WO2010110771A2

WO2010110771A2 - Substituted thiol-containing alkyl fatty acids and process for synthesizing derivatives thereof

Info

Publication number: WO2010110771A2
Application number: PCT/US2009/001857
Authority: WO
Inventors: Frank Gibson; Lakmal W. Boteju; Robert Shorr; Robert Rodriguez
Original assignee: Frank Gibson; Boteju Lakmal W; Robert Shorr; Robert Rodriguez
Priority date: 2009-03-25
Filing date: 2009-03-25
Publication date: 2010-09-30
Also published as: WO2010110771A3

Abstract

An improved process for the synthesis of derivatives of substituted thiol-containing alkyl fatty acids, such as but not limited to lipoic acid, generates intermediate compounds and occurs in a temperature-controlled optimal sequence of monitored events to generate increased yield and purity of final product.

Description

SUBSTITUTED THIOL-CONT AINING ALKYL FATTY ACIDS AND PROCESS FOR SYNTHESIZING DERIVATIVES THEREOF

Field of the Invention This invention relates to the synthesis of pharmaceutical compositions, and more particularly to the improved synthesis, yield, and purity on a multigram scale of derivatives of substituted thiol-containing alkyl fatty acids, such as but not limited to lipoic acid, in a reproducible temperature-controlled optimal sequence of process steps under monitored conditions, and novel structures synthesized thereby.

Background of the Invention

Both thiol-containing alkyl fatty acids such as octanoic acid and substituted thiol- containing alkyl fatty acids such as lipoic acid play an important role in cellular function and disease. Enzymes interacting with such molecules, as well as receptors for such molecules, may represent novel drug targets for the treatment of a variety of diseases such as cancer, inflammation, diabetes, and other metabolic disorders, among others. Hence, for example, US Patents 6,331,559 and 6,951,887 to Bingham et al, as well as US Patent Application No. 12/105,096 by Bingham et al, all herein incorporated by reference, disclose a novel class of lipoic acid derivatives useful for the treatment of cancer. The derivatives disclosed in Bingham et al have been manufactured according to a two-step process. In the first step, NaHCO₃ is added to an aqueous solution of D,L-α-lipoic acid and stirred at ambient temperature for five minutes. This suspension is flushed with argon gas and cooled to 5⁰C. Subsequently, NaBH₄ is added in small portions at a temperature between 5-10⁰C. This solution is next stirred for a further thirty minutes between 5-10⁰C, then for an additional thirty minutes at 20-25⁰C. The solution is then both cooled to under 10°C and the pH adjusted to ~1 using 2N HCl, which vigorously generates hydrogen gas. The resultant oily mixture is then extracted using methylene chloride under constant presence of argon, with these extracts eluted through anhydrous Na₂SO₄. The filtrate is washed with additional methylene chloride with solvent removed by evaporation using a 35°C bath. The resulting isolated oil containing 6,8-disulfanyloctanoic acid is further dried under these conditions for three hours.

In the second step, benzyl bromide is added to the 6,8-disulfanyl octanoic acid, this mixture then dissolved in absolute ethanol and placed under positive pressure of nitrogen. Separately, a solution of sodium ethoxide in ethanol was prepared by reacting sodium metal with absolute ethanol. This solution was added to the octanoic acid solution drop- wise with stirring over thirty minutes, with continued stirring at ambient temperature for thirty minutes and then at 35-40°C for an additional two hours. Upon precipitation, the mixture is cooled and the ethanol removed using a rotary evaporator with temperature maintained at 60°C. The pH is adjusted to ~1 with 2N HCl, and methylene chloride is added. Upon stirring between five to ten minutes, the mixture separates into two layers, with the methylene chloride layer collected. The aqueous layer is extracted again with methylene chloride and again stirred between five to ten minutes, with a second resulting methylene chloride layer collected and combined with the first extracted layer. This combined methylene chloride solution is extracted with saturated NaCl solution, then separated and eluted over a bed of anhydrous Na₂SO₄. The Na₂SO₄ is washed with additional methylene chloride, and this methylene chloride is removed by evaporation from a 35⁰C water bath. The product is dissolved in methylene chloride and subsequently filtered, with the methylene chloride removed by evaporation from a 35⁰C water bath. Upon transfer, the crude product is mixed with ethyl acetate, then dissolved and warmed to 40⁰C. This solution is treated with heptane added over fifteen minutes, then cooled to 23⁰C and subsequently to 0°C. The solvent is removed and the crude product re-dissolved in methylene chloride while maintained under argon atmosphere. IN HCl is stirred into the methylene chloride solution for thirty minutes, adjusting the pH to ~1. The layers are separated, with the methylene chloride layer extracted with water. These layers are also separated, and the methylene chloride layer is concentrated by evaporation from a 35⁰C water bath. The crude material is re-dissolved in ethyl acetate and filtered. The ethyl acetate is removed by evaporation from a 35° C water bath. The product is re-dissolved in ethyl acetate at room temperature, and heptane is added over eighteen minutes, with crystallization starting after six minutes. The product is cooled to 0°C for sixteen hours, and the resulting 6,8-bis-benzylsulfanyl octanoic acid product is collected by filtration and then rinsed with a solution of 25 % ethyl acetate in heptane. The final crystalline product is placed in a vacuum oven set at 25⁰C for sixteen hours.

