C and heated to 80-85°C for 1 hour. The mixture was then slowly cooled to 25-30°C, stirred for 2 hours, chilled to 0-5°C for 2 hours. The precipitated solid was filtered and dried under vacuum at 70- 75°C to yield Type I crystals of prulifloxacin (55 gms, 99.5 % HPLC purity).

Example 5

Preparation of prulifloxacin

(55 gms, 0.371 moles) of 4-(chloromethyl)-5-methyl-1 ,3-dioxol-2-one is taken in 5.0 ml of DMF at 25-30°C. (77 gms, 0.748 moles) of sodium bromide is added and slowly heated the reaction mass to 40-45°C. The contents are stirred at 40-45°C for 2 hours, 500 ml of acetone is added at 40-45°C and stirred for 3 hours. The reaction mass is clarified over hyflo, and the bed washed with 100 ml of acetone to yield a solution of 4- (bromomethyl)-5-methyl-1 ,3-dioxol-2-one (formula V).

To a solution of 6-fluoro-1-methyl-4-oxo-7-(1-piperazinyl)-4H-[1 ,3]-thiazeto-[3,2-a]- quinoline-3-carboxylic acid of formula IV (100 gms, 0.286 moles) in 3.5 Its of acetone was at room temperature DIPEA (70 ml, 0.402 moles) and stirred for 10 minutes. The reaction mass was cooled to 10-15°C and a solution of 4-(bromomethyl)-5-methyl-1 ,3- dioxol-2-one (formula V) in acetone was slowly added to the reaction mass at 10-15°C over a period of 1 hour. The contents were further stirred at 25-30°C for 20 hour, filtered over hyflo and the bed washed with 200 ml of acetone. The solvent was distilled off completely under vacuum below 50°C. Acetonitrile (100 ml) was added at 50°C and the contents were stirred for 30-60 minutes. The reaction mass was further chilled to 0- 5°C and stirred for 2 hours. The precipitated solid was filteredand dried to yield prulifloxacin.

………………….

http://www.google.com/patents/WO2008059512A1?cl=en

novel process for preparing 6-fluoro-1-methyl-4-oxo-7-(1-piperazinyl)-4H-[1 ,3]thiazeto [3,2-a]quinoline-3-carboxylic acid of formula I:

which comprises: a) reacting the difluoro-quinoline compound of formula

wherein R represents hydrogen atom or alkyl containing 1 to 4 carbon atoms; with boric acid of formula III:

in presence of acetic anhydride and acetic acid to give borane compound of formula IV:

b) reacting the borane compound of formula IV with piperazine of formula V:

HN NH V

to give piperazine compound of formula Vl:

c) treating the compound of formula Vl with an alkaline metal hydroxide, carbonate or bicarbonate to obtain the compound of formula I.

Prulifloxacin and pharmaceutically acceptable acid addition salts of prulifloxacin can be prepared by using the compound of formula I by known methods for example as described in the European Patent No. 315828. Borane compound of the formula IV and Vl are novel and forms part of the invention. Preferably the reaction in step (a) is carried out at about 30⁰C to reflux temperature more preferably at about 80⁰C to reflux temperature and still more preferably at reflux temperature.

Example 1 Step-I:

Acetic anhydride (24 ml) and acetic acid (11 ml) are added to boric acid (3.5 gm) under stirring at 25 – 30⁰C, the contents are heated to reflux and then stirred for 3 hours at reflux. The reaction mass is cooled to 100⁰C, ethyl 6,7- difluoro-1-methyl-4-oxo-4H-[1 ,3]thiazeto[3,2-a]quinoline-3-carboxylate (20 gm) is added at 100⁰C, the contents are heated to reflux and then refluxed for 2 hours. The reaction mass is cooled to 25 – 35⁰C, toluene (200 ml) is added under stirring, the reaction mass is cooled to 5⁰C and then stirred for 1 hour at 5 – 10⁰C. Filtered the solid, washed with 20 ml of toluene and then dried to give 25.5 gm of 6,7-difluoro-1-methyl-4-oxo-4H-[1 ,3]thiazeto[3,2-a]quinoline-3-carboxylate -O³,0⁴/bis/acetato-0/-borone. Step-I I: Acetonitrile (125 ml), dimethylsulfoxide (125 ml) and piperazine (13.8 gm) are added to 6,7-difluoro-1-methyl-4-oxo-4H-[1 ,3]thiazeto[3,2-a]quinoline-3- carboxylate-0³,0⁴/bis/acetato-0/-borone (25.5 gm, obtained in step-l) under stirring at 25 – 35⁰C, the contents are heated to 85⁰C and then stirred for 3 hours at 80 – 85⁰C to form a clear solution. The solution is cooled to 10⁰C and then stirred for 1 hour at 10 – 15⁰C. Filtered the solid, washed with 25 ml of acetonitrile and then dried to give 26 gm of 6-fluoro-1-methyl-4-oxo-7-(1- piperazinyl)-4H-[1,3]thiazeto[3,2-a]quinoline-3-carboxylate-0³,0⁴/bis/acetato-0/- borone. Step-Ill: Water (155 ml), potassium hydroxide (17 gm) are added to 6-fluoro-1- methyl-4-oxo-7-(1-piperazinyl)-4H-[1 ,3]thiazeto[3,2-a]quinoline-3-carboxylate- O³,O⁴/bis/ acetato-0/-borone (26 gm, obtained in step-ll) under stirring at 25 – 35⁰C, the contents are heated to 65⁰C and then stirred for 4 hours at 60 – 65⁰C. The reaction mass is cooled to 25⁰C, filtered the undesired solid on hi-flow bed and then pH of the resulting filtrate is adjusted to 7 – 7.5 with 50% HCI solution at 25 – 30⁰C. The separated solid is stirred for 1 hour at 25 – 30⁰C, filtered the solid, washed with 35 ml of water and then dried to give 17 gm of 6-fluoro-1- methyl-4-oxo-7-(1 -piperazinyl)-4H-[1 ,3]thiazeto [3,2-a]quinoline-3-carboxylic acid (HPLC Purity: 98.5%). Example 2 Step-I:

Acetic anhydride (12 ml) and acetic acid (5.5 ml) are added to boric acid (1.25 gm) under stirring at 25 – 30⁰C, the contents are heated to reflux and then stirred for 3 hours at reflux. The reaction mass is cooled to 100⁰C, 6,7-difluoro-1- methyl-4-oxo-4H-[1 ,3]thiazeto[3,2-a]quinoline-3-carboxylic acid (10 gm) is added at 100⁰C, the contents are heated to reflux and then refluxed for 3 hours. The reaction mass is cooled to 50⁰C, toluene (100 ml) is added under stirring at 50⁰C, the resulting mass is cooled to 10⁰C and then stirred for 1 hour at 10 – 15⁰C. Filtered the solid, washed with 20 ml of toluene and then dried to give 10 gm of 6,7-difluoro-1-methyl-4-oxo-4H-[1 ,3]thiazeto[3,2-a]quinoline-3-carboxylate -O³ , 0⁴/bis/acetato-0/-borone . Step-I I:

Acetonitrile (50 ml), dimethylsulfoxide (50 ml) and piperazine (5.5 gm) are added to 6,7-difluoro-1-methyl-4-oxo-4H-[1 ,3]thiazeto[3,2-a]quinoline-3- carboxylate-0³,0⁴/bis/acetato-0/-borone (10 gm, obtained in step-l) under stirring at 25 – 35⁰C, the contents are heated to 85⁰C and then stirred for 3 hours at 80 – 85⁰C to form a clear solution. The solution is cooled to 10⁰C and then stirred for 1 hour at 10 – 15⁰C. Filtered the solid, washed with 10 ml of acetonitrile and then dried to give 10.4 gm of 6-fluoro-1-methyl-4-oxo-7-(1- piperazinyl)-4H-[1 ,3]thiazeto[3,2-a]quinoline-3-carboxylate-0³,0⁴/bis/acetato-0/- borone. Step-Ill :

Water (62 ml), potassium hydroxide (7 gm) are added to 6-fluoro-1-methyl-4- oxo-7-(1-piperazinyl)-4H-[1 ,3]thiazeto[3,2-a]quinoline-3-carboxylate-0³,0⁴/bis/ acetato-OAborone (10.4 gm, obtained in step-ll) under stirring at 25 – 35⁰C, the contents are heated to 65⁰C and then stirred for 4 hours at 60 – 65⁰C. The reaction mass is cooled to 25⁰C, filtered the undesired solid on hi-flow bed and then pH of the resulting filtrate is adjusted to 7 – 7.5 with 50% HCI solution at 25 – 30⁰C. The separated solid is stirred for 30 minutes at 25 – 30⁰C, filtered the solid, washed with 20 ml of water and then dried to give 68 gm of 6-fluoro-1- methyl-4-oxo-7-(1-piperazinyl)-4H-[1 ,3]thiazeto [3,2-a]quinoline-3-carboxylic acid (HPLC Purity: 98.6%). Example 3

Acetonitrile (560 ml) and potassium bicarbonate (8 gm) are added to 6- fluoro-i-methyM-oxo-y-CI-piperazinyO^H-CI .SKhiazetofS^-alquinoline-S- carboxylic acid (14 gm, obtained as per the processes described in examples 1 and 2) under stirring at 25 – 30⁰C, the contents are cooled to 15⁰C and then the solution of 4-bromomethyl-5-methyl-1 ,3-dioxolen-2-one (10 gm) in acetonitrile (140 ml) is added at 15 – 20⁰C for 30 to 45 minutes. The contents are stirred for 25 hours at 25 to 30⁰C, filtered and the resulting filtrate is distilled under vacuum. To the residue added acetonitrile (70 ml), cooled the mass to 20⁰C and then stirred for 1 hour to 1 hour 30 minutes at 20 – 25⁰C. Filtered the solid, washed the solid with 15 ml of chilled acetonitrile and then dried to give 16 gm of prulifloxacin crude (HPLC Purity: 98.8%).

To the prulifloxacin crude (obtained above) added acetonitrile (200 ml) at 25 – 30⁰C, the contents are heated to reflux and then refluxed for 30 minutes. To the reaction mass added activated carbon (5 gm) and refluxed for 15 minutes. The reaction mass is filtered on hi-flo bed, the resulting filtrate is cooled to 20⁰C and then stirred for 3 – 4 hours at 20 – 25⁰C. Filtered the solid, washed with 20 ml of acetonitrile and then dried to give 14 gm of prulifloxacin (HPLC Purity: 99.9%).

…………………

http://www.google.com/patents/WO2008111016A1?cl=en

In a first aspect, a process for the preparation of prulifloxacin is provided, the process comprising: a) reacting a compound of Formula II with a compound of Formula III to obtain prulifloxacin;

FORMULA III

FORMULA II

b) contacting the prulifloxacin obtained in step a) with an acid in a biphasic solvent system, wherein the biphasic solvent system comprises water and a water- immiscible organic solvent; c) separating the aqueous layer from the reaction mixture obtained in step b); d) treating the aqueous layer with a base; and e) isolating prulifloxacin.

The process described in steps b – e above may be carried out with prulifloxacin made from any process however.

The compounds of Formula II and Formula III may be prepared according to the methods provided in U.S. Patent No. 5,086,049.

Example 1: Process for the Preparation of Prulifloxacin:

Step A): A solution of 4-(bromomethyl)-5-methyl-l,3-dioxol-2-one (35.5 g, 0.184 mole) in N,N-dimethylformamide (200 ml) was added dropwise at 0 to 5° C to a stirred solution of 6-fluoro-l-methyl-4-oxo-7-piperazin-l-yl-4H-[l,3]thiazeto[3,2-α]quinoline-3- carboxylic acid (50 g, 0.143 mole and potassium bicarbonate (15.8 g, 0.1578 mole) in N,N-dimethylformamide (200 ml). The resulting mixture was stirred at 25° to 28°C for 3 to 4 hours. After the completion of the reaction, the reaction mixture was poured into water (1250 ml). The solid obtained was filtered, washed with water (100 ml), and subsequently dissolved in a mixture of chloroform: methanol (7:3; 1250 ml). The lower organic layer was separated and water (500 ml) was added to the organic layer. A dilute aqueous solution of hydrochloric acid was added to the biphasic reaction mixture to adjust pΗ to 0.8 to 1.0. The reaction mixture was stirred for 15 minutes, allowed to settle and the upper aqueous layer was separated. The process was repeated twice and the aqueous layers were combined. Activated charcoal (10%) was added to the combined aqueous layer and stirred for 30 minutes, filtered and cooled to 20° to 25° C. The pΗ of the reaction mixture was adjusted to 6.5 to 7.0 by adding an aqueous solution of sodium bicarbonate. The solid obtained was extracted with chloroform (375 ml), stirred for 15 minutes and the organic layer was separated. The aqueous layer was further extracted with a mixture of chloroform: methanol (7:3 ratio; 50 ml). The combined organic layer was distilled under vacuum at 35° to 40° C to recover the solvent up to 125 ml. The reaction mass so obtained was stirred for 3 to 4 hours at 28° to 30° C, filtered and washed with chilled chloroform (50 ml). The wet cake obtained was dried at 45° C for 12 hours to obtain the title compound. Step B): The prulifloxacin (30 g) obtained in Step A) was suspended in a mixture of chloroform: ethanol (10:1, v/v, 585 ml: 58.5 ml) and heated to reflux temperature. Activated carbon (3.9 gm) was added to the partially cleared solution and refluxed for 30 minutes, followed by filtration through Celite bed. The bed was further washed with chloroform: ethanol (10:1, v/v, 585 ml: 58.5 ml). The filtrate so obtained was distilled at atmospheric pressure till to partially remove the solvent. The concentrate so obtained was stirred at about 25° C for 1 hour, and filtered. The solid obtained was washed with chloroform: ethanol (39 ml X 2), dried under vacuum at 45° C for 12 hours to obtain the title compound. Yield: 22 g

HPLC Purity: 99%

………………………….

SEE

Studies on pyridonecarboxylic acids. 1. Synthesis and antibacterial evaluation of 7-substituted-6-halo-4-oxo-4H-[1,3]thiazeto[3, 2-a]quinoline-3-carboxylic acids
J Med Chem 1992, 35(25): 4727

http://pubs.acs.org/doi/pdf/10.1021/jm00103a011

The reaction of 3,4-difluoroaniline (I) with carbon disulfide and triethylamine gives triethylammonium N-(3,4-difluorophenyl)dithiocarbamate (II), which by reaction with ethyl chloroformate and triethylamine in chloroform is converted into 3,4-difluorophenyl isothiocyanate (III). The reaction of (III) with diethyl malonate and KOH in dioxane affords the potassium salt (IV), which is treated with chloromethyl methyl ether in DMF to give the corresponding methoxymethylsulfanyl compound (V). The cyclization of (V) at 240 C in diphenyl ether affords 6,7-difluoro-4-hydroxy-2-(methoxymethylsulfanyl)quinoline-3-carboxylic acid ethyl ester (VI), which by treatment with HCl in ethanol gives the corresponding mercapto compound (VII). The cyclization of (VII) with 1,1-dibromoethane by means of K2CO3 and KI in hot DMF yields 5,6-difluoro-1-methyl-4-oxo-4H-[1,3]thiazeto[3,2-a]quinoline-3-carboxylic acid ethyl ester (VIII), which is condensed with piperazine (IX) in DMF to afford the corresponding piperazino-derivative (X). The hydrolysis of (X) with KOH in hot tert-butanol gives the corresponding free acid (XI) , which is finally condensed with 4-(bromomethyl)-5-methyl-1,3-dioxol-2-one (XII) by means of KHCO3 in DMF.

………………….

