INDUCTION OF APOPTOSIS BY ORGANOTIN COMPOUNDS IN-VITRO - NEURONAL PROTECTION WITH ANTISENSE OLIGONUCLEOTIDES DIRECTED AGAINST STANNIN

Immortalized cell lines and primary neuronal cultures were used to cha racterize the selective toxicity of trimethyltin (TMT), triethyltin (T ET) and tributyltin (TBT). TBT and TET were cytotoxic at similar conce ntrations in the immortalized cell lines tested; the 50% toxic concent ration (TC50) was 1 to 11 mu M. In contrast, immortalized cell lines v aried considerably in their sensitivity to TMT, with sensitive cell li nes (neuroblastomas, T-, B-cell lines) showing TC50 values of 2 to 8 m u M, whereas insensitive cells (NIH-3T3 fibroblast, HTB-14 glioma, TC- 7 kidney cells) had TC50 values > 100 mu M. Primary neuronal cell cult ures were very sensitive to organotins (TC50 similar to values, 1-10 n M), and showed patterns of selective toxicity with respect to neuronal and glial cells. Because organotin toxicity evolves over 24 to 48 hr, we determined whether these compounds induced apoptosis in primary cu ltures. TMT increased (P < .05) the fraction of apoptotic cells 6 and 12 hr after treatment with TMT at TC50 concentrations. Prior studies s uggested that a protein, stannin, was localized in cells sensitive to organotins, Stannin was expressed in several TMT-sensitive cell lines (PC12, T, B cells) and in primary neurons in culture. Stannin was abse nt in the resistant HTB-14 glioma cell line, The role of stannin in me diating TMT toxicity in primary cultures was investigated by blocking stannin expression with specific antisense oligonucleotides. Treatment of primary cultures with antisense oligonucleotides for 48 hr before and during TMT treatment significantly protected neurons from the neur otoxic and apoptotic effects of TMT, This effect was not observed with scrambled oligonucleotide controls. Thus, TMT may induce apoptosis in sensitive cells, which is partly mediated by stannin. Based on the av ailable data, we conclude that stannin expression is necessary, but no t sufficient for TMT toxicity.