Relative refractoriness of breast cancer cells to tumour necrosis factor-α induced necroptosis

Necroptosis, a recently identified programmed cell death pathway, has attracted attention as an alternative route to target apoptosis-resistant cancer cells. The status of the necroptosis pathway in different subtypes of breast cancer has not been well explored. Stimulating the cells by TNF-α can trigger cell survival or death depending on the combination of downstream players involved. In this work, we attempted to induce necroptosis in them using a combination of TNF-α and Z-VAD-FMK with and without chemotherapy. Cell viability, apoptosis, and necroptosis were assessed using MTT and Annexin-V/PI assays, respectively. Gene and protein expression was analysed by qPCR and immunophenotyping. Both the cell lines were resistant to induction of cell death by necroptosis. There was no enhancement in cell death when chemotherapeutic drugs were combined with necroptosis induction. Expression studies showed reduced translational expression of key necroptosis molecules like RIP kinases and MLKL in breast cancer cells compared to positive control cell line L929. Also, cell survival molecules were expressed more in MDA-MB-231 in contrast to death pathway molecules which were expressed more in T47D cells. In this work, the two breast cancer cell lines were observed to be resistant to TNF-α induced necroptosis with or without chemotherapy. Expression of key necroptosis players revealed relative insufficiency of the molecular machinery involved in the above pathway. In our opinion this may be the cause for resistance to necroptosis and novel strategies to upregulate these molecules need to be developed to sensitize the breast cancer cells towards cell death by necroptosis.