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Patent Review of Manufacturing Routes to Fifth Generation Cephalosporin Drugs. Part 2, Ceftaroline Fosamil and Ceftobiprole Medocaril David L Hughes Org. Process Res. Dev., Just Accepted Manuscript • Publication Date (Web): 12 May 2017 Downloaded from http://pubs.acs.org on May 14, 2017

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Organic Process Research & Development

Patent Review of Manufacturing Routes to Fifth Generation Cephalosporin Drugs. Part 2, Ceftaroline Fosamil and Ceftobiprole Medocaril David L. Hughes Cidara Therapeutics, Inc., 6310 Nancy Ridge Dr., Suite 101, San Diego, California 92121, United States ABSTRACT: Ceftaroline fosamil and ceftobiprole medocaril are recently approved fifth generation cephalosporins that are active against serious Gram negative and Gram positive bacterial infections, including methicillin-resistant Staphylococcus aureus (MRSA). This article provides a brief background on the mechanisms of β-lactam antibacterial resistance and reviews the patent literature on synthetic routes and final forms of ceftaroline fosamil and ceftobiprole medocaril. GRAPHICAL ABSTRACT: H2N S

ceftobiprole medocaril N OH sodium Patent

N N O

N

N

Literature

NH

O

ceftaroline fosamil acetate

Manufacturing Routes & Final Forms

S

O O

N

Na

AcOH N

O N

O

O N

N O O O

O

H

O

N

HN O

S

S

S

O

N S

N O P OH OH

O

1 ACS Paragon Plus Environment


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____________________________________________________________________________ This article is part two of a series of reviews on the patent literature on synthetic routes and API forms of recently approved antibacterial drugs.1 Part one included a short history of the development of cephalosporin drugs and a review of ceftolozane. The current article reviews ceftaroline fosamil and ceftobiprole medocaril, two other recently approved cephalosporins. All three drugs are fifth generation cephalosporins with broad spectrum antibacterial activity against Gram negative and Gram positive organisms, including methicillin-resistant Staphylococcus aureus (MRSA). Regarding future cephalosporin drugs, cefiderocol from Shionogi is the only cephalosporin drug candidate that is currently in either Phase 2 or Phase 3 development as of Dec 2016, according to Pew Trusts.2 Resistance to antibacterial drugs is a growing concern as pathogenic bacteria, the so-called “superbugs,” are emerging that are resistant to all known antibacterial drugs. The Center for Disease Control and Prevention (CDC) estimates that each year about 2 million individuals in the U.S. contract infections caused by resistant bacteria, resulting in an estimated 23 000 deaths in the U.S.3 Worldwide, resistant infections are estimated to result in 200 0004a to 700 0004b deaths and $20 billion4a in healthcare spending annually. As outlined in our previous article, while the threat of resistant bacterial has increased, antibacterial R&D has declined over the past 20 years, leading to a diminished flow of new drugs to treat the emerging resistant strains.1 A number of government initiatives have been undertaken to spur new investment and R&D in antibacterial research as it has become clear that bold measures and coordinated actions will be required to address the mounting “superbug” threat.


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1. Bacterial Resistance to β-Lactam Antibiotics 1.1 Bacterial resistance - background Bacteria have thrived on planet earth for some three billion years by rapidly adapting to abruptly changing conditions. Some adaptations occur via the normal, but slow, process of random genetic mutations during reproduction, but stress conditions can trigger more rapid evolution.5 Bacteria can also evolve by transferring genetic material with other bacteria via three mechanisms known as transduction, conjugation, and transformation. This transfer of DNA generally occurs between bacteria of the same species, but sometimes species to species transmission occurs, a process termed horizontal gene transfer. As discussed below, this process provides a way of transferring genes conferring bacterial resistance from one pathogenic species to another; that is, an individual species does not have to develop resistance on its own but can borrow it from another.6 This mode of gene transmission has contributed to the rapid rise of resistance across many bacterial species. The introduction of β-lactam antibacterial drugs in the 1930s presented bacteria with a new threat. Harnessing their suite of adaptive mechanisms, bacteria have countered the challenges of β-lactam drugs in at least three ways: (1) production of enzymes that hydrolyze the β-lactam ring, thereby inactivating β-lactam drugs; (2) modification of the structure of enzymes that are required for cell wall construction, called penicillin-binding proteins or PBPs, making them less susceptible to the drugs; and (3) development of efflux pumps that remove antibacterial drugs from the cell.7 The speed with which bacterial resistance can emerge became evident soon after research began on the development of β-lactam antibiotics. In 1940 the Oxford team of Abraham and Chain



