Design And Synthesis Of Antitumor Agents Croptophycin Analogues

The cryptophycins are a family of macrocyclic depsipeptides isolated from terrestrial blue-green algae (Nostoc sp), which are a potent tubulin-binding anti-mitotic agents with excellent activities against multi-drug resistant (MDR) cancer cell lines and mammary derived tumors. Cryptophycin-1 (Cr-1) is the key cytotoxin in Nostoc sp and displays IC50 values in the pM range. More importantly, compared to vinblastine, colchicine, and paclitaxel, the cryptophycin-1 presents reduced susceptibility to P-glycoprotein-mediated multiple drug resistance .By 2005, twenty eight cryptophycin analogues had been isolated, while hundreds of synthetic analogues had been synthesized for the structure-activity relationship (SAR) studies in search for more potent compounds. Some synthetic cryptophycin analogues have demonstrated superior activities and drugable properties to their natural compounds. The closely related synthetic analogue cryptophycin-52 ( LY355703) was identified by Eli Lilly as the pharmacologically most promising first-generation clinical candidate, and is currently in clinical studies as an anti-tumor agent. The low bioavailability in vivo cytoxic assays of cryptophycin-1 causing by their poor stability and poor solubility led some researchers to initiate an attempt to improve it through designing new analogues.Cryptophycins blocks the cell cycle at the G2/M phase apparently through inhibition of tubulin polymerization into microtubules. This compouds binds to a tubulin site distinct from the colchicine site, but one that may overlap with the vinblastine site. Cr-1 induced microtubule depolymerization in cancer cell was blocked by pretreatment with paclitaxel (taxol). It was found that Cr-1 inhibits microtubule formation and dissociates from a tubulin-cryptophycin complex at a very low rate. It is a highly potent stabilizer of microtubule dynamics at low concentrations that have no effect on net microtubule polymerization, an effect that has been demonstrated in many other tubulin binding antimitotic agents and is no doubt the more clinically relevant one. Its extremely low picomolar potency has led to additional studies investigating other possible modes of action.In this work, we planed to resolve the poor stability through introducing a lactam on C4, and introducing polar groups on C6 (for instants: sulfur element of Met; OH of Ser; lactam of Asp et al.) to improve the solubility. Choosing Cr-1 as the leading compound, three series analogues have been synthesized and their in vitro cytotoxicties were evaluated by the MTT assay. A preliminary in vivo evaluation of the cytotoxic activity of analogues was performed at BALB/C-nu mice which implanted SPCA-1. About the synthesis, new tripeptide structures were introduced as the southern hemisphere unit and the northern hemisphere (A unit) was also synthesized through asymmetry method. Brown's crotylboration between crotyl borane and protected aldehyde and Sharpless oxidation were used to building the chiral center of A unit. The reaction conditions and the advance or shortcoming of the two methods were investigated and discussed. Several classical reactions, including Arndt-Eistert rearrangment, Sharpless Oxidation, Swern Oxidation, Wittig reaction, Grigand reaction, Heck reaction and Crotylborylation et al, were applied in this preparation route and investigated sufficiently. The total synthesis include 24 to 27 steps and was interesting and difficult.In addition, other analogues with simpler structure than the leading compoud, which were designed through substituting the A unit byδ-hydroxy-(E)-tert-butyl -pen-2-enoate orδ-hydroxy-tert-butyl 2-acetamido-penanoate were synthesized. The motivation of the design was to optimize the tough synthetic burden and to modify the complex structure of leading compound with simpler unit. By using convenient staring materials and reagents, the analogues have been obtained and the synthetic process was predigested in a certain extent. The reaction conditions and the antitumor activities were also studied. Though the activity of these analogues were not proleptic, the result indicate that the 16 macro-cycle of leading compound is necessary.In summary, the new analogues of Cryptophycin in this paper were exhibited broader cytotoxicity than Taxol and MTX in both solid tumor and leukemia cell lines studied. The results of pharmacological test indicated that the analogues were inhibitors in vivo assay and can delay the growth of tumor. In most cell lines tested, compound NCA-2 was slightly more active than compound NCA-1, and the thio-substitute on C6 of compound NCA-2 was believed to produce a positive effect on the cytotoxic activity. Though more evidence is needed, this encourage observation has led us to design potentially more potent compounds for the anti-cancer screening.