Total synthesis of a key series of vinblastines modified at C4 that define the importance and surprising trends in activity

Abstract

The total synthesis and evaluation of a key systematic series of vinblastines that incorporate the first deep-seated changes to the substituent at C4 are detailed. The synthetic approach features an expanded and redefined scope of a 1,3,4-oxadiazole [4 + 2]/[3 + 2] cycloaddition cascade in which electronically mismatched electron-deficient trisubstituted alkenes and unactivated trisubstituted alkenes were found to productively initiate the cycloaddition cascade with tethered electron-deficient 1,3,4-oxadiazoles. Such cycloaddition cascades were used to directly introduce altered C4 substituents, providing the basis for concise total syntheses of a series of C4 modified vindolines and their subsequent single-step incorporation into the corresponding synthetic vinblastines in routes as short as 8–12 steps. Evaluation of the synthetic vinblastines revealed a surprisingly large impact and role of the C4 substituent on activity even though it was previously not thought to intimately interact with the biological target tubulin. Only the introduction of a C4 methyl ester, a constitutional isomer of vinblastine in which the carbonyl carbon and ester oxygen of the C4 acetate are transposed, provided a synthetic vinblastine that matched the potency of the natural product. In contrast, even introduction of a C4 acetamide or N-methyl carboxamide, which incorporate single heavy atom exchanges (amide NH for ester oxygen) in vinblastine or the C4 methyl ester, provided compounds that were ≥10-fold less active than vinblastine. Other C4 acetate replacements, including a C4 amine, carboxylic acid, hydroxymethyl or acetoxymethyl group, led to even greater reductions in potency. Even replacement of the C4 acetoxy group or its equally active C4 methyl ester with an ethyl or isopropyl ester led to 10-fold or more reductions in activity. These remarkable trends in activity, which correlate with relative tubulin binding affinities, retrospectively may be ascribed to the role the substituent serves as a H-bond acceptor for α-tubulin Lys336 and Asn329 side chains at a site less tolerant of a H-bond donor, placing the methyl group of the C4 acetate or C4 methyl ester in a spatially restricted and well-defined hydrophobic half pocket created by a surrounding well-ordered loop. This remarkable impact of the C4 substituent, its stringency, and even the magnitude of its effect are extraordinary, and indicate that its presence was selected in Nature to enhance the effects of vinblastine and related natural products.

Inverse Electron Demand Diels–Alder Reactions of Heterocyclic Azadienes, 1-Aza-1,3-Butadienes, Cyclopropenone Ketals, and Related Systems. A Retrospective

“…In the course of these efforts, we comprehensively defined the importance and role of nearly every atom and substituent in vinblastine and vincristine, including the vindoline C4 acetate, C5 ethyl substituent, C6–C7 double bond, and the vindoline core structure itself, and have systematically explored the upper catharanthine-derived C20′ ethyl substituent, C16′ methyl ester, added C10′ or C12′ indole substitutions, and, most recently, the C20′ alcohol . We have shown that the latter is remarkably tolerant to replacement, resulting in the discovery of both ultrapotent analogues of the natural products as well as those that match the potency of the natural products but do not suffer from overexpressed Pgp-derived resistance (Figure ).…”

“…Our efforts focused on the use of ( Z )- and ( E )- 178 , bearing isomeric trisubstituted electron-rich enol ethers as the tethered initiating dienophiles. Trisubstitution of the dienophile generally slows initiation of the intramolecular [4 + 2]/[3 + 2] cycloaddition cascade, although recent studies have defined reaction conditions that permit the use of even more difficult trisubstituted electron-deficient and unactivated alkenes . The exceptions to this generalization are trisubstituted olefins bearing an electron-donating substituent that serves to activate the dienophile for participation in an 1,3,4-oxadiazole inverse electron demand Diels–Alder reaction.…”

Vinblastine 20′ Amides: Synthetic Analogues That Maintain or Improve Potency and Simultaneously Overcome Pgp-Derived Efflux and Resistance

“…28–30 As a result of these developments, we have prepared several series of key analogs, systematically exploring and defining the impact individual structural features and substituents have on tubulin binding affinity and cancer cell growth inhibition. 10,19 Complementary to the studies detailed herein, we have systematically probed the impact and role of the vindoline C4 acetate, 31,32 C5 ethyl substituent, 33 C6–C7 double bond, 34–36 and the vindoline core structure itself, 36 and have systematically explored the upper catharanthine-derived subunit C20’ ethyl substituent, 37,38 C16’ methyl ester, 39 and added C10’ or C12’ indole substitutions. 40 In addition and in preceding studies, we have shown that replacement of the C20'-OH with 20’ ureas was possible, 41 that substantial 42 and even remarkable 43 potency enhancements were obtainable with such 20’ ureas, and that some exhibited further improvements in activity against vinblastine-resistant cancer cell lines.…”

Ultra-potent vinblastine analogues improve on-target activity of the parent microtubulin-targeting compound

“…10 Due to recent advances in the total synthesis of vinblastine, a large number of analogues exploiting strategic changes in the upper catharanthine subunit have been synthesized, evaluated, and disclosed in our efforts. [11][12][13][14][15][16][17][18][19][20][21][22][23][24] Among them, several C20' urea vinblastine analogues (2, 3, 4) displayed remarkable potency against vinblastine-sensitive cell lines (IC 50 50-75 pM; L1210, HCT116) and decreased susceptibility to Pgp efflux-derived resistance in the vinblastine-resistant cell line HCT116/VM46 ( Figure 2). 23 The extended C20' ureas of these analogues continue to bind along and further disrupt the protein-protein interaction at the tubulin / head-to-tail dimer-dimer interface.…”