Multi-hallmark long noncoding RNA maps reveal non-small cell lung cancer vulnerabilities [ASOseries]. Multi-hallmark long noncoding RNA maps reveal non-small cell lung cancer vulnerabilities [ASOseries]

Long noncoding RNAs (lncRNAs) are widely dysregulated in cancer, yet their functional roles in cellular disease hallmarks remain unclear. Here we employ pooled CRISPR deletion to perturb 831 lncRNAs detected in KRAS-mutant non-small cell lung cancer (NSCLC), and measure their contribution to proliferation, chemoresistance and migration across two cell backgrounds. Integrative analysis of this data outperforms conventional “dropout” screens in identifying cancer genes, while prioritising disease-relevant lncRNAs with pleiotropic and background-independent roles. Altogether 80 high-confidence oncogenic lncRNAs are active in NSCLC, the majority identified here for the first time, and which tend to be amplified and overexpressed in tumours. A follow-up antisense oligonucleotide (ASO) screen shortlisted two candidates, Cancer Hallmarks in Lung LncRNA 1 (CHiLL 1) and GCAWKR, whose knockdown consistently suppressed cancer hallmarks in a variety of two- and three-dimension tumour models. Molecular phenotyping reveals that CHiLL 1 and GCAWKR control cellular-level phenotypes via distinct transcriptional networks converging on common oncogenic pathways. In summary, this work reveals a multi-dimensional functional lncRNA landscape underlying NSCLC that contains potential therapeutic vulnerabilities. Overall design: RNA sequencing of A549 and NCI-H460 cells treated with antisense oligonucleotides targeting two top lncRNA candidates.

长链非编码RNA（long noncoding RNAs）在癌症中广泛存在表达失调现象，但其在细胞疾病特征中的功能作用仍不明晰。本研究采用混合CRISPR敲除筛选技术，对KRAS突变型非小细胞肺癌（non-small cell lung cancer，以下简称NSCLC）中检测到的831个lncRNAs进行功能扰动，并在两种细胞背景下定量评估其对细胞增殖、化学耐药性及迁移能力的影响。本研究对该数据的整合分析在鉴定癌症基因方面优于传统的"dropout"筛选，同时优先筛选出具有多效性且不受细胞背景影响的疾病相关lncRNAs。总计有80个高置信度致癌lncRNAs在NSCLC中发挥活性，其中绝大多数为本次研究首次鉴定，且这类lncRNAs在肿瘤中多存在拷贝扩增与过表达现象。后续通过反义寡核苷酸（antisense oligonucleotide，以下简称ASO）筛选得到两个候选靶点：肺lncRNA癌症特征1（Cancer Hallmarks in Lung LncRNA 1，以下简称CHiLL 1）与GCAWKR，其敲低可在多种二维及三维肿瘤模型中持续抑制癌症特征表型。分子表型分析显示，CHiLL 1与GCAWKR通过不同的转录网络调控细胞水平表型，这些网络最终汇聚于共同的致癌通路。综上，本研究揭示了NSCLC背后存在的多维度功能性lncRNA调控图谱，其中蕴含潜在的治疗脆弱靶点。整体实验设计：对靶向两种顶级候选lncRNAs的ASO处理后的A549与NCI-H460细胞进行RNA测序。