Genome-scale CRISPR-Cas9 knockout screening in hepatocellular carcinoma with lenvatinib resistance

TM4SF1, a member of the transmembrane 4 superfamily, is crucial for both healthy and malignant human tissues. The significant function of TM4SF1 in the incidence and progression of cancer has been widely recognized in recent years. Although some achievements have been made in the study of TM4SF1, the effect of TM4SF1 on cancer stemness in hepatocellular carcinoma (HCC) and its molecular basis are yet to be reported. We found through abundant in vitro and in vivo experiments which the expression of TM4SF1 was positively correlated with the progression and cancer stemness of HCC. We identified the downstream protein MYH9 of TM4SF1 and its final regulatory target NOTCH pathway using bioinformatics analysis and protein mass spectrometry. We cultivated a lenvatinib-resistant strain from HCC cells to examine the relationship between cancer stemness and tumor drug resistance. The study confirmed that TM4SF1 could regulate the NOTCH pathway by upregulating MYH9, thus promoting cancer stemness and lenvatinib resistance in HCC. This study not only provided a new idea for the pathogenesis of HCC but also confirmed that TM4SF1 might become a new intervention point to improve the clinical efficacy of lenvatinib in treating HCC.

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“…However, it still cannot prevent patients from developing drug resistance, resulting in a poor prognosis [24] .…”

Section: Introductionmentioning

confidence: 99%

TM4SF1 Upregulates MYH9 to Activate the NOTCH Pathway to Promote Cancer Stemness and Lenvatinib Resistance in HCC

Yang

Jin

Zhou

et al. 2022

Preprint

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“…Xue Sui et al identi ed a novel cisplatin-resistant esophageal oncogene, ERCC8, based on genome-scale CRISPR/Cas9 screening [21]. In Lu Yonggang's study, they performed a genome-wide CRISPR/Cas9 knockout library screen using the HCC cell line Huh7 [22]. They identi ed two key resistance genes, neuro brin 1 (NF1) and bi-speci c phosphatase 9 (DUSP9), and demonstrated that they induce Lenvatinib resistance by activating PI3K/AKT and MEK/ERK signaling pathways in vivo and in vitro.…”

Section: Discussionmentioning

confidence: 99%

“…They identi ed two key resistance genes, neuro brin 1 (NF1) and bi-speci c phosphatase 9 (DUSP9), and demonstrated that they induce Lenvatinib resistance by activating PI3K/AKT and MEK/ERK signaling pathways in vivo and in vitro. They also screened out trametinib, a small-molecule pathway inhibitor that can be used to reverse Lenvatinib resistance in HCC [22]. Shanzhou Huang et al identi ed six genes associated with Lenvatinib resistance in HCC HepG2 and Huh7 cell lines, including DUSP4, CCBL1, DHDH, CNTN2, NOS3 and TNF.…”

Section: Discussionmentioning

confidence: 99%

CRISPR/Cas9 screening reveals that ASB2 inhibits p53-dependent ferroptosis to mediates Lenvatinib resistance in hepatocellular carcinoma

Sun

Chen

Wang

et al. 2023

Preprint

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Background Lenvatinib is in a first-line therapy for advanced hepatocellular carcinoma (HCC), but its resistance is one of the main obstacles to treatment failure. The molecular mechanism of Lenvatinib resistance has not been well explored. Methods Genome-wide CRISPR/Cas9 knockout screening system was developed and bioinformatic analysis was used to identify key genes associated with Lenvatinib resistance in HCC. Whole transcriptome sequencing including coding and non-coding RNAs has also been performed in Lenvatinib resistance and sensitive HCC cells. Co-immunoprecipitation, confocal localization, western blot, immunofluorescence and other experiments were employed to assess the role of ASB2 in Lenvatinib resistance. Results ASB2 was found to be significantly increased at the mRNA and protein levels in Lenvatinib resistant HCC cells. ASB2 knockdown inhibited HCC Lenvatinib resistance cell proliferation, invasion and migration. Mechanistically, ASB2 activated NF-κB pathway by promoting IκBα ubiquitination and inhibited ferroptosis by promoting P53 ubiquitination and then mediated Lenvatinib resistance in HCC. Interestingly, NOTCH1 was shown to transcriptionly promote ASB2 expression and regulate NF-κB as well as ferroptosis pathways to induce Lenvatinib resistance in HCC. In further clinical translation, we found that Venetoclax could bind to ASB2 through a virtual screen of protein potential binding small molecules, and confirmed that Venetoclax and Lenvatinib combined significantly inhibited the progression of HCC, and the efficacy was better than Lenvatinib alone in vitro and vivo. Conclusion This study reveals that ASB2 which was transcriptionly promoted by NOTCH1, activated NF-κB pathway by promoting IκBα ubiquitination and inhibited ferroptosis by promoting P53 ubiquitination and mediated HCC Lenvatinib-resistance based on CRISPR/Cas9 screening. Venetoclax can potentially inhibit the function of ASB2 and the combination of Venetoclax and Lenvatinib can significantly inhibit the progression of HCC, which provides new targets and specific strategies for the treatment of HCC Lenvatinib-resistance, bringing new hope and benefits to HCC patients.

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“…Moreover, using CRISPR/Cas9 library screening method, two key resistance genes, neurofibromin 1(NF1) and dual specificity phosphatase 9 (DUSP9), are screened out as critical drivers for Lenvatinib resistance in HCC cells. Trametinib, an inhibitor of MEK pathway, can be used to reverse Lenvatinib resistance mediated by NF1 and DUSP9 loss in HCC cells ( Lu et al, 2021 ). These findings suggest that targeting the resistance-associated molecules or pathway will improve the anti-tumor effect of Lenvatinib therapy for HCC.…”

Section: Discussionmentioning

confidence: 99%

Combination of Vitamin C and Lenvatinib potentiates antitumor effects in hepatocellular carcinoma cells in vitro

Wang

Qian

Wang

et al. 2023

PeerJ

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Lenvatinib has become a first-line drug in the treatment of advanced hepatocellular carcinoma (HCC). Investigating its use in combination with other agents is of great significance to improve the sensitivity and durable response of Lenvatinib in advanced HCC patients. Vitamin C (L-ascorbic acid, ascorbate, VC) is an important natural antioxidant, which has been reported to show suppressive effects in cancer treatment. Here, we investigated the effect of the combination of VC and Lenvatinib in HCC cells in vitro. We found that treatment of VC alone significantly inhibited the proliferation, migration and invasion in HCC cells. Additionally, VC was strongly synergistic with Lenvatinib in inhibition of the proliferative, migratory and invasive capacities of HCC cells in vitro. In conclusion, our results demonstrate that the combination of VC and Lenvatinib has synergistic antitumor activities against HCC cells, providing a promising therapeutic strategy to improve the prognosis of HCC patients.

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Genome-scale CRISPR-Cas9 knockout screening in hepatocellular carcinoma with lenvatinib resistance

Cited by 51 publications

References 33 publications

TM4SF1 Upregulates MYH9 to Activate the NOTCH Pathway to Promote Cancer Stemness and Lenvatinib Resistance in HCC

TM4SF1 Upregulates MYH9 to Activate the NOTCH Pathway to Promote Cancer Stemness and Lenvatinib Resistance in HCC

CRISPR/Cas9 screening reveals that ASB2 inhibits p53-dependent ferroptosis to mediates Lenvatinib resistance in hepatocellular carcinoma

Combination of Vitamin C and Lenvatinib potentiates antitumor effects in hepatocellular carcinoma cells in vitro

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