Treatment of 3,4-difluoroaniline (I) with CS2 and Et3N gives triethylammonium dithiocarbamate (II), which reacts with ethyl chloroformate in chloroform to yield (III). Isothiocyanate (III) is converted into the potassium salt (IV) by reaction with diethyl malonate and KOH in dioxane and then transformed into methoxymethyl thioether (VI) by means of reagent (V) and Et3N in toluene. Cyclization of (VI) by heating in diphenyl ether affords quinoline (VII), which then reacts with benzoyl chloride (VIII) in pyridine to furnish (IX). Benzoyloxy derivative (IX) is converted into (X) by means of HCl in EtOH, and its reaction with 1-bromo-2-fluoroethane (XI) and NaHCO3 yields compound (XII). Chlorination of (XII) with SO2Cl2 in hexane provides (XIII), which by simultaneous hydrolysis and intramolecular cyclization by means of Et3N /H2O in THF provides the mixture of isomers (XIV). (+)-(XV) is obtained by HPLC chromatography of (XIV) on a chiral stationary phase. Treatment of (+)-(XV) with 1-methylpiperazine (XVI) in DMF provides ethyl ester (+)-(XVII), which is finally hydrolyzed by means of H2SO4 in H2O.

INTERMEDIATES

154330-67-3

Ethyl 6,7-difluoro-2-ethylmercapto-4-hydroxyquinoline-3-carboxylate

154330-68-4

Ethyl 4-acetoxy-6,7-difluoro-2-(ethylthio)quinoline-3-carboxylate

113046-72-3

Ethyl 6,7-difluoro-1-methyl-4-oxo-4H-[1,3]thiazeto[3,2-a]quinoline-3-carboxylate

113028-17-4

Ethyl 6-fluoro-1-methyl-4-oxo-7-(1-piprazinyl)-4H-[1,3]thiazeto[3,2-a]quinoline-3-carboxylate

112984-60-8

6-Fluoro-1-methyl-4-oxo-7-(1-piperazinyl)-4H-[1,3]thiazeto[3,2-a]quinoline-3-carboxylic acid

REFERENCES

Nelson, Jennifer M.; Chiller, Tom M.; Powers, John H.; Angulo, Frederick J. (2007). “Food Safety: Fluoroquinolone‐Resistant Campylobacter Species and the Withdrawal of Fluoroquinolones from Use in Poultry: A Public Health Success Story”. Clinical Infectious Diseases 44 (7): 977–80. doi:10.1086/512369. PMID 17342653.
Kawahara S (1998). “[Chemotherapeutic agents under study]”. Nippon Rinsho (in Japanese) 56 (12): 3096–9. PMID 9883617.
Fritsche, T. R.; Biedenbach, D. J.; Jones, R. N. (2008). “Antimicrobial Activity of Prulifloxacin Tested against a Worldwide Collection of Gastroenteritis-Producing Pathogens, Including Those Causing Traveler’s Diarrhea”. Antimicrobial Agents and Chemotherapy 53 (3): 1221–4. doi:10.1128/AAC.01260-08. PMC 2650572.PMID 19114678.
Giannarini, Gianluca; Tascini, Carlo; Selli, Cesare (2009). “Prulifloxacin: clinical studies of a broad-spectrum quinolone agent”. Future Microbiology 4 (1): 13–24.doi:10.2217/17460913.4.1.13. PMID 19207096.
JP patent 1294680, Kise Masahiro; Kitano Masahiko; Ozaki Masakuni; Kazuno Kenji; Matsuda Masato; Shirahase Ichiro; Segawa Jun, “Quinolinecarboxylic Acid Derivative”, issued November 28, 1989
Prulifloxacin. Drugfuture.com. Retrieved on 2010-11-03.
Anonymous (2002). “Prulifloxacin [‘Quisnon’; Nippon Shinyaku] has been approved in Japan”. Inpharma 1 (1362): 22.
Research and Development Department of Angelini. Angelinipharma.com. Retrieved on 2010-11-03.
Nippon Shinyaku, Annual Report 2007
“Prulifloxacin. NAD-441A, NM 441, Quisnon”. Drugs in R&D 3 (6): 426–30. 2002.PMID 12516950.
Annual Report 2008, p. 34

Segawa,J,Mashiko kitano, Kenji Kazuno et al, Studies on Pyridonecarboxylic acids,1.Sythesis and antibacterial Evaluation of 7-substituted-6-halo-4-oxo-4H-[1,3]thiazeto [3,2-]quionoline- 3-caroboxylic acids[J].J Med Chem. 1992,35(25):4727-4738.

Masato Matsuoka, Jun Segawa, Yoshihiko.et al, Studies on Pyridone Carb oxylic acids. V.A Practial synthesis of Ethyl 6,7–Difuoro-1-methyl-4-oxo-[1,3] Thiazeto [3,2-a]quinoline-3- Caroboxylate a key intermediate for the new tricyclic quinolone, prulifloxacin (NM441) and Versatile new syntheses of the 2-thioquinoline Skeleton[J].J Heterocyclic Chem.1997,34,1773-1779.

US5498422	3-13-1996	Sustained release capsule
US5456923	10-11-1995	Method of manufacturing solid dispersion
US5086049	2-5-1992	7(4-(5 METHYL-2-OXO-1,3-DIOXALEN-4-YL)METHYL 1-PIPERZINYL)-4-OXO-4H-(1,3)THIAZETO(3,2-A)QUINOLINE-3-CARBOXYLIC ACIDS


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US2011034690	2-11-2011	PROCESS FOR THE PREPARATION OF PURE PRULIFLOXACIN
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US2010087409	4-9-2010	COMPOSITION COMPRISING AN ANTIBIOTIC AND A CORTICOSTEROID
US2009306035	12-11-2009	Compounds and Methods for modulating the Silencing of a Polynucleotide of Interest
US2007196504	8-24-2007	PHARMACEUTICAL COMPOSITION
US2007148235	6-29-2007	PHARMACEUTICAL COMPOSITION
US2005152975	7-15-2005	Pharmaceutical composition
US2004022848	2-6-2004	Medicinal composition

WO2008059512A1	Nov 17, 2006	May 22, 2008	Hetero Drugs Ltd	Process for preparation of prulifloxacin using novel intermediates
WO2008111016A1	Mar 14, 2008	Sep 18, 2008	Ranbaxy Lab Ltd	Process for the preparation of pure prulifloxacin
WO2008111018A2	Mar 14, 2008	Sep 18, 2008	Ranbaxy Lab Ltd	Process for the preparation of crystals of prulifloxacin
WO2010084508A2	Dec 10, 2009	Jul 29, 2010	Elder Pharmaceuticals Ltd.	Process for the preparation of type i, type ii and type iii crystalline prulifloxacin
EP0315828A1 *	Oct 26, 1988	May 17, 1989	Nippon Shinyaku Company, Limited	Quinolinecarboxylic acid derivatives
EP1626051A1	Apr 28, 2004	Feb 15, 2006	Nippon Shinyaku Co., Ltd.	Crystals of quinolinecarboxylic acid derivative solvate
US5086049	Apr 8, 1991	Feb 4, 1992	Nipponshinyaku Co., Ltd.	7[4-(5 methyl-2-oxo-1,3-dioxalen-4-yl)methyl 1-piperzinyl]-4-oxo-4H-[1,3]thiazeto[3,2-a]quinoline-3-carboxylic acids
US20070149540	Apr 28, 2004	Jun 28, 2007	Nippon Shinyaky Co., Ltd.	Crystals of quinolinecarboxylic acid derivative solvate

EXTRA INFO

http://www.google.com/patents/EP2524922A1?cl=en

formula 1 is S-(-)-6-fluoro-1-methyl-7-[4-(5-methyl-2-oxo-1,3-dioxolen-4-yl)methyl-1-piperazinyl]-4-oxo-4H-[1,3]thiazet o[3,2-α]quinoline-3-carboxylic acid (levo-prulifloxacin for short); its stereo configuration is S configuration; it has optical property of levorotatory polarized light:
S-(-) ulifloxacin (as shown in formula 2 below) as raw material and the compound as shown in the following formula 3 are reacted in organic solvent in the presence of alkaline material. The reaction formula is shown below:
- S-(-)-ulifloxacin and R-(+)-ulifloxacin are prepared according to the method disclosed in CN101550142A .
- Japanese scholars Masato Matsuoka et al. have proved the absolute configuration of optically pure prulifloxacin. The study (see the publication Chem. Pharm. Bull. 43(7) 1238-1240 (1995)) verifies that (-)-ulifloxacin is S configuration while (+)-ulifloxacin is the enantiomer of R configuration by applying chemical methods together with single-crystal X-ray diffraction.
- Accordingly, R-prulifloxacin can be prepared from R-(+)-ulifloxacin and the compound of formula 3 by the method described hereinbefore.
- [0022]
  
  The reaction formula is depicted below:
- S-prulifloxacin prepared in accordance with the present invention is determined to be laevorotatory by optical rotation measurement, so it is S-(-)-prulifloxacin. R-prulifloxacin prepared in accordance with the present invention is determined to be dextrorotatory by optical rotation measurement, so it is R-(+)-prulifloxacin.
- The present invention studied the absorption features of S-(-)-prulifloxacin and R-(+)-prulifloxacin on circular polarized light by circular dichroism spectroscopy. The two spectrograms are mirror images of each other, which proves that S-(-)-prulifloxacin and R-(+)-prulifloxacin are enantiomer of each other.
- Comparing the circular dichroism spectrogram as depicted in figure 4 with the circular dichroism spectrogram of analogue of the similar structure with known absolute configuration as disclosed in the publication Chem. Pharm. Bull. 47(12) 1765-1773 (1999), it is found that (-)-prulifloxacin has similar Cotton effect to the two analogues reported in the publication, ethyl S-(-)-6,7-difluoro-1-methyl-4-oxo-4H-[1,3] thiazeto[3,2-α]quinoline-3-carboxylate and ethyl S-(-)-6, 7-difluoro-1-fluoromethyl-4-oxo-4H-[1,3]thiazeto[3,2-α]quinoline-3-carboxylate; so does (+)-prulifloxacin. The results also verify on the other hand that the absolute configuration of levo-prulifloxacin of the present invention is S type while the absolute configuration of dextro-prulifloxacin is R type.
- The compound of the present invention and physiologically acceptable acid can be prepared to salts: dissolving or suspending S-(-)-prulifloxacin in solvent such as chloroform, DMF and the like; adding into acid or acid solution (for example, hydrochloric acid or hydrogen chloride-methanol solution and the like) while stirring; precipitating and filtering to obtain solid salt from the solvent solution, or alternatively removing solvent from the salt solution directly by concentration, spray drying and the like to obtain the salt of S-(-)-prulifloxacin. The obtained solid may be further recrystallized.
Example 1 Preparation of (S)-(-)-uliflourxacin
- 105 g of racemic uliflourxacin was dissolved in 1,500 mL of dimethyl sulfoxide. 27 g of D-tartaric acid was dissolved in 405 mL of dimethyl sulfoxide dropwise while stirring. After stirring at room temperature for 20 hours, the precipitate was filtrated. The collected solid was dried under vacuum to obtain 86 g solid, which was recrystallized in dimethyl sulfoxide to obtain 37 g of levoulifloxacin-D-tartrate, with C49.08%, H5.06%, N9.50%, S7.44% shown by elemental analysis (molecular formula: C₁₆H₁₆FN₃O₃S·1/2C₄H₆O₆·H₂O, calculated values: C48.86%, H4.78, N9.50%, S7.25%). Said salt was added into water to obtain a suspension, and the pH value was adjusted to 7-8 with 2% NaOH aqueous solution while stirring. After precipitation, filtration, and drying, 24.5 g of (S)-uliflourxacin was obtained, having a chemical name (S)-(-)-6-fluoro-1-methyl-4-oxo-(1-piperazinyl)-1H,4H-[1,3]thiazeto [3,2-α]quinoline-3-carboxylic acid.
- Specific rotation [α]²⁰ _D= -133° (c=0.5, 0.1 mol/L methanesulfonic acid); ¹H-NMR (DMSO-d₆) ^δ2.11 (3H, d, j=6.2 Hz), 2.87 (4H, m), 3.19 (4H, m), 6.40 (1H, q, j=6.2 Hz), 6.89 (1H, d, j=7.4Hz), 7.79 (1H, d, j=13.9Hz), optical purity e.e. 96%.
Example 2 Preparation of (R)-(+)-uliflourxacin
- 105 g of racemic uliflourxacin was dissolved in 1,500 mL of DMSO. 27 g of L-tartaric acid was dissolved in 405 mL dimethyl sulfoxide dropwise while stirring to allow that the solution became turbid and the precipitation occurred. The solution was stirred at room temperature for 20 hours and then filtered. The collected solid was dried under vacuum to obtain 82 g solid which was recrystallized in dimethyl sulfoxide to obtain 34 g of dextrouliflourxacin-L-tartarte. Said salt was added into water to obtain a suspension, and the pH value was adjusted to 7-8 with 2% NaOH aqueous solution while stirring. After filtration and drying, 22 g of (R)-uliflourxacin was obtained, having a chemical name (R)-(+)-6-fluoro-1-methyl-4-oxo-(1-piperazinyl)-1H,4H-[1,3]thiazeto[3,2-a]quinoline -3-carboxylic acid.
- Specific rotation [α]²⁰ _D= +132.4° (c=0.5, 0.1 mol/L methanesulfonic acid), optical purity e.e. 96%.
Example 3 Preparation of S-(-)-prulifloxacin
- 3.49 g (0.01 mol) of S-(-)-uliflourxacin prepared in Example 1, 2.02 g (0.02 mol) of triethylamine and 20 ml of dimethylformamide (hereinafter referred to as DMF) were mixed and stirred. After the solution was cooled to -5∼5 °C, 0.012 mol of 4-bromomethyl-5-methyl-1,3-dioxolen-2-one (hereinafter referred to as DMDO-Br) in DMF (5 ml) solution was added thereinto, followed by stirring at -5∼5 °C for 3 hours. The reaction solution was poured into 100 ml of ice water, stirred for 30 minutes, and then filtered. The filter residue was washed with water. The solid was collected and dried under vacuum. After recrystallization from acetonitrile, 2.9 g of S-(-)-prulifloxacin was obtained, having a chemical name: S-(-)-6-fluoro-1-methyl-7-[4-(5-methyl-2-oxo-1,3-dioxolen-4-yl)methyl-1-piperazinyl ]-4-oxo-4H-[1,3]thiazeto[3,2-α]quinoline-3-carboxylic acid, with a purity of 98% and a yield rate of 63%. Specific rotation [α]²⁰ _D= -108° (c=0.5, 0.1 mol/L methanesulfonic acid)
Example 4 Preparation of R-(+)-prulifloxacin
- R-(+)-prulifloxacin prepared in Example 2 was used as raw material to prepare 2.7 g of target product R-(+)-prulifloxacin in accordance with the method as described in Example 3, with a yield rate of 60.7% and a purity of 98%. Specific rotation [α]²⁰ _D= +108° (c=0.5, 0.1 mol/L methanesulfonic acid).
- Comparing the circular dichroism spectrogram as depicted in Figure 4 with the circular dichroism spectrogram of analogue of the similar structure with known absolute configuration as disclosed in the publication Chem. Pharm. Bull. 47(12) 1765-1773 (1999), it was found that (-)-prulifloxacin has similar Cotton effect with the two analogues reported in the publication, ethyl S-(-)-6,7-difluoro-1-methyl-4-oxo-4H-[1,3] thiazeto[3,2-α]quinoline-3-carboxylate and ethyl S-(-)-6, 7-difluoro-1-fluoromethyl-4-oxo-4H-[1,3]thiazeto[3,2-α]quinoline-3-carboxylate; so does (+)-prulifloxacin. The results also verify on the other hand that the absolute configuration of levo-prulifloxacin of the present invention is S type while the absolute configuration of dextro-prulifloxacin is R type.
- Conclusion: The absolute configuration of the sample prepared in Example 3 is S configuration, as shown in the formula below:
Example 5 Preparation of S-(-)-prulifloxacin
- 3.49 g (0.01 mol) of S-(-)-uliflourxacin, 1.2 g (0.012 mol) of anhydrous potassium bicarbonate and 20 ml of dimethylsulfoxide were mixed and stirred. 0.012 mol of DMDO-Br in DMSO (5 mL) solution was added dropwise at -20 °C. Stirring proceeded at -20 °C for 3 hours. The reaction solution was poured into 100 ml of ice water, and the pH value was adjusted to 7 with 20% acetic acid. The solution was filtered after stirring for 30 minutes. The filter residue was washed with water. The solid was collected and dried under vacuum. After recrystallization from acetonitrile, 2.5 g of the target product levo-prulifloxacin was obtained with a purity of 98% and a yield rate of 54%.
  Specific rotation [α]²⁰ _D= -108° (c=0.5, 0.1 mol/L methanesulfonic acid)
Example 6 Preparation of S-(-)-prulifloxacin
- 3.49 g (0.01 mol) of S-(-)-uliflourxacin, 1.04 g (0.008 mol) of N,N-diisopropylethylamine and 20 mL of N,N-dimethylformamide (DMF) was mixed and stirred, 0.008 mol of DMDO-Br in DMF (5 mL) solution was added thereinto. The solution was heating to 60 °C and reacted for 15 minutes. The reaction solution was poured into 100 ml of ice water, and the pH value was adjusted to 7 with 20% acetic acid. The solution was filtered after stirring for 30 minutes. The filter residue was washed with water. The solid was collected and dried under vacuum. After recrystallization from acetonitrile, 2.0 g of the target product levo-prulifloxacin was obtained with a purity of 98% and a yield rate of 43%.
  Specific rotation [α]²⁰ _D= -108° (c=0.5, 0.1 mol/L methanesulfonic acid)
Example 7 Preparation of S-(-)-prulifloxacin
- 10 g (0.029 mol) of S-(-)-uliflourxacin, 30 ml of N,N-dimethylacetylamide and 14.7 g (0.145mol) of triethylamine was mixed and cooled to 5~10 °C. 8.5 g (0.03 mol) 4-(p-toluenesulfonic acid-1-methyl ester)-5-methyl-1,3-dioxolen-2-one in 25 ml of N,N-dimethylacetylamide solution was added dropwise while stirring. After addition, the solution was reacted at room temperature for 10 hours. The reaction solution was poured into 200 ml of ice water, and the pH value was adjusted to 7 with 20% acetic acid. The solution was filtered after stirring for 30 minutes. The filter residue was washed with water. The solid was collected and dried under vacuum. After recrystallization from acetonitrile, 7.46 g of the target product levo-prulifloxacin was obtained with a purity of 98% and a yield rate of 57%. Specific rotation [α]²⁰ _D= -108° (c=0.5, 0.1 mol/L methanesulfonic acid).
Example 8 Preparation of S-(-)-prulifloxacin
- 3.49 g (0.01 mol) of S-(-)-uliflourxacin, 0.79 g (0.05 mol) of potassium carbonate and 20 ml of dimethylformamide (DMF) was mixed and stirred. 0.012 mol of DMDO-Br in DMF (5ml) solution was added at -10 °C. At the same temperature, the solution was reacted for 2 hours. The reaction solution was poured into 100 ml of ice water, and the pH value was adjusted to 7 with 20% acetic acid. The solution was filtered after stirring for 30 minutes. The filter residue was washed with water. The solid was collected and dried under vacuum. After recrystallization from acetonitrile, 2.2 g of the target product levo-prulifloxacin was obtained with a purity of 98% and a yield rate of 48%. Specific rotation [α]²⁰ _D= -108° (c=0.5, 0.1 mol/L methanesulfonic acid).
Example 9 Preparation of S-(-)-prulifloxacin
- 3.49 g (0.01 mol) of S-(-)-uliflourxacin, 0.79 g (0.02 mol) of diisopropylamine and 20 ml of dimethylformamide (DMF) was mixed and stirred. 0.02 mol of DMDO-Br in DMF (5ml) solution was added at 0 °C. At the same temperature, the solution was reacted for 2 hours. The reaction solution was poured into 100 ml of ice water, and the pH value was adjusted to 7 with 20% acetic acid. The solution was filtered after stirring for 30 minutes. The filter residue was washed with water. The solid was collected and dried under vacuum. After recrystallization from acetonitrile, 2.5 g of the target product levo-prulifloxacin was obtained with a purity of 98% and a yield rate of 54%. Specific rotation [α]²⁰_D= -108° (c=0.5, 0.1 mol/L methanesulfonic acid).
Example 10 Preparation of R-(+)-prulifloxacin
- In accordance with the method as described in Example 5, the raw material R-(+)-prulifloxacin was prepared to 2.5 g of the target product R-(+)-prulifloxacin with a purity of 98% and a yield rate of 54%. Specific rotation [α]²⁰ _D= +108° (c=0.5, 0.1 mol/L methanesulfonic acid).
Example 11 Preparation of levo-prulifloxacin hydrochloride
- 0.5 g of S-(-)-prulifloxacin was dissolved in 15 mL of chloroform and then 0.5 mL of 33% (v/v) hydrochloric acid- methanol solution was added while stirring. The solution was filtered and the filtration residue was washed with methanol. The collected solid was dried to obtain 450 mg said compound with a yield rate of 83%. The melting point of the product is higher than 220 °C (the sample became darker during the test).
Example 12 Preparation of levo-prulifloxacin mesylate
- 0.5 g of S-(-)-prulifloxacin was dissolved in 15 mL of chloroform and then 0.5 mL of 50% methanesulfonic acid- methanol solution was added while stirring. The solution was filtered and the filtration residue was washed with methanol. The collected yellow solid was dried with calcium chloride under vacuum for 24 hours and further dried with calcium chloride at 80 °C under vacuum for 5 hours to obtain 470 mg said compound with a yield rate of 78%. The melting point of the product is higher than 220 °C (the sample became darker during the test).
Example 13 Preparation of levo-prulifloxacin hydrochloride
- 0.5 g of S-(-)-prulifloxacin was dissolved in 15 mL of chloroform and then 0.5 mL of 33% (v/v) hydrochloric acid- methanol solution was added while stirring. The solution was dried by evaporation. Methanol was added to the residue and stirred for 10 minutes. The solution was filtered and the filtration residue was washed with methanol. The collected solid was dried to obtain 460 mg said compound with a yield rate of 85%.

