A Review on Platensimycin: A Selective FabF Inhibitor

Emerging resistance to existing antibiotics is an inevitable matter of concern in the treatment of bacterial infection. Naturally occurring unique class of natural antibiotic, platensimycin, a secondary metabolite from Streptomyces platensis, is an excellent breakthrough in recent antibiotic research with unique structural pattern and significant antibacterial activity. β-Ketoacyl-(acyl-carrier-protein (ACP)) synthase (FabF) whose Gram-positive bacteria need to biosynthesize cell membranes is the target of inhibition of platensimycin. So, isolation, retrosynthetic analysis, synthesis of platensimycin, and analogues of platensimycin synthesized till today are the objectives of this review which may be helpful to further investigate and to reveal untouched area on this molecule and to obtain a potential antibacterial lead with enhanced significant antibacterial activity.


Introduction
The majority of the "illness" lies in the fact that when immune system is defeated, it is in war with pathogens. Development starting from ethnic to modern synthetic approaches in drug discovery mainly provides better weapons to combat and survive against pathogens. In contrast to medical development, pathogens also have acquired protection called "resistance." More or less all classes of antibiotics are resistant to bacteria; hence a novel process to the discovery of antibiotic with new mechanism of action is essential [1,2]. Microbial and chemical groups at Merck in the year 2006 have found three novel chemical classes, using antisense technology, from their older microbial screening library [3]. Two compounds, platensimycin and platencin, were found as potent inhibitors of fatty acid biosynthesis. Platensimycin selectively inhibits fatty acid acyl carrier protein synthase II (FabF). Platensin is a balanced dual inhibitor of both FabF and FabH (fatty acid acyl carrier protein synthase III) [4]. The third compound lucensimycin A was found to inhibit ribosomal protein synthesis. FabF is one of the enzymes which catalyze the biosynthesis of fatty acids in bacteria. It makes FabF an essential target for inhibiting bacterial growth in resistant bacteria. Previously two classes of inhibitor, cerulenin [5] and thiolactomycin [6][7][8], were reported, but inhibitory activity was poor (IC 50 ranges within 1.3-13 g/mL) with poor antimicrobial activity (Streptococcus aurous, MIC-64 g/mL) [9]. On the other hand, platensimycin had shown selective FabF inhibitory activity on S. aurous and E. coli with IC 50 value of 48 nM and 68 nM [3]. The ability of this class of compounds to bind and inhibit FabF enzyme has given a new class of antibiotics. Among them platensimycin and platencin are most promising and need further investigation. There might be an argument that if fatty acid synthesis is an attractive target for antibacterials, why there have not many drugs or natural inhibitors targeting this pathway been isolated? One of the reasons may be that the Streptomyces and related Actinomyces, the organisms that have delivered most of the existing antibiotics, are constrained in their ability to produce fatty acid synthesis inhibitors by the adjacent relationship between the synthetic pathways of fatty acids and polyketides. These pathways share many chemical, mechanistic, and structural features, and, thus, fatty acid synthesis inhibitors recurrently inhibit polyketide synthesis (e.g., cerulenin is a powerful inhibitor of polyketide synthesis). An organism producing a fatty acid synthesis inhibitor must retain not only a resistant form of the fatty acid synthetic enzyme but also a resistant form of the antibiotic-producing polyketide synthase. Furthermore, if several polyketides are needed for survival of the organism in its ecological niche, then resistant forms of each of these  pathways would be prerequisite. Such precincts seem likely to severely narrow the opportunities to evolve fatty acid synthesis inhibitors and may account for the scarceness of such antibiotics. However, various Streptomyces produce analogues of thiolactomycin, which has not been reported to block polyketide synthesis, thus demonstrating that development of fatty acid synthesis inhibitors by this organism is conceivable [10].

Isolation of Platensimycin
Platensimycin was isolated by Wang et al. in 2006 at concentration of 2 to 4 mg/L from fermentation broth of S. platensis (MA7327 and MA7331) using SephadexLH20 liquid chromatography medium by reversed-phase HPLC chromatography [3]. In a subsequent study, three-step isolation was modified by Singh et al. and they established two-step method eliminating the SephadexLH20 step [11]. It was also isolated from Streptomyces platensis (MA7327), recovered from soil samples collected in Eastern Cape, South Africa [12].

Structure of Platensimycin
Platensimycin ( Figure 1) consists of a benzoic acid moiety substituted at ortho and para with hydroxyl group and in meta position is conjugated with a unique pentacyclic ketolide by an amide linkage [13]. The structure was established by combination of DQ-COSY and TOCSY correlations (2D NMR), UV, and IR spectroscopy and confirmed by X-ray crystallography [14].