There are numerous problems and issues of practicality with the aforementioned process. First, it is evident from the aforementioned process that there is a multitude of extractions and isolations which must occur in each step. This involves a great deal of labor and cost and necessitates the process taking place over an extended period of time. Also, there is similarly a wide range of temperature over which the reactions of the process occur, from 0-40⁰C depending on the step performed. Additionally, issues of safety exist with both the generation of hydrogen gas in the first step of the process and the use of metallic sodium to produce the sodium ethoxide required in the second step. Indeed, the use of sodium ethoxide can lead to potential impurities in the final product. Furthermore, the addition of sodium ethoxide after the addition of benzyl bromide in the second step of the process may be expected to decrease final product yield: sodium ethoxide can react directly with benzyl bromide, leaving less of the latter to react with the 6,8-disulfanyl octanoic acid produced during after the first step. Finally, high-performance liquid chromatography (HPLC) monitoring for purity did not occur during the process.

The ability to produce such derivatives as previously disclosed by Bingham et ah, as well as other substituted thiol-containing alkyl fatty acids and derivatives, in sufficient quantities to provide of active pharmaceutical ingredient (API) material for clinical investigation and to meet regulatory and pharmacopeia guidelines is essential for both commercial development and sufficient drug supply to meet worldwide needs. Furthermore, it is essential that the purity of such API material be reproducible and cost-effective.

Given the above, then, it has been demonstrated that there is a need for the synthesis of derivatives of substituted thiol-containing alkyl fatty acids, such as but not limited to lipoic acid, to occur according to a shorter, safer, better-monitored, more cost-effective process using reagents in an optimal sequence of addition under temperature-controlled conditions to produce an increased yield of API material on a multigram scale.

Objects of the Invention and Industrial Applicability

Consequently, it is an object of the present invention to provide a process for the synthesis of derivatives of substituted thiol-containing alkyl fatty acids, such as but not limited to lipoic acid, which demonstrates higher safety during the process, is of shorter duration, and is of lower cost. It is a further object of the present invention to provide a process for the synthesis of derivatives of substituted thiol-containing alkyl fatty acids, such as but not limited to lipoic acid, which uses reagents in an reproducible optimal sequence of addition to produce an increased yield of desired API material on a multigram scale. It is a still further object of the present invention to provide a process for the synthesis of derivatives of substituted thiol-containing alkyl fatty acids, such as but not limited to lipoic acid, which permits monitoring of reagents and products by HPLC during the process.

It is a still further object of the present invention to provide a process for the synthesis of derivatives of substituted thiol-containing alkyl fatty acids, such as but not limited to lipoic acid, which runs under temperature-controlled conditions.

Summary of the Invention

In accordance with the aforementioned aims, the present invention broadly provides an improved process for the synthesis of derivatives of substituted thiol-containing alkyl fatty acids, such as but not limited to lipoic acid, including but not limited to those described in US

Patents 6,331,559 and 6,951,887 to Bingham et ah, as well as US Patent Application No.

12/105,096 by Bingham et al., by a novel temperature-controlled reproducible optimal sequence of process steps under constant HPLC monitoring. This sequence results in an improved synthesis, yield, and purity of such derivatives as may be desired on a multigram scale for API material, all leading to substantial improvement over the synthesis process previously described.

The novel combination of process steps broadly comprises:

Mixing a thiol-containing alkyl fatty acid with a deprotonating reagent at an initial temperature;

Adding a hydride-donating reagent to the reaction mixture at a second temperature, Said second temperature maintained throughout the rest of the process; Adding a second deprotonating reagent to the reaction mixture; and Adding an alkyl- or acyl-donating reagent to the reaction mixture to form desired final product. In a preferred embodiment, the novel combination of process steps further comprises:

Crystallization of the final product through use of an organic solvent system at a temperature cooler than that of the second temperature.