Clinafloxacin from kyorin

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Apr 052014

File:Clinafloxacin.png

Clinafloxacin

7-(3-Aminopyrrolidin-1-yl)-8-chloro-1-cyclopropyl-6-fluoro-4-oxoquinoline-3-carboxylic acid

7-(3-Amino-1-pyrrolidinyl)-8-chloro-1-cyclopropyl-6-fluoro-l,4-dihydro-4-oxo-3-quinolinecarboxylic acid

(±)-7-(3-Amino-1-pyrrolidinyl)-8-chloro-1-cyclopropyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid

105956-99-8 cas no

Clinafloxacin (INN) is a fluoroquinolone antibiotic. Its use is associated with phototoxicity and hypoglycaemia.^[1]

Clinafloxacin is a novel quinolone with wide activity against the plethora of microorganisms encountered in intraabdominal infections.

Clinafloxacin is a chlorofluoroquinolone with excellent bioavailability and activity against gram-positive, gram-negative, and anaerobic pathogens . Typical MICs for α-streptococci are 0.06–0.12 µg/mL . MIC₉₀ values for methicillin-resistant Staphylococcus aureus (MRSA) average 1.0 µg/mL. The MIC₉₀ for enterococci is typically 0.5 µg/mL . Both intravenous and oral formulations have been developed . Several studies have demonstrated the efficacy of clinafloxacin monotherapy for serious infections Clinafloxacin was also active in animal models of endocarditis, including endocarditis due to ciprofloxacin-resistant S. aureus infection .

Clinafloxacin HCl, CI-960 HCl, 105956-99-8, Clinafloxacin hydrochloride (USAN), Clinafloxacin hydrochloride [USAN], AC1L1SJB,

Molecular Formula: C₁₇H₁₈Cl₂FN₃O₃ Molecular Weight: 402.247523

……………………………………..

EP 0195316

http://www.google.com/patents/EP0195316A1?cl=en

preparation process for the compound of the invention.

A mixture of 8-chloro-1-cyclopropyl-6,7-difluoro-1,4-di- hydro-4-oxo-3-quinolinecarboxylic acid (0.6 g), anhydrous acetonitrile (6 ml), 3-aminopyrrolidine (0.35 g) and DBU (0.31 g) was refluxed for an hour. Then, 3-aminopyrrolidine (0.2 g) was more added and further refluxed for 2 hours. After cooling, the resulting precipitate was collected by filtration, dissolved in water (9 ml) containing sodium hydroxide (0.12 g) and neutralized with acetic acid. The resulting precipitate was collected by filtration and washed with water and acetonitrile successively to give the title compound (0.52 g) as colorless powder, mp 237-238 °C (decompd.).
Analysis (%) for C₁₇H₁₇ClFN₃O₃·H2O, Calcd. (Found): C, 53.20 (52.97); H, 4.99 (4.62); N, 10.95 (10.83).

Example 29 7-(3-Amino-1-pyrrolidinyl)-8-chloro-1-cyclopropyl-6-fluoro-l,4-dihydro-4-oxo-3-quinolinecarboxylic acid hydrochloride

To a suspension of 7-(3-amino-1-pyrrolidinyl)-8-chloro-1-cyclopropyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid (100 mg) in ethanol (2 ml) was added 0.2 ml of ethanol solution of hydrogen chloride (7.0 mmol HC1/ml) and then the mixture was concentrated. The resulting residue was recrystallized from methanol to give the title compound (79 mg) as light yellow prisms, mp 263-265 °C (decompd.).
Analysis (%) for C₁₇H₁₇ClFN₃O₃.HCl, Calcd. (Found): C, 50.76 (50.50); H, 4.51 (4.44); N, 10.45 (10.38).

…………………..

J. Med. Chem., 23, 1358 (1980)

structural formula D

may be readily prepared from the known starting material methyl 5-oxo-l-(phenylmethyl)-3-pyrrolidinecarboxylate, A, [J. Org. Chem., 26, 1519 (1961)] by the following reaction sequence.
The compound wherein R₃ is hydrogen, namely 3-pyrrolidinemethanamine, has been reported in J. Org. Chem., 26, 4955 (1961).

Journal of Medicinal Chemistry, 1988 , vol. 31, p. 983 – 991

References

Rubinstein, E. (2001). “History of quinolones and their side effects.”. Chemotherapy. 47 Suppl 3: 3–8; discussion 44–8.doi:10.1159/000057838. PMID 11549783.

EP0106489A2 *	Sep 6, 1983	Apr 25, 1984	Warner-Lambert Company	Antibacterial agents
EP0153163A2 *	Feb 15, 1985	Aug 28, 1985	Warner-Lambert Company	7-Substituted-1-cyclopropyl-6,8-difluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acids; 7-substituted-1-cyclopropyl-1,4-dihydro-6-fluoro-4-oxo-1,8-naphthyridine-3-carboxylic acids; their derivatives; and a process for preparing the compounds
BE899399A1 *				Title not available
GB2057440A *				Title not available




	Examples of
	reported trade
	names for products
	containing the 6-
6-Fluoroquinolin-	fluoroquinolin-
4(1H)-one	4(1H)-one	Structure

amifloxacin

balofloxacin

ciprofloxacin	Cipro®, Ciprobay, & Ciproxin

clinafloxacin

danofloxacin	Advocin & Advocid

difloxacin	Dicural® & Vetequinon

enrofloxacin	Baytril®

fleroxacin	Megalone

flumequine	Flubactin

garenoxacin

gatifloxacin	Tequin® & Zymar®

grepafloxacin	Raxar

ibafloxacin

levofloxacin	Levaquin®, Gatigol, Tavanic, Lebact, Levox, & Cravit

lomefloxacin	Maxaquin®

marbofloxacin	Marbocyl® & Zenequin

moxifloxacin	Avelox® & Vigamox®

nadifloxacin	Acuatin, Nadoxia, & Nadixa

norfloxacin	Noroxin®, Lexinor, Quinabic, & Janacin

ofloxacin	Floxin®, Oxaldin, & Tarivid

orbifloxacin	Orbax® & Victas

pazufloxacin

pefloxacin

pradofloxacin

prulifloxacin

rufloxacin	Uroflox

sarafloxacin	Floxasol, Saraflox, Sarafin

sitafloxacin

sparfloxacin	Zagam

temalioxacin	Omniflox





enoxacin	Penetrex & Enroxil

gemifloxacin	Factive

tosufloxacin

trovafloxacin	Trovan

Cenicriviroc in Phase 2 for HIV by Takeda/Tobira

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Apr 032014

Cenicriviroc

TAK-652; TBR-652

1-Benzazocine-5-carboxamide, 8-[4-(2-butoxyethoxy)phenyl]-1,2,3,4-tetrahydro-1-(2-methylpropyl)-N-[4-[[(1-propyl-1H-imidazol-5-yl)methyl]sulfinyl]phenyl]-, (5E)-

(-)-(S)-8-[4-(2-Butoxyethoxy)phenyl]-1-isobutyl-N-[4-(1-propyl-1H-imidazol-5-ylmethylsulfinyl)phenyl]-1,2,3,4-tetrahydro-1-benzazocine-5-carboxamide

(S)–(−)-8-{4-[2-(Butoxy)ethoxy]phenyl}-1-isobutyl-N-(4-{[(1-propyl-1H-imidazol-5-yl)methyl]sulfinyl}phenyl)-1,2,3,4-tetrahydro-1-benzazocine-5-carboxamide methanesulfonate

497223-25-3, Molecular Formula: C41H52N4O4S Molecular Weight: 696.94098

497223-28-6 (mesylate)^{C41 H52 N4 O4 S . C H4 O3 S, 793.047}

Cenicriviroc, Cenicriviroc (USAN/INN), TAK652, TBR652, , 497223-25-3, D09878

Cenicriviroc (TAK-652, TBR-652) is an experimental drug candidate for the treatment of HIV infection.^[1] It is being developed by Takeda Pharmaceutical and Tobira Therapeutics.

TBR-652 (formerly TAK-652) is a highly potent and orally active CCR5 antagonist in phase II clinical trials at Takeda for the treatment of HIV infection. Tobira Therapeutics is evaluating the compound in preclinical studies for the treatment of rheumatoid arthritis.

TBR-652 binds CCR5 receptors to interfere with the entry of the HIV-1 virus into macrophages and activated T-cells by inhibiting fusion between viral and cellular membranes. This mechanism of action differs from currently used HIV treatments such as nucleoside reverse transcriptase inhibitors and protease inhibitors.

In 2007, Takeda entered into an agreement with Tobira pursuant to which Tobira obtained exclusive worldwide rights to develop, manufacture and commercialize TBR-652 for the treatment of HIV infection.

Cenicriviroc is an inhibitor of CCR2 and CCR5 receptors,^[2] allowing it to function as an entry inhibitor which prevents the virus from entering into a human cell. Inhibition of CCR2 may have an anti-inflammatory effect.

A double-blind, randomized, placebo-controlled clinical study to assess the antiviral activity, safety, and tolerability of cenicriviroc was conducted in 2010. HIV-infected patients taking cenicriviroc had significant reductions in viral load, with the effect persisting up to two weeks after discontinuation of treatment.^[3] Additional Phase II clinical trials are underway.^[4]

Phase IIb data presented at the 20th Conference on Retroviruses and Opportunistic Infections (CROI) in March 2013 showed similar viral suppression rates of 76% for patients taking 100 mg cenicriviroc, 73% with 200 mg cenicriviroc, and 71% with efavirenz. Non-response rates were higher with cenicriviroc, however, largely due to greater drop-out of patients. A new tablet formulation with lower pill burden may improve adherence. Looking at immune and inflammatory biomarkers, levels of MCP-1 increased and soluble CD14 decreased in the cenicriviroc arms.^[5]

Although HIV has been largely rendered a chronic infection, there remains a need for new drugs because of the virus’s propensity to develop resistance to the drugs used to keep it at bay.

