PO.MCB02.01 · 分子与细胞生物学
靶向BCL2通路以增强ALK+ NSCLC的免疫原性
Targeting BCL2 pathway to enhance immunogenicity in ALK + NSCLC
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
背景：间变性淋巴瘤激酶（ALK）基因易位发生于3-5%的非小细胞肺癌（NSCLC）患者中，并采用ALK酪氨酸激酶抑制剂（TKI）治疗。然而，对ALK TKI的耐药通过靶点突变和旁路信号传导而产生。对ALK TKI的治疗反应还依赖于凋亡——一种由BCL2家族蛋白调控的复杂机制。因此，该通路的失调促进肿瘤进展和治疗耐药。据报道，癌细胞依赖BCL-2家族的抗凋亡蛋白（如BCL-2、MCL-1、BCL-xL）来存活。这促成了BH3模拟物的开发，即抗凋亡蛋白的抑制剂。ALK+ NSCLC被认为是一种免疫“冷”肿瘤，免疫原性低，表现为新抗原较少和缺乏细胞毒性肿瘤浸润淋巴细胞（TIL），导致免疫检查点靶向疗法在这些患者中疗效有限。据报道，一些TKI可通过释放损伤相关分子模式（DAMP）诱导免疫原性细胞死亡（ICD），从而激发持久的抗肿瘤免疫。然而，这一点在ALK+ NSCLC背景下尚未得到研究。我们假设ALK导向治疗与BH3模拟物联合将通过释放DAMP启动ICD，并增加对ALK TKI的敏感性。
方法：我们评估了ALK TKI（Lorlatinib、Alectinib）单独使用以及与BH3模拟物navitoclax（BCL-2/BCL-xL抑制剂）、venetoclax（BCL-2抑制剂）和s63845（MCL-1抑制剂）联合使用，在ALK+ NSCLC细胞系中诱导损伤相关分子模式（DAMP）的效果。
结果：ALK+癌细胞系表现出MCL-1和BCL-xL蛋白的高表达，表明它们依赖BCL-2家族蛋白来存活。与ALK TKI单药治疗相比，加入BH3模拟物增强了凋亡诱导（表现为PARP切割增加），并进一步降低了集落形成效率。ALK TKI单药和与BH3模拟物的联合治疗均诱导了对ICD至关重要的关键DAMP，包括：（1）通过IFN-beta（3-7倍）和CXCL10（2-3倍）增强I型干扰素信号；（2）胞外ATP在处理7小时时达到释放峰值；以及（3）与对照相比eIF2alpha磷酸化增强。
结论：ALK导向治疗有潜力诱导免疫原性细胞死亡（ICD）并激发抗肿瘤免疫反应。将BH3模拟物与ALK TKI联合可能是一种有前景的治疗策略，以增强抗肿瘤免疫并克服ALK+ NSCLC中的ALK TKI耐药。
查看英文原文 English abstract
Background : Translocations of the Anaplastic Lymphoma Kinase (ALK) gene occur in 3-5% of Non-Small Cell Lung Cancer (NSCLC) patients and are treated with ALK tyrosine kinase inhibitors (TKIs). However, resistance to ALK TKIs develop through on-target mutations and bypass signaling. Treatment response to ALK TKIs also relies on apoptosis, a complex mechanism regulated by BCL2 family proteins. Therefore, dysregulation of this pathway promotes tumor progression and therapy resistance. It has been reported that cancer cells depend on anti-apoptotic proteins of BCL-2 family (e.g. BCL-2, MCL-1, BCL-xL) for their survival. This has led to the development of BH3 mimetics, which are inhibitors of anti-apoptotic proteins. ALK+ NSCLC is considered an immune “cold” tumor with low immunogenicity, demonstrated by fewer neoantigens and lack of cytotoxic tumor-infiltrating lymphocytes (TILs), leading to limited efficacy of immune checkpoint-targeting therapies in these patients. Some TKIs have been reported to elicit durable antitumor immunity by inducing immunogenic cell death (ICD) through the release of damage-associated molecular patterns (DAMPs). However, this has not been studied in the context of ALK+ NSCLC.We hypothesized that ALK-directed therapy in combination with BH3 mimetics would initiate ICD through the release of DAMPs and increase sensitivity to ALK TKIs.
Methods: We evaluated the efficacy of ALK TKIs (Lorlatinib, Alectinib) alone and in combination with the BH3 mimetics navitoclax (BCL-2/BCL-xL inhibitor), venetoclax (BCL-2 inhibitor) and s63845 (MCL-1 inhibitor) to induce damage-associated molecular patterns (DAMPs) in ALK+ NSCLC cell lines.
Results: ALK+ cancer cell lines exhibited high expression of MCL-1 and BCL-xL proteins, indicating their dependency on BCL-2 family proteins for survival. The addition of BH3 mimetics enhanced apoptosis induction, as evidenced by increased PARP cleavage, and further reduced colony formation efficiency compared to ALK TKI single-agent treatment. Both ALK TKI single agent and combination treatment with BH3 mimetics induced key DAMPs essential for ICD, including: (1) increased type I interferon signaling via IFN-beta (3-7-fold) and CXCL10 (2-3-fold); (2) peak release of extracellular ATP at 7 hours of treatment; and (3) enhanced phosphorylation of eIF2alpha compared to the control.
Conclusions: ALK-directed therapy has the potential to induce immunogenic cell death (ICD) and elicit antitumor immune responses. Combining BH3 mimetics with ALK TKIs can be a promising therapeutic strategy to enhance anti-tumor immunity and overcome ALK TKI resistance in ALK+ NSCLC
利益披露 Disclosure
B. Munkhjargal, None.