In a preferred embodiment of the present invention, the initial temperature is between 20-25° C; the deprotonating reagent is NaOH; the second temperature is between 40-45° C; the hydride-donating reagent is NaBH₄; the alkyl- or acyl-donating reagent is benzyl bromide; and the organic solvent system is ethyl acetate and heptane.

In a further preferred embodiment of the present invention, the thiol-containing alkyl fatty acid is octanoic acid or analogs, salts, congeners, or derivatives thereof. In a still further preferred embodiment of the present invention, the thiol-containing alkyl fatty acid is lipoic acid.

The present invention also discloses novel compounds produced as key intermediates of the final product using the process described herein.

Brief Description of the Drawings

FIGURE 1 depicts a chromatogram showing the generation of a key intermediate of the final product produced by the novel combination of process steps of the present invention.

Detailed Description of the Invention In accordance with the aforementioned aims, the present invention broadly provides an improved process for the synthesis of derivatives of substituted thiol-containing alkyl fatty acids, such as but not limited to lipoic acid, including but not limited to those described in US Patents 6,331,559 and 6,951,887 to Bingham et al., as well as US Patent Application No. 12/105,096 by Bingham et al., from reagents by a novel temperature-controlled reproducible optimal sequence of process steps, concomitantly generating increased yield and purity of API material product on a multigram scale. Additionally, there is less evolution of volatile hydrogen gas, thereby increasing the safety of the process. The novel sequence of process steps broadly comprises:

Adding a hydride-donating reagent to the reaction mixture at a second temperature,

Said second temperature maintained throughout the rest of the process;

Adding a second deprotonating reagent to the reaction mixture; and

Adding an alkyl- or acyl-donating reagent to the reaction mixture to form desired final product.

In a preferred embodiment, the novel combination of process steps further comprises: Crystallization of the final product through use of an organic solvent system at a temperature cooler than that of the second temperature.

In a preferred embodiment, the synthesis process to generate the lipoic acid derivatives disclosed in Bingham et al. is conducted in two steps, according to the following reaction schemes:

Step 1 :

1. lequiv NaBH₄, 1 equiv 1 M NaOH

40 C, 30 minutes

"100%" conversion lipoic acid dithiol intermediate by HPLC not isolated CPI-611 pure by HPLC CPI-612

Step 2: 1.2 equivNa0H

rt, 20 minutes

CPI-612

CPI-613

In Step 1, one equivalent of α-lipoic acid is dissolved by stirring same into one equivalent of IM NaOH (5 mL of IM NaOH/g α-lipoic acid) at between 20-25°C. After dissolution, 1 equivalent of NaBH₄ (s) is added to the solution to reduce the disulfide to the corresponding dithiol, with the reaction mixture raised to between 40-45 °C, remaining at this temperature for the remainder of the process. The reaction is monitored by HPLC for the conversion of α-lipoic acid to dihydrolipoic acid (DHLA). The conversion to DHLA is essentially 100%, and there is no need for isolation of any intermediate reagents or products, permitting direct turnover of DHLA into Step 2 in an expedient manner. In Step 2, 2 equivalents of 2M NaOH is added to the DHLA formed at the end of Step

1 (5 mL of 2M NaOH/g DHLA) to deprotonate the DHLA, followed thereafter by 2 equivalents of benzyl bromide to benzylate both of the thiols of the DHLA. By adding the NaOH prior to adding the benzyl bromide, reaction between the NaOH and the benzyl bromide is minimized, allowing essentially 100% benzylation of the DHLA. Furthermore, through the use of NaOH as the deprotonating agent, and not sodium ethoxide, since metallic sodium is not used in this step to produce sodium ethoxide, the process avoids the hazards of handling metallic sodium. Precipitation of the desired product occurs almost immediately after addition of benzyl bromide. The reaction is easily monitored by HPLC and conversion of DHLA to the desired dibenzylated product is typically complete within thirty minutes. At the conclusion of the synthesis process, the reaction mixture is diluted with 10 mL of organic solvent/g lipoic acid derivative. In a preferred embodiment, this solvent is ethyl acetate, although methyl tert-butyl ether (MTBE) may also be used. The reaction mixture is then made acidic with the addition of concentrated HCl until the pH of the aqueous layer is < 2, with the lipoic acid derivative redissolving into the organic solvent layer. The resulting phases are split, and the remaining top organic solvent layer is reserved. The aqueous layer is extracted with 5 mL of organic solvent/g lipoic acid derivative, and the two extracted layers are combined. The organic solvent layer is then washed with one portion of 0.1 M HCl (5 mL 0.1 M HCl/g lipoic acid derivative), with the resulting lower aqueous layer discarded. At this point, the final product in solution should be > 95% pure by HPLC analysis. The product can then be crystallized from the organic layer by the addition of heptanes.

