PO.CL01.03 · 临床研究
KRAS突变癌症中的myCAF与ECM格局：利用液体生物标志物追踪KRAS诱导的纤维化和KRAS抑制剂疗效
The myCAF and ECM landscape in KRAS-mutated cancer: Utilizing liquid biomarkers to track KRAS-induced fibrosis and KRAS inhibitor efficacy
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
背景：KRAS驱动的癌症约占所有癌症的25%，在胰腺癌（PDAC）、结直肠癌（CRC）和非小细胞肺癌（NSCLC）中比例较高。突变的KRAS和TGF-beta信号激活癌症相关成纤维细胞（CAFs），形成纤维化的细胞外基质（ECM）。纤维化ECM促进免疫抑制和肿瘤生长，但有可能通过KRAS抑制剂（KRASi）逆转。我们最近证实，KRAS突变PDAC中的肌成纤维细胞CAFs（myCAFs）高表达III、V、VIII、XI和XII型胶原。在本研究中，我们探讨了跨KRAS突变癌症的KRAS诱导myCAF激活和ECM改变的格局。我们利用这些改变来识别KRASi疗效的生物标志物。
方法：对KRAS突变PDAC（n=141）、CRC（n=223）或NSCLC（n=74）患者进行胶原表达评估，并采用基因集变异分析（GSVA）评估hallmark KRAS和TGF-beta信号。使用Spearman相关分析评估KRAS和TGF-beta信号以及跨癌种胶原表达之间的相关性。使用Seurat分析来自KRASi处理的KPPC小鼠的单细胞RNA-seq数据。对CAFs进行亚聚类，并根据标志基因识别亚型（myCAF、炎性CAF和抗原递呈CAF）。分别使用Fisher精确检验和Wilcoxon秩和检验比较KRASi与载体之间的CAF亚型和胶原表达。在健康对照及PDAC、CRC或NSCLC患者的血清中测定I型（PRO-C1）、III型（PRO-C3）、V型（PRO-C5）、VIII型（PRO-C8）、XI型（PRO-C11）和XII型（PRO-C12）胶原形成的生物标志物。采用Wilcoxon秩和检验评估癌症与对照之间的差异。
结果：在KRAS突变的PDAC、CRC和NSCLC患者中，KRAS信号与TGF-beta信号以及COL3A1、COL5A2和COL8A1的表达显著相关（R>0.4，p<0.05）。与载体相比，KRASi处理的PDAC中成纤维细胞的Col3a1、Col5a2、Col8a1、Col11a1和Col12a1表达显著升高（p<0.001）。KRASi处理的PDAC中myCAFs显著减少（p<0.001），提示KRAS抑制减少了myCAF来源的ECM。与健康对照相比，myCAF胶原形成的血清生物标志物（PRO-C3、PRO-C5、PRO-C8、PRO-C11和PRO-C12）在PDAC、CRC和NSCLC患者中显著上调（p<0.001）。I型胶原形成未受影响（PRO-C1，p=0.37），提示存在KRAS特异性的胶原形成。
结论：KRAS突变肿瘤具有富含myCAF的微环境和独特的胶原表达谱，且该谱可被KRAS抑制所改变。在KRAS驱动癌症患者血清中升高的、无创的myCAF来源胶原生物标志物，有可能量化KRAS信号并追踪KRASi疗效，以指导临床决策。
查看英文原文 English abstract
Background: KRAS-driven cancer represents ~25% of cancers, with high rates in pancreatic cancer (PDAC), colorectal cancer (CRC), and non-small cell lung cancer (NSCLC). Mutated KRAS and TGF-beta signaling activate cancer-associated fibroblasts (CAFs), creating a fibrotic extracellular matrix (ECM). The fibrotic ECM promotes immunosuppression and tumor growth but is potentially reversible with KRAS inhibitors (KRASi). We have recently demonstrated that myofibroblast CAFs (myCAFs) in KRAS-mutated PDAC express high levels of collagens III, V, VIII, XI, and XII. In this study, we investigated the landscape of KRAS-induced myCAF activation and ECM change across KRAS-mutated cancers. We utilized these changes to identify biomarkers for KRASi efficacy.
Methods: Patients with KRAS-mutated PDAC (n=141), CRC (n=223), or NSCLC (n=74) were evaluated for collagen expression and gene-set variation analysis (GSVA) was used for hallmark KRAS and TGF-beta signaling. Correlations of KRAS and TGF-beta signaling, and collagen expressions across cancers, were evaluated using Spearman correlation. Single-cell RNA-seq data from KRASi treated KPPC mice was analyzed using Seurat. CAFs were subclustered and subtypes identified from marker genes (myCAF, inflammatory CAF, and antigen-presenting CAF). CAF subtype and collagen expression were compared between KRASi and vehicle with Fisher's exact test and Wilcoxon rank-sum test, respectively. Biomarkers for formation of collagens I (PRO-C1), III (PRO-C3), V (PRO-C5), VIII (PRO-C8), XI (PRO-C11), and XII (PRO-C12) were measured in serum from healthy controls and patients with PDAC, CRC, or NSCLC. Differences between cancer and control were evaluated by Wilcoxon rank-sum test.
Results: KRAS signaling was significantly correlated with TGF-beta signaling and expression of COL3A1, COL5A2, and COL8A1 across patients with KRAS-mutated PDAC, CRC, and NSCLC (R>0.4, p<0.05). Significantly increased expression of Col3a1, Col5a2, Col8a1, Col11a1, and Col12a1 was observed from fibroblasts in KRASi treated PDAC compared to vehicle (p<0.001). Significantly fewer myCAFs in KRASi treated PDAC (p<0.001) suggested reduced myCAF-derived ECM with KRAS inhibition. Serum biomarkers for formation of myCAF collagens (PRO-C3, PRO-C5, PRO-C8, PRO-C11, and PRO-C12) were significantly upregulated in patients with PDAC, CRC, and NSCLC compared to healthy controls (p<0.001). Type I collagen formation was unaffected (PRO-C1, p=0.37), suggesting KRAS-specific collagen formation.
Conclusion: KRAS-mutated tumors have a myCAF-enriched microenvironment with a unique collagen expression profile that is altered by KRAS inhibition. Non-invasive myCAF-derived collagen biomarkers that are elevated in serum from patients with KRAS-driven cancers, can potentially quantify KRAS signaling and track KRASi efficacy to guide clinical decision-making.
利益披露 Disclosure
M. B. Rasmussen,
Nordic Bioscience Employment.
R. S. Pedersen,
Nordic Bioscience Employment.
N. Willumsen,
Nordic Bioscience Employment.
M. Karsdal,
Nordic Bioscience g., Board of Directors, non-salaried role).