弥漫性大B细胞淋巴瘤（DLBCL）表现出显著的基因异质性，这导致了药物抗性，需要开发新的治疗方法。环依赖性激酶（CDK）的药理学抑制剂在DLBCL的临床前研究中表现出活性，但许多都停滞于临床发展。在这里，我们显示AZD4573，一种选择性CDK9抑制剂，限制了DLBCL细胞的生长。CDK9抑制（CDK9i）导致了转录组和蛋白质组的快速变化，多个致癌蛋白（例如MYC、Mcl-1、JunB、PIM3）被下调，磷酸肌醇-3激酶（PI3K）和细胞衰老途径失调。在RNAPII暂停引起的初步转录抑制后，我们观察到了几个致癌基因，包括MYC和PIM3的转录恢复。ATAC-Seq和ChIP-Seq实验证明CDK9i引起了表观遗传重塑，具有双向的染色质可及性变化，抑制了启动子激活，并导致超增强子景观的持续重编程。CRISPR文库筛选表明介导复合物中的SE相关基因以及AKT1能够对CDK9i产生抗性。与此一致，sgRNA介导的MED12敲除使细胞对CDK9i敏感。根据我们的机制发现，我们将AZD4573与PIM激酶或PI3K抑制剂结合使用。这两种组合在DLBCL和原发淋巴瘤细胞中均减少了增殖并诱导了凋亡，在体内的小鼠移植模型中结果为肿瘤进展延迟和生存延长。因此，CDK9i引起表观遗传景观的重编程，超增强子驱动的选择性致癌基因恢复可能对CDK9i的抗性有贡献。PIM和PI3K代表了在DLBCL异质性景观中规避CDK9i抗性的潜在目标。©2023年作者。

Diffuse large B-cell lymphoma (DLBCL) exhibits significant genetic heterogeneity which contributes to drug resistance, necessitating development of novel therapeutic approaches. Pharmacological inhibitors of cyclin-dependent kinases (CDK) demonstrated pre-clinical activity in DLBCL, however many stalled in clinical development. Here we show that AZD4573, a selective inhibitor of CDK9, restricted growth of DLBCL cells. CDK9 inhibition (CDK9i) resulted in rapid changes in the transcriptome and proteome, with downmodulation of multiple oncoproteins (eg, MYC, Mcl-1, JunB, PIM3) and deregulation of phosphoinotiside-3 kinase (PI3K) and senescence pathways. Following initial transcriptional repression due to RNAPII pausing, we observed transcriptional recovery of several oncogenes, including MYC and PIM3. ATAC-Seq and ChIP-Seq experiments revealed that CDK9i induced epigenetic remodeling with bi-directional changes in chromatin accessibility, suppressed promoter activation and led to sustained reprograming of the super-enhancer landscape. A CRISPR library screen suggested that SE-associated genes in the Mediator complex, as well as AKT1, confer resistance to CDK9i. Consistent with this, sgRNA-mediated knockout of MED12 sensitized cells to CDK9i. Informed by our mechanistic findings, we combined AZD4573 with either PIM kinase or PI3K inhibitors. Both combinations decreased proliferation and induced apoptosis in DLBCL and primary lymphoma cells in vitro as well as resulted in delayed tumor progression and extended survival of mice xenografted with DLBCL in vivo. Thus, CDK9i induces reprogramming of the epigenetic landscape, and super-enhancer driven recovery of select oncogenes may contribute to resistance to CDK9i. PIM and PI3K represent potential targets to circumvent resistance to CDK9i in the heterogeneous landscape of DLBCL.© 2023. The Author(s).