The Role Of Mutations In Non-sarcomeric Genes In The Pathogenesis And Prognosis Of Hypertrophic Cardiomyopathy

Objective:The genetic basis of a considerable fraction of hypertrophic cardiomyopathy（HCM）cases remains unknown.Likewise,little was known about the contribution of non-sarcomeric genes,as well as their role in the pathogenesis and prognosis of HCM.Whether the gene encoding RNA Binding Motif Protein 20（RBM20）and the missense variants in alpha-protein kinase 3（ALPK3）are implicated in HCM and the correlation of clinical characteristics of these non-sarcomeric gene heterozygotes with HCM remain unresolved.Additionally,prior studies revealed that patients with hypertrophic cardiomyopathy（HCM）exhibited differences in severity and prognosis,indicating that there were potential HCM subtypes in these patients.We aimed at proposing a systemic subtyping classification frame to illustrate the relationship between phenotype and genotype.Methods:This study compared rare variants in the RBM20 gene by whole-exome sequencing in 793 HCM patients and 414 healthy controls.And the frequency of rare deleterious variants in ALPK3 in the HCM cohort was compared to that in the Genome Aggregation Database（gnom AD）.Based on a case-control approach,we used optimal sequence kernel association test（SKAT-O）to explore whether RBM20 and ALPK3 are both associated with HCM.The pathogenicity of these variants was further validated by family co-segregation studies.The clinical characteristics and prognoses of these non-sarcomeric gene carriers were compared with those of non-carriers.And finally,consensus clustering based on echocardiography features was performed in these patients with HCM to identify potential subtypes with an average of 32.78±27.58 month follow-up.Analyses driven by machine-learning modeling and interactome network detection techniques were subsequently conducted to explore the genetic spectrum underlying each subtype.Another independent cohort consisting of 414 patients with HCM was recruited to replicate the findings.Results:Gene-based association analysis implicated RBM20 as a susceptibility gene for developing HCM,the result of SKAT-O revealed an enrichment of RBM20 heterozygotes in the HCM population(19/701 vs.3/414,P _SKAT-O=0.0332)compared with normal controls.Patients with RBM20 variants had higher incidences of resuscitated cardiac arrest（HR 7.52,95%CI:1.94～29.08,P<0.001）,recurrent non-sustained ventricular tachycardia（NSVT）（HR 4.45,95%CI:1.24～15.97,P=0.012）and malignant arrhythmias（including SCD,SCA,recurrent NSVT and recurrent syncope）（HR 2.70,95%CI:1.14～6.40,P=0.019）,relative to those non-carriers.Rare deleterious variants in ALPK3 were significantly enriched in HCM compared with gnom AD controls（total:1.26%vs.0.29%,P=1.65e-4;missense:0.63%vs.0.15%,P=0.02;truncating:0.63%vs.0.13%,P=5.16e-4）.Further comparison and survival analysis revealed that ALPK3 heterozygotes displayed more severe hypertrophy in left ventricular segments,namely interventricular septum（IVS）（20.60 mm vs.17.14 mm,P=0.012）,left ventricular posterior wall（LVPW）（14.60 mm vs.12.11 mm,P=0.01）,and apex（14.10 mm vs.10.95 mm,P<0.001）,and a higher incidence of sudden cardiac death（SCD）（HR=9.38,95%CI:1.19～74.26,P=0.01）.Moreover,two subtypes characterized by different clinical outcomes were identified in the HCM cohort.Patients with subtype 2 presented with asymmetric septal hypertrophy that were associated with a stable course,whereas subtype 1 displayed LV systolic dysfunction and aggressive progression.Machine-learning modeling based on personal whole exome identified 46 non-sarcomeric genes whose mutation burden could accurately predict subtype propensities.Furthermore,patients in another cohort predicted as subtype1 by the 46-gene model presented with increased left ventricular end-diastolic diameter（LVEDD）and reduced left ventricular ejection fraction（LVEF）.Conclusion:This study identified that heterozygous rare deleterious variants in RBM20and ALPK3,both missense and truncating variants,are associated with HCM and suggested the two genes as potential causal genes of HCM.In addition,variants in RBM20are associated with increased risk for SCA in HCM,and variants in ALPK3 confer an increased risk for SCD.By employing echocardiography and genetic screening for the 46non-sarcomeric genes,HCM can be classified into two subtypes with distinct clinical outcomes.