Synthesis of Platensimycin
Platensimycin consists of an aromatic acid conjugated with aliphatic moiety by an amide linkage. The effective synthetic strategy is to synthesize the aromatic and aliphatic parts separately and then combine them by amide linkage. Many synthetic methods are available for the synthesis of those two building blocks.

Retrosynthetic Analysis of Platensimycin.
Using retrosynthetic analysis (Scheme 1) they separated the aromatic amine 2 and the carboxylic acid 3 by disconnection of amide linkage. The carboxylic acid was further simplified to a simplified enyne in successive three retrosynthetic steps. This resulted in two target molecules to synthesize the tetracyclic carboxylic acid and the aromatic amine from simplified starting material.

Synthesis of Tetracyclic
Cage. The simplified enone generated from 3-ethoxycyclohex-2-enone which was used as a starting material (Scheme 2). Allylic bromide 9 [27] (LDA, 92%) and propargyl bromide (LDA, 97%) were used as reagents of choice to generate the bis-alkylated enone 10 from 8. Reduction followed by acidic hydrolysis and reintroduction of the TBS ether produced enone 11 from enone 10 (yield 84%). Spirocycle 12 was generated by cycloisomerization of 11 [28,29]. Oxidation of 12 produced bis-enone 13 [30] which upon acid hydrolysis gave desired aldehyde 6. Secondary alcohol 14 was prepared by addition of samarium(II) iodide in a dilute solution of aldehyde 6 HFIP in THF/HMPA followed by NH 4 Cl solution. Esterification of 14 with TEA resulted in the formation of cage-like structure 4 which on treatment with KHMDS and MeI followed by KHMDS and allyl iodide produced olefin 16. Vinyl pinacol boronate and 16 reacted in presence of the Grubbs second generation catalyst to produce vinyl boronate 19 which on reacting with trimethylamine Noxide gave 20. Following Pinnick protocol 20 was converted to desired carboxylic acid 3.

Synthesis of Aromatic
Amine. The synthesis of the aromatic amine 2 was started from 2-nitroresorcinol 21 by protecting with MOM ether, followed by catalytic hydrogenation, 24 was formed (Scheme 3). Again protecting the amino group and followed by silylation, lithiation, and quenching with methyl cyanoformate 24 was carboxylated and by unprotecting amino group using thermolysis, desired aniline 2 was synthesized.

Synthesis of Platensimycin
Core. The total synthesis of platensimycin was completed by the coupling of carboxylic acid 3 with aniline 2 which was achieved by treatment with HATU followed by hydrolysis with LiOH (Scheme 3).

Another Approach for Synthesizing Tetracyclic Cage by Nicolaou et al. .
After successfully reporting the first total synthetic strategy of platensimycin, Nicolaou et al. [31] report a new synthetic strategy that starts from the readily available and inexpensive (R)-(−)-carvone to the tetracyclic enone 2.

Retrosynthetic Strategy of Tetracyclic Enone.
A fivestep retrosynthetic disconnection approach was shown to generate commercially available (R)-(−)-carvone 31 from 26 (Scheme 4). (i)  33 was treated with Hg(OAc) 2 and later on with NaBH 4 to generate approximately 1 : 1 mixture of exo-and endotertiary alcohol, which on dehydration using Martin's sulfurane reagent produced exocyclic alkene 35 which was treated with TMSCl and HMDS and followed by an electrophilic quench with PhSeCl and subsequent oxidative elimination (H 2 O 2 ) to give enone 36. Enone 37 was treated with SmI 2 for radical cyclization, and by adding mentioned reagent (Scheme 5) in subsequent 8 steps of tetracyclic cage 26 was synthesized.

Synthesis of Tetracyclic Cage. O-TBS and O-benzoyl-
protected enones 47a and 47b were prepared from 1,3cyclohexadiene 49 through allylboration, selective oxidation, and utilizing the Dess-Martin oxidation, with the yield of (97%). Diels-Alder reaction in between 47a and 47b and siloxydienes produced two stereogenic mixtures 46a, 46b and 46c, 46d. The mixture of two inseparable diastereomers (46b and 46d) was employed in the next step. Following Noyori's procedure [26] followed by hydrolysis of the benzoyl group and separation of diastereomers 53 was given which upon catalytic oxidation using palladium(II) chloride and copper(II) acetate produced 54a and 54b in 10 : 1 ratio. Vinyl triflate 55 was produced from 54 and reduced to 56 which was transformed to 57. Transannular radical cyclization of monothioacetal 57 gave desired 4 (Scheme 7).  aldehyde 68 containing oxoethylene group was synthesized from alcohols 66 by Mandai protocol [39]. Using Ohira-Bestmann reagent 74 [40,41], enyne 69 was formed which was treated with AIBN in tert-BuOH and PPTS to initiate the radical cyclization resulting formation of tetracyclic cage 72. L-Selectride, THF, and TFA/CH 2 Cl 2 were consecutively added with 72 to produce the desired enone 58 (Scheme 9).  [47] used an intramolecular Robinson annulation approach [48] in the retrosynthetic analysis presented in (Scheme 11). The bicyclic compound 84 to give the tetracyclic core structure 4 is key step in the retrosynthesis by Robinson annulation event.

