The p53 tumor suppressor gene is thought to be required for the induction of programmed cell death (apoptosis) initiated by DNA damage. We show here, however, that the human promyelocytic leukemia cell line HL-60, which is known to be deficient in p53 because of large deletions in the p53 gene, can be induced to undergo apoptosis following X-irradiation. We demonstrate that the decision to undergo apoptosis in this cell line appears to be made at a G 2 checkpoint. In addition, we characterize an HL-60 variant, HCW-2, which is radioresistant. HCW-2 cells display DNA damage induction and repair capabilities identical to those of the parental HL-60 cell line. Thus, the difference between the two cell lines appears to be that X-irradiation induces apoptosis in HL-60, but not in HCW-2, cells. Paradoxically, HCW-2 cells display high levels of expression of bax, which enhances apoptosis, and no longer express bcl-2, which blocks apoptosis. HCW-2 cells' resistance to apoptosis may be due to the acquisition of expression of bcl-x L , a bcl-2-related inhibitor of apoptosis. In summary, apoptosis can be induced in X-irradiated HL-60 cells by a p53-independent mechanism at a G 2 checkpoint, despite the presence of endogenous bcl-2. The resistance shown by HCW-2 cells suggests that bcl-x L can block this process.

We have recently shown that the plant alkaloid 20(S)-camptothecin and its derivatives 9-nitro-20(S)-camptothecin(9NC) and 9-amino-20(S)-camptothecin(9AC) inhibit the growth of a variety of human tumors xenografted in nude mice. In this report, we demonstrate that 9NC and 9AC effectively inhibit growth, and subsequently induce regression, of human ovarian tumors grown in nude mice. Tumor regression is accompanied by degenerative changes in the tumor cells as assessed by microscopic observations of histological sections prepared from the tumors. Parallel experiments in vitro show that 9NC inhibits in a similar manner the growth of human ovarian carcinoma cells, regardless of their ability to induce tumors when xenografted in nude mice, and induces similar morphological changes in both non-tumorigenic and tumorigenic cells, as assessed by microscopic observation. Flow cytometry studies show that 9NC-induced growth inhibition of the non-tumorigenic cells is associated with accumulation of these cells in G2. In contrast, 9NC-induced growth inhibition of the tumorigenic cells is associated with the generation of cells containing a reduced DNA content, that is, cells programmed to die. In conclusion, camptothecins appear to be cytostatic for non-tumorigenic, but cytotoxic for tumorigenic cells, an important finding from viewpoints of cell biology, pharmacology and cancer chemotherapy.

The cytotoxicity of camptothecin (CPT) esters 1-6 was measured. Like parental camptothecin, esters 2 and 3, but not 1, 4, 5, and 6, inhibited proliferation of human leukemia cells in culture and induced programmed cell death as assessed by flow cytometry studies. Exhibition of similar levels of antiproliferative activities of CPT 2 and 3 required different incubation time periods in cell cultures, with CPT and 3 requiring the shortest and longest periods, respectively. Both 2 and 3 were inactive against cells resistant to the semisynthetic CPT derivative 9-nitrocamptothecin and unable to stabilize DNA-topoisomerase I (Topo I) "cleavable complexes" in a cell-free system, suggesting that Topo I activity was required but insufficient for the mechanism of action of 2 and 3. Mouse liver homogenate converted esters to parental CPT, but the conversion rates were different with different esters. Of four tested esters in this experiment, ester 2 had the fastest conversion rate. In vivo studies showed that ester 2 had an exceptional lack of toxicity in nude mice, even at enormous doses, and demonstrated extensive activity against human breast and colon tumors grown as xenografts in immunodeficient nude mice, whereas no antitumor activity was observed for the other esters. In conclusion, ester 2 is a prodrug of the antitumor compound CPT, and it can be administered at very high doses in mice with no appearance of toxicity. This study provides a basis for further evaluation of CPT ester 2 as an investigational anticancer agent.