Cytotoxic Mechanisms of FK317, a New Class of Bioreductive Agent with Potent Antitumor Activity

Abstract

FK317 is a member of a new class of bioreductive agents that exhibit strong cytotoxicity against various human cancer cells. The effect of FK317 was found to be stronger than that of mitomycin C (MMC), adriamycin (ADR) or cisplatin (CDDP). Alkaline elution analysis indicated that FK317 formed interstrand DNA-DNA and DNA-protein cross-links in cells. On the other hand, no DNA single-strand breaks were observed in the cells treated with FK317. In a cell-free system the deacetylated metabolites produced cross-linked DNA under reductive conditions, though FK317 itself did not form DNA-DNA cross-links. In order to elucidate the metabolic activation mechanisms, we established an FK317-resistant subline from human non-small cell lung cancer cells (Lu99) by stepwise and brief exposure (1 h) to FK317. The resistant subline (Lu99/317) showed cross-resistance to MMC and carboquone (CQ), but not to ADR or CDDP. DT-diaphorase, which is one of the activation enzymes of MMC and CQ, was deficient in Lu99/317 cells as determined by enzyme activity assay. However, the levels of NADPH:cytochrome P450 reductase, which is another activation enzyme for MMC and CQ, were comparable in resistant and parent cell lines. Treatment of the cells with dicumarol, an inhibitor of DT-diaphorase, reduced the cytotoxicity of FK317 to Lu99 cells, but not to Lu99/317 cells. These results indicate that deacetylation of FK317 is necessary for its reductive activation, and deacetylated FK317 is reduced by DT-diaphorase to form an active metabolite, which produces DNA-DNA interstrand and DNA-protein cross-links that lead to cell death.Key words: FK317 -Antitumor effect -Bioreductive agent -DT-diaphorase -Deacetyl metabolite FK317 is a new analog of FK973, which we previously showed to exhibit strong antitumor activities in a wide variety of animal tumor models and human xenografts, and has strong cytotoxic effects against in vitro-cultured tumor cell lines.1) FK973 has also been shown to have high therapeutic efficacy in clinical studies. However we were unable to develop this compound because it induces vascular leak syndrome (VLS) as a major side effect. 2, 3)In attempts to synthesize new FK973 derivatives which retain the antitumor activity, without the VLS side effect, FK317, 11-acetyl-8-carbamoyloxymethyl-4-formyl-6-methoxy-14-oxa-1,11-diazatetracyclo[7.4.1.0 2, 7 .0 10, 12 ]tetradeca-2,4,6,-trien-9-yl acetate, was identified. This agent has a unique bicyclic hydroxylamine hemiketal ring system, and its antitumor activity is more potent than, or equivalent to, that of mitomycin C (MMC), adriamycin (ADR), cisplatin (CDDP), taxol or irinotecan hydrochloride against murine tumors and human xenografts in mice. Moreover, its hematotoxic and myelosuppressive effects were weaker than those of MMC in mice. These preclinical data suggested that FK317 might represent a major advance in the treatment of cancer.The mechanisms by which FK317 induces these potent antitumor effects have not yet been defined, but if more clearly understood, would offer valuable in...

Enantioselective Total Synthesis of FR900482

“…This compound possesses potent antitumor activity against a number of tumor cell lines, including mitomycin C- and vincristine-resistant P388 leukemia cells . In extensive efforts to develop more potent and less toxic agents, FK973 ( 3 ) and FK317 ( 4 ), semisynthetic derivatives from 1 , were found to be promising clinical candidates. The mode of action of this class of compounds has recently been proven through detailed mechanistic studies .…”

Effects of Photochemically Activated Alkylating Agents of the FR900482 Family on Chromatin

“…6897, is structurally and functionally related to MMC [9,10] but was dropped from clinical development in Phase I studies due to the development of vascular leak syndrome in patients. More recently, a semi-synthetic derivative, FK317, has exhibited promising antitumor activity [10][11][12][13][14][15] and has advanced from Phase I to Phase II human clinical trials. FK317 (4, Figure 1A) shares structural similarities and properties with FR900482 (2, Figure 1A), but exhibited fewer side effects, in particular with respect to vascular leak syndrome, when compared to FR900482 [11,16].…”

Synthesis of 7-Epi (+)-FR900482: An Epimer of Comparable Anti-Cancer Activity

“…Naoe has shown a connection between the metallo-enzyme DT-diaphorase and cytotoxicity for FK317 (Scheme ). In this case, exposure of cells to dicumarol, an inhibitor of this enzyme significantly lowers cytotoxicity; furthermore, FK317 is more active against cells expressing this enzyme.…”