We report a series of new in vitro and in vivo data proving the selective antitumor activity of our somatostatin structural derivative, TT-232. In vitro, it inhibited the proliferation of 20 different human tumor cell lines in the range of 50-95% and induced a very strong apoptosis. In vivo was not toxic at a dose of 120 mg/kg of b.w. in mice. Long-term incubation (24 h) of tumor cells with TT-232 caused significant inhibition of tyrosine kinases in good correlation with the apoptosis-inducing effect. The level of p53 or KU86 did not change following TT-232 treatment, suggesting a p53-independent apoptotic effect. Preincubation of human breast cancer cells (MDA-MB-453) with TT-232 for 2 h decreased the growth factor receptor autophosphorylation. All of these data suggest that TT-232 is a promising and selective antitumor agent.

A prospective study on 16 patients with advanced (stage III and IV) prostate cancer was carried out. TNM stage, general clinical status, serum PSA level, the histological type and Gleason’s grade of the tumor were registered. Total androgen blockade or single-drug therapy (flutamide) was performed. On average, 4.81 months after the start of therapy rebiopsy, serum PSA determination and general clinical examination were performed. Histologic examination before and after treatment of HE-stained slides, as well as apop-tag reaction to show apoptotic cells, p53, bcl₂, and Ki-67 immunostaining. Clinical improvement manifested by regression or lack of progression was observed in 10 patients. Increase of the apoptotic index and decrease of the mitotic index was detected in these cases. Serum PSA level decreased in all patients except in 3 fatal cases. The 6 clinically nonresponders who died after the second biopsy did not show an increased apoptotic or decreased mitotic index. Ki-67 positivity correlated well with the mitotic activity. Mutant p53 expression was higher in patients in whom antiandrogen therapy was ineffective. The bcl₂ expression was a characteristic of the tumors of patients who later died. These results show that the degree of induction of apoptosis in prostate carcinoma by hormonal therapy varies from case to case. A given prostate cancer patient’s response to therapy may be predicted by following apoptotic and mitotic activity, as well as Ki-67 and p53 expression in repeated biopsies.

The programmed cell death inducing effect of the EGF receptor tyrosine kinase inhibitor alpha-cyano-3,4-dihydroxycinnamthioamide (AG213) was investigated in vitro on HT-29 human colon tumor. AG213 at concentrations between 45 to 450 microM blocks the proliferation of HT-29 cells. Morphological findings suggest that the selective tyrosine kinase inhibitor AG213 induces Clarke III type (non-lysosomal vesiculate cytoplasmic) programmed cell death; unlike ATP analog non-selective tyrosine kinase inhibitors like Genistein which were found to induce apoptosis. Cycloheximide and Actinomycin-D reduced the effect of AG213 pointing to the fact that protein and RNA synthesis are also needed for this form of cell death. Acid phosphatase activity was found in the Golgi and in the newly formed intracytoplasmic vacuoles 3 hours after AG213 treatment which disappeared by 6 hours. The induction of Clarke III cell death by tyrosine kinase inhibitors may open a new modality to selective killing of tumor cells.