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isolated an extract from E. coli that was capable of inactivating penicillin.8 They termed this enzyme penicillinase, which was later determined to be a β-lactamase enzyme that hydrolyzed the β-lactam ring. This resistance to the antibacterial action of penicillin surfaced even before penicillin was introduced for commercial use. Making use of the horizontal gene transfer process, staphylococci with penicillinase became widespread in bacterial infections found in hospitalized patients by the early 1950s, rendering the early penicillin antibiotics ineffective against these resistant strains. The rapidity with which bacteria could overcome antibacterial drugs led to major R&D programs within academia and the pharmaceutical industry to better understand the mechanisms of resistance and to design the next generation of drugs. The introduction of cephalosporin drugs in the 1960s was an important next step in building an effective antibacterial tool box since the cephalosporins were not inactivated by penicillinases. However, resistance soon emerged with bacteria that encoded a gene for cephalosporinase AmpC, a β-lactamase capable of hydrolyzing penicillins, cephalosporins, and monobactams.7 Eleven classes of β-lactamase enzymes now exist to contend with the structural variety of β-lactam drugs that have been developed.9 As fast as scientists have found ways to design drugs to attack a particular bacterial vulnerability, bacteria have quickly developed countermeasures. 1.2 Design of β -Lactam Drugs to Overcome Resistance Bacterial cell walls are primarily composed of glycosylated oligopeptides termed peptidoglycans. The resilient lattice structure of the cell wall is created by cross-linking these oligopeptide chains, a process catalyzed by membrane-associated transpeptidase enzymes known as penicillin-binding proteins (PBPs).10


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The mechanism of action of β-lactam antibiotics, proposed by Tipper and Strominger in 1965 and now well documented,11 involves irreversible binding of the drug to the PBP enzyme, inactivating the PBP and ultimately leading to deterioration of the cell wall and cell rupture. The substrate for PBP is the acyl-D-ala-D-ala dipeptide terminus of the peptidoglycan, which is structurally similar to the β-lactam moiety (Figure 1). As such, the β-lactam drug is able to bind to the active site of the PBP. After binding, a serine residue (Ser 403) of the enzyme in the active site reacts with the β-lactam, forming a covalent ester with the enzyme that is resistant to further hydrolysis (deacylation), thereby occupying the active site and deactivating the PBP enzyme.12-14 Figure 1. Structural Similarity of Penicillin and D-Ala-D-Ala PBP Terminus

Bacteria have 4 PBPs (PBP1a, 1b, 2 and 3) that are inactivated by covalent inhibition by βlactam antibiotics. Resistant bacteria have developed structurally modified PBPs that are no longer susceptible to early generation β-lactam drugs. Resistant strains of Streptococcus pneumoniae encode three additional PBPs, termed PBP2x, PBP2b and PBP1a, while the βlactam resistance of methicillin-resistant Staphylococcus aureus (MRSA) strains results from PBP2a, acquired via horizontal gene transfer.12-14 PBP2a reacts with early generation β-lactam drugs 10- to 1000-fold slower than the native PBPs and the cell is therefore able to maintain catalytic peptidoglycan crosslinking and cell wall construction even in the presence of high concentrations of β-lactam antibiotics that inactivate the other PBPs in the cell.12-14



