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Functional single-cell analysis of the stem cell plasticity associated with drug-tolerant persister cells in colorectal cancer

与结直肠癌耐药持久细胞相关的干细胞可塑性的功能单细胞分析

基本信息

批准号：
22K15539
负责人：
COPPO Roberto
金额：
$ 2.91万
依托单位：
Kyoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Early-Career Scientists
财政年份：
2022
资助国家：
日本
起止时间：
2022-04-01 至 2023-03-31
项目状态：
已结题

项目摘要

Using patient-derived colorectal cancer (CRC) organoids, we previously generated an analysis platform consisting of slow-growing CRC cells isolated from different human samples to represent the tumor heterogeneity. In this study, we tracked the fate of each cell through a clonogenic growth assay and found that the CRC cells showed a wide range of growth ability. Further rounds of the clonogenic growth assay revealed that the spheroid forming cells in CRC organoids consisted of distinct subpopulations; the cells generating large spheroids (L-cells) and the cells generating small spheroids (S-cells). The cells derived from the small spheroids gave rise to only small spheroids, consisting of slow-growing cells (S-pattern). While the cells derived from the large spheroids gave rise to both small and large spheroids, showing a dual-growing phenotype (D-pattern). Although the S-pattern spheroids never gave rise to large spheroids once isolated, transition to the D-pattern occurred by various extrinsic triggers, in which Musashi-1 (MSI1) played a key role. We revealed that the suppression of MSI1 in large spheroids, by using the CRISPR/Cas9 system, induced a transition from the D- to the S-pattern. We also found that the S-pattern spheroids were resistant to chemotherapy and transited to the D-pattern upon drug treatment. In conclusion, the isolated S-cells could be a novel platform for investigating drug-tolerant persister cells (DTPs) and developing the DTP targeting treatment. As the transition is linked to the drug resistance, it can be a therapeutic target.

利用患者来源的结直肠癌（CRC）类器官，我们之前建立了一个分析平台，该平台由从不同人类样本中分离的生长缓慢的CRC细胞组成，以代表肿瘤的异质性。在这项研究中，我们通过克隆生长试验追踪了每个细胞的命运，发现CRC细胞表现出广泛的生长能力。进一步的克隆生长试验显示，CRC类器官中的球状形成细胞由不同的亚群组成；产生大球体的细胞（l细胞）和产生小球体的细胞（s细胞）。由小球体衍生的细胞只产生小球体，由生长缓慢的细胞组成（s型）。而来源于大球体的细胞可同时产生小球体和大球体，呈现双生长表型（d型）。虽然s型球体一旦被分离就不会产生大的球体，但向d型的转变是由各种外在触发因素引起的，其中Musashi-1 （MSI1）起了关键作用。我们发现，通过使用CRISPR/Cas9系统，在大球体中抑制MSI1诱导了从D-模式到s模式的转变。我们还发现s型球体对化疗有耐药性，并在药物治疗后转变为d型。综上所述，分离的s细胞可作为研究耐药持久性细胞（DTP）和开发DTP靶向治疗的新平台。由于这种转变与耐药性有关，因此可以作为治疗靶点。