Pfizer’s maraviroc was the first drug that acted on the cells to prevent viral entry by antagonising the CCR5 co-receptor. Several others have been investigated and have failed; another that is undergoing clinical trials is Takeda’s cenicriviroc, which has been licensed to Tobira Therapeutics. Unlike maraviroc, the new agent also acts at the CCR2 co-receptor, which is implicated in cardiovascular and metabolic diseases.

In a Phase I double blind, placebo controlled trial designed to study safety, efficacy and pharmacokinetics, treatment-experienced but CCR5 antagonist-naïve patients with HIV-1 were given doses of 25, 50, 75, 100 or 150mg of the drug, or placebo once a day for 10 days.² The maximum median reductions in HIV-1 RNA values were 0.7, 1.6, 1.8 and 1.7 log10 copies/ml for the respective doses, with a median time to nadir of 10 to 11 days. The effect on CD-4 cell counts was negligible. There was also a significant reduction in levels of monocyte chemotactic protein 1, suggesting that CCR2 was also being blocked. The drug was both generally safe and well tolerated, and no patients withdrew from the trial due to adverse events.

In another Phase I trial, designed to look at pharmacokinetics and pharmacodynamics and carried out in a similar patient population, subjects were given the drug as oral monotherapy for 10 days, again in doses of 25, 50, 75, 100 and 150mg, or placebo.³ The drug was well absorbed into the systemic circulation, and the concentration levels declined slowly, with meant elimination half-lives of one to two days. Potent, dose-dependent reductions in viral load were seen, and again it was generally safe and well tolerated across all levels.

In June 2011, Tobira initiated a multi-centre, double blind, double dummy, 48-week comparative Phase IIb trial in 150 patients with HIV-1 infection. Subjects are being given 100 or 200mg once-daily doses of the drug to evaluate its efficacy, safety and tolerability.

PATENTS

WO 2003014105

WO 2003076411

WO 2005116013

WO 2007144720

WO 2011163389

US 20130079233

WO 2013167743

References

Klibanov, Olga M.; Williams, Shannon H.; Iler, Cameron A (2010). “Cenicriviroc, an orally active CCR5 antagonist for the potential treatment of HIV infection”. Current Opinion in Investigational Drugs 11 (8): 940–950. PMID 20721836.
Baba, Masanori; Takashima, Katsunori; Miyake, Hiroshi; Kanzaki, Naoyuki; Teshima, Koichiro; Wang, Xin; Shiraishi, Mitsuru; Iizawa, Yuji (2005). “TAK-652 inhibits CCR5-mediated human immunodeficiency virus type 1 infection in vitro and has favorable pharmacokinetics in humans”. Antimicrobial Agents and Chemotherapy 49 (11): 4584–4591. doi:10.1128/AAC.49.11.4584-4591.2005. PMC 1280155. PMID 16251299.
C. Reviriego (2011). Drugs of the Future 36 (7): 511–517. doi:10.1358/dof.2011.36.7.1622066.
“Tobira Therapeutics Initiates Phase 2b Trial of Cenicriviroc”. The Body. July 5, 2011.
CROI 2013: CCR5/CCR2 Inhibitor Cenicriviroc Has Both Anti-HIV and Anti-inflammatory Effects. Highleyman, Liz. HIVandHepatitis.com. 7 March 2013.


22931505	11-26-2012	Chemokine receptor antagonists.	Journal of medicinal chemistry
21317794	6-1-2011	Safety, efficacy, and pharmacokinetics of TBR-652, a CCR5/CCR2 antagonist, in HIV-1-infected, treatment-experienced, CCR5 antagonist-naive subjects.	Journal of acquired immune deficiency syndromes (1999)
20721836	8-1-2010	Cenicriviroc, an orally active CCR5 antagonist for the potential treatment of HIV infection.	Current opinion in investigational drugs (London, England : 2000)
19064629	3-1-2009	The relative activity of “function sparing” HIV-1 entry inhibitors on viral entry and CCR5 internalization: is allosteric functional selectivity a valuable therapeutic property?	Molecular pharmacology
17116673	2-1-2007	Isolation and characterization of human immunodeficiency virus type 1 resistant to the small-molecule CCR5 antagonist TAK-652.	Antimicrobial agents and chemotherapy
17037325	9-10-2006	[Progress in AIDS therapy].	Nihon Naika Gakkai zasshi. The Journal of the Japanese Society of Internal Medicine
16539392	3-23-2006	Highly potent and orally active CCR5 antagonists as anti-HIV-1 agents: synthesis and biological activities of 1-benzazocine derivatives containing a sulfoxide moiety.	Journal of medicinal chemistry
16251299	11-1-2005	TAK-652 inhibits CCR5-mediated human immunodeficiency virus type 1 infection in vitro and has favorable pharmacokinetics in humans.	Antimicrobial agents and chemotherapy
15658852	1-27-2005	Stereoselective synthesis of [L-Arg-L/D-3-(2-naphthyl)alanine]-type (E)-alkene dipeptide isosteres and its application to the synthesis and biological evaluation of pseudopeptide analogues of the CXCR4 antagonist FC131.	Journal of medicinal chemistry
16430202	1-1-2005	TAK-652, a novel CCR5 inhibitor, has favourable drug interactions with other antiretrovirals in vitro.	Antiviral therapy

……………….

Chemical structures of selected small molecule CCR5 inhibitors. A. Maraviroc (MVC, Selzentry), B. Vicriviroc (VCV), C. Cenicriviroc (TBR-652), D. PF-232798.

http://www.intechopen.com/books/immunodeficiency/chemokine-receptors-as-therapeutic-targets-in-hiv-infection

RANBEZOLID FROM RANBAXY

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Apr 022014

Ranbezolid

392659-39-1 hydrochloride

392659-38-0 (free base)

N-{[(5S)-3-(3-Fluoro-4-{4-[(5-nitro-2-furyl)methyl]-1-piperazinyl}phenyl)-2-oxo-1,3-oxazolidin-5-yl]methyl}acetamide

(S)-N-[[3-fluoro-4-[N-1[4-{2-furyl-(5-nitro)methyl}]piperazinyl]-phenyl]-2-oxo-5-oxazolidinyl]-methyl]acetamide

AC1LAX1P, RBx7644 (*Hydrochloride*),RBx-7644

Molecular Formula: C₂₁H₂₄FN₅O₆ Molecular Weight: 461.443563

Ranbaxy Lab Ltd ORIGINATOR

Ranbezolid is a novel oxazolidinone antibacterial. It competitively inhibits monoamine oxidase-A (MAO-A).^[1]

Infections due to Gram-positive bacteria such as methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococcus faecium (VRE), and penicillin-resistant Streptococcus pneumoniae(PRSP) are the leading cause of morbidity and mortality in hospital settings and community today. Oxazolidinones are a new class of totally synthetic antibacterial agents active against Gram-positive infections. Linezolid (Zyvox™, Pharmacia/Pfizer, is a drug in this class, approved in the United States and Europe for treatment of Gram-positive nosocomial and community-acquired pneumoniae and skin infections. Oxazolidinones inhibit the bacterial protein synthesis prior to the chain initiation step, by binding to the 23S rRNA of 50S ribosomal subunit, and interfering with the initiator fMet–tRNA binding to the P-site of the ribosomal peptidyltransferase centre

Ranbezolid hydrochloride, RBx-7644

US2005209248

9-23-2005

Plymorphic forms of phenyl oxazolidinone derivatives

The title compound is prepared by reductive alkylation of the known piperazinyl oxazolidinone derivative (I) with 5-nitro-2-furfural (II) in the presence of NaBH(OAc)3, followed by conversion to the corresponding hydrochloride salt.

EP 1303511; US 2002103186; WO 0206278; WO 0307870; WO 0308389

…………….

synthesis

The antibacterial activity of RBx-7644 is due to the 5(S)-acetamidomethyl configuration at the oxazolidinone ring, and thus, asymmetric synthesis of only the 5(S)-enantiomer was desirable: 3,4-Difluoronitrobenzene (I) is condensed with piperazine in acetonitrile to give 4-(2-fluoro-4-nitrophenyl)-piperazine (II) as a light yellow compound. Compound (II) is dissolved in dichloromethane and triethylamine, followed by the addition of Boc-anhydride, to provide compound (III). 4-(tert-Butoxycarbonyl)-1-(2-fluoro-4-nitrophenyl)piperazine (III), upon hydrogenation with H2 over Pd/C in methanol at 50 psi, yields 4-(tert-butoxycarbonyl)-1-(2-fluoro-4-aminophenyl)piperazine (IV) as a dark solid. Compound (IV) reacts with benzylchloroformate in dry THF in the presence of solid sodium bicarbonate to afford the desired compound (V). 4-(tert-Butoxycarbonyl)-1-[2-fluoro-4-(benzyloxycarbonylamino)phenyl]piperazine (V), upon treatment with n-BuLi and (R)-glycidyl butyrate at -78 癈, gives the desired (R)-(-)-3-[3-fluoro-4-[4-(tert-butoxycarbonyl)piperazin-1-yl]phenyl]-5-(hydroxymethyl)-2-oxazolidinone (VI). The hydroxymethyl compound (VI) is treated with methanesulfonyl chloride in dichloromethane in the presence of triethylamine to give (R)-(-)-3-[3-fluoro-4-[4-(tert-butoxycarbonyl)piperazin-1-yl]phenyl]-5-(methylsulfonyloxymethyl)-2-oxazolidinone (VII). The sulfonyl derivative (VII) is treated with sodium azide in dimethylformamide to provide the azide (VIII) as a white solid. (R)-(-)-3-[3-Fluoro-4-[4-(tert-butoxycarbonyl)piperazin-1-yl)phenyl]-5-(azidomethyl)-2-oxazolidinone (VIII), upon hydrogenation with H2 over Pd/C at 45 psi, gives (S)-(-)-3-[3-fluoro-4-[4-(tert-butoxycarbonyl)-piperazin-1-yl]phenyl]-5-(aminomethyl)-2-oxazolidinone (IX). The aminomethyl compound (IX), upon treatment with acetic anhydride in dichloromethane in the presence of triethylamine, affords the acetamide derivative (X). The acetamidomethyl-oxazolidinone derivative (X), upon treatment with trifluoroacetic acid, gives (S)-(-)-3-[3-fluoro-4-(1-piperazinyl)phenyl]-5-(acetamidomethyl)-2-oxazolidinone, which, without isolation, is treated with 5-nitro-2-furaldehyde in the presence of sodium triacetoxy borohydride to provide compound (XI). Compound (XI), upon treatment with ethanolic HCl, affords RBx-7644 as a light yellow crystalline solid.

………………….

polymorphs

http://www.google.com/patents/US20050209248

(S)-N-[[3-fluoro-4-[N-1[4-{2-furyl-(5-nitro)methyl}]piperazinyl]-phenyl]-2-oxo-5-oxazolidinyl]-methyl]acetamidehydrochloride having the Formula I.

The compound of Formula I, namely, (S)-N-[[3-fluoro-4-[N-1 [4-{2-furyl-(5-nitro)methyl}] piperazinyl]-phenyl]-2-oxo-5-oxazolidinyl]-methyl]acetamide hydrochloride is a phenyl oxazolidinone derivative, as disclosed in PCT application WO 02/06278. It is said to be useful as antimicrobial agent, effective against a number of human and veterinary pathogens, including gram-positive aerobic bacteria, such as multiply resistant staphylococci, streptococci and enterococci as well as anaerobic organisms such as Bacterioides spp. andClostridia spp. species, and acid fast organisms such as Mycobacterium tuberculosis, Mycobacterium avium and Mycobacterium spp.

The PCT application WO 02/06278 describes the preparation of compounds of Formula I. The products of Formula I obtained by following the cited methods tend to be hygroscopic and difficult to filter. These types of disadvantageous properties have proven to be serious obstacles to the large-scale manufacture of a compound. Further, handling problems are encountered during the preparation of pharmaceutical compositions comprising the hygroscopic compound of Formula I obtained by following the method disclosed in WO 02/06278.

EXAMPLE 1 Preparation of Polymorphic ‘Form A’ of the Compound of Formula I

50 gm of free base of Formula I was dissolved in ethanol (750 ml) by heating at about 60° C. and to this solution was added ethanolic HCl (13.36 ml, 8.9 N) at about 45-50° C. The reaction mixture was cooled to about 10° C., and stirred for about 4 hours. The separated solid was filtered off and dried under vacuum at 60° C. The solid was then digested in ethanol (150 ml) at 70-80° C. for about 4 hours. It was then cooled to about 10° C., the solid was filtered and dried under vacuum at 60-65° C. to give 30 gm of the pure polymorphic ‘Form A’ of compound of Formula I.

………………

Synthesis and SAR of novel oxazolidinones: Discovery of ranbezolid

Bioorg Med Chem Lett 2005, 15(19): 4261

http://www.sciencedirect.com/science/article/pii/S0960894X05008310

Synthesis and SAR of novel oxazolidinones: Discovery of ranbezolid

Pages 4261-4267
Biswajit Das, Sonali Rudra, Ajay Yadav, Abhijit Ray, A.V.S. Raja Rao, A.S.S.V. Srinivas, Ajay Soni, Suman Saini, Shalini Shukla, Manisha Pandya, Pragya Bhateja, Sunita Malhotra, Tarun Mathur, S.K. Arora, Ashok Rattan, Anita Mehta

Graphical abstract

Novel oxazolidinones were synthesized containing a number of substituted five-membered heterocycles attached to the ‘piperazinyl–phenyl–oxazolidinone’ core of eperezolid. Further, the piperazine ring of the core was replaced by other diamino-heterocycles. These modifications led to several compounds with potent activity against a spectrum of resistant and susceptible Gram-positive organisms, along with the identification of ranbezolid (RBx 7644) as a clinical candidate.

Substitution of five-membered heterocycles on to the ‘piperazinyl–phenyl–oxazolidinone’ core structure led to the identification of ranbezolid as a clinical candidate. Further replacement of piperazine ring with other diamino-heterocycles led to compounds with potent antibacterial activity.

Scheme 5.