Synthesis of Tetracyclic
Core. The bicyclic ketone 84 can be synthesized from known lactone 85 which could be generated from ketone 86, through a Baeyer-Villiger oxidation/rearrangement sequence [49,50] and by utilizing Bronsted acid assisted chiral Lewis acid catalyzed Diels-Alder reaction, and subsequent N-nitrosoaldol addition/decarboxylation, ketone 86 could be easily prepared from inexpensive, commercially available starting materials (Scheme 12). Diels-Alder reaction between methyl acrylate 87 and methyl cyclopentadiene 88 produced adduct 89. The reaction was catalyzed by BLA and carbon-based Bronsted acid [51]. Ketone 90 was obtained in one pot reaction from adduct 89 using lithium enolate and lithium hydroxide in dioxane. Baeyer-Villiger oxidation in basic condition [52] of ketone 90 gave lactone 85. Vinyl lactone 91 was obtained from lactone 85 using cuprate reagent [53] and trifluoromethanesulfonimide [54,55]. DIBAL-H reduction followed by cyanation produced cyanide 92a, 92b, which was reduced and subjected to Wadsworth-Emmons conditions [56] to give enone 93; ruthenium-catalyzed oxidation [57] produced aldehyde 84. Using L-proline as the chiral control element followed by sodium hydroxide treatment gave the desired tetracyclic core structure 4.

Pharmacology of Platensimycin
Bacterial cell wall synthesis, protein synthesis, and DNA replication are the predominant targets for widely used antibiotics. But the emergence of resistance to antibiotics   demands new antibacterial targets. Fatty acid synthase (FAS) pathway is now an attractive target for antibacterial agents because, as a new target, FAS inhibition will not suffer from bacterial resistance immediately and biosynthesis pathway of bacteria, plants, and parasites (FAS II in which component proteins are dissociated) is different from mammals (FAS I in which component proteins are generally single-chain, multidomain homodimers or two-chain heterodimers carrying all proteins of the pathway) in subcellular organization of components which demonstrate a target specificity for the FAS II inhibitors. A general scheme for type II fatty acid biosynthesis is shown in Figure 2 [9]. A recent development in finding inhibitors of fatty acid biosynthesis is the discovery of platensimycin which shows broad-spectrum Grampositive antibacterial activity (Staphylococcus aureus (MRSA) and Enterococci (VRE), MIC < 1.0 g mL −1 ) by selectively inhibiting cellular lipid biosynthesis [58]. The mechanism of action is the selective inhibition of elongation of FabF    bacteria incorporates exogenous fatty acids into their cell [61,62]. Platensimycin exhibited antibacterial activity against efflux-negative Escherichia coli (tolC), but not against wildtype E. coli, specifying that efflux mechanisms, and not compound specificity, limit the effectiveness of platensimycin in E. coli and possibly other Gram-negative bacteria [3].

Platensimycin Analogues
The major drawback to most natural products, including platensimycin, is poor pharmacokinetic properties and negligible oral bioavailability. Only the continuous infusion of a high platensimycin dose showed effectiveness in mice   MIC of (+)adamantaplatensimycin against S. aureus (MSSA), >88 g/mL [25] infected with S. aureus [3]. For the improvement of pharmacokinetic profile, many researchers have developed platensimycin analogues (Table 1); however potent antimicrobial activity than that of platensimycin is yet to get.

Conclusion
Development of bacterial resistance to the existing antibiotics is an alarming situation in the 20th century. Finding new target for bacterial demolition is essential. Platensimycin serving as a potential antibiotic interacting with FabF may bypass bacterial resistance. It should be noted that starting from 2006 till now a huge number of scientists provided many synthetic strategies and derivatives which are very encouraging.
Though some evidence like incorporation of exogenous fatty acid inside bacteria when supplied makes the hope regarding platensimycin is uncertain, overall, isolation of platensimycin as a selective FabF inhibitor, complex synthesis of tetracyclic cage, and enhancement of its pharmacokinetic properties by its derivative synthesis are the excellent works in the era of medicinal chemistry. It is expected that this review might be helpful for the medicinal chemist.