Concise, three-step enantioselective total synthesis of (4 S ,5 S )-4-hydroxy-5- octylpyrrolidin-2-one, a colibactin pathway metabolite

We report the first synthesis of (4 S ,5 S )-4-hydroxy-5-octylpyrrolidin-2-one, one of the colibactin pathway metabolites isolated and characterized in 2021. Our synthesis started from ( S )-malimide, a chiral building block previously developed from our laboratory. The first five-step approach consists of a highly regioselective Grignard addition at C-2 of ( S )-malimide, catalytic dehydration, catalytic cis -selective hydrogenation of enamide, and successive cleavages of N -PMB and O -benzyl groups. By telescoping three reactions into one step, an even concise, three-step enantioselective total synthesis of the title compound has been achieved. Through this first enantioselective total synthesis, full physical and spectral data of this natural product were obtained.


Introduction
Natural products have long been recognized as the most important resources for developing new medicines. 1owever, since the early 1990', with the emergence of novel technologies such as combinatory chemistry and high throughput screening (HTS), the natural product-based drug discovery strategy has been abandoned by the major pharmaceutical companies.About fifteen years late, although the renaissance of drug discovery from natural products has had been expected, it is challenging for several reasons.Among them, the availability of natural products and analogs remains the major challenge.Thus, the synthetic efficiency is becoming one of the major pursuits of total synthesis of natural products. 2raditionally, natural products refer to secondary metabolites of plants, animals, and microorganisms, 1 which are considered to be the chemicals useful for their survival in the related ecological systems.Investigation of natural products from humans is rare.However, the investigation on steroids and prostaglandins have resulted in a series of important medicines in modern society.On the other hand, during recent decades, it has been recognized that the human microbiota is a dynamic collection of microbes that have existed within and on us, throughout our evolutionary history, 3 and microbe-derived natural products are mediators of many human disease. 4Colibactins are a group of genotoxic secondary metabolites produced by some members of the gut microbiota, including Escherichia coli and other Enterobacteriaceae, which show genotoxicity relevant to colorectal cancer.In 2021, the teams of Kim and Crawford reported 10 new colibactin pathway metabolites (1 -10).These compounds were revealed to be produced from a new biosynthetic crosstalk relationship between secondary polyketide and primary fatty acid pathways featuring an αaminomalonate-derived lactam ring. 5Among them the two most abundant metabolites were isolated and structurally characterized as (4S,5S)-5-octyl-4-hydroxypyrrolidin-2-one (1) and (4S,5S)-5-[(Z)-dec-3-en-1-yl]-4hydroxypyrrolidin-2-one (2).Although the absolute configuration of natural lactam 1 has been determined as 4S,5S, its optical rotation has not been reported, neither biological activities, possibly due to the paucity of the natural product.
][8][9][10] For example, streptopyrrolidine (11) was isolated as an angiogenesis inhibitor from the fermentation broth of a marine Streptomyces sp.isolated from the deep-sea sediment, 11 levetiracetam ( 12) is applied as a medication for epilepsy with a global sale of 770 M€ in 2018, 12 while (−)-clausenamide (13), a natural product isolated from traditional Chinese medicinal plant, is under clinical trial as a potential drug for treatment of Alzheimer's disease. 13© AUTHOR(S) Figure 1.Representative -lactam-containing bioactive natural products and medicinal agents.
0][21] As a biomimetic approach, 2,22 we have developed several chiral building blocks and related selective reactions.One of such building block-based methodology is protected (S)malimide (4) 23 and the regio-and trans-diastereoselective reductive alkylation.In 2009, the report of (−)streptopyrrolidine (11) as a natural product 11 prompted us to develop a method for the cis-selective reductive alkylation of malimide and achieved the first total synthesis of this natural product. 23We report herein the first enantioselective total synthesis of natural product 1, which allowed revealing its optical rotation as levorotatory.