Reagents and conditions: (a) Method A: TFA, CH₂Cl₂, 0 °C → rt; 5-chloromethyl-2-furaldehyde, potassium carbonate, DMF, rt; or (b) Method B: TFA, CH₂Cl₂, 0 °C → rt; 5-nitrofuran-2-carboxaldehyde, sodiumtriacetoxyborohydride, THF, molecular sieves 3 Å, rt. 7 = ranbezolid

Synthesis of compound 7: (S)-N-[[3-[3-Fluoro-4-(N-4-tert-butoxycarbonyl-piperazin-1-yl)phenyl]-2-oxo-5-oxa-zolidinyl]-methyl]acetamide (28a, 3.65 kg, 8.37 mol) was dissolved in dichloromethane (30.86 L) and cooled to 5 °C. To it trifluoroacetic acid (6.17 L) added dropwise and stirred for 14 h allowing the reaction mixture to warm to rt. The reaction mixture was evaporated in vacuo and the residue dissolved in tetrahydrofuran (58 L) followed by addition of molecular sieves 4 Å (4.2 kg). To the resulting mixture 5-nitro-2-furaldehyde (1.5 kg, 10.77 mol) was added followed by sodium triacetoxyborohydride (5.32 kg, 25.1 mol) and stirred for 14 h. The reaction mixture was filtered over Celite and filtrate evaporated in vacuo. The residue was dissolved in ethylacetate (85.6 L) and washed with satd sodium bicarbonate solution (36 L) and water (36 L). The organic layer was dried over anhyd sodium sulfate (3 kg) and evaporated in vacuo. The crude residue was purified by column chromatography (1–3% methanol in ethylacetate) to obtain (S)-N-[[3-[3-fluoro-4-[N-4-(5-nitro-2-furylmethyl)-piperazin-1-yl]phenyl]-2-oxo-5-oxa-zolidinyl]methyl]acetamide (39, 2.6 kg, yield 67%). Mp: 136 °C. ¹H NMR (CDCl₃): δ 7.42 (dd, 1H, phenyl–H), 7.29 (m, 2H, furyl–H), 7.07 (d, 1H, phenyl–H), 6.92 (t, 1H, phenyl–H), 6.51 (d, 1H, furyl–H), 6.11 (t, 1H, –NHCO–), 4.77 (m, 1H, oxazolidinone ring C₅–H), 4.01 (t, 1H), 3.85–3.45 (m, 5H), 3.09 (m, 4H, piperazine–H), 2.72 (m, 4H, piperazine–H), 2.02 (s, 3H, –COCH₃). MS m/z (rel. int.): 462.1 [(M+H)⁺, 100%], 484 [(M+Na)⁺, 25%], 500.2 [(M+K)⁺, 20%]. HPLC purity: 98%.
Compound 39(3.6 kg, 7.81 mol) was dissolved in abs ethanol (53.8 L) by heating to 60 °C. The resulting solution was cooled to 45 °C and ethanolic hydrochloride (1.48 L, 7.9 N) was added dropwise in 10 min. The mixture was then cooled to 10 °C and stirred for 4 h and the precipitate formed was filtered and washed with ethanol and dried to obtain (S)-N-[[3-[3-fluoro-4-[N-4-(5-nitro-2-furylmethyl)-piperazin-1-yl]phenyl]-2-oxo-5-oxazolidinyl]-methyl]acetamide hydrochloride, ranbezolid (7, 3.2 kg, yield from 39: 82%, yield from 28a: 55%).
Ranbezolid
Mp: 207–209 °C.
¹H NMR (DMSO, 300 MHz): δ 8.30 (t, 1H, –NHCO–), 7.75 (d, J = 3.3 Hz, 1H, furyl–H), 7.52 (dd, 1H, phenyl–H), 7.3–7.0 (m, 3H, phenyl–H, furyl–H), 4.70 (m, 1H, oxazolidinone ring C₅–H), 4.63 (s, 2H), 4.08 (t, J = 8.8 Hz, 1H, –CH₂–), 3.73 (t, J = 7.5 Hz, 1H), 3.43 (br m, piperazine–H merged with H₂O in DMSO), 1.83 (s, 3H, –COCH₃).
HPLC purity: 98%. Anal. Calcd for C₂₁H₂₅ClN₅O₆·0.5H₂O: C, 50.76; H, 5.48; N, 14.09. Anal. Found: C, 50.83; H, 5.17; N, 13.83.

References

European Journal of Pharmacology. 2006. 545, 167–172
US2005209248, 9-23-2005
Plymorphic forms of phenyl oxazolidinone derivatives

23142085	1-1-2013	Anti-anaerobic potential of ranbezolid: insight into its mechanism of action against Bacteroides fragilis.	International journal of antimicrobial agents
19811911	11-15-2009	Synthesis and biological activity of novel oxazolidinones.	Bioorganic & medicinal chemistry letters
19075051	4-1-2009	Mode of action of Ranbezolid against staphylococci and structural modeling studies of its interaction with ribosomes.	Antimicrobial agents and chemotherapy
18676219	8-1-2008	Effect of oxazolidinone, RBx 7644 (ranbezolid), on inhibition of staphylococcal adherence to plastic surfaces.	Journal of chemotherapy (Florence, Italy)
18622683	4-1-2008	Utilization of Bombyx mori larvae as a surrogate animal model for evaluation of the anti-infective potential of oxazolidinones.	Journal of infection and chemotherapy : official journal of the Japan Society of Chemotherapy
17624776	9-15-2007	Synthesis and in vitro antibacterial activity of novel methylamino piperidinyl oxazolidinones.	Bioorganic & medicinal chemistry letters
16899241	9-18-2006	Ranbezolid, a novel oxazolidinone antibacterial: in vivo characterisation of monoamine oxidase inhibitory potential in conscious rats.	European journal of pharmacology
16054358	10-1-2005	Synthesis and SAR of novel oxazolidinones: discovery of ranbezolid.	Bioorganic & medicinal chemistry letters
15885988	6-1-2005	Activity of RBx 7644 and RBx 8700, new investigational oxazolidinones, against Mycobacterium tuberculosis infected murine macrophages.	International journal of antimicrobial agents
15380263	10-1-2004	In vitro activity of RBx 7644 (ranbezolid) on biofilm producing bacteria.	International journal of antimicrobial agents

12604558	3-1-2003	Antianaerobe activity of RBX 7644 (ranbezolid), a new oxazolidinone, compared with those of eight other agents.	Antimicrobial agents and chemotherapy
12604559	3-1-2003	Antipneumococcal and antistaphylococcal activities of ranbezolid (RBX 7644), a new oxazolidinone, compared to those of other agents.	Antimicrobial agents and chemotherapy

MK 2048 an HIV integrase inhibitor from Merck

Uncategorized Comments Off

Apr 022014

MK 2048

Molecular Formula: C₂₁H₂₁ClFN₅O₄ Molecular Weight: 461.873943

869901-69-9, 3oyl, 3oyn

Merck & Co., Inc.

(6S)-2-(3-chloro-4-fluorobenzyl)-8-ethyl-10-hydroxy-N,6-dimethyl-1,9-dioxo-1,2,6,7,8,9-hexahydropyrazino‌[1′,2′:1,5]‌pyrrolo‌[2,3-d]‌pyridazine-4-carboxamide

6(S)-2-(3-Chloro-4-fluorobenzyl)-8-ethyl-10-hydroxy-N,6-dimethyl-l,9-dioxo-l,2,6,7,8,9- hexahydropyrazino[r,2′:l,5]pyrrolo[2,3-d]pyridazine-4-carboxamide

US7538112

5-27-2009

Hiv Integrase Inhibitors

MK-2048 is a second generation integrase inhibitor, intended to be used against HIV infection. It is superior to the first available integrase inhibitor,raltegravir, in that it inhibits the HIV enzyme integrase 4 times longer. It is being investigated for use as part of pre-exposure prophylaxis (PrEP). ^[1]

It is being developed by Merck & Co.^[2]

MK-2048 is a second generation integrase inhibitor for HIV-1 integrase. MK-2048 inhibits subtype B and subtype C integrase activities. MK-2048 inhibits R263K mutants slightly more effectively than G118R mutants.

MK-2048 inhibits S217H intasome and, by contrast, MK2048 remains fully active against the N224H intasome. MK2048 displays substantially lower dissociation rates compared with raltegravir, another integrase inhibitor.

MK-2048 is active against viruses resistant to RAL and EVG. MK-2048 exposure leads to the selection of G118R as a possible novel resistance mutation after 19 weeks. MK-2048, with continued pressure, subsequently leads to an additional substitution, at position E138K, after 29 weeks, within the IN gene.

Although the G118R mutation alone confers only slight resistance to MK-2048 but not to RAL or EVG, its presence arouses a dramatic reduction in viral replication capacity compared to wild-type NL4-3. E138K both partially restores viral replication capacity and also contributes to increased levels of resistance against MK-2048.

Structure of MK-2048 with important pharmacophore highlighted

…………………..

Synthesis

WO2005110415A1

http://www.google.as/patents/WO2005110415A1?cl=en

EXAMPLE 62 6(S)-2-(3-Chloro-4-fluorobenzyl)-8-ethyl-10-hydroxy-N,6-dimethyl-l,9-dioxo-l,2,6,7,8,9- hexahydropyrazino[r,2′:l,5]pyrrolo[2,3-d]pyridazine-4-carboxamide

Step 1: te rt-Butyl[( 1 S)-2-(ethylamino)- 1 -methylethyl] carbamate To a cold (0 °C) solution of N-(tø/ -butoxycarbonyl)-L-alanine N’-methoxy-N’- methylamide (15.6 g, 67.2 mmol) in anhydrous THF (150 mL) and diethyl ether (400 mL), solid lithium aluminum hydride (5.1 g, 134.3 mmol) was added portionwise over a period of 30 minutes. The mixture was stirred at room temperature for 3 hours and cooled back to 0 °C. The reaction was treated carefully with an aqueous solution of potassium hydrogen sulfate (250 mL, 1M). The resultant mixture was diluted with diethyl ether.

The organic extract was washed successively with dilute hydrochloric acid, and brine, dried over anhydrous sodium sulfate, filtered, and concentrated under vacuum to provide the corresponding aldehyde as colorless solid. Without further purification, a cold (0 °C), stirred solution of the intermediate aldehyde (10.7 g, 61.8 mmol) and ethylamine hydrogen chloride (10.1 g, 123.5 mmol) in methanol (72 mL) was treated with sodium triacetoxyborohydride (17.2 g, 80.9 mmol) in one portion. The mixture was allowed to warm up to room temperature.

After stirring at room temperature overnight, the solution was concentrated under vacuum. The residue was partitioned between diethyl ether and cold aqueous sodium hydroxide (1.5 M). The ethereal extract was washed with brine, dried over anhydrous sodium sulfate, filtered, and concentrated under vacuum to provide the titled compound. lH NMR (400 MHz, CDCI3) δ 4.68 (br s, IH), 3.75 (br t, IH), 2.62 (m, 5 H), 1.13 (d, J = 6.7 Hz, 3H),

1.09 (t, J = 7.0 Hz, 3H). ES MS M+l = 203

Step 2: ført-Butyl { ( 1 S)-2-[(bromoacetyl)ethylamino] – 1 -methylethyl } carbamate To a cold (0 °C) stirred solution of ?ert-butyl[(lS)-2-(ethylamino)-l- methylethyl]carbamate (11.0 g, 54.6 mmol) in a mixture of ethyl acetate (107 mL) and saturated aqueous sodium bicarbonate (65 mL), bromoacetyl bromide (12.1 g, 60.0 mmol) was added portionwise under an atmosphere of nitrogen. The mixture was allowed to warm up to room temperature over a period of 3.5 hours. The organic phase was separated, washed successively with saturated aqueous sodium bicarbonate, and brine. The organic extract was dried over anhydrous sodium sulfate, filtered, and concentrated under vacuum. The residue was concentrated as a solution in toluene under vacuum to afford the title compound. ES MS M+l = 323, 325.

Step 3: fe7 -Butyl (2S)-4-ethyl-2-methyl-5-oxopiperazine-l-carboxylate To a stirred slurry of sodium hydride (1.7 g, 69.8 mmol) in anhydrous THF (800 mL), a solution of tert-butyl{(lS)-2-[(bromoacetyl)ethylamino]-l-methylethyl}carbamate (17.4 g, 53.7 mmol) in anhydrous THF (100 mL) was added dropwise over a period of 1 hour under an atmosphere of nitrogen. The reaction mixture was stirred at room temperature for two hours, cooled in an ice-water bath, and quenched with dropwise addition of aqueous citric acid (80 mL, 1M). The mixture was concentrated under vacuum. The residue was partitioned between chloroform and saturated aqueous sodium bicarbonate. The organic extract was dried over anhydrous sodium sulfate, filtered, and concentrated under vacuum. The residue was subjected to column chromatography on silica gel eluting with a gradient of 0-15% acetonitrile in chloroform. Collection and concentration of appropriate fractions provided the title compound. lH NMR (400 MHz, CDCI3) δ 4.46 (br s, IH), 4.24 (d, J = 18.4 Hz, 1 H), 3.78 (d, J = 18.4 Hz, 1 H),

3.64 (dd, J = 12.3, 4.2 Hz, 1 H), 3.54 (heptet, J = 7.1 Hz, 1 H), 3.38 (heptet, J = 7.1 Hz, 1 H), 2.99 (dd, J = 12.3, 1.8 Hz, 1 H), 1.47 (s, 9H), 1.21 (d, J = 6.8 Hz, 3H), 1.14 (t, J = 7.1 Hz, 3H). ES MS M+l = 243.

Step 4: (5S)-l-Ethyl-5-methylpiperazin-2-one hydrochloride Anhydrous hydrogen chloride gas was bubbled into a cold (-20 °C) solution of tert-butyl (2S)-4-ethyl-2-methyl-5-oxopiperazine-l-carboxylate (10.5 g, 43.4 mmol) in ethyl acetate (250 mL) under nitrogen. After the solution was saturated with hydrogen chloride, the reaction mixture was stirred in an ice-water bath for 30 minutes. The product mixture was purged with nitrogen, concentrated under vacuum to provide the title hydrogen chloride salt as pale yellow solid. lH NMR (400 MHz, DMSO-d6) δ 10.00 (br d, 2H), 3.72 (d, J = 16.6 Hz, 1 H), 3.62(d, J = 16.6 Hz, 1 H),

3.49-3.35 (m, 5 H), 3.29 (heptet, /= 7.3 Hz, 1 H), 1.31 (d, / = 6.6 Hz, 3H), 1.05 (t, J = 7.1 Hz, 3H).

Step 5: Ethyl (4S)-2-ethyl-8-hydroxy-4-methyl-l-oxo-l,2,3,4-tetrahydropyrrolo[l,2-a]pyrazin-7- carboxy late Anhydrous ammonia gas was bubbled into a cold (0 °C) solution of (5S)-l-Ethyl-5- methylpiperazin-2-one hydrochloride (5.8 g, 32.3 mmol) in chloroform for 30 minutes. The resultant slurry was filtered and concentrated under vacuum. The residual oil was concentrated as a solution in toluene under vacuum, redissolved in toluene (120 mL) and treated with diethyl ethoxymethylenemalonate (7.0 g, 32.3 mmol) and heated in a sealed flask in an oil bath at 100 °C overnight. The resultant solution was concentrated under vacuum. The residual oil was concentrated as a solution in toluene under vacuum to provide the corresponding diethyl { [(2S)-4-ethyl-2-methyl-5- oxopiperazin-l-yl]methylene}malonate. Without further purification, to a solution of the malonate (10.5 g, 33.5 mmol) in anhydrous THF (330 mL) warmed with an external oil bath at 65 °C under an atmosphere of nitrogen, a solution of lithium bis(trimethylsilyl)amide (35.1 mL, 1 M, 35.1 mmol) was added. The solution was heated at the same temperature for one hour and concentrated under vacuum. The residue was partitioned between dichloromethane and hydrochloric acid (1M). The organic extract was washed with brine, dried over anhydrous magnesium sulfate, filtered, and concentrated under vacuum. The residue was triturated with diethyl ether. The solid precipitated was filtered, washed with diethyl ether to provide the title compound as pale brown solid. lH NMR (400 MHz, CDCI3) δ 8.43 (s, IH), 7.11 (s, IH), 4.32 (q, J = 7.1 Hz, 2H), 4.24 (m, IH), 3.65-

3.35 (m, 4H), 1.51 (d, J = 6.4 Hz, 3H), 1.36 (t, J = 7.0 Hz, 3H), 1.19 (t, J = 7.0 Hz, 3H). ES MS M+l = 267

Step 6: Ethyl (4S)-2-ethyl-8-methoxy-4-methyl-l-oxo-l,2,3,4-tetrahydropyrrolo[l,2-a]pyrazin-7- carboxylate A mixture of ethyl (4S)-2-ethyl-8-hydroxy-4-methyl-l -oxo- 1,2,3, 4-tetrahydropyrrolo[ 1,2- a]pyrazin-7-carboxylate (6.6 g, 24.8 mmol), anhydrous potassium carbonate (13.7 g, 99.1 mmol, 325 mesh), and iodomethane (4.2 g, 29.7 mmol) in anhydrous DMF (123 mL) was stirred at room temperature overnight. The mixture was filtered and concentrated under vacuum. The residue was partitioned between chloroform and dilute hydrochloric acid. The organic extract was washed with brine, dried over anhydrous sodium sulfate, filtered, and concentrated under vacuum. The residue was subjected to column chromatography on silica gel eluting with a gradient of 0-3% methanol in chloroform. Collection and concentration of appropriate fractions provided the title compound. Residual methanol was removed by concentrating from its solution in toluene under vacuum. lH NMR (400 MHz, CDCI3) δ 7.19 (s, IH), 4.29 (q, J = 7.1 Hz, 2 H), 4.24 (m, IH), 4.03 (s, 3H), 3.70-

3.32 (m, 4 H), 1.52 (d, J = 6.6 Hz, 3H), 1.35 (t, J = 7.0 Hz, 3H), 1.19 (t, J = 7.2 Hz, 3H). ES MS M+l = 281

Step 7: Ethyl (4S)-6-bromo-2-ethyl-8-methoxy-4-methyl-l-oxo-l,2,3,4-tetrahydropyrrolo[l,2- a]pyrazin-7-carboxylate To a mixture of ethyl (4S)-2-ethyl-8-(methoxy)-4-methyl-l-oxo-l,2,3,4- tetrahydropyrrolo[l,2- ]pyrazine-7-carboxylate (6.2 g, 22.1 mmol) and sodium bicarbonate (20.0 g, 238.0 mmol) in dichloromethane (500 mL) at 0 °C, a solution of bromine in dichloromethane (24.2 mmol, 0.5 M) was added over a period of 60 minutes. The reaction mixture was stirred at room temperature for 2 h, filtered, and concentrated under vacuum. The residue was subjected to column chromatography on silica gel eluted with ethyl acetate. Collection and concentration of appropriate fractions provided the corresponding bromide. Residual ethyl acetate was removed by concentrating from its solution in benzene under vacuum. lH NMR (400 MHz, CDCI3) δ 4.58 (br m, IH), 4.34 (m, IH), 3.99 (s, 3H), 3.92 (dd, J = 13.0, 4.0 Hz,

IH), 3.67 (heptet, J = 7.1 Hz, 1 H), 3.49 (heptet, J = 7.1 Hz, 1 H), 3.23 (d, J = 13.0 Hz, IH), 1.40 (d, J = 7.1 Hz, 3H), 1.38 (t, 7 = 7.0 Hz, 3H), 1.20 (t, J = 7.0 Hz, 3H). ES MS M+l = 359, 361.

