Dolastatin 10 (1) is a potent antineoplastic pentapeptide. Novel dolastatin 10 analogs each modified at one of the constituent amino acid derivatives, were synthesized and their antitumor activity was evaluated against P388 leukemia in mice. The structural requirements for antitumor activity are discussed. Some of the analogs, 31c, 35c, 38b, and 50c showed excellent activity in vivo. Highly active 50c, which lacks the thiazole group of 1, was selected for further development as an antitumor agent.

Dolastatin 10, a pentapeptide isolated from the marine mollusk Dolabella auricularia, has antitumor activity. TZT‐1027, a dolastatin 10 derivative, is a newly synthesized antitumor compound. We evaluated its antitumor activity against a variety of transplantable tumors in mice. Intermittent injections of TZT‐1027 were more effective than single or repeated injections in rake with P388 leukemia and B16 melanoma. Consequently, TZT‐1027 shows schedule dependency. TZT‐1027 was effective against P388 leukemia not only when administered i.p., but also when given i.v. However, although TZT‐1027 given i.v. was active against murine solid tumors, TZT‐1027 administered i.p. was ineffective against all the tumors tested with the exception of colon 26 adenocarcinoma. The i.v. injection of TZT‐1027 at a dose of 2.0 mg/Ag remarkably inhibited the growth of three murine solid tumors; colon 26 adenocarcinoma, B16 melanoma and M5076 sarcoma, with T/C values of less than 6%. The antitumor activities of TZT‐1027 against these tumors were superior or comparable to those of the reference agents; dolastatin 10, cisplatin, vincristine, 5‐fluorouracil (5‐FU) and E7010. In experiments with drug‐resistant P388 leukemia, TZT‐1027 showed good activity against cisplatin‐resistant P388 and moderate activity against vincristine‐ and 5‐fluorouracil‐resistant P388, but no activity against adriamycin‐resistant P388. TZT‐1027 was also effective against human xenografts, that is, tumor regression was observed in mice bearing MX‐1 breast and LX‐1 lμng carcinomas. TZT‐1027 at 10 μM almost completely inhibited the assembly of porcine brain microtubules. Therefore, its mechanism of antitumor action seems to he, at least in part, ascrihable to the inhibition of microtubule assembly. Because of its good preclinical activity, TZT‐1027 has been entered into phase I clinical trials.

TZT-1027, a dolastatin 10 derivative, is an antimicrotubule agent with potent antitumor activity both in vitro and in vivo. In this study, we performed biochemical and histopathological examinations, and evaluated TZT-1027-induced tumoral vascular collapse and tumor cell death in an advanced tumor model, murine colon 26 adenocarcinoma. In addition, we studied the effects of TZT-1027 on cultured human umbilical vein endothelial cells (HUVEC). Tolerable doses of TZT-1027 induced tumor-selective hemorrhage within 1 h. This hemorrhage occurred mainly in the peripheral area of the tumor mass. Measurements of tumoral hemoglobin content and dye permeation revealed that the hemorrhage occurred firstly and tumor blood flow stopped secondarily. The vascular damage was followed by continuous induction of apoptosis of the tumor cells, tumor tissue necrosis, and tumor regression. In cultured HUVEC, TZT-1027 induced marked cell contraction with membrane blebbing in 30 min. These cell changes were completely inhibited by K252a, a broad-spectrum inhibitor of protein kinases. These effects of TZT-1027 on both tumor vasculature and HUVEC were greater than those of vincristine. In conclusion, TZT-1027 quickly attacked the well-developed vascular system of advanced tumors by a putative protein kinase-dependent mechanism, and then blocked tumor blood flow. Therefore, TZT-1027 has both a conventional antitumor activity and a unique anti-tumoral vascular activity, making it a potentially powerful tool for clinical cancer therapy.