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Comparison of the crystal structures of native PBP2 with resistant PBP2a reveals structural differences near serine 403 in the active site. In PBP2a this serine is now located in a much narrower groove than in PBP2.14 The early generation β-lactam drugs generally bind much more poorly to the PBP2a active site. Further, once the drug binds to the active site, the serine is no longer aligned for nucleophilic attack, thus reducing the rate of acylation. The β-lactam drug can only act as a weak reversible inhibitor of PBP2a which is not sufficient to disable enzyme function.14 The effectiveness of the three fifth generation cephalosporin drugs (ceftaroline, ceftobiprole, and ceftolozane, Figure 2) against MRSA appears to be a combination of factors: (1) the C-3 side chains bind more effectively to the narrowed groove of the PBP2a active site, primarily due to the permanent positive charge in the C-3 side chains of ceftaroline and ceftolozane and the strongly basic pyrrolidine of ceftobiprole which will be protonated at physiological pH; (2) upon binding in active site, the β-lactam is aligned in an orientation that allows an increased rate of acylation; and (3) the acylated PBP2a has a lower deacylation rate that prevents enzyme turnover.12-14 Figure 2. Fifth Generation Cephalosporin Drugs


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2. Ceftaroline fosamil Sold under the brand names Teflaro (U.S.) and Zinforo (Europe), ceftaroline fosamil was approved in the U.S. in Oct 2010 and Europe in Aug 2012 for the treatment of communityacquired bacterial pneumonia and acute bacterial skin infections. Ceftaroline maintains the C-7 dithiazole E-alkoxyimino side chain from earlier generation cephalosporin drugs. The thiadiazole group provides broad spectrum activity against Gram positive and Gram negative organisms while the E-alkoxyimino group minimizes hydrolysis by β-lactamases. The C-3 side chain has been optimized for binding to PBP2a, specific for MRSA activity, and contains a 2-thioazolythio linker with a N-methylpyridinium group. While having improved stability to β-lactamases over early generation cephalosporins, ceftaroline is inactivated by several classes of β-lactamases and therefore is not indicated for Gram-negative



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bacteria with extended spectrum beta-lactamases (ESBLs) having the mutant strains derived from TEM, SHV or CTX-M families, 390 serine carbapenemases, metallo-beta-lactamases, or AmpC cephalosporinases. The parent drug, ceftaroline, has poor water solubility (2.3 mg/mL), precluding formulation as an IV drug. A prodrug approach culminated in the selection of the N-phosphono prodrug which has excellent water solubility at pH 7 (>100 mg/mL) as well as adequate stability in aqueous solution and as a crystalline solid.15,16 Ceftaroline fosamil is rapidly converted into bioactive ceftaroline in plasma by phosphatase enzymes. Measurable levels of the prodrug can only be detected while the drug is being infused.17 2.1 Medicinal Chemistry Route to Ceftaroline Fosamil18 The Medicinal Chemistry route to ceftaroline fosamil starting from the commercially available cepham triflate 1a is outlined in Scheme 1.16,19 Synthesis of the cepham core structure is described in Section 2.5. Triflate 1a was reacted with the sodium salt of 4-(4-pyridyl)thiazole-2thiol (2) in THF in an ice bath for 2 h. Work up consisted of concentration, then addition of EtOAc and water, resulting in crystallization of the THF solvate of 3a, isolated in 79% yield. The authors report that some double bond isomerization (from ∆3 to ∆2 in the 6-membered ring) occurred during this reaction, although the extent of isomerization was not indicated.

