N-acetyltransferase 10 Enhances Doxorubicin Resistance In Hepatocellular Carcinoma Cell Lines By Promoting The Epithelial-to-mesenchymal Transition

Hepatocellular carcinoma(HCC)is one of the major malignant tumors worldwide.Although the diagnosis and treatment of HCC patients have achieved considerable progress and many patients have been given the opportunity to prolong their lives,the recurrence and metastasis of HCC patients after operation and the resistance to chemotherapeutic drugs will still affect the survival of patients.Among the widely used classification system of hepatocellular carcinoma,TNM classification involves lymph nodes and distant metastasis,which are related to the prognosis of patients.However,the classification of TNM does not include morphological,histological and molecular changes,which have important implications for prognosis.Moreover,the pathological morphology and molecular markers of tumors do not depend on TNM classification.Therefore,screening out the best combination of information related to the prognosis of hepatocellular carcinoma can help clinicians to make the most accurate prognostic judgment for patients,and thus to adopt the optimal treatment scheme.Machine learning algorithm,which can automatically analyses and obtains rules from large data,uses rules to predict unknown data.mRMR algorithm is a kind of machine learning,which is suitable for selecting the best combination of features from high-dimensional data.We used mRMR feature selection algorithm to combine clinicopathological features with immunohistochemistry features to construct a model that can predict the prognosis of 5-year survival and 3-year recurrence of tumors patients.Resistance to doxorubicin in HCC patients is the main cause of recurrence of hepatocellular carcinoma after operation.However,the cellular and molecular mechanisms of chemotherapeutic resistance in hepatocellular carcinoma remain unclear.Studies have found that epithelial-mesenchymal transition(EMT)can transform chemotherapy-resistant cells into chemosensitivity cells.At present,N-acetyltransferase 10(NAT10)plays a certain role in the process of cell biology and the occurrence and development of malignant tumors.However,the role of NAT 10 in EMT and chemoresistance in hepatocellular carcinoma has been less studied.Part? Prediction of hepatocellular carcinoma prognosis and recurrence after surgical resection using clinicopathologic and immunohistochemical characteristicsAimsApproximately 50%hepatocellular carcinoma(HCC)patients die within 5-year after surgical resection.The present staging systems do not fully allow to accurately predict the prognosis.This study aimed to identify markers to establish classifiers to predict the 5-year prognosis and the 3-year tumor recurrence in HCC patients post-operatively.Patients and MethodsIn this retrospective multicenter study,we used three independent cohorts of HCC patients,underwent surgical resection to develop and validate a prognostic classifier and a tumor recurrence classifier.We investigated the protein expression of 29 biomarkers by Immunohistochemical(IHC)analysis of 406 patients from the Institution I.The prognostic classifier was developed using the maximum relevance minimum redundancy algorithm jointly with the multivariable regression method.The classifier was further validated by an internal validation,136 patients from the Institution I and an independent validation,105 patients from the Institution II.A recurrent classifier was developed and validated using the same methods as the prognostic classifier.ResultsThe prognostic classifier integrated three clinicopathologic characteristics of age,tumor number,tumor diameter,and seven immunomarkers,including MMP-9,E-cadherin,CD34,CEA,S100A9,TYSY,and BAX.The prognostic classifier distinguished HCC patients into high-and low-probability survival groups with significant differences in all cohorts(AUC:the training cohort:0.726,95%CI:0.680-0.769;the internal validation cohort:0.731,95%CI:0.648-0.804;the independent validation cohort:0.753,95%CI:0.659-0.832).The 3-year HCC recurrence classifier also demonstrated a good discrimination in all cohorts.ConclusionThis study presented a prognostic classifier and a recurrent classifier using clinicopathologic and IHC characteristics.The developed classifiers stratified HCC patients into high-and low-probability survival groups,which would further enhance individualized care.Part ? N-acetyltransferase 10 enhances doxorubicin resistance in hepatocellular carcinoma cell lines by promoting the epithelial-to-mesenchymal transitionAimsN-acetyltransferase 10(NAT 10)has been reported to be expressed at high levels in hepatocellular carcinoma(HCC);however,its role in chemoresistance is unclear.This study aimed to investigate whether NAT 10 regulates the epithelial mesenchymal transition(EMT)and chemoresistance in HCC.MethodsHCC cell lines(Huh-7,Bel-7402,SNU387,SNU449)were treated with remodelin,an inhibitor of NAT 10 or transfected with small inhibitory RNAs(siRNAs)targeting NAT 10 or Twist.The EMT was induced by hypoxia.The CCK-8 assay was used to quantify cell viability,the EdU incorporation assay to assess cell proliferation.SiRNA knockdown efficiency and epithelial/mesenchymal marker expression were assessed by Western blotting.ResultsKnockdown of NAT 10 using the siRNA or inhibition of NAT 10 using remodelin increased the sensitivity of HCC cell lines to doxorubicin;similar effects were observed in cells transfected with the Twist siRNA.Inhibition of NAT10 using remodelin also reversed the ability of doxorubicin to induce the EMT in HCC cells.Furthermore,inhibiting NAT 10 reversed the hypoxia-induced EMT.Finally,we confirmed combining doxorubicin with remodelin delayed tumor growth and reduced tumor cell proliferation in a mouse xenograft model of HCC.ConclusionNAT 10 may contribute to chemoresistance in HCC by regulating the EMT.The mechanism by which NAT 10 regulates the EMT and doxorubicin sensitivity in HCC cells merits further investigation.