TZT-1027 (Soblidotin), an antimicrotubule agent, has been demonstrated to show potent antitumor effects, though the relationships among antitumor effect, cytotoxicity and anti-vascular effect of TZT-1027 have not been studied. We established in vivo human lung vascular-rich tumor models using a vascular endothelial growth factor-secreting tumor (SBC-3/VEGF). SBC-3/VEGF tumors exhibited a high degree of angiogenesis in comparison with the mock transfectant (SBC-3/Neo) tumors in a dorsal skinfold chamber model and grew much faster and larger than SBC-3/Neo tumors in the tumor growth study. The antitumor activity of antimicrotubule agents, including TZT-1027, was evaluated in both early-and advanced-stage SBC-3/Neo and SBC-3/VEGF tumor models to elucidate the relationship between the antitumor activity and anti-vascular effect of these agents. TZT-1027 exhibited potent antitumor activity against both early-and advanced-stage SBC-3/Neo and SBC-3/VEGF tumors, whereas combretastatin A4 phosphate did not. Vincristine and docetaxel exhibited potent antitumor activity against early-stage SBC-3/Neo and SBC-3/VEGF tumors, and advanced-stage SBC-3/Neo tumors, but did not exhibit activity against advanced-stage SBC-3/VEGF tumors. The difference in antitumor activity between these agents could be ascribed to differences in direct cytotoxicity and anti-vascular effect. Furthermore, a prominent accumulation of erythrocytes in the tumor vasculature, followed by leakage and scattering of these erythrocytes from the tumor vasculature, was observed after TZT-1027 administration to mice bearing advanced-stage SBC-3/VEGF tumors. These findings strongly suggest that TZT-1027 has a potent anti-vascular effect, in addition to direct cytotoxicity. (Cancer Sci 2003; 94: 826-833) ngiogenesis is the process whereby capillaries sprout from pre-existing blood vessels. The overall process is complicated, involving many cell types and biological functions, including endothelial cell activation, migration and proliferation.1) Previous studies have demonstrated that tumor angiogenesis is required for the growth and metastasis of primary solid tumors.2) This angiogenesis is influenced by the balance between the activation of promoting and inhibiting factors. The activation of promoting factors, such as basic fibroblast growth factor (bFGF) or vascular endothelial growth factor (VEGF), released from hypoxic tumor cells, leads to neovascularization, while the activation of inhibiting factors generally causes tumors to undergo necrotic cell death.3) The importance of angiogenesis in human tumors was proved by recent reports demonstrating the vessel density in the tumor mass to be a prognostic factor for survival in people with various types of tumor. [4][5][6][7][8]

TZT‐1027, a derivative of dolastatin 10 isolated from the Indian Ocean sea hare Dolabella auricularia in 1987 by Pettit et al. , is a potent antimicrotubule agent. We have compared the activity of TZT‐1027 with that of dolastatin 10 as well as the vinca alkaloids vinblastine (VLB), vincristine (VCR) and vindesine (VDS). TZT‐1027 and dolastatin 10 inhibited microtubule polymerization concentration‐dependently at 1–100 μ M with IC 50 values of 2.2±0.6 and 2.3±0.7 μ M , respectively. VLB, VCR and VDS inhibited microtubule polymerization at 1–3 μ M with IC 50 values of 2.7±0.6, 1.6±0.4 and 1.6±0.2 μ M , respectively, but showed a slight decrease in inhibitory effect at concentrations of 10 μ M or more. TZT‐1027 also inhibited monosodium glutamate‐induced tubulin polymerization concentration‐dependently at 0.3–10 μ M , with an IC 50 of 1.2 μ M , whereas VLB was only effective at 0.3–3 μ M , with an IC 50 of 0.6 μ M , and caused so‐called “aggregation” of tubulin at 10 μ M. Scatchard analysis of the binding data for [ 3 H]VLB suggested one binding site (K d 0.2±0.04 μ M and B max 6.0±0.26 n M /mg protein), while that for [ 3 H]TZT‐1027 suggested two binding sites, one of high affinity (K d 0.2±0.01 μ M and B max 1.7±0.012 n M /mg protein) and the other of low affinity (K d 10.3±1.46 μ M , and B max 11.6±0.83 n M /mg protein). [ 3 H]TZT‐1027 was completely displaced by dolastatin 10 but only incompletely by VLB. [ 3 H]VLB was completely displaced by dolastatin 10 and TZT‐1027. Furthermore, TZT‐1027 prevented [ 3 H]VLB from binding to tubulin in a non‐competitive manner according to Lineweaver‐Burk analysis. TZT‐1027 concentrationdependently inhibited both [ 3 H]guanosine 5′‐triphosphate (GTP) binding to and GTP hydrolysis on tubulin. VLB inhibited the hydrolysis of GTP on tubulin concentration‐dependently to a lesser extent than TZT‐1027, but no inhibitory effect of VLB on [ 3 H]GTP binding to tubulin was evident even at 100 μ M . Thus, TZT‐1027 affected the binding of VLB to tubulin, but its binding site was not completely identical to that of VLB. TZT‐1027 had a potent inhibitory effect on tubulin polymerization and differed from vinca alkaloids in its mo...