The isolated thiol-containing alkyl fatty acid product can be recrystallized from solution as necessary from methanol, ethanol, isopropanol, acetone, acetonitrile using water as an antisolvent, or ethyl acetate and heptanes at reduced temperatures. (See, e.g., Wright AE (1998). Isolation of marine natural products. In RJP Cannell (Ed.), Methods in Biotechnology: Natural Products Isolation (pp. 365-408). Totowa, NJ: Humana Press, herein incorporated by reference.) The present invention also discloses novel compounds produced during the process described herein. As seen in the chromatogram illustrated in FIGURE 1, during Step 2, 6- benzylsulfanyl octanoic acid or 8-benzylsulfanyl octanoic acid, seen at 8.966 minutes on the chromatogram, is necessarily produced as a key monobenzylated intermediate upon adding benzyl bromide to the reaction mixture, starting at 8.056 minutes. While the process has been initially illustrated here through synthesis of those lipoic acid derivatives disclosed by Bingham et al. , it must be stressed that the synthesis process is completely applicable to any derivative of a thiol-containing alkyl fatty acid where both sulfides are equally substituted by alkyl or acyl groups. "Alkyl" as used herein includes both saturated and unsaturated branched- or straight-chain aliphatic hydrocarbon groups having at least three to eighteen carbon atoms. In a preferred embodiment of the present invention, the alkyl fatty acid is octanoic acid, or analogs, salts, congeners, or derivatives thereof, and in a more preferred embodiment, the alkyl fatty acid is lipoic acid.

Additionally, different reagents may be used throughout the process without detriment to the formation of the desired final product. Thus, for example, benzyl chloride may be used in place of benzyl bromide as the the alkyl- or acyl-donating reagent; other reagents will be well known to those skilled in the art of the present invention. Also, by way of example, common bases other than NaOH, such as but not limited to KOH or NaHCO₃, may be used as the deprotonating agent. Finally, other hydride-donating reagents such as but not limited to LiAlH₄, NaAlH₂(OCH₂CH₂OCH₃)₂, or NaAlH₂(C₂Hs)₂ may be used without detriment to the formation of the desired final product.

Furthermore, combinations of substituents and/or variables are permissible only if such combinations result in stable compounds or structures. "Stable compound" or "stable structure" as used herein consequently means a compound or structure that is sufficiently robust to survive from a reaction mixture to a useful degree of purity. It is thus contemplated that such stable compounds or structures may include monosubstituted derivatives of substituted alkyl fatty acids as well as disubstituted derivatives thereof.

The substituted thiol-containing alkyl fatty acid derivatives to be synthesized by the process of the present invention described herein may have asymmetric centers. All chiral, diastereomeric, and racemic forms are included in the present invention. It will be appreciated that certain compounds to be synthesized according to the present invention contain an asymmetrically substituted carbon atom, and may be isolated in optically active or racemic forms. It is well known in the art how to prepare optically active forms, such as by resolution of racemic forms or by synthesis, from optically-active starting materials. All chiral, diastereomeric, racemic forms and all geometric isomeric forms of a structure are intended, unless the specific stereochemistry or isomer form is specifically indicated.

The two-step synthesis process is expected to take no longer to complete than a single day in a manufacturing facility, with the crystallization process taking another day. The present invention is contemplated to be practiced on at least a multigram scale, kilogram scale, multikilogram scale, or industrial scale. "Multigram scale," as used herein, is preferably the scale wherein at least one starting material is present in ten grams or more, more preferably at least fifty grams or more, even more preferably at least one hundred grams or more. "Multikilogram scale," as used herein, is intended to mean the scale wherein more than one kilogram of at least one starting material is used. "Industrial scale," as used herein is intended to mean a scale which is other than a laboratory scale and which is sufficient to supply product sufficient for either clinical tests or distribution to consumers.

The following non-limiting examples are provided both to facilitate understanding of the process of the present invention and to exemplify the invention and are not to be construed as limiting the invention's scope.

EXAMPLE 1 Preparation of 6.8-to(benzylthio)octanoie acid dimethyl analog

Lipoic acid (I g, 0.000485 mol) was dissolved with stirring in 5 mL (1 equiv) of IM NaOH. Solid NaBH₄ (0.183 g, 1 equiv) was added in one portion, and the resulting reaction mixture was heated to 40° C. After thirty minutes, the reaction solution had become completely colorless. 5 mL of 2M NaOH was then added, followed by 2 equivalents of methyl iodide (0.604 mL). The reaction solution was stirred at room temperature for 45 minutes. The reaction mixture was diluted with 10 mL of MTBE and acidified to pH < 2 with concentrated HCl. The top organic layer was separated and concentrated on a rotary evaporator (rotovap). 6,8-dimethylthio-octanoic acid was isolated as a mobile oil, 1.1 g, 96%.