The methylation of the pyridine was conducted using MeI in DMF at room temperature, with the product 4a isolated as a solid from Et2O. The amide side chain was cleaved using PCl5/pyridine to form a presumptive imino chloride (not isolated) which was converted to the C-7 amine 5a by the addition of MeOH at – 20 oC. The amine 5a was concentrated to an oil. Deprotection of the PMB ester was carried out with TFA and anisole in CH2Cl2 to afford carboxylic acid 6, which 8 ACS Paragon Plus Environment

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was purified by SP-207 resin chromatography, eluting with 15% EtOH:85% water, and isolated by lyophilization of the rich cuts. Yield across the 3-steps from 3a was 45%. The amidation with the phosphono acid chloride side chain 7 was carried out with NaHCO3 in water/THF to afford crude ceftaroline fosamil, which was purified by SP-207 resin chromatography, eluting with 10% EtOH:90% water, in overall 62% after lyophilization. Scheme 1. Medicinal Chemistry Route to Ceftaroline Fosamil Acetate



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2.2 Process Chemistry Route to Ceftaroline Fosamil Acetate The process route to ceftaroline (Scheme 2)16,20 focused on optimization of the Medicinal Chemistry route. The protecting group of the carboxylic acid was changed from PMB to benzhydryl, which provided several intermediates that were crystalline and could be purified without chromatography. The process described in U.S. patent 7,419,973 starts with hydroxy-cepham 1b, which is converted to mesylate 1c in 90% yield using MsCl and i-Pr2NEt in either acetone or acetonitrile.20b For the next step, sodium methoxide was used for deprotonation of 2 instead of NaH which was used in the Medicinal Chemistry route. The product was isolated directly from the reaction mixture by addition of MeOH/water to afford solid 3b in 81% yield. Methylation with MeI in DMF afforded the N-methylpyridinium salt 4b, which was isolated in 97% yield by adding the reaction mixture to EtOAc to crystallize the product. Cleavage of the side chain was accomplished with PCl5 in CH2Cl2 as in the Medicinal Chemistry route to generate the imino chloride intermediate, but MeOH was replaced with i-BuOH for release of the amine group. Use of i-BuOH resulted in precipitation of 5b from the reaction mixture, which was isolated in 93% yield. Deprotection of benzyhydryl intermediate 5b was accomplished with aq. HCl in MeCN affording 6 as a crystalline bis-HCl salt, thereby avoiding the resin chromatography and lyophilization sequence of the Medicinal Chemistry route.


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Coupling of side chain 7 was carried out in water using NaOAc as base. At the end of the reaction, EtOAc was added, the layers were separated, then EtOH was added to the EtOAc layer to crystallize ceftaroline fosamil as the disodium salt, which was isolated and dried. Purification was carried out using SP-207 resin chromatography using 10% EtOH/water to elute the product. The rich cuts were concentrated, then aq. HCl was added to adjust the pH to 0.5, which resulted in crystallization of zwitterionic ceftaroline fosamil in an isolated yield of 77%. In another example, purification was accomplished by dissolution of the disodium salt in water, then addition of HOAc, HCl, and EtOH to precipitate the zwitterion in 80% yield and 91% purity. The acetic acid solvate was prepared by suspending the zwitterion in water, addition of aq. NaOAc to give a clear solution, then addition of HOAc and H2SO4 to induce crystallization of ceftaroline fosamil acetate, which was isolated in 57% yield. When the crystallization was carried out in the presence of D-mannitol (1 equiv), the yield improved to 74% on a kilogram scale. Since no further patent applications or journal articles related to the synthesis have been published by the innovator companies, this route is likely the manufacturing route, although further development has no doubt been carried out to improve yields and streamline processing. According to the European Public Assessment Report (EPAR),21 the manufacturing process of ceftaroline fosamil consists of six chemical transformations and five purification steps, which is consistent with Scheme 2, except each of the six reaction products are isolated by crystallization as described in the Takeda patent and publication. The Takeda patents do not include composition of matter claims for any intermediates. Scheme 2. Process Route to Ceftaroline Fosamil Acetate