Results and Discussion
Because 1 is an analog of (−)-streptopyrrolidine (11), hence the synthetic route developed for the synthesis of 11 23 was adopted.Thus, our synthesis started with Grignard addition (Scheme 1).Treatment of the known N,O-bis-protected (S)-malimide 14 23 with n-octyl magnesium bromide in CH2Cl2 at −15 C for 45 minutes yielded regioselectively (4S)-hemiaminal 15 as a diastereomeric mixture.Because the subsequent dehydration reaction was supposed to run through N-acyliminium intermediate A, the diastereomeric mixture, without separation, was subjected to the TsOH•H2O-catalyzed (5 mol %) dehydration reaction (CH2Cl2, 0 °C to RT) to give enamide 16 in 62% yield over two steps.Compound 16 was obtained as the sole observable regio and (E)geometric isomer.The latter was tentatively assigned by analogy with our previous results. 23Subjecting enamide 16 to 10% Pd/C catalyzed hydrogenation in the presence of NH4OAc afforded N,O-bis-protected hydroxylactam 17 as a single diastereomer (dr > 20:1, determined by 1 H NMR spectroscopy of the crude reaction products) in 73% yield.Exposing 17 to ceric ammonium nitrate (CAN) in a mixed solvent system [MeCN/H2O = 9:1 (v/v)] allowed cleaving the N-protecting group (PMB) to give 18 in 85% yield.Finally, under 10% Pd/C-catalyzed hydrogenolysis conditions (H2, 2 M HCl), 18 was debenzylated to produce the target molecule 1 in 92% yield {white solid, mp: 82 -83 C; [α]D 20 −31.3 (c 1.0, CHCl3)}.Because our synthesis started from (S)-malimide 14, the absolute configuration of our synthetic product is the natural one: (4S,5S)-1.The © AUTHOR(S) spectral data of our synthetic compound 1 are identical with those reported for the natural product 1.Thus, we have achieved a five-step total synthesis of natural -lactam (4S,5S)-1 from the known chiral building block (S)-14, and revealed the sense of this natural product's optical rotation as levorotatory.Scheme 1. Five-step enantioselective total synthesis of the title natural product.

Conclusions
To summarize, we have disclosed the first enantioselective total synthesis of (4S,5S)-4-hydroxy-5octylpyrrolidin-2-one, achieved in five steps with an overall yield of 35.4% from the known chiral building block (S)-malimide (14).By telescoping three steps, a shorter, three-step total synthesis [five steps from commercially available (S)-malic acid] was accomplished with an overall yield of 29.3%.This work demonstrated once more the versatility of (S)-malimide for the enantioselective synthesis of functionalized, chiral -lactams which are ready precursors of other classes of natural products such as 2,5-disubstituted pyrrolidines [24][25][26][27] and -amino -hydroxy carboxylic acids. 5The work is ongoing in our laboratory, and results will be reported in due course.

Experimental Section
General.All commercially available reagents were used without further purification unless indicated otherwise.Column chromatography was performed on silica gel (300 -400 mesh).Melting points were determined on a Büchi M560 Automatic Melting Point apparatus and are uncorrected.Optical rotation data were measured on an Anton Paar MCP 500 polarimeter at 589 nm.The information of chemical bonds or functional groups contained in the molecule were obtained by infrared spectroscopy on a Nicolet Avatar 330 FT-IR spectrometer.NMR spectra were recorded on a Bruker Avance III 400 with TMS as internal standard at room temperature.The samples were dissolved in CDCl3 or DMSO-d6.Chemical shifts were given in values of δH and δC referenced to residual solvent signals (δH 0.00 ppm for 1 H, δC 77.0 ppm for 13 C in CDCl3, δH 2.50 ppm © AUTHOR(S) for 1 H, δC 13.8 ppm for 13 C in DMSO-d6).High resolution mass spectra (HRMS) were recorded using a Bruker Dalton Esquire 3000 plus mass spectrometer by the ESI method.