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Genome-wide CRISPR screening identifies PHF8 as an effective therapeutic target for KRAS- or BRAF-mutant colorectal
Zhao Liu1,2, Yiqi Li3, Simeng Wang1
1Department of Endocrinology and International Joint Research Center for Tumor Precision Medicine of Shaanxi Province, The First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, 710061, P.R. China.
Background:
Mutations in KRAS and BRAF genes are prevalent in colorectal cancer (CRC), which strikingly promote tumorigenesis and lead to poor response to a variety of treatments including immunotherapy by activating the MAPK/ERK pathway. Thus, there is an urgent need to discover effective therapeutic targets and strategies.
Methods:
CRISPR-Cas9 lentiviral knockout library was used to screen the suppressors of anti-PD1 immunotherapy. Bioinformatic analysis was used to analyze the correlation between PHF8 expression and immune indicators in CRC. In vitro and in vivo experiments were utilized to determine the effects of PHF8 on the immune indexes and malignant phenotypes of CRC cells. qRT-PCR, western blotting, immunohistochemical (IHC) staining, and chromatin immunoprecipitation (ChIP)-qPCR assays were used to determine the regulatory effects of PHF8 on PD-L1, KRAS, BRAF, and c-Myc and the regulatory effect c-Myc/miR-22-3p signaling axis on PHF8 expression in CRC cells.
Results:
This study identified histone lysine demethylase PHF8 as a negative regulator for the efficacy of anti-PD1 therapy and found that it was highly expressed in CRCs and strongly associated with poor patient survival. Functional studies showed that PHF8 played an oncogenic role in KRAS- or BRAF-mutant CRC cells, but not in wild-type ones. Mechanistically, PHF8 up-regulated the expression of PD-L1, KRAS, BRAF, and c-Myc by increasing the levels of transcriptional activation marks H3K4me3 and H3K27ac and decreasing the levels of transcriptional repression mark H3K9me2 within their promoter regions, promoting immune escape and tumor progression. Besides, our data also demonstrated that PHF8 was up-regulated by the c-Myc/miR-22-3p signaling axis to form a positive feedback loop. Targeting PHF8 substantially improved the efficacy of anti-PD1 therapy and inhibited the malignant phenotypes of KRAS- or BRAF-mutant CRC cells.
Conclusion:
Our data demonstrate that PHF8 may be an effective therapeutic target for KRAS- or BRAF-mutant CRCs.
Insights
Histone lysine demethylase PHF8 negatively regulates anti-PD1 immunotherapy efficacy in colorectal cancer (CRC) by promoting immune escape. Targeting PHF8 offers a promising therapeutic strategy for KRAS- or BRAF-mutant CRC patients.
Area of Science:
- Oncology
- Molecular Biology
- Immunotherapy
Background:
- KRAS and BRAF mutations are common in colorectal cancer (CRC), driving tumorigenesis and immunotherapy resistance via the MAPK/ERK pathway.
- Effective therapeutic targets are urgently needed for KRAS/BRAF-mutant CRC to overcome treatment resistance.
Purpose of the Study:
- To identify novel therapeutic targets that can overcome resistance to anti-PD1 immunotherapy in colorectal cancer.
- To investigate the role of PHF8 in regulating anti-PD1 therapy efficacy and tumor progression in CRC.
Main Methods:
- CRISPR-Cas9 lentiviral knockout library screening to identify suppressors of anti-PD1 immunotherapy.
- Bioinformatic analysis of PHF8 expression and immune indicators in CRC.
- In vitro and in vivo experiments, qRT-PCR, western blotting, IHC, and ChIP-qPCR to elucidate PHF8 functions and regulatory mechanisms.
Main Results:
- PHF8 was identified as a negative regulator of anti-PD1 therapy efficacy, highly expressed in CRC and associated with poor survival.
- PHF8 promotes oncogenesis in KRAS/BRAF-mutant CRC by up-regulating PD-L1, KRAS, BRAF, and c-Myc expression through epigenetic modifications.
- A positive feedback loop involving c-Myc/miR-22-3p signaling up-regulates PHF8, enhancing immune escape and tumor progression.
Conclusions:
- PHF8 plays a critical oncogenic role in KRAS/BRAF-mutant colorectal cancer, contributing to immune evasion.
- Targeting PHF8 significantly enhances anti-PD1 therapy efficacy and inhibits tumor cell malignancy.
- PHF8 represents a potential therapeutic target for KRAS/BRAF-mutant colorectal cancer.