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H N

Ph

S

O

N O Ph

Ph

N

OMs

O

i-Pr2NEt acetone 90%

O

MeONa, MeOH, THF, -5 oC

S

O

MsCl

OH

O

1b

H N

Ph

O

1c

N

O

Ph

N

Ph HS

Ph

S

O

N

S

O

O

Ph

S

O

DMF rt 97%

S

H N

Ph

MeI

N

O

N

N

S

O

1) PCl5, CH2Cl2 pyridine, 5 oC

N

O

S

2) i-BuOH -10 oC

O

Ph

Ph

3b Cl

2

S 81%

I

N H N

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93%

Ph

4b

N

N 2 HCl

H2N

S N

S

O O

H2N

Conc HCl

N

N

CH3CN, rt 76%

S

O

1) NaOAc, H2O, rt

N S

O

O

Ph

S

N

S

Cl

N

OH

O HN P Cl Cl

6

Ph

O

5b

S N

O

7

2) purify by SP-207 resin 77%

N H N

O P HO

OH

N

H N

N S N

N

O

O

S

N

N

S

O O

1) Aq. NaOAc

O

ceftaroline fosamil

S

H N

O P

2) Aq. AcOH H2SO4 57%

HO

OH

H N

N S N

N

O

O

S

N

N

S

O O

O

S AcOH

ceftaroline fosamil acetate

2.3 Alternate Routes to Ceftaroline Fosamil 12 ACS Paragon Plus Environment

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Patents and patent applications have been published for four modified routes to ceftaroline fosamil, all starting with the same core cepham structure. 2.3.1

Alternate methylating agent

Chinese patent application CN 102977124 from Shandong Chengchuang Pharmaceutical Technology Development Company reports methylation of 3b with dimethyl carbonate, a greener and cheaper alternative to MeI.22 The reaction was carried out in 3.3 vol DMF with Et3N as base, keeping the reaction temperature 250 mg/mL) was developed as the active pharmaceutical ingredient (Figure 2). This ester is rapidly converted to the carboxylic acid, ceftobiprole, by plasma esterases as well as nonenzymatic hydrolysis.44


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3.1 Route 1. Medicinal Chemistry Synthesis45 The initial route to ceftobiprole started with 3-formyl-cepham 25, which was coupled with phosphonium salt 26 to afford alkene 27 (Scheme 10).46 According to an earlier patent, the double bond of 3-formyl-cepham 25 is prone to isomerization such that the product of this reaction has the double bond in the ∆3 position regardless of whether the starting material is ∆2 or ∆3.47 The ∆3 double bond of 27 was then isomerized to desired ∆2 position by mCPBA oxidation to the sulfoxide 28, which was reduced back to the sulfide 29 with PBr3. The Boc and benzhydryl groups were deprotected with TFA to afford amine 30 followed by introduction of the C-7 side chain via coupling with thioester 31 to yield protected ceftobiprole 32. The allyloxycarbonyl protecting group was removed with (PPh3)2PdCl2 to generate the free pyrrolidine 33 followed by deprotection of the trityl group using TFA and triethylsilane, affording ceftobiprole. The medocaril prodrug was synthesized by reaction with carbonate 34 in DMSO.48 Once the reaction was complete, sodium 2-ethylhexanoate in acetone was added to precipitate the sodium salt of ceftobiprole medocaril. Scheme 10. Initial Route to Ceftobiprole



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3.2 Second Generation Medicinal Chemistry Route to Ceftobiprole Medocaril An improved route to ceftobiprole is described in U.S. patent 6,504,025 and European patent 1,067,131 granted to Basilea (Scheme 11).49,50 This route avoids the undesired isomerization of the cepham double bond to the ∆3 position under the basic conditions used to synthesize aldehyde 25. The order of addition of the side chains is reversed relative to the first generation route, which allows for the introduction of the C-3 side chain as a penultimate step with the medocaril group already incorporated. The second generation route starts with the unprotected primary alcohol 36, a commercially available material that is prepared by deacylation of 7-aminocephalosporanic acid (7-ACA). Reaction with activated thioester 31 mediated by tetramethylguanidine afforded oxime 37, which was not isolated but directly protected with diphenyldiazomethane (35) to furnish benzhydryl product 38, which was isolated by crystallization from hexane in 90% yield for the two steps. Oxidation to the aldehyde 39 was carried out with TEMPO/NaOCl in biphasic aq./CH2Cl2 in 74% yield with no isomerization of the ∆2 double bond. Oxidation with MnO2 suspended in THF/ CH2Cl2 was also described, affording aldehyde 39 in 52 % yield. The free amine of the side chain and the divalent sulfur in the cepham moiety were unaffected by these oxidation conditions as long as the reagent amounts, temperature, and reaction time were controlled.