Step 8: Ethyl (4S)-2-ethyl-8-(methoxy)-6-[methoxy(oxo)acetyl]-4-methyl-l-oxo-l,2,3,4- tetrahydropyrrolo[ 1 ,2- ]pyrazine-7-carboxylate To a cold (-78 °C) solution of ethyl (4S)-6-bromo-2-ethyl-8-methoxy-4-methyl-l-oxo- l,2,3,4-tetrahydropyrrolo[l,2-a]pyrazin-7-carboxylate (8.51 g, 23.7 mmol) in anhydrous THF (800 mL) under an atmosphere of dry nitrogen, a solution of n-BuLi in hexane (10.5 mL, 26.3 mmol, 2.5 M) was added. The resultant mixture was stirred at -78 °C for 20 minutes. A solution of dimethyl oxalate (6.4 g, 53.8 mmol; dried from concentration from benzene under vac) in anhydrous THF (30 mL) was added. The reaction mixture was stirred at -78 °C for 1 hour and cannulated into a mixture of aqueous sulfuric acid (240 mL, 2M) and THF (200 mL) maintained between at -5 to -35 °C. The mixture was extracted with ethyl acetate (3 times). The organic extracts were combined, washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated under vacuum. The residue was subjected to column chromatography on silica gel eluted with 40 to 100% ethyl acetate- hexane gradient. Collection and concentration of appropriate fractions provided the titled compound. lH NMR (400 MHz, CDCI3) δ 5.07 (m, IH), 4.29 (q, J = 7.2 Hz, 2H), 4.00 (s, 3H), 3.99-3.93 (m, IH), 3.89 (s, 3H), 3.74-3.66 (m, IH), 3.53-3.48 (m, IH), 3.23 (dd, J = 1.3, 13.2 Hz, IH), 1.46 (d, J = 6.6 Hz, 3H), 1.36 (t, J = 7.2 Hz, 3H), 1.22 (t, 7= 7.1 Hz, 3H). ES MS M+l = 367

Step 9: (6S)-8-Ethyl-10-methoxy-6-methyl-l,9-dioxo-l,2,6,7,8,9- hexahydropyrazino[r,2′:l,5]pyrrolo[2,3-d]pyridazine-4-carbohydrazide A mixture of ethyl (4S)-2-ethyl-8-(methoxy)-6-[methoxy(oxo)acetyl]-4-methyl-l-oxo- l,2,3,4-tetrahydropyrrolo[l,2-α]pyrazine-7-carboxylate (3.3 g, 8.9 mmol) and anhydrous hydrazine (1.7 mL, 53.7 mmol) in methanol (400 mL) was stirred at room temperature for one hour. The reaction mixture was concentrated under vacuum. The residue was concentrated from toluene. The resultant gummy solid was treated with methanol (20 mL). Diethyl ether was added to the resultant slurry which was filtered to provide the title compound as white solid. lH NMR (400 MHz, CDCI3) δ 8.99 (br s, 2H), 5.54 (br m, IH), 4.12 (m, IH), 4.10 (s, 3H), 3.81 (m, IH),

3.39 (m, IH), 3.21 (d, 7 = 12.6 Hz, IH), 1.44 (d, 7 = 6.4 Hz, 3H), 1.23 (t, 7 = 7.3 Hz, 3H). ES MS M+l =

335

Step 10: (6S)-8-Ethyl-10-methoxy-N,6-dimethyl-l,9-dioxo-l,2,6,7,8,9- hexahydropyrazino[r,2′:l,5]pyrrolo[2,3-d]pyridazine-4-carboxamide To a solution of (6S)-8-ethyl-10-methoxy-6-methyl-l,9-dioxo-l,2,6,7,8,9- hexahydropyrazino[r,2′:l,5]pyrrolo[2,3-d]pyridazine-4-carbohydrazide (0.39 g, 1.2 mmol) and methylamine (5.9 mL, 11.8 mmol; 2 M in THF) in anhydrous dichloromethane (25 mL) in a water bath at room temperature, a solution of iodine (0.60 g, 2.4 mmol) in dichloromethane was added dropwise.

After the addition was completed, an aqueous solution of sodium sulfite was added and the mixture was stirred vigorously for 10 minutes. The organic phase was separated, diluted with chloroform, and washed with brine. The organic extract was dried over anhydrous sodium sulfate, filtered, and concentrated under vacuum. The residue was triturated with a mixture of ethanol (7 mL) and diethyl ether (25 mL). The white solid precipitated was obtained by filtration and dried from its solution in toluene under vacuum. 1H NMR (400 MHz, CDCI3) δ 11.57 (s, IH), 7.38 (m, IH), 5.95 (br m, IH), 4.17 (s, 3H), 4.03 (dd, 7 =

13.4, 3.8 Hz, 1 H), 3.76 (heptet, 7 = 7.1 Hz, 1 H), 3.50 (heptet, 7 = 7.1 Hz, 1 H), 2.99 (dd, 7 = 12.9, 1.0 Hz, 1 H), 3.03 (d, 7 = 5.0 Hz, 3H), 1.44 (d, 7 = 6.6 Hz, 3H), 1.23 (t, 7 = 7.2 Hz, 3H). ES MS M+l = 334 Step 11: (6S)-2-(3-Chloro-4-fluorobenzyl)-8-ethyl-10-methoxy-N,6-dimethyl-l,9-dioxo- l,2,6,7,8,9-hexahydropyrazino[r,2′: l,5]pyrrolo[2,3-d]pyridazine-4-carboxamide To a cold (0 °C) solution of (6S)-8-ethyl-10-methoxy-N,6-dimethyl-l,9-dioxo- l,2,6,7,8,9-hexahydropyrazino[l’,2′: l,5]pyrrolo[2,3-d]pyridazine-4-carboxamide (1.58 g, 4.73 mmol) in anhydrous DMF (50 mL), a solution of lithium bis(trimethylsilyl)amide (4.97 mL, 4.97 mmol, 1 M in THF) was added. After stirring at the same temperature for 25 minutes, 3-chloro-4-fluorobenzyl bromide (1.27 g, 5.68 mmol) was added. The reaction mixture was stirred at room temperature for 10 minutes and concentrated under vacuum. The residue was partitioned between chloroform and brine. The organic extract was dried over anhydrous sodium sulfate, filtered, and concentrated under vacuum. The residue was subjected to column chromatography on silica gel eluting with a 1-5% methanol in ethyl acetate gradient. Collection and concentration of appropriate fractions provided the title compound. lH NMR (400 MHz, CDCI3) δ 7.46 (dd, 7 = 6.9, 2.2 Hz, IH), 7.32 (m, IH), 7.09 (t, 7 = 7.6 Hz, IH), 7.03

(br signal, IH), 5.92 (m, IH), 5.32 (d, 7 = 14.1 Hz, IH), 5.26 (d, 7= 14.1 Hz, IH), 4.14 (s, 3H), 3.97 (dd, 7 = 13.2, 3.7 Hz, IH), 3.73 (heptet, 7 = 7.2 Hz, 1 H), 3.51 (heptet, 7 = 7.1 Hz, IH), 3.21 (dd, 7= 13.2, 1.7 Hz, IH), 3.03 (d, 7 = 5.0 Hz, 3H), 1.42 (d, 7 = 6.6 Hz, 3H), 1.23 (t, 7 = 7.1 Hz, 3H). ES MS M+l = 476

Step 12:

(6S)-2-(3-Chloro-4-fluorobenzyl)-8-ethyl-10-hydroxy-N,6-dimethyl-l,9-dioxo- l,2,6,7,8,9-hexahydropyrazino[r,2′:l,5]pyrrolo[2,3-d3pyridazine-4-carboxamide

To a solution of (6S)-2-(3-chloro-4-fluorobenzyl)-8-ethyl-10-methoxy-N,6-dimethyl-l,9- dioxo-l,2,6,7,8,9-hexahydropyrazino[r,2′:l,5]pyrrolo[2,3-d]pyridazine-4-carboxamide (1.15 g, 2.41 mmol) in anhydrous dichloromethane (800 mL), a solution of boron tribromide in dichloromethane (3.14 mL, 3.14 mmol; 1 M) was added. After stirring at room temperature for 5 minutes, the reaction mixture was treated with anhydrous methanol, stirred for 30 minutes, and concentrated under vacuum. The procedure was repeated twice. The residue was dissolved in a mixture of methanol and acetonitrile and treated with aqueous sodium hydroxide. The mixture was subjected to purification on preparative reverse phase high pressure column chromatography. Collection and lyophilization of appropriate fractions provided the title compound as white amorphous solid.

MK 2048

lH NMR (400 MHz, CDCI3) δ 7.48 (dd, 7 = 7.0, 2.2 Hz, IH), 7.33 (m, IH), 7.09 (t, 7 = 8.7 Hz, IH), 6.01 (m, IH), 5.33 (d, 7= 14.1 Hz, IH), 5.27 (d, 7 = 14.1 Hz, IH), 3.99 (dd, 7= 12.8, 4.0 Hz, 1 H), 3.71(heptet, 7 = 7.1 Hz, 1 H), 3.49 (heptet, 7 = 7.1 Hz, 1 H), 3.24 (dd, 7 = 13.2, 1.5 Hz, 1 H), 3.03 (d, 7 = 5.1 Hz, 3H), 1.42 (d, 7 = 6.6 Hz, 3H), 1.24 (t, 7 = 7.3 Hz, 3H). ES MS M+l = 462

The amorphous product was dissolved in boiling methanol (1.4 g/200 mL). Upon cooling in an ice-water bath, a precipitate formed which was separated by obtained by filtration to afford a white crystalline solid.

MK 2048sodium salt

The corresponding sodium salt was prepared by treatment of a solution of (6S)-2-(3- chloro-4-fluorobenzyl)-8-ethyl- 10-hydroxy-N,6-dimethyl-l ,9-dioxo- 1 ,2,6,7,8,9- hexahydropyrazino[r,2′:l,5]pyrrolo[2,3-d]pyridazine-4-carboxamide (920 mg, 1.99 mmol) in aqueous acetonitrile with aqueous sodium hydroxide (1.03 equivalent), followed by lyophilization of the resultant solution.

ChemSpider 2D Image | (5S)-1-Ethyl-5-methylpiperazin-2-on | C7H14N2O

(5S)-1-ethyl-5-methylpiperazin-2-one

1,5-Cyclooctadiene-iridium(I) chloride dimer, Chloro(1,5-cyclooctadiene)iridium(I) dimer, Di-μ-chlorobis[(1,2,5,6-η)-1,5-cyclooctadiene]diiridium, Iridium(I) chloride 1,5-cyclooctadiene complex dimer, [Ir(1,5-cod)Cl]2, [Ir(1,5-cod)Cl]₂, [Ir(cod)Cl]2

(S)-1-[(R)-2-Di-(4-methoxy-3,5-dimethylphenyl-phosphino)ferrocenyl]-ethyl-dicyclohexylphosphine

SL-J006-2

(5S)-l-Ethyl-5-methylpiperazin-2-one was alternatively prepared as follows:

Step 1: N^rf-Butoxycarbonyl-N^ethylglycinamide Ethylamine (37 g, 0.82 mol) was condensed into a pressure vessel at 0 °C. N-(tert- butoxycarbonyl)glycine methyl ester (50 mL, 0.34 mol) was added. The vessel was sealed and the mixture was stirred at room temperature overnight. The product mixture was concentrated under vacuum and the residue was passed through a pad of silica gel eluting with ethyl acetate. The solution was concentrated under vacuum to provide the title compound as a clear oil. lH NMR (400 MHz, CDCI3) δ 6.11 (br s, IH), 5.18 (br s, IH), 3.77 (d, 7 = 5.7 Hz, 2H), 3.31 (q, 7 = 7.1

Hz, 2H), 1.15 (t, 7 = 7.1 Hz, 3H).

Step 2: l-Ethyl-5-methylpyrazin-2(lH)-one A cold (0 °C) solution of N^tø^butoxycarbonyl-N^ethylglycinamide (68.0 g, 0.33 mol) in anhydrous dichloromethane (500 mL) was saturated with anhydrous hydrogen chloride gas. After stirring at the same temperature for 1.5 hours, the solution was recharged with more hydrogen chloride gas and stirred for additional 15 minutes. The reaction mixture was concentrated under vacuum. The residue was dissolved in methanol, diluted with toluene, and concentrated under vacuum to afford the intermediate N-ethylglycinamide HCI salt.

This was stored under vacuum overnight and used without further purification. A solution of N-ethylglycinamide HCI salt (44.2 g, 0.32 mol), aqueous sodium hydroxide (640 mL, 1M), water (350 mL), pyruvic aldehyde (20.9 mL, 40% solution in water) was heated in an oil bath at 120 °C for one hour. The reaction mixture was cooled and saturated with solid sodium chloride. The mixture was extracted with chloroform (4×250 mL).

The combined organic extract was dried over anhydrous sodium sulfate, filtered, and passed through a plug of silica gel. The silica gel was rinsed successively with ethyl acetate and then 2% methanol in ethyl acetate. The eluted fractions were combined and concentrated under vacuum. The residual solid was recrystallized from diethyl ether to afford the title compound as pale yellow solid. lH NMR (400 MHz, CDCI3) δ 8.11 (s, IH), 6.92 (s, IH), 3.92 (q, 7 = 7.2 Hz, 2H), 2.28 (s, 3H), 1.37 (t, 7 = 7.2 Hz, 3H).

