mTOR inhibition amplifies the anti-lymphoma effect of PI3Kβ/δ blockage in diffuse large B-cell lymphoma. mTOR inhibition amplifies the anti-lymphoma effect of PI3Kβ/δ blockage in diffuse large B-cell lymphoma

Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous diagnostic category that can be stratified in molecular subtypes based on gene expression profiles and genetic alterations. As roughly one-third of DLBCL patients do not sustainably respond to the current standard chemo-immunotherapy, novel treatment options targeting oncogenic pathways might improve their outcome. Chronic B-cell receptor signaling or PTEN deficiency drive the constitutive activation of the phosphoinositide 3-kinase (PI3K). Since pan-PI3K inhibitors cause severe side effects, we investigated the anti-lymphoma efficacy of the specific PI3Kβ/δ inhibitor AZD8186. We identified a subset of DLBCL models within activated B cell-like (ABC) and germinal center B cell-like (GCB) DLBCL that were sensitive to AZD8186 treatment. On the molecular level, PI3Kβ/δ inhibition decreased the pro-survival nuclear factor kappa-B (NF-κB) and activator protein 1 (AP-1) activity or led to downregulation of the oncogenic transcription factor MYC. In AZD8186-resistant models, we detected a feedback activation of the PI3K/AKT/mTOR pathway following PI3Kβ/δ inhibition. The combined treatment with AZD8186 and the mTOR inhibitor AZD2014 overcame resistance to PI3Kβ/δ inhibition and completely prevented outgrowth of lymphoma cells in vivo in cell line- and patient-derived xenograft mouse models. Collectively, our study reveals that subsets of DLBCLs are addicted to PI3Kβ/δ signaling and thus identifies a previously unappreciated role of the PI3Kβ isoform in DLBCL survival. Furthermore, our data demonstrate that combined targeting of PI3Kβ/δ and mTOR is effective in all major DLBCL subtypes supporting the evaluation of this strategy in a clinical trial setting. Overall design: For gene expression profiling, we performed RNA-sequencing of two AZD8186-sensitive ABC DLBCL cell lines (OCI-Ly10 and TMD8) and two sensitive GCB DBLCL cell lines (HT and K422) after 6, 12 (not TMD8), 18, and 24 hours of AZD8186 treatment, compared to untreated controls. Each time point was measured once, resulting in 30 samples in total. Statistics were computed over all time points and the two cell lines of the same DLBCL subtype.

弥漫性大B细胞淋巴瘤（Diffuse large B-cell lymphoma, DLBCL）是一类异质性诊断类别，可基于基因表达谱与遗传变异分为分子亚型。约三分之一的DLBCL患者无法从当前标准化学免疫治疗中获得持续应答，靶向致癌通路的新型治疗方案或可改善其预后。慢性B细胞受体信号通路异常或PTEN缺失会驱动磷脂酰肌醇3-激酶（phosphoinositide 3-kinase, PI3K）的组成性激活。由于泛PI3K抑制剂会引发严重不良反应，我们探究了特异性PI3Kβ/δ抑制剂AZD8186的抗淋巴瘤疗效。我们在活化B细胞样（activated B cell-like, ABC）与生发中心B细胞样（germinal center B cell-like, GCB）DLBCL中鉴定出对AZD8186治疗敏感的DLBCL模型亚群。在分子层面，PI3Kβ/δ抑制会降低促存活核因子κB（nuclear factor kappa-B, NF-κB）与激活蛋白1（activator protein 1, AP-1）的活性，或导致致癌转录因子MYC的表达下调。在AZD8186耐药模型中，我们检测到PI3Kβ/δ抑制后出现了PI3K/AKT/mTOR通路的反馈激活。联合使用AZD8186与mTOR抑制剂AZD2014可克服PI3Kβ/δ抑制的耐药性，并在细胞系及患者来源的异种移植小鼠模型中完全阻断体内淋巴瘤细胞的增殖。综上，本研究揭示了部分DLBCL依赖PI3Kβ/δ信号通路，从而明确了PI3Kβ亚型在DLBCL存活中此前未被重视的作用。此外，我们的数据表明，同时靶向PI3Kβ/δ与mTOR对所有主要DLBCL亚型均有效，支持在临床试验中评估该治疗策略。整体实验设计：为进行基因表达谱分析，我们对2株AZD8186敏感的ABC DLBCL细胞系（OCI-Ly10与TMD8）及2株敏感GCB DLBCL细胞系（HT与K422）进行了RNA测序，处理组分别接受6、12（TMD8未设置该时间点）、18、24小时的AZD8186治疗，以未处理的细胞作为对照。每个时间点仅检测1次，共计获得30份样本。统计分析基于所有时间点及同一DLBCL亚型的2株细胞系进行。