Role of PCIF1‐mediated 5′‐cap N6‐methyladeonsine mRNA methylation in colorectal cancer and anti‐PD‐1 immunotherapy

• Published in: The EMBO journal 2023-01, Vol.42 (2), p.e111673-n/a
• Main Authors: Wang, Lingling, Wu, Lujing, Zhu, Zhouting, Zhang, Qiong, Li, Wanyu, Gonzalez, Gwendolyn Michelle, Wang, Yinsheng, Rana, Tariq M
• Format: Article
• Language: English
• Subjects: Adaptor Proteins, Signal Transducing - metabolism; Adenine; Adenosine; Animal models; Animals; Antibodies; anti‐PD‐1 treatment; Cancer; Cell migration; Colorectal cancer; Colorectal carcinoma; Colorectal Neoplasms - drug therapy; Colorectal Neoplasms - genetics; Context; CRISPR; Depletion; EMBO03; EMBO36; Gene silencing; Humans; Immunotherapy; Life Sciences; Lipids; m6Am methylation; Malignancy; Membrane Proteins - metabolism; Methylation; Methyltransferase; Mice; mRNA; N6-methyladenosine; Nanoparticles; Natural killer cells; NK cells; Nuclear Proteins - metabolism; PCIF1; PD-1 protein; RNA modification; RNA, Messenger - genetics; RNA-Binding Proteins - metabolism; Silence; Stat1 protein; Survival; Transforming Growth Factor beta - metabolism; Transforming growth factor-b; Tumor necrosis factor; Tumor-infiltrating lymphocytes; Tumorigenesis; Tumors
• ISSN: 0261-4189; 1460-2075
• DOI: 10.15252/embj.2022111673

Adenosine N6‐methylation (m6A) and N6,2′‐O‐dimethylation (m6Am) are regulatory modifications of eukaryotic mRNAs. m6Am formation is catalyzed by the methyl transferase phosphorylated CTD‐interacting factor 1 (PCIF1); however, the pathophysiological functions of this RNA modification and PCIF1 in cancers are unclear. Here, we show that PCIF1 expression is upregulated in colorectal cancer (CRC) and negatively correlates with patient survival. CRISPR/Cas9‐mediated depletion of PCIF1 in human CRC cells leads to loss of cell migration, invasion, and colony formation in vitro and loss of tumor growth in athymic mice. Pcif1 knockout in murine CRC cells inhibits tumor growth in immunocompetent mice and enhances the effects of anti‐PD‐1 antibody treatment by decreasing intratumoral TGF‐β levels and increasing intratumoral IFN‐γ, TNF‐α levels, and tumor‐infiltrating natural killer cells. We further show that PCIF1 modulates CRC growth and response to anti‐PD‐1 in a context‐dependent mechanism with PCIF1 directly targeting FOS, IFITM3, and STAT1 via m6Am modifications. PCIF1 stabilizes FOS mRNA, which in turn leads to FOS‐dependent TGF‐β regulation and tumor growth. While during immunotherapy, Pcif1‐Fos‐TGF‐β, as well as Pcif1‐Stat1/Ifitm3‐IFN‐γ axes, contributes to the resistance of anti‐PD‐1 therapy. Collectively, our findings reveal a role of PCIF1 in promoting CRC tumorigenesis and resistance to anti‐PD‐1 therapy, supporting that the combination of PCIF1 inhibition with anti‐PD‐1 treatment is a potential therapeutic strategy to enhance CRC response to immunotherapy. Finally, we developed a lipid nanoparticles (LNPs) and chemically modified small interfering RNAs (CMsiRNAs)‐based strategy to silence PCIF1 in vivo and found that this treatment significantly reduced tumor growth in mice. Our results therefore provide a proof‐of‐concept for tumor growth suppression using LNP‐CMsiRNA to silence target genes in cancer. Synopsis m6Am formation in eukaryotic mRNAs is catalyzed by PCIF1, a cap‐specific adenine N6‐methyltransferase; however, the function and mechanism of m6Am modification and PCIF1 in colorectal cancer remain unclear. Here, we reveal that PCIF1 modulates colorectal cancer malignancy and response to anti‐PD‐1 immunotherapy in a context‐dependent manner. PCIF1 is overexpressed in colorectal cancer (CRC) and its increased expression negatively correlates with patient survival. Disruption of PCIF1 inhibits tumor malignancy and sensitizes colorectal cancer to i