Antisense-mediated downregulation of anti-apoptotic proteins induces apoptosis and sensitizes head and neck squamous cell carcinoma cells to chemotherapy

Abstract

We have earlier reported that the inhibition of apoptosis in head and neck squamous cell carcinomas (HNSCC) is because of upregulated expression of Bcl-2, Bcl-X L and Survivin. Hence, we addressed the question whether antisense approach towards these inhibitors of apoptosis could restore the apoptosis in HNSCC. Further, we wanted to see whether chemotherapeutic efficacy of Cisplatin and Etoposide could be enhanced by using these drugs in combination with antisense oligonucleotides in human laryngeal carcinoma HeP2 and tongue carcinoma Cal27 cells. The effect of these antisense oligonucleotides was examined on the mRNA expression by RT-PCR and on protein expression by Western blotting. Apoptosis was measured by flowcytometry, TUNEL assay and caspase-3 activity assay. Treatment of HeP2 and Cal27 cells with 400nM antisense oligonucleotides against Bcl-2, Bcl-X L and Survivin for 48 hrs decreased their expression both at the mRNA as well as at the protein level, resulting in the induction of apoptosis. Treatment of HeP2 and Cal27 cells with these antisense oligonucleotides augmented Cisplatin and Etoposide induced apoptosis. Our findings emphasize the importance of Bcl-2, Bcl-X L and Survivin as survival factors in HNSCC cells. Antisense treatment against these survival factors in combination with lower doses of chemotherapy offers potential as a less toxic chemoadjuvant therapy.

Apoptosis in cancer: from pathogenesis to treatment

“…It was shown that transfection of anti-sense Survivin into YUSAC-2 and LOX malignant melanoma cells resulted in spontaneous apoptosis in these cells [90]. The anti-sense approach has also been applied in head and neck squamous cell carcinoma and reported to induce apoptosis and sensitise these cells to chemotherapy [91] and in medullary thyroid carcinoma cells, and was found to inhibit growth and proliferation of these cells [92]. Another approach in targeting Survivin is the use of siRNAs, which have been shown to downregulate Survivin and diminish radioresistance in pancreatic cancer cells [93], to inhibit proliferation and induce apoptosis in SPCA1 and SH77 human lung adenocarcinoma cells [94], to suppress Survivin expression, inhibit cell proliferation and enhance apoptosis in SKOV3/DDP ovarian cancer cells [95] as well as to enhance the radiosensitivity of human non-small cell lung cancer cells [96].…”

Survivin as a Preferential Target for Cancer Therapy

“…This led to decreased cell proliferation and increased caspase-dependent apoptosis in different tumor (lung, sarcomas, lymphomas, thyroid, head, and neck) cell lines [92–96]. Furthermore, down-regulation of survivin via AO was shown to enhance sensitivity to cytotoxic agents such as TRAIL [61,63], cisplatin [93], taxol [97], imatinib [98], etoposide [93,99,100], as well as to cytotoxicity induced by radiation therapy [101]. Moreover, treatment with survivin AO resulted in inhibition of tumor growth in murine models [102].…”

Survivin-directed Anticancer Therapies – A Review of Pre-clinical Data and Early-phase Clinical Trials

“…Transfection with the ASO triggered spontaneous apoptosis, and two-colour flow cytometry confirmed decreased endogenous survivin expression. Other pre-clinical studies confirmed these results 47,48 and found that survivin downregulation sensitises certain human cancer cell lines to cisplatin and etoposide, 49 as well as to radiotherapy. 50 LY-2181308 is a second-generation ASO.…”