Mitomycin A

A set of 30 mitomycin C and mitomycin A analogues, including five new compounds, was screened against three different solid human tumor cell lines using the MTT tetrazolium dye assay. A statistically significant correlation among antitumor activity, quinone reduction potential (E1/2), and the logarithm of the partition coefficient (log P) was obtained, with the most easily reduced and the most lipophilic compounds being the most potent. When these analogues were separated into mitomycin C and mitomycin A subsets, the former gave a correlation only with E1/2, whereas the latter (which differ little in their E1/2 values) gave a correlation only with log P. These correlations are in contrast to those made in the P388 leukemia assay in mice wherein the most active mitomycin C and mitomycin A analogues were the most hydrophilic ones. When the same compounds were tested against P388 leukemia cells in the MTT assay, the results were the same as those of the solid tumor assays. Thus, the substantial differences in relative potencies of mitomycins are related not to the kind of tumor cell, but to the type of assay performed, cell culture versus whole animal. No correlation was found between antitumor potency in the cell culture systems and calculated relative DNA binding strengths, probably because the limiting factors in antitumor potency of mitomycins appear to be tumor cell uptake (log P) and/or bioreductive activation (E1/2).

Over 600 analogues of mitomycin C (MMC) have been made in the past and more recently a number of Mitomycin A (MMA) derivatives have been prepared. Since many of the MMA type had the same organic side chain at the 7-position as previously prepared MMC analogues it was of interest to see if the biological effects of MMCs could predict for those of MMAs. Using the P388 leukemia model it was possible to compare the activity of 27 matched pairs and the potency of 24 pairs. It was found that antitumor effects did not correlate but that MMAs were significantly more potent than MMCs. These findings were duplicated in tests of 7 pairs against subcutaneously implanted B16 melanoma. We conclude that any MMA derivative would have a high likelihood of being more potent than its MMC equivalent but that its antitumor effects must be independently determined since they cannot be predicted from the results with MMC analogs.

Dimeric DNA alkylating agents have drawn significant interest because these compounds are expected to provide at least two reactive sites and as a result, generate enhanced levels of DNA interstrand cross-link (DNA ISC) adducts compared to their monomeric agents. We report the synthesis and mechanistic studies of a novel mitomycin dimer, 7-N,7'-N'-(1″,2″-dithiocanyl-3″,8″-dimethylenyl)bismitomycin C (8) connected by an eight-membered cyclic disulfide. Mitomycins require prior activation (i.e., transformation to a good electrophile) for DNA adduction and therefore, 8 was aimed to undergo facile nucleophilic activation and produce enhanced levels of DNA ISC. At the core of this function lies a cyclic disulfide in 8. It was expected that disulfide cleavage by an appropriate nucleophile would successively produce two thiols that may trigger activation of two mitomycin rings in a dimer through intramolecular cyclization to quinine rings. Compound 8 was synthesized from mitomycin A (1) and the key intermediate, cyclic disulfide (11), along with the reference diol mitomycin 7-N,7'-N'-(2″,7″-dihydroxy-1″,8″-octanediyl)bismitomycin C (23) which does not contain the disulfide unit. We found that 8 underwent significantly enhanced nucleophilic activation in the presence of Et(3)P compared with 23, and that the disulfide unit in 8 played a key role for the nucleophilic activation. Based on these findings, we proposed a mechanism for nucleophilic activation of 8. We further demonstrated that 8 generated much higher levels of DNA ISC (94%) compared with 23 (4%) and 2 (3%) in the presence of Et(3)P (and L-DTT) leading to the conclusion that 8 is more efficient for DNA ISC processes than 23 and 2 due to the role of disulfide unit.