未知原发癌（CUP）发展所需的遗传变化仍然不清楚。对14个经过严格筛选的CUP样本进行全外显子组分析未发现已知驱动基因中特定的经常性突变。然而，通过将CUP的突变谱与大多数其他人类肿瘤类型进行比较，发现基于轴突导向KEGG通路的基因的变化持续富集。特别是，PlexinB2（PlxnB2）的G842C突变被预测为活化。实际上，在CUP干细胞中敲除突变但不敲除野生型PlxnB2导致在培养中自我更新和增殖以及在小鼠中的肿瘤形成能力受损。相反，将G842C-PlxnB2遗传转移足以促进小鼠中CUP干细胞的增殖和肿瘤形成。值得注意的是，CUP细胞中的G842C-PlxnB2表达与基础EGFR磷酸化有关，EGFR阻断剂可降低依赖于突变受体的CUP细胞的活力。此外，突变的PlxnB2诱导CUP细胞侵袭性，经EGFR抑制剂治疗可被阻断。总之，我们发现一个轴突引导基因PLXNB2的新的活化突变可以支持来自未知原发癌的干细胞的增殖自主性，并以EGFR依赖的方式赋予侵袭性。©2023作者。根据CC BY 4.0许可证条件发布。

The genetic changes sustaining the development of cancers of unknown primary (CUP) remain elusive. The whole-exome genomic profiling of 14 rigorously selected CUP samples did not reveal specific recurring mutation in known driver genes. However, by comparing the mutational landscape of CUPs with that of most other human tumor types, it emerged a consistent enrichment of changes in genes belonging to the axon guidance KEGG pathway. In particular, G842C mutation of PlexinB2 (PlxnB2) was predicted to be activating. Indeed, knocking down the mutated, but not the wild-type, PlxnB2 in CUP stem cells resulted in the impairment of self-renewal and proliferation in culture, as well as tumorigenic capacity in mice. Conversely, the genetic transfer of G842C-PlxnB2 was sufficient to promote CUP stem cell proliferation and tumorigenesis in mice. Notably, G842C-PlxnB2 expression in CUP cells was associated with basal EGFR phosphorylation, and EGFR blockade impaired the viability of CUP cells reliant on the mutated receptor. Moreover, the mutated PlxnB2 elicited CUP cell invasiveness, blocked by EGFR inhibitor treatment. In sum, we found that a novel activating mutation of the axon guidance gene PLXNB2 sustains proliferative autonomy and confers invasive properties to stem cells isolated from cancers of unknown primary, in EGFR-dependent manner.© 2023 The Authors. Published under the terms of the CC BY 4.0 license.