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The side chain 40, incorporating the medocaril group, was prepared from 26 in two steps. First, the allyloxycarbonyl group was deprotected with (PPh3)2PdCl2 and n-Bu3SnH, then coupling with carbonate 34 afforded the side chain 40. The Wittig reaction of 39 was then conducted with 40 at -78 oC to afford olefin 41 in 74% yield.49,50 Deprotection of the benzhydryl and trityl groups of 41 with TFA and triethylsilane afforded ceftobiprole medocaril. To isolate the product, the reaction mixture was evaporated to dryness, then the residue was titurated with Et2O to afford a suspension which was filtered and dried to afford a beige solid in 92% overall yield. Compounds 38 and 39 are claimed in the patent application.49 Scheme 11. Second Generation Route to Ceftobiprole Medocaril


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3.3 Process Chemistry Route to Ceftobiprole Medocaril While the convergent aspect of the second generation route is attractive, isolation of the API in sufficient purity could be problematic since it is an amorphous solid that cannot be purified by crystallization.



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More recent patents from Basilea suggest the manufacturing route from alcohol 38 may proceed as presented in Scheme 12, with ceftobiprole as the final intermediate that can be isolated as a pure crystalline material prior to appending the medocaril group in the final step. An improved oxidation procedure of alcohol 38 is described in two Basilea patents, U.S. 9,006,422 and U.S. 9,499,566, using (diacetoxyiodo)benzene as oxidant with 5-10 mol % TEMPO catalysis in THF, MeCN, EtOAc, CH2Cl2 or mixtures of these solvents.51 Aldehyde 39 was isolated by precipitation upon addition of a hydrocarbon anti-solvent such as hexane. Yields up to 84% were obtained with the purity of 39 ranging from 93-98%.51 The Wittig olefination was carried out with phosphonium salt 42 having the pyrrolidine protected as a Boc group, which allows for a global TFA deprotection at the end of the synthesis.49,52,53 In the first iteration of this process, olefin product 43 was purified by slurrying the solid in CH2Cl2/EtOAc, which produced fine brown particles of 90% purity that were difficult to filter and contained 1.4 equiv EtOAc. In U.S. patents 8,865,697 and 9,163,034 assigned to Basilea, olefin product 43 was purified by crystallization as a DMSO solvate.53 Crude 43 was dissolved in EtOAc at 23-27 oC followed by addition of DMSO, which resulted in crystallization of 43 DMSO solvate. The product was recrystallized by dissolution at ambient temperature in CH2Cl2 followed by addition of an equal volume of EtOAc to crystallize the DMSO solvate (approx. 2 moles of DMSO) with a purity of 97%. The DMSO solvate 43 is claimed in the granted patents.53 Compound 43 was globally deprotected using Et3SiH and TFA to generate ceftobiprole. In an international patent application, Basilea describes five polymorphs of crystalline ceftobiprole, with form D identified as the most stable form.54 Form D was isolated as follows. Once the


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deprotection reaction was complete, EtOAc, CH3CN, water, and TFA were added to create a two-phase mixture, with the product distributing to the aq. phase. The aq. phase was loaded onto a non-functionalized polystyrene-based resin and eluted with 9% CH3CN/water containing 0.4% TFA. The rich cuts were concentrated to remove CH3CN, then the pH was adjusted to 3, which resulted in crystallization of ceftobiprole form D in about 95% purity. This form was shown to be more stable than amorphous ceftobiprole, with a decrease in HPLC wt % of 3% over one month at 40 oC vs a 40% decrease with amorphous material.54 In the final step, the medocaril side chain was appended using carbonate 34 in DMSO. Ceftobiprole medocaril was isolated as the sodium salt via precipitation with sodium 2ethylhexanoate in acetone, then further purified by a reslurry in acetone.48 Scheme 12. Third Generation Route to Ceftobiprole Medocaril



