From pathogenesis to new therapeutic targets in diffuse large B cell lymphoma

项目总结 弥漫性大B细胞淋巴瘤(DLBCL)在约30%的患者中是无法治愈的，尽管最近在治疗方面取得了进展。 T细胞疗法，仍然是一个重大的临床挑战。合理定位新疗法的一个障碍是 这些肿瘤具有显著的异质性，这使得多达20%-50%的病例无法根据 起源细胞分类或更新的基于遗传的分类。这可能部分是由于目前的情况 分类学仅限于对编码区的分析，仅占基因组的3%，而进一步 致病相关的遗传复杂性可能存在于基因组的非编码调控部分。至 为此，我们最近调查了关键的监管领域，如增强剂和超级增强剂 (SES)可能是DLBCL功能相关突变的部位。我们发现与之相对应的区域 在97%的DLBCL病例中，活跃的SES高度和特异地过度突变，而相同的基因座在 作为SE未处于活动状态。这种异常的SE超突变表现出AID活性的特征，并与基因有关 编码B细胞调节因子和成熟的癌基因。作为致癌相关性的证据，我们展示了 与BCL6、BCL2和CXCR4原癌基因连锁的高突变SE阻止了结合和 转录抑制物对相应靶基因的转录下调，包括BLIMP1 (BCL6)和类固醇受体NR3C1(BCL2和CXCR4)。值得注意的是，CRISPR/Cas9调解的 SE热点突变恢复了靶基因的调控并损害了细胞的生长，表明SE在其中起着关键作用 保持转化表型的突变(Bal等人，《自然》2022)。总体而言，这些发现确定了一个 高度普遍的、与病理相关的突变机制，可能会显著影响 DLBCL体细胞遗传格局的研究现状。这项研究计划的总体目标是 将：i)识别和机械地解剖顶端经常性突变/功能相关的SE和相关的 靶基因；ii)了解糖皮质激素受体途径的作用，这似乎是常见的 以SE高突变机制和直接编码突变为靶点，在正常B细胞生物学和 淋巴肿大。我们预计，这一新的基因改变层将确定新的机制 已知癌基因的失调，以及新的失调基因和通路，对 DLBCL的精确分型和治疗靶点。

PROJECT SUMMARY Diffuse Large B cell Lymphoma (DLBCL) is incurable in ~30% of patients and, despite recent advances in CAR- T cell therapies, remains a significant clinical challenge. One barrier to rationally targeted new therapies is the remarkable heterogeneity of these tumors, which leaves as many as 20-50% of cases unclassified based on cell-of-origin or more recent genetic-based classifications. This may be due in part to the fact that current taxonomies are limited to the analysis of coding regions, representing only 3% of the genome, while further genetic complexity of pathogenetic relevance may reside in the non-coding regulatory portion of the genome. To this end, we recently investigated whether critical regulatory domains such as enhancers and super-enhancers (SEs) could be the site of functionally relevant mutations in DLBCL. We found that regions corresponding to active SEs are highly and specifically hypermutated in 97% of DLBCL cases, as compared to the same loci when not active as SE. Such aberrant SE hypermutation displays signatures of AID activity and is linked to genes encoding B cell regulators and well-established oncogenes. As evidence of oncogenic relevance, we showed that the hypermutated SEs linked to the BCL6, BCL2, and CXCR4 proto-oncogenes prevent the binding and transcriptional downregulation of the corresponding target gene by transcriptional repressors, including BLIMP1 (BCL6) and the steroid-receptor NR3C1 (BCL2 and CXCR4). Of note, CRISPR/Cas9-mediated correction of the SE hotspot mutation restored target gene regulation and impaired cell growth, indicating a key role for the SE mutation in maintaining the transformed phenotype (Bal et al., Nature 2022). Overall, these findings identify a highly pervasive, pathogenetically relevant, mutational mechanism that is likely to significantly influence the current understanding of the somatic genetic landscape of DLBCL. The overall goal of this research program will be to: i) identify and mechanistically dissect the top recurrently mutated/functionally relevant SEs and associated target genes; ii) understand the role of the glucocorticoid receptor pathway, which appears to be commonly targeted by the SE hypermutation mechanism as well as by direct coding mutations, in normal B cell biology and lymphomagenesis. We anticipate that this new layer of genetic alterations will identify novel mechanisms of dysregulation for known oncogenes, as well as new dysregulated genes and pathways, with implications for precision classification and therapeutic targeting of DLBCL.