Student Research Paper
Cisplatin is the chemotherapeutic of choice to treat several cancers, notably cervical and esophageal, which are commonly induced by the tumor virus HPV. Cisplatin’s mechanism of action is to create intra-strand and inter-strand linkages between DNA; however some tumors have been shown to have developed resistance. It has been postulated that resistance to cisplatin can be linked to BIN1 expression levels or the isoforms created as the result of splicing by SRSF1. BIN1 (bridging integrator-1) is involved in many functions of the cell, but most importantly it regulates the cell cycle and apoptosis through its tumor suppressor characteristics. To help elucidate the mechanisms involved in cancer induction by tumor viruses and help further chemotherapeutic treatment options, this research was undertaken to determine if cisplatin was effective against cells transformed with the Simian Virus 40 (SV40) T antigen and to determine if the mechanism of action is linked to BIN1 expression levels. To accomplish this, human diploid fibroblast immortalize with telomerase [HDF(tert)] were transfected with a plasmid encoding the SV40 T antigen; and, two distinct clones were expanded [HDF(tert)+T Clone 1 and HDF(tert)+T Clone 2]. All cells were exposed to cisplatin and growth was assessed. It was found that cells expressing the oncoprotein were more sensitive to cisplatin than the parental cell line. To determine if the parental line resistance to cisplatin correlated to BIN1, expression level and isoform production was assessed. This research found that there is no difference in the accumulated levels of BIN1 protein being expressed among the 3 cell lines as determined by Western blotting. However, RT-PCR revealed multiple isoforms being expressed. The predominant form in all cell lines appeared to be isoform 10 which correlates to the 456 base pair band and the 45 kDa protein band. Interestingly, a new, ninth unpublished isoform was isolated which was present in both the immortalized parental line [HDF(tert)] and the transfected cell lines [HDF(tert)+T Clone 1 and HDF(tert)+T Clone 2]. Overall this data indicated that BIN1 expression is not solely responsible for cisplatin effectiveness. T-antigen-oncoprotein-expressing-cells were killed up to 90%; yet, BIN1 levels were equal to that of the non-transformed line. Whereas the non-transformed cells exhibited 50% survival. It is important to determine the mechanism for cisplatin sensitivity in order to apply it to make cancer cells less resistant to treatment.
Monostra, Carli, "Chemotherapeutic sensitivity and the expression of BIN1 in Human Diploid Fibroblasts expressing SV40 T-antigen" (2020). Biology: Student Scholarship & Creative Works. 8.