Abstract：
BACKGROUND: In the last years, several efforts have been made to classify colorectal cancer (CRC) into well-defined molecular subgroups, representing the intrinsic inter-patient heterogeneity, known as Consensus Molecular Subtypes (CMSs).
METHODS: In this work, we performed a meta-analysis of CRC patients stratified into four CMSs. We identified a negative correlation between a high level of anaplastic lymphoma kinase (ALK) expression and relapse-free survival, exclusively in CMS1 subtype. Stemming from this observation, we tested cell lines, patient-derived organoids and mice with potent ALK inhibitors, already approved for clinical use.
RESULTS: ALK interception strongly inhibits cell proliferation already at nanomolar doses, specifically in CMS1 cell lines, while no effect was found in CMS2/3/4 groups. Furthermore, in vivo imaging identified a role for ALK in the dynamic formation of 3D tumor spheroids. Consistently, ALK appeares constitutively phosphorylated in CMS1, and it signals mainly through the AKT axis. Mechanistically, we found that CMS1 cells display several copies of ALKAL2 ligand and ALK-mRNAs, suggesting an autocrine loop mediated by ALKAL2 in the activation of ALK pathway, responsible for the invasive phenotype. Consequently, disruption of ALK axis mediates the pro-apoptotic action of CMS1 cell lines, both in 2D and 3D and enhanced cell-cell adhesion and e-cadherin organization. In agreement with all these findings, the ALK signature encompassing 65 genes statistically associated with worse relapse-free survival in CMS1 subtype. Finally, as a proof of concept, the efficacy of ALK inhibition was demonstrated in both patient-derived organoids and in tumor xenografts in vivo.
CONCLUSIONS: Collectively, these findings suggest that ALK targeting may represent an attractive therapy for CRC, and CMS classification may provide a useful tool to identify patients who could benefit from this treatment. These findings offer rationale and pharmacological strategies for the treatment of CMS1 CRC.
METHODS: In this work, we performed a meta-analysis of CRC patients stratified into four CMSs. We identified a negative correlation between a high level of anaplastic lymphoma kinase (ALK) expression and relapse-free survival, exclusively in CMS1 subtype. Stemming from this observation, we tested cell lines, patient-derived organoids and mice with potent ALK inhibitors, already approved for clinical use.
RESULTS: ALK interception strongly inhibits cell proliferation already at nanomolar doses, specifically in CMS1 cell lines, while no effect was found in CMS2/3/4 groups. Furthermore, in vivo imaging identified a role for ALK in the dynamic formation of 3D tumor spheroids. Consistently, ALK appeares constitutively phosphorylated in CMS1, and it signals mainly through the AKT axis. Mechanistically, we found that CMS1 cells display several copies of ALKAL2 ligand and ALK-mRNAs, suggesting an autocrine loop mediated by ALKAL2 in the activation of ALK pathway, responsible for the invasive phenotype. Consequently, disruption of ALK axis mediates the pro-apoptotic action of CMS1 cell lines, both in 2D and 3D and enhanced cell-cell adhesion and e-cadherin organization. In agreement with all these findings, the ALK signature encompassing 65 genes statistically associated with worse relapse-free survival in CMS1 subtype. Finally, as a proof of concept, the efficacy of ALK inhibition was demonstrated in both patient-derived organoids and in tumor xenografts in vivo.
CONCLUSIONS: Collectively, these findings suggest that ALK targeting may represent an attractive therapy for CRC, and CMS classification may provide a useful tool to identify patients who could benefit from this treatment. These findings offer rationale and pharmacological strategies for the treatment of CMS1 CRC.
摘要：
背景：在过去的几年中，已经做出了一些努力来将结直肠癌（CRC）分类为明确定义的分子亚组，代表内在的患者间异质性，称为共有分子亚型(CMSs)。
方法：在这项工作中，我们对CRC患者进行了荟萃分析,分为4个CMS.我们发现高水平的间变性淋巴瘤激酶（ALK）表达与无复发生存率之间存在负相关，仅在CMS1子类型中。从这一观察开始，我们测试了细胞系，患者来源的类器官和具有强效ALK抑制剂的小鼠，已经批准用于临床。
结果：在纳摩尔剂量下，ALK拦截已经强烈抑制细胞增殖，特别是在CMS1细胞系中，而CMS2/3/4组无效果。此外，体内成像确定了ALK在3D肿瘤球体动态形成中的作用。始终如一，ALK在CMS1中表现为组成型磷酸化，并且主要通过AKT轴发出信号。机械上,我们发现CMS1细胞显示几个拷贝的ALKAL2配体和ALK-mRNA,提示ALKAL2介导的自分泌环激活ALK途径，负责侵袭表型。因此,ALK轴的破坏介导CMS1细胞系的促凋亡作用，在2D和3D中都增强了细胞-细胞粘附和e-cadherin组织。与所有这些发现一致，包含65个基因的ALK信号在统计学上与CMS1亚型无复发生存率较差相关.最后,作为概念的证明，ALK抑制的功效在患者来源的类器官和体内肿瘤异种移植物中均得到证实.
结论：总的来说，这些结果表明，ALK靶向可能是一种有吸引力的CRC治疗方法，和CMS分类可能提供了一个有用的工具来识别哪些患者可以从这种治疗中获益.这些发现为CMS1CRC的治疗提供了理论基础和药理学策略。
方法：在这项工作中，我们对CRC患者进行了荟萃分析,分为4个CMS.我们发现高水平的间变性淋巴瘤激酶（ALK）表达与无复发生存率之间存在负相关，仅在CMS1子类型中。从这一观察开始，我们测试了细胞系，患者来源的类器官和具有强效ALK抑制剂的小鼠，已经批准用于临床。
结果：在纳摩尔剂量下，ALK拦截已经强烈抑制细胞增殖，特别是在CMS1细胞系中，而CMS2/3/4组无效果。此外，体内成像确定了ALK在3D肿瘤球体动态形成中的作用。始终如一，ALK在CMS1中表现为组成型磷酸化，并且主要通过AKT轴发出信号。机械上,我们发现CMS1细胞显示几个拷贝的ALKAL2配体和ALK-mRNA,提示ALKAL2介导的自分泌环激活ALK途径，负责侵袭表型。因此,ALK轴的破坏介导CMS1细胞系的促凋亡作用，在2D和3D中都增强了细胞-细胞粘附和e-cadherin组织。与所有这些发现一致，包含65个基因的ALK信号在统计学上与CMS1亚型无复发生存率较差相关.最后,作为概念的证明，ALK抑制的功效在患者来源的类器官和体内肿瘤异种移植物中均得到证实.
结论：总的来说，这些结果表明，ALK靶向可能是一种有吸引力的CRC治疗方法，和CMS分类可能提供了一个有用的工具来识别哪些患者可以从这种治疗中获益.这些发现为CMS1CRC的治疗提供了理论基础和药理学策略。
