The MTD of infusional flavopiridol is 50 mg/m2/d x 3 with dose-limiting secretory diarrhea at 62.5 mg/m2/d x 3. With ADP, 78 mg/m2/d x 3 was the MTD, with dose-limiting hypotension at 98 mg/m2/d x 3. Based on chronic tolerability, 50 mg/m2/d x 3 is the recommended phase II dose without ADP. Antitumor effect was observed in certain patients with renal, prostate, and colon cancer, and non-Hodgkin's lymphoma. Concentrations of flavopiridol (200 to 400 nM) needed for cyclin-dependent kinase inhibition in preclinical models were achieved safely.

2-(4-Aminophenyl)benzothiazoles 1 and their N-acetylated forms have been converted to C- and N-hydroxylated derivatives to investigate the role of metabolic oxidation in the mode of action of this series of compounds. 2-(4-Amino-3-methylphenyl)benzothiazole (1a, DF 203, NSC 674495) is a novel and potent antitumor agent with selective growth inhibitory properties against human cancer cell lines. Very low IC(50) values (<0.1 microM) were encountered in the most sensitive breast cancer cell lines, MCF-7 and T-47D, whereas renal cell line TK-10 was weakly inhibited by 1a. Cell lines from the same tissue origin, MDA-MB-435 (breast), CAKI-1 (renal), and A498 (renal), were insensitive to 1a. Accumulation and metabolism of 1a were observed in sensitive cell lines only, with the highest rate of metabolism occurring in the most sensitive MCF-7 and T-47D cells. Thus, differential uptake and metabolism of 1a by cancer cell lines may underlie its selective profile of anticancer activity. A major metabolite in these sensitive cell lines has been identified as 2-(4-amino-3-methylphenyl)-6-hydroxybenzothiazole (6c). Hydroxylation of 1a was not detected in the homogenate of previously untreated MCF-7, T-47D, and TK-10 cells but was readily observed in homogenates of sensitive cells that were pretreated with 1a. Accumulation and covalent binding of [(14)C]1a derived radioactivity was observed in the sensitive MCF-7 cell line but not in the insensitive MDA-MB-435 cell line. The mechanism of growth inhibition by 1a, which is unknown, may be dependent on the differential metabolism of the drug to an activated form by sensitive cell lines only and its covalent binding to an intracellular protein. However, the 6-hydroxy derivative 6c is not the 'active' metabolite since, like all other C- and N-hydroxylated benzothiazoles examined in this study, it is devoid of antitumor properties in vitro.

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The papain-family cathepsins are cysteine proteases that are emerging as promising therapeutic targets for a number of human disease conditions ranging from osteoporosis to cancer. Relatively few selective inhibitors for this family exist, and the in vivo selectivity of most existing compounds is unclear. We present here the synthesis of focused libraries of epoxysuccinyl-based inhibitors and their screening in crude tissue extracts. We identified a number of potent inhibitors that display selectivity for endogenous cathepsin targets both in vitro and in vivo. Importantly, the selectivity patterns observed in crude extracts were generally retained in vivo, as assessed by active-site labeling of tissues from treated animals. Overall, this study identifies several important compound classes and highlights the use of activity-based probes to assess pharmacodynamic properties of small-molecule inhibitors in vivo.