Step 3: (5 S)- 1 -Ethyl-5-methylpiperazin-2-one

A mixture of chloro-l,5-cyclooctadiene iridium (I) dimer (34 mg, 51 μmol) and (S)-l-[(R)-2-di-(3,5-bis(trifluoromethyl)phenyl)phosphino)ferrocenyl]ethyldicyclohexylphosphine (44 mg, 51 μmol; Solvias AG, SL-J006-2) in a mixture of 1:2 toluene and methanol (100 mL; purged with nitrogen for 15 minutes) was sonicated under an atmosphere of nitrogen for 15 minutes. To the resultant mixture, iodine (0.39 g, 1.52 mmol) and l-ethyl-5-methylpyrazin-2(lH)-one (7.0 g, 50.66 mmol) was added. The resultant mixture was heated in an oil bath at 50 °C under an atmosphere of hydrogen gas at 800 psi for 48 hours. The product mixture was filtered through a pad of Celite. The filtrate was concentrated under vacuum. The residue was treated with chloroform saturated with ammonia gas (100 mL). The resultant suspension was filtered through a pad of Celite, which was the rinsed with chloroform saturated with ammonia gas. The combined filtrate was concentrated under vacuum. The residue was concentrated as a solution in toluene for subsequent reaction. lH NMR (400 MHz, CDCI3) δ 3.58 (d, 7 = 17.2 Hz, IH), 3.53(d, 7 = 17.2 Hz, IH), 3.49-3.35 (m, 2H),

1.19 (d, 7 = 5.9 Hz, 3H), 1.14 (t, 7 = 7.2 Hz, 3H).

……………..

US 7538112

http://www.google.com/patents/US7538112

Step 12: (6S)-2-(3-Chloro-4-fluorobenzyl)-8-ethyl-10-hydroxy-N,6-dimethyl-1,9-dioxo-1,2,6,7,8,9-hexahydropyrazino[1′,2′:1,5]pyrrolo[2,3-d]pyridazine-4-carboxamide

To a solution of (6S)-2-(3-chloro-4-fluorobenzyl)-8-ethyl-10-methoxy-N,6-dimethyl-1,9-dioxo-1,2,6,7,8,9-hexahydropyrazino[1′,2′:1,5]pyrrolo[2,3-d]pyridazine-4-carboxamide (1.15 g, 2.41 mmol) in anhydrous dichloromethane (800 mL), a solution of boron tribromide in dichloromethane (3.14 mL, 3.14 mmol; 1 M) was added. After stirring at room temperature for 5 minutes, the reaction mixture was treated with anhydrous methanol, stirred for 30 minutes, and concentrated under vacuum. The procedure was repeated twice. The residue was dissolved in a mixture of methanol and acetonitrile and treated with aqueous sodium hydroxide. The mixture was subjected to purification on preparative reverse phase high pressure column chromatography. Collection and lyophilization of appropriate fractions provided the title compound as white amorphous solid.

¹H NMR (400 MHz, CDCl₃) δ 7.48 (dd, J=7.0, 2.2 Hz, 1H), 7.33 (m, 1H), 7.09 (t, J=8.7 Hz, 1H), 6.01 (m, 1H), 5.33 (d, J=14.1 Hz, 1H), 5.27 (d, J=14.1 Hz, 1H), 3.99 (dd, J=12.8, 4.0 Hz, 1 H), 3.71 (heptet, J=7.1 Hz, 1 H), 3.49 (heptet, J=7.1 Hz, 1 H), 3.24 (dd, J=13.2, 1.5 Hz, 1 H), 3.03 (d, J=5.1 Hz, 3H), 1.42 (d, J=6.6 Hz, 3H), 1.24 (t, J=7.3 Hz, 3H). ES MS M+1=462

The corresponding sodium salt was prepared by treatment of a solution of (6S)-2-(3-chloro-4-fluorobenzyl)-8-ethyl-10-hydroxy-N,6-dimethyl-1,9-dioxo-1,2,6,7,8,9-hexahydropyrazino[1′,2′:1,5]pyrrolo[2,3-d]pyridazine-4-carboxamide (920 mg, 1.99 mmol) in aqueous acetonitrile with aqueous sodium hydroxide (1.03 equivalent), followed by lyophilization of the resultant solution.

References

Keith Alcorn. Ralvetgravir shows potential for use as PrEP drug AIDSmap.com. 28 April 2009. Accessed 8 Nov 2009.
Mark Mascolini. Merck Offers Unique Perspective on Second-Generation Integrase Inhibitor. 10th International Workshop on Clinical Pharmacology of HIV Therapy, April 15–17, 2009, Amsterdam. Accessed 8 Nov 2009.

WO2011121105A1	1 Apr 2011	6 Oct 2011	Tibotec Pharmaceuticals	Macrocyclic integrase inhibitors
EP1756114A2 *	3 May 2005	28 Feb 2007	Merck and Co., Inc.	Hiv integrase inhibitors
US7517929	3 Dec 2004	14 Apr 2009	Momentive Performance Materials Inc.	Star-branched silicone polymers as anti-mist additives for coating applications

US5693640 *	6 Jun 1995	2 Dec 1997	Merck, Sharp & Dohme, Ltd.	Pyridazino-indole derivatives
US5756501 *	3 Dec 1996	26 May 1998	American Home Products Corporation	Saturated and unsaturated pyridazino 4,5-B! indolizines useful as antidementia agents

SEROTONIN..makes me feel happy

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Mar 302014

Serotonin /ˌ s ɛr ə ˈ t oʊ n ɨ n / or 5-hydroxytryptamine (5-HT) is a monoamine neurotransmitter. Biochemically derived from tryptophan, serotonin is primarily found in the gastrointestinal (GI) tract, platelets, and the central nervous system (CNS) of animals, including humans. It is popularly thought to be a contributor to feelings of well-being and happiness.^[6]

In 1935, Italian Vittorio Erspamer showed an extract from enterochromaffin cells made intestines contract. Some believed it contained adrenaline, but two years later, Erspamer was able to show it was a previously unknown amine, which he named “enteramine”. In 1948, Maurice M. Rapport, Arda Green, and Irvine Page of the Cleveland Clinic discovered a vasoconstrictor substance inblood serum, and since it was a serum agent affecting vascular tone, they named it serotonin.

In 1952, enteramine was shown to be the same substance as serotonin, and as the broad range of physiological roles was elucidated, the abbreviation 5-HT of the proper chemical name 5-hydroxytryptamine became the preferred name in the pharmacological field. Synonyms of serotonin include: 5-hydroxytriptamine, thrombotin, enteramin, substance DS, and 3-(β-Aminoethyl)-5-hydroxyindole.In 1953, Betty Twarog and Page discovered serotonin in the central nervous system.

Approximately 90% of the human body’s total serotonin is located in the enterochromaffin cells in the alimentary canal (gut), where it is used to regulate intestinal movements.^[7]^[8] The remainder is synthesized in serotonergic neurons of the CNS, where it has various functions. These include the regulation of mood, appetite, and sleep. Serotonin also has some cognitive functions, including memory and learning. Modulation of serotonin at synapses is thought to be a major action of several classes of pharmacological antidepressants.

Serotonin secreted from the enterochromaffin cells eventually finds its way out of tissues into the blood. There, it is actively taken up by bloodplatelets, which store it. When the platelets bind to a clot, they release serotonin, where it serves as a vasoconstrictor and helps to regulatehemostasis and blood clotting. Serotonin also is a growth factor for some types of cells, which may give it a role in wound healing.

Serotonin is metabolized mainly to 5-HIAA, chiefly by the liver. Metabolism involves first oxidation by monoamine oxidase to the correspondingaldehyde. This is followed by oxidation by aldehyde dehydrogenase to 5-HIAA, the indole acetic acid derivative. The latter is then excreted by the kidneys. One type of tumor, called carcinoid, sometimes secretes large amounts of serotonin into the blood, which causes various forms of thecarcinoid syndrome of flushing (serotonin itself does not cause flushing. Potential causes of flushing in carcinoid syndrome include bradykinins, prostaglandins, tachykinins, substance P, and/or histamine.), diarrhea, and heart problems. Because of serotonin’s growth-promoting effect on cardiac myocytes,^[9] persons with serotonin-secreting carcinoid may suffer a right heart (tricuspid) valve disease syndrome, caused by proliferation of myocytes onto the valve.

In addition to animals, serotonin is found in fungi and plants.^[10] Serotonin’s presence in insect venoms and plant spines serves to cause pain, which is a side-effect of serotonin injection. Serotonin is produced by pathogenic amoebae, and its effect on the gut causes diarrhea. Its widespread presence in many seeds and fruits may serve to stimulate the digestive tract into expelling the seeds.

Serotonin system, contrasted with thedopamine system

Introduction

Serotonin was first recognised as a powerful vasoconstrictor in blood serum. It was isolated in 1948 by Page and was later found to be associated with the central nervous system.

The chemical name for serotonin is 5-hydoxytryptamine which is often abbreviated to 5-HT.

Serotonin is naturally produced in the Pineal gland which lies deep at the centre of the human brain. The average adult human possesses only 5 to 10 mg of serotonin, 90 % of which is in the intestine and the rest in blood platelets and the brain.

One role of this ‘wonder drug’ is as a neurotransmitter, allowing numerous functions in the human body including the control of appetite, sleep, memory and learning, temperature regulation, mood, behaviour, cardiovascular function, muscle contraction, endocrine regulation and depression. Subsequent to his discovery of Serotonin, Page commented that no physiological substance known possesses such diverse actions in the body as does serotonin.

5-HT is also found in wasp stings and scorpion venom where its function is of an irritant, since intravenous injection of serotonin in humans leads to pain, gasping, coughing, a tingling and prickling sensation, nausea, cramps and other unpleasant symptoms.

Serotonin is manufactured in the human brain using the essential amino acid tryptophan which is found in foods such as bananas, pineapples, plums, turkey and milk.

The enzyme tryptophan hydroxylase adds a hydroxyl group to tryptophan’s benzene ring at position 5, creating 5-hydroxytryptophan. Another enzyme, amino acid decarboxylase, then removes a carboxyl group from 5-hydroxytryptophan, forming 5-hydroxytryptamine which is more commonly known as serotonin.

In animals including humans, serotonin is synthesized from the amino acid L–tryptophan by a short metabolic pathway consisting of two enzymes: tryptophan hydroxylase (TPH) and amino acid decarboxylase (DDC). The TPH-mediated reaction is the rate-limiting step in the pathway. TPH has been shown to exist in two forms: TPH1, found in several tissues, and TPH2, which is a neuron-specific isoform.

Serotonin can be synthesized from tryptophan in the lab using Aspergillus niger and Psilocybe coprophila as catalysts. The first phase to 5-hydroxytryptophan would require letting tryptophan sit in ethanol and water for 7 days, then mixing in enough HCl (or other acid) to bring the pH to 3, and then adding NaOH to make a pH of 13 for 1 hour. Asperigillus niger would be the catalyst for this first phase. The second phase to synthesizing tryptophan itself from the 5-hydroxytryptophan intermediate would require adding ethanol and water, and letting sit for 30 days this time. The next two steps would be the same as the first phase: adding HCl to make the pH = 3, and then adding NaOH to make the pH very basic at 13 for 1 hour. This phase uses the Psilocybe coprophila as the catalyst for the reaction.

Serotonin taken orally does not pass into the serotonergic pathways of the central nervous system, because it does not cross theblood–brain barrier. However, tryptophan and its metabolite 5-hydroxytryptophan (5-HTP), from which serotonin is synthesized, can and do cross the blood–brain barrier. These agents are available as dietary supplements, and may be effective serotonergic agents. One product of serotonin breakdown is 5-hydroxyindoleacetic acid (5-HIAA), which is excreted in the urine. Serotonin and 5-HIAA are sometimes produced in excess amounts by certain tumors or cancers, and levels of these substances may be measured in the urine to test for these tumors.

Serotonin is a neurotransmitter involved in the transmission of nerve impulses. Neurotransmitters are chemical messengers within the brain that allow the communication between nerve cells.

Packets of serotonin (vesicles) are released from the end of the presynaptic cell www.thebrain.mcgill.ca/flash/i/i_01/i_01_m/i_01_m_ana/i_01_m_ana.html into the synaptic cleft. The serotonin molecules can then bind to receptor proteins within the postsynaptic cell, which causes a change in the electrical state of the cell. This change in electrical state can either excite the cell, passing along the chemical message, or inhibit it. Excess serotonin molecules are taken back up by the presynaptic cell and reprocessed.

www.thebrain.mcgill.ca/flash/i/i_01/i_01_m/i_01_m_ana/i_01_m_ana.html

The neurons in the brain that release serotonin are found in small dense collections of neurons called Raphe Nuclei. The Raphe Nuclei are found in the medulla, pons and midbrain which are all located at the top of the spinal cord. Serotonergic neurons have axons which project to many different parts of the brain, therefore serotonin affects many different behaviors.

Low serotonin levels are believed to be the cause of many cases of mild to severe depression which can lead to symptoms such as anxiety, apathy, fear, feelings of worthlessness, insomnia and fatigue. The most concrete evidence for the connection between serotonin and depression is the decreased concentrations of serotonin metabolites in the cerebrospinal fluid and brain tissues of depressed people.

http://www.depression.org/

If depression arises as a result of a serotonin deficiency then pharmaceutical agents that increase the amount of serotonin in the brain should be helpful in treating depressed patients. Anti-depressant medications increase serotonin levels at the synapse by blocking the reuptake of serotonin into the presynaptic cell. Anti-depressants are one of the most highly prescribed medications despite the serious side-effects they can cause.

If depression is mild enough it can sometimes be managed without prescribed medications. The most effective way of raising serotonin levels is with vigorous exercise. Studies have shown that serotonin levels are increased with increased activity and the production of serotonin is increased for some days after the activity. This is the safest way of increasing serotonin levels and many other benefits result from regular exercise.

Serotonin levels can also be controlled through the diet. A diet deficient in omega-3 fatty acids may lower brain levels of serotonin and cause depression. Complex carbohydrates raise the level of tryptophan in the brain resulting in a calming effect. Vitamin C is also required for the conversion of tryptophan into serotonin.

Lysergic acid diethylamide, more commonly known as LSD, is a non-toxic, non-addictive molecule which mimics serotonin in the brain. The body ‘mistakes’ LSD for serotonin and shoots it across the synaptic cleft. LSD has a higher affinity for 5-HT receptors than serotonin, thus the presence of LSD prevents

serotonin from sending neural messages in the brain. Once the LSD molecule is bound to the receptor proteins the message is not carried any further. Instead the impulse is redirected to the older parts of the brain, where the bloodstream then takes it to the sense interpretive centres and the motor areas.

There are many similarities between the molecules of serotonin and LSD which allows this process to occur, the most obvious being their close structural similarities, particularly the indole ring shown highlighted in blue.

Serotonin Lysergic acid diethylamide

Another close similarity between LSD and serotonin is the electron density of the highest occupied molecular orbital. The electron density is lowest in the areas around the indole ring in both molecules. This is indicated by the blue areas in the diagrams.

Serotonin LSD

The dipole moment of the two molecules are very close. Serotonin is 2.98 debye and LSD is 3.04 debye, with the dipole moment going towards the NH₂ group in both molecules. The close similarity in dipole moment is key to the ability of LSD to fit into the same receptors as serotonin.