EXAMPLE 2 Preparation of 6,8-6/s(benzylthio)octanoic acid di-n-propyl analog

Lipoic acid (1 g, 0.000485 mol) was dissolved with stirring in 5 mL (1 equiv) of IM NaOH. Solid NaBH₄ (0.183 g, 1 equiv) was added in one portion, and the resulting reaction mixture was heated to 40⁰C. After thirty minutes, the reaction solution had become completely colorless. 5 mL of 2M NaOH was then added, followed by 2 equivalents of n- propyl bromide (0.88 mL). The reaction solution was stirred at room temperature for 45 minutes. The reaction mixture was diluted with 10 mL of MTBE and acidified to pH < 2 with concentrated HCl. The top organic layer was separated and concentrated on a rotovap. 6,8-di-n-propylthio-octanoic acid was isolated as a mobile oil, 1.3 g, 92%.

EXAMPLE 3

Preparation of 6.8-6fo(benzylthio)octanoic acid di-isopropyl analog

Lipoic acid (1 g, 0.000485 mol) was dissolved with stirring in 5 mL (1 equiv) of IM NaOH. Solid NaBH₄ (0.183 g, 1 equiv) was added in one portion, and the resulting reaction mixture was heated to 40⁰C. After thirty minutes, the reaction solution had become completely colorless. 5 mL of 2M NaOH was then added, followed by 2 equivalents of isopropyl iodide (0.96 mL). The reaction solution was heated to 50⁰C then stirred at room temperature for four hours. The reaction mixture was diluted with 10 mL of MTBE and acidified to pH < 2 with concentrated HCl. The top organic layer was separated and concentrated on a rotovap. 6,8-di-isopropylthio-octanoic acid was isolated as a mobile oil, 1.3 g, 92%. The foregoing discussion discloses and describes merely specific exemplary embodiments of the present invention. One skilled in the art will readily recognize from such discussion, and from the accompanying claims, that various changes, modifications, and variations can be made therein without departing from the spirit and scope of the inventive concept as defined in the following claims. Furthermore, while exemplary embodiments have been expressed herein, others practiced in the art may be aware of other designs or uses of the present invention. Also, all patent applications, patents, and other publications cited herein are incorporated by reference in their entirety. Thus, while the present invention has been described in connection with exemplary embodiments thereof, it will be understood that many modifications in both design and use will be apparent to those of ordinary skill in the art, and this application is intended to cover any changes, modifications, adaptations, or variations thereof. It is therefore manifestly intended that this invention be limited only by the claims and the equivalents thereof.

Claims

The invention to be claimed is:

1. A process for the synthesis of derivatives of substituted thiol-containing alkyl fatty acids, said process comprising:

Adding a hydride-donating reagent to the reaction mixture at a second temperature, Said second temperature maintained throughout the rest of the process; Adding a second deprotonating reagent to the reaction mixture; and Adding an alkyl- or acyl-donating reagent to the reaction mixture to form the desired final product.

2. The process of claim 1 , wherein the initial temperature is between 20-25 ° C.

3. The process of claim 1, wherein the second deprotonating agent is identical to the first deprotonating agent.

4. The process of claim 1 , wherein the deprotonating reagent is NaOH, KOH, or NaHCO₃.

5. The process of claim 1 , wherein the second temperature is between 40-45⁰C.

6. The process of claim 1, wherein the hydride-donating reagent is NaBH₄, LiAlH₄, NaAlH₂(OCH₂CH₂OCH₃)2, or NaAlH₂(C₂Hs)₂.

7. The process of claim 1 , wherein the alkyl- or acyl-donating reagent is benzyl bromide or benzyl chloride.

8. The process of claim 1, further comprising:

9. The process of claim 8, wherein the organic solvent system is ethyl acetate and heptane.

10. The process of claim 1, wherein the alkyl fatty acid has a chain length of at least three to eighteen carbons.

11. The process of claim 10, wherein the alkyl fatty acid is octanoic acid or analogs, congeners, salts, or derivatives thereof.

12. The process of claim 11, wherein the alkyl fatty acid is lipoic acid.

13. An intermediate formed during a process for the synthesis of derivatives of substituted thiol-containing alkyl fatty acids.

14. The intermediate of claim 13, wherein the intermediate is 6-benzylsulfanyl octanoic acid.

15. The intermediate of claim 13, wherein the intermediate is 8-benzylsulfanyl octanoic acid.