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OTr H N

N N H2N

S N

N

O

N

OTr H N

S N

O

N S H2 N

TEMPO

OH

O

PhI(OAc)2 O

38

H2 N

N

N

OTr H N

O

S N

O

N

N

N

OH H N

S N

O

N H2N

O

Ph

O

TFA

O

S

O

34

N

O

Ot-Bu Et3SiH CH2Cl2

43

O

N

Ph

Ph

1) O2N

O O

NH

O

DMSO

O O

N

O O

Ot-Bu

42

O

S

O

N

Ph3P

O Ph

O Br

O

N O

Ph

N t-BuOK,CH2Cl2 Toluene, THF

S

39

O

Ph

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2) Sodium ethylcaproate, acetone

OH

ceftobiprole

N

OH H N

S N

O

N H2 N

S N

N O

N

O

O O

O

O Na

ceftobiprole medocaril sodium

O O O

3.4 Alternate Routes to Ceftobiprole Medocaril 3.4.1 Fourth Generation Route without Carboxylic Acid Protection


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A concise three step route to ceftobiprole has been described by Sandoz in U.S. patent 9,096,610 (Scheme 13).55 In this route, the Wittig reaction is carried out first using the fully elaborated medocaril side chain 40. Scheme 13. Fourth Generation Route to Ceftobiprole Medocaril

The Wittig reaction was carried out using unprotected aldehyde 44 with phosphonium salt 40 in a mixture of bis(trimethylsilyl)acetamide (BSA) and propylene oxide. The BSA was used for in situ formation of the trimethylsilyl ester of the carboxylic acid. The role of propylene oxide was



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not specified in the patent, but it apparently serves to activate the phosphonium salt by a rarely used method described by Buddrus in 1968 in which the epoxide activates the phosphonium salt through dehydrohalogenation, as indicated in Scheme 14, resulting in formation of the active alkylidene phosphorane intermediate.56 Using these reagents, the Wittig reaction was conducted at 1 oC in contrast to the -78 oC conditions required under the strong base conditions described in the Roche and Basilea patents.48-50,53 The crude product 45 was isolated by adding 2-PrOH to the reaction mixture, resulting in precipitation of the product. Further purification could be accomplished by crystallization of the p-TsOH salt from MeOH/2-PrOH.55 Scheme 14. Activation of Phosphonium Salts via Dehydrohalogenation with an Epoxide

In step 2, the oxime side chain 46, as its acid chloride, is reacted with the free base of 45 in CH2Cl2 solvent, again using BSA for in situ protection of the carboxylic acid. After aq. workup, the organic layer was concentrated to dryness to afford crude 47. In the final step, cleavage of the trityl group was carried out using Et3SiH and TFA in CH2Cl2. After complete reaction, the solution was added to Et2O, resulting in precipitation of ceftobiprole medocaril.55 The major downside of this route is use of propylene oxide, a low boiling liquid (bp 34 oC) that is a known animal carcinogen and a suspected human carcinogen.57 The U.S. National Institute for Occupational Safety and Health (NIOSH) recommends that closed systems should be used for working with propylene oxide.57 This may require use of propylene oxide in a facility equipped for handling potent compounds. 36 ACS Paragon Plus Environment

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3.4.2 Fifth Generation Route Similar to the fourth generation route, the fifth generation route (Scheme 15) published in U.S. patent 9,139,597 from Sandoz starts with unprotected aldehyde 44 with the Wittig reaction conducted as the first step.58 While the fourth generation route used the fully elaborated phosphonium side chain 40 in the Wittig reaction, this route used the Boc-protected salt, 42, in the solvent mixture of BSA and propylene oxide, to afford olefin 48 which was purified as its crystalline dicyclohexylamine salt. Addition of side chain 46 afforded bis-protected ceftobiprole 49, which was deprotected with using Et3SiH and TFA in CH2Cl2 to afford ceftobiprole. The medocaril prodrug was prepared with 34 in DMSO as outlined in Scheme 12. As noted above, isolation of ceftobiprole allows for purification of a crystalline final intermediate before appending the medocaril group and generating the amorphous final product.