The combination of all of these chemical similarities allows LSD to imitate serotonin and cause psychedelic hallucinations and visions.

serotonin that is ingested will be broken down by the metabolic enzyme MAO. The second point is that even if large quantities of serotonin were to survive the MAO metabolism, it would not directly increase serotonin levels in the brain. The third part of the answer is that simply increasing serotonin levels does not lead to MDMA-like effects.Serotonin, as a molecule, has two particular characteristics that makes it unlikely to be able to be taken orally and reach the brain with a significant concentration to cause a noticeable effect. A key characteristic for this type of molecule is how the nitrogen dangling off the end (see the structure link above and look for the N that is not in the ring structure) behaves with respect to the number of hydrogens that is bound to it and the electrical charge it has (positive or neutral) depending on its environment. The so-called “terminal nitrogen” (on the end) is a base and will change its state (three hydrogens/positive charge vs. two hydrogens/neutral charge) depending on the pH of the solution it is in (acid/base chemistry). At any given time, in most physiological environments (stomach, blood, inside cells, etc.), most serotonin molecules will be positively charged.Now, to get into the brain, molecules have to pass through a layering of cells that surround the brain’s blood vessels commonly referred to as the “blood-brain barrier”. These cells, as with all the cells in your body, have a membrane made up of fat-like, hydrophobic (water-disliking), “non-polar” chains of molecules stacked up on each other (a bi-layer). While the outside and inside of this membrane bi-layer is hydrophilic (water-liking), charged, and/or “polar”, in order to pass into the cell(s) and/or through the membrane(s), a molecule has to pass through this fat-like region. If the molecule itself is charged or has any groups on it that are water-liking/polar, this is unlikely to happen. Think of trying to push a droplet of water to the bottom of a cup of oil with a fork. It can also be thought of as having to get the chemical to dissolve in the fat-layer to get through it: charged / polar chemicals simply will not “dissolve” and pass through this layer.There are ‘active transporters’ or ‘active carriers’ that act as gateways through the blood-brain barrier for molecules of particular shapes. The normal amino acids (building blocks for the neurotransmitters) such as tryptophan, phenylalanine, tyrosine, etc get through the BBB by having special systems for pushing them through. As a side note, there are fewer of these transporters than there are possible molecules and the same transporters work on many different chemicals, so there is competition for transport into the brain between different chemicals, however this does not really impact the question at hand.

Pietra, S.;Farmaco, Edizione Scientifica 1958, Vol. 13, pp. 75–9.
Mazák, K.; Dóczy, V.; Kökösi, J.; Noszál, B. (2009). “Proton Speciation and Microspeciation of Serotonin and 5-Hydroxytryptophan”. Chemistry & Biodiversity 6 (4): 578–90.doi:10.1002/cbdv.200800087. PMID 19353542.
Erspamer, Vittorio (1952). Ricerca Scientifica 22: 694–702.
Tammisto, Tapani (1968). Annales Medicinae Experimentalis et Biologiea Fenniae 46 (3, Pt. 2): 382–4.
Young SN (2007). “How to increase serotonin in the human brain without drugs”. Rev. Psychiatr. Neurosci. 32(6): 394–99. PMC 2077351. PMID 18043762.
King MW. “Serotonin”. The Medical Biochemistry Page. Indiana University School of Medicine. Retrieved 1 December 2009.
Berger M, Gray JA, Roth BL (2009). “The expanded biology of serotonin”. Annu. Rev. Med. 60: 355–66.doi:10.1146/annurev.med.60.042307.110802.PMID 19630576.
Bianchi, P. (2005). “A new hypertrophic mechanism of serotonin in cardiac myocytes: Receptor-independent ROS generation”. The FASEB Journal. doi:10.1096/fj.04-2518fje.
Kang K, Park S, Kim YS, Lee S, Back K (2009). “Biosynthesis and biotechnological production of serotonin derivatives”. Appl. Microbiol. Biotechnol. 83 (1): 27–34.doi:10.1007/s00253-009-1956-1. PMID 19308403.

www.encyclopedia.com/html/s1/serotoni.asp

www.angelfire.com/hi/TheSeer/seratonin.html

www.findthelight.net/Depression/the_chemistry_of_dep.htm

http://www.cmste.uncc.edu/Document%20Hold/Sawsun-%20Serotonin%20FINAL%20PAPER.doc

www.totse.com/en/technology/science_technology/seroton.html

www.macalester.edu/~psych/whathap/ubnrp/mdma/serotonin.html

www.serendipity.li/mcclay/pineal.html#a1.6

www.serendip.brynmawr.edu/bb/neuro/neuro98/202s98-paper3/Frederickson3.html

www.serendip.brynmawr.edu/bb/neuro/neuro99/web1/Byrd.html

BERAPROST….Stable prostacyclin analog.

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Mar 242014

BERAPROST

https://www.ama-assn.org/resources/doc/usan/beraprost.pdf

2,3,3a,8b-tetrahydro-2-hydroxy-1-(3-hydroxy-4-methyl-1-octen-6-ynyl)-1H-cyclopenta(b)benzofuran-5-butanoic acid

(±)-(IR*,2R*,3aS*,8bS*)-2,3,3a,8b-tetrahydro-2-hydroxy-1-[(E)-(3S*)-3-hydroxy-4-methyl-1-octene-6-inyl]-1H-cyclopenta[b]benzofuran-5-butyric acid

rac-4-{(1R,2R,3aS,8bS)-2-hydroxy-1-[(1E,3S,4RS)-3-hydroxy-4-methyloct-1-en-6-ynyl]-2,3,3a,8b-tetrahydro-1H-cyclopenta[b][1]benzofuran-5-yl}butanoic acid

88430-50-6 88475-69-8

Beraprost
Beraprostum
Beraprostum [INN-Latin]
MDL 201229
MDL-201229
ML 1229
ML-1229
UNII-35E3NJJ4O6

Beraprostum, Beraprostum [INN-Latin], ML 1229, MDL 201229, 88430-50-6

Molecular Formula: C₂₄H₃₀O₅

Molecular Weight: 398.492

CLINICAL TRIALS..http://clinicaltrials.gov/search/intervention=Beraprost

Beraprost is a synthetic analogue of prostacyclin, under clinical trials for the treatment of pulmonary hypertension. It is also being studied for use in avoiding reperfusion injury.

As an analogue of prostacyclin PGI₂, beraprost effects vasodilation, which in turn lowers the blood pressure. Beraprost also inhibits plateletaggregation, though the role this phenomenon may play in relation to pulmonary hypertension has yet to be determined.

Beraprost …sodium salt

ML 1129; Procyclin; TRK 100 (CAS 88475-69-8)

Beraprost is an analog of prostacyclin in which the unstable enol-ether has been replaced by a benzofuran ether function. This modification increases the plasma half-life from 30 seconds to several hours, and permits the compound to be taken orally. Doses of 20-100 µg in humans, given 1 to 3 times per day, have been demonstrated to improve clinical end points in diseases responsive to prostacyclin. Oral beraprost therapy improved the survival and pulmonary hemodynamics of patients with primary pulmonary hypertension.1 Beraprost inhibits platelet aggregation in healthy subjects and in diabetic patients at similar doses.2,3

Synonyms	ML 1129 Procyclin TRK 100
Formal Name	2,3,3a,8b-tetrahydro-2-hydroxy-1-(3-hydroxy-4-methyl-1-octen-6-ynyl)-1H-cyclopenta[b]benzofuran-5-butanoic acid, monosodium salt
CAS Number	88475-69-8
Molecular Formula	C24H29O5 · Na
Formula Weight	420.5

Beraprost sodium is a prostacyclin analog and an NOS3 expression enhancer that was first launched in 1992 in Japan pursuant to a collaboration between Astellas Pharma and Toray for the oral treatment of peripheral vascular disease (PVD), including Raynaud’s syndrome and Buerger’s disease. In 2000, the drug was commercialized for the treatment of pulmonary hypertension. Development for the oral treatment of intermittent claudication associated with arteriosclerosis obliterans (ASO) was discontinued at Kaken and United Therapeutics after the product failed to demonstrate statistically significant results in a phase III efficacy trial.

In terms of clinical development, beraprost sodium is currently in phase II clinical trials at Kaken for the treatment of lumbar spinal canal stenosis and at Astellas Pharma for the oral treatment of primary chronic renal failure. The company is also conducting phase III trials for the treatment of nephrosclerosis. The drug has also been studied through phase II clinical trials at Kaken for the oral treatment of diabetic neuropathy, but recent progress reports for this indication have not been made available.

Beraprost is an oral form of prostacyclin, a member of the family of lipid molecules known as eicosanoids. Prostacyclin is produced in the endothelial cells from prostaglandin H2 by the action of the enzyme prostacyclin synthase. It has been shown to keep blood vessels dilated and free of platelet aggregation.

Beraprost sodium was originally developed at Toray in Japan, and rights to the drug were subsequently acquired by Astellas Pharma. A 1972 alliance between Toray and Kaken Pharmaceutical to develop and commercialize prostaglandin led to a later collaboration agreement for the development of beraprost. In 1990, Toray granted the right to market the drug to Sanofi (formerly known as sanofi-aventis), a licensing agreement that was later expanded to include Canada, the U.S., South America, Africa, Southeast Asia, South Asia, Korea and China. In September 1996, Bristol-Myers Squibb entered into separate agreements with Sanofi and Toray to acquire all development and marketing rights to beraprost in the U.S. and Canada. In January 1999, United Therapeutics and Toray agreed to cooperatively test the drug in North America, and in July 2000, a new agreement was signed pursuant to which United Therapeutics gained exclusive North American rights to develop and commercialize sustained-release formulations of beraprost for all vascular and cardiovascular diseases. In 1999, orphan drug designation was received in the U.S. for the treatment of pulmonary arterial hypertension associated with any New York Heart Association classification (Class I, II, III, or IV). In 2011, orphan drug designation was assigned in the U.S. for the treatment of pulmonary arterial hypertension.

The compound name of beraprost which is used as an antimetastasis agent of malignant tumors according to the present invention is (±)-(IR*,2R*,3aS*,8bS*)-2,3,3a,8b-tetrahydro-2-hydroxy-1-[(E)-(3S*)-3-hydroxy-4-methyl-1-octene-6-inyl]-1H-cyclopenta[b]benzofuran-5-butyric acid. This compound has the following structure.

Beraprost is described in Japanese Laid-open Patent Application (Kokai) Nos. 58-32277, 57-144276 and 58-124778 and the like as a PGI₂ derivative having a structure in which the exoenol moiety characteristic to beraprost is converted to inter-m-phenylene structure. However, it is not known that beraprost has an activity to inhibit metastasis of malignant tumors.
The beraprost which is an effective ingredient of the agent of the present invention includes not only racemic body, but also d-body and l-body. Beraprost can be produced by, for example, the method described in the above-mentioned Japanese Laid-open Patent Application (Kokai) No. 58-124778. The salts of beraprost include any pharmaceutically acceptable salts including alkaline metal salts such as sodium salt and potassium salt; alkaline earth metal salts such as magnesium salt and calcium salt; ammonium salt; primary, secondary and tertiary amine salts; and basic amino acid salts.

EP0623346A1

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US7005527

EXAMPLE 6 Beraprost of the Formula (I)

0.246 g (0.6 mmol) of compound of the general formula (II) obtained in Example 5 is dissolved in 1 ml of methanol and 1 ml of 1 M aqueous sodium hydroxide solution is added dropwise slowly thereto. After stirring for an hour the methanol is distilled off from the reaction mixture in vacuum. The aqueous residue is diluted with 10 ml of water extracted with methyl-tert.butyl-ether and the combined organic phase is washed with saturated NaCl solution, dried on Na₂SO₄and evaporated. The residue of evaporation is crystallized from ethylacetate-hexane mixture and the pure above mentioned title compound is obtained as colourless crystals.

Yield: 0.21 g (87%)

TLC-R_f(toluene-dioxan-acetic acid 20:10:1)=0.41

Melting point: 98–112° C.

¹H NMR (400 MHz, CDCl₃), δH (ppm): 1.00d, 1.03d [3H; J=6.8 Hz; 21-H₃]; 1.79m [1H; 16-H]; 1.80t, 1.81t [3H, J=2.5,2.4 Hz; 20-H₃]; 2.3–1.9m [5H, 3-H₂, 10H_b, 17-H₂]; 2.34t [1H; J=7.4 Hz; 2-H₂]; 2.43m [1H; 12-H]; 2.64m [3H; 10-H_a, 4-H₂]; 3.43t, 3.44t [1H, J=8.7,8.5 Hz; 8-H]; 3.92m [1H; 11-H]; 4.07t, 4.17t [1H, J=7.3,5.6 Hz; 15-H]; 4.3b [2H; OH]; 5.09m [1H, 9-H]; 5.58dd, 5.61dd [1H; J=15.3,6.5 Hz; 14-H]; 5.67dd, 5.68dd [1H; J=15.3,8.0 Hz; 13-H]; 6.77m [1H; 2′-H]; 6.95m [2H; 1′-H,3′-H]¹³C NMR (100 MHz, CDCl₃), δC (ppm): 3.5, 3.6 [C-20]; 14.7, 15.8 [C-21]; 22.3, 22.6 [C-17]; 24.6 [C-2]; 29.1 [C-4]; 33.1 [C-3]; 38.2, 38.3 [C-16]; 41.2 [C-10]; 50.4 [C-8]; 58.8 [C-12]; 75.8, 76.3, 76.4 [C-11, C-15]; 77.2, 77.4 [C-18, C-19]; 84.5, 84.6 [C-9]; 120.6 [C-2′]; 121.9 [C-3′]; 123.2 [C-5]; 129.0 [C-1′]; 129.7 [C-7]; 132.3, 133.0, 133.8, 134.0 [C-13, C-14]; 157.2 [C-6]; 178.3 [C-1].

EXAMPLE 7 Beraprost Sodium Salt (The Sodium Salt of the Compound of Formula (I)

0.199 g of beraprost is dissolved in 2 ml of methanol, 0.5 ml of 1 M aqueous solution of sodium hydroxide is added thereto and after their mixing the solvent is evaporated in vacuum and thus the above title salt is obtained as colourless crystals.

Yield: 0.21 g (100%)

Melting point: >205° C.

¹H NMR (400 MHz, DMSO-d₆), δH (ppm): 0.90d, 0.92d [3H; J=6.7 Hz; 21-H₃]; 1.75–1.55m [7H; 10H_b, 16-H, 3-H₂, 20-H₃]; 1.89t [2H, J=7.6 Hz; 2-H₂]; 1.94m [1H; 17-H_b]; 2.16q [1H, J=8.5 Hz; 12-H]; 2.25m [1H; 17-H_a]; 2.44t [2H; J=7.5 Hz; 4-H₂]; 2.50o [1H; 10-H_a]; 3.39t [1H, J=8.5 Hz; 8-H]; 3.72td [1H; J=8.5,6.1 Hz; 11-H]; 3.84t 3.96t [1H, J=6.5,6.0 Hz; 15-H]; 4.85b [2H, OH]; 5.01dt [1H, J=8.5,6.6 Hz; 9-H]; 5.46dd, 5.47dd [1H; J=15.4,6.5 Hz, J=15.4,6.0 Hz; 14-H]; 5.65dd, 5.66dd [1H; J=15.4,8.5 Hz; 13-H]; 6.71m [1H; 2′-H]; 6.92m [2H; 1′-H, 3′-H] During the above thin layer chromatography (TLC) procedures we used plates MERCK Kieselgel 60 F₂₅₄, thickness of layer is 0.2 mm, length of plates is 5 cm.

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Reaction Scheme A.
The starting material of bromocarboxylic acid, Compound 1, and the process for the preparation thereof are disclosed in Japanese Patent Application No. 29637/81.

Scheme B.

REACTION SCHEME B

https://www.google.com/patents/EP0084856A1

Org Lett 2012, 14(1): 299

http://pubs.acs.org/doi/abs/10.1021/ol203060v

http://pubs.acs.org/doi/suppl/10.1021/ol203060v/suppl_file/ol203060v_si_001.pdf

EP0024943A1	Sep 2, 1980	Mar 11, 1981	Toray Industries, Inc.	5,6,7-Trinor-4,8-inter-m-phenylene PGI2 derivatives and pharmaceutical compositions containing them
EP0084856A1	Jan 19, 1983	Aug 3, 1983	Toray Industries, Inc.	5,6,7-Trinor-4, 8-inter-m-phenylene prostaglandin I2 derivatives
JP3069909B				Title not available