Radiosensitization by a novel Bcl-2 and Bcl-XL inhibitor S44563 in small-cell lung cancer

Radiotherapy has a critical role in the treatment of small-cell lung cancer (SCLC). The effectiveness of radiation in SCLC remains limited as resistance results from defects in apoptosis. In the current study, we investigated whether using the Bcl-2/Bcl-X_L inhibitor S44563 can enhance radiosensitivity of SCLC cells in vitro and in vivo. In vitro studies confirmed that S44563 caused SCLC cells to acquire hallmarks of apoptosis. S44563 markedly enhanced the sensitivity of SCLC cells to radiation, as determined by a clonogenic assay. The combination of S44563 and cisplatin-based chemo-radiation showed a significant tumor growth delay and increased overall survival in mouse xenograft models. This positive interaction was greater when S44563 was given after the completion of the radiation, which might be explained by the radiation-induced overexpression of anti-apoptotic proteins secondary to activation of the NF-κB pathway. These data underline the possibility of combining IR and Bcl-2/Bcl-X_L inhibition in the treatment of SCLC as they underscore the importance of administering conventional and targeted therapies in an optimal sequence.Identifying the mechanisms leading to radioresistance including resistance to apoptosis is essential to improve clinical outcome in cancer patients. Disabled apoptosis has been catalogued among the fundamental hallmarks of cancer¹ and the proteins of the Bcl-2 family play a fundamental role in regulating this modality of cell death. The Bcl-2 family comprises both pro- and anti-apoptotic members; the latter (Bcl-2, Bcl-X_L and Mcl-1) are often overexpressed in cancer cells to facilitate the survival of cells that under normal circumstances should have undergone apoptosis.² The molecular interactions between pro- and anti-apoptotic Bcl-2 family members determine cellular sensitivity to multiple lethal triggers, including many standard chemotherapeutic agents and ionizing radiation (IR).^{3, 4} Overexpression of Bcl-2 is known to increase clonogenic survival and inhibit IR-induced apoptosis.^{3, 4} Bcl-X_L expression also shows a strong correlation with resistance to cytotoxic anticancer therapies including IR.^{5, 6}Lung cancer is the leading cause of cancer deaths in western countries.⁷ Small-cell lung cancer (SCLC) accounts for 15% of all lung cancer cases and is distinguished from non-SCLC by its characteristic cytomorphology, rapid proliferation and early dissemination to metastatic sites.⁸ The standard of care to patients with limited-stage SCLC and good performance status is based on a combination of IR and cisplatin-based chemotherapy, resulting in a complete response rate as high as 50–80% coupled to a deceptive 12–20% 5-year survival.⁹ Initially, SCLC is responsive to chemo- and radiotherapy. However, SCLC recurs within the first 12 months.¹⁰ To date, the pathways mediating chemo- and radioresistance in SCLC are largely unknown.Deletion of pro-apoptotic gene and amplification of anti-apoptotic gene are frequently observed in SCLC, especially amplification of the BCL2L1 and BCL2L2 genes.¹¹ At the protein level, increased expression of Bcl-2 has been reported in up to 90% of metastatic SCLC. Bcl-2 overexpression, downregulation of the pro-apoptotic Bcl-2 antagonist Bax and a shift in the Bcl-2/Bax ratio to levels >1 are correlated with lower apoptotic index in tumors¹² and are associated with chemotherapeutic resistance in SCLC cell lines.¹³ In contrast with most solid tumor cell lines, where apoptosis does not appear as a predominant cell death mechanism after IR,¹⁴ overexpression of Bcl-2 can abrogate chemotherapy-induced apoptosis in SCLC cell lines.¹³ Apoptosis may be one of the mechanisms that cause SCLC cells to die in response to radiotherapy.^{15, 16}Recently, a small synthetic compound ABT-737 and its orally bioavailable form ABT-263 (Navitoclax) were shown to efficiently antagonize Bcl-2 and Bcl-X_L by binding to their BH3 receptor domain. ABT737 or its derivatives mediate antitumoral effects in chronic lymphocytic leukemia (CLL) and SCLC in preclinical and early clinical trials.^{17, 18} However, there is no published study that evaluates the combination of new Bcl-2/Bcl-X_L inhibitors, IR and chemo-radiotherapy.