Scheme 15. Fifth Generation Route to Ceftobiprole



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3.5 Ceftobiprole - Side Chain Syntheses 3.5.1 Phosphonium salt 42 Basilea has patented two routes to phosphonium salt 42. In the first route (Scheme 16)46,59 N-Cbz-dimesylate 50 was reacted with hydroxylamine hydrochloride to afford N-hydroxypyrrolidine 51 which was isolated in 80-85% yield after crystallization from hexane. The hydroxyl group was reductively cleaved with Raney nickel with in situ Boc protection to furnish bis-protected pyrrolidine 52, which was isolated as an oil after silica treatment. The Cbz group was deprotected by hydrogenation to generate free amine


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53, isolated as a crude oil, which was reacted with 2-bromo-4-chlorobutyryl chloride (54) in a 2phase mixture of 50% NaOH/CH2Cl2 to afford pyrrolidinone 55. The phosphonium salt 42 was then generated using an excess of triphenylphosphine in dichloromethane. Scheme 16. Route One to Phosphonium Salt 42

The second route starts with unnatural Cbz-(R)-asparagine methyl ester (56) (Scheme 17).60 Cyclization was carried out with NaH followed by protection with benzyl bromide to afford dioxo-pyrrolidine 57. The Cbz group was selectively deprotected by hydrogenation in HOAc with the resulting amine 58 isolated as the crystalline bis-acetate salt/solvate from EtOAc/hexane. After salt break, reduction was carried out with Vitride® in toluene at 80 oC to produce pyrrolidine 59 which was isolated as a crude oil. Treatment of 59 with 2,4dibromobutyryl chloride (60) in MeCN using Na3PO4 as base afforded pyrrolidinone 61, isolated as a crude solid. The phosphonium salt 62 was generated by treatment with PPh3 in toluene at 110 oC. The workup consisted of dilution with EtOAc, extraction of the product into aq. NaBr,



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washing with EtOAc to remove O=PPh3, then extraction of the aq. layer 7 times with CH2Cl2 and concentration of the organic layer to a foam. Protecting group switch was accomplished in one pot via hydrogenation at 60 oC in MeOH in the presence of Boc2O to afford phosphonium salt 42 as a foam after filtration and concentration.60 Scheme 17. Route Two to Phosphonium Salt 42

3.5.2

Ceftobiprole Thiadiazole Side Chain Acid 67

An improved route to the ceftobiprole thiazole side chain acid 67 has been published by Zhong and co-workers (Scheme 18) in 6 steps, 4 isolations, and 19% overall yield.61 Earlier routes to


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this side chain are not discussed here since they are summarized in the Zhong publication and the routes are similar to those discussed above for the ceftaroline side chain. Scheme 18. Route to Ceftobiprole Thiadiazole Acid Side Chain 67

Malononitrile was reacted with hydroxylamine hydrochloride and MeONa in MeOH and then selectively acylated in the same pot. Addition of water resulted in crystallization of isoxazole 63, which was isolated in 71% yield. Isoxazole 63 was added to a mixture of benzoyl chloride and KSCN in CH3CN, forming the presumptive intermediate 64, which then rearranged to the thiadiazole 65, which crystallized directly from the reaction mixture and was isolated in 76% yield. Oxime formation was carried out with isopropyl nitrite and conc. HCl in THF to selectively form the desired Z-isomer with

Patent Review of Manufacturing Routes to Fifth ... - ACS Publications

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