Genomic Profiling Of Rare Tumors:Implications For Precisely Targeted And Immunetherapy

Purpose: There are huge differences in the definition of rare tumors worldwide: in the United States,according to data from the FDA and the National Cancer Institute(NCI),rare tumors are defined as tumors with an annual incidence of less than 15 per 100,000.In Europe,the European Society for Medical Oncology(ESMO)and the European Medicines Agency(EMA)define cancers with an annual incidence rate of less than 6 per 10,000 as rare tumors.There is no clear definition of rare tumors in our country.The emergence of precise targeted therapy guided by genotyping and immune checkpoint inhibitors has brought significant curative effects and improved survival to some patients with metastatic common malignant tumors.However,the overall five-year survival rate for malignant tumors in my country is only about35%,which is still quite different from 45% in developed countries in Europe and the United States.Part of the reason is that China does not pay enough attention to rare tumors,and patients with rare tumors lack diagnosis and treatment guidelines,clinical trials,and the latest updates.The experience of targeted and immunotherapy has resulted in the overall treatment effect of this part of patients being inferior to that of Europe and the United States.Therefore,this project aims to apply a comprehensive analysis based on the epidemiological data of rare tumors at home and abroad and standard treatment guidelines to recommend the definition of rare tumors in China.At the same time,it uses high-throughput second-generation sequencing technology to perform high-resolution analysis of biological samples from patients with rare advanced tumors.In this study we would like to provide effective reference for subsequent clinical research on targeted therapy and immunotherapy from throughput sequencing to analyze the incidence of target mutations in rare tumors and clarifying the status of biomarkers related to immunotherapy efficacy in rare tumors.Methods: Based on a comprehensive analysis of the epidemiological data of rare tumors as well as the standard treatment guidelines regarding the rare tumors inside and outside of China.This study recommends that solid tumors with an annual incidence rate of less than2.5/100,000 in China be defined as rare tumors,including 141 pathological subtypes in 12 systems.According to the new definition,among the 141 rare tumors of pathological subtypes,we conducted NGS Panel testing on the biological samples of 1312 patients to analyze the incidence of their target points,and at the same time collect the genomes of rare tumors in the c Bio Portal database Data,comparing and summarizing China's data with global data.Finally,the data of the Chinese population was compared with the data in the c Bio Portal database.TGA(Targeted gene alterations)is defined as ALK,ATM,BRAF,BRCA1,BRCA2,CDKN2 A,EGFR,ERBB2,FGFR1/2/3,KIT,MET,NF1,NTRK1/2/3,PIK3 CA,The mutations of PTEN,RET,and ROS1 are based on the target classification based on evidencebased medicine evidence in the Onco KB knowledge database.These gene mutations all have valid evidence of level 1 to 4.In addition,the subject analyzed the results of molecular markers related to the efficacy of immunotherapy on tumor samples from 852 rare tumors in China.Among them,136 patients provided tissue sections for the detection of programmed cell death ligand-1(PD-L1)expression,821 Patients with tissue samples underwent tumor mutation burden(TMB)analysis,705 patients underwent microsatellite instability(MSI)analysis,and355 patients underwent human leukocyte antigen class I(HLA-I)heterozygosity testing.The study analyzed the positive rates of the above indicators and clarified the consistency of TMB and PD-L1,TMB and MSI,and HLA-I and PD-L1 in rare tumors.Finally,the tumor tissues of 13 patients with oral squamous cell carcinoma(OTSCC)and 30 patients with salivary adenoid cystic carcinoma(ACC)were sequenced with whole exome,and the mononuclear tumors of two rare tumors were systematically reviewed,as well as the characteristics of nucleotide variation copy number variation and chromosomal variation.Results: Based on the epidemiological investigations combined with the US WHO/NCI and European ESMO research,as well as China's large population and special characteristics of tumor types,it is recommended to define tumor types with an annual incidence rate of less than 2.5/100,000 as rare tumors in China.The results showed that 515 tumor types with incidence of less than 2.5 per 100,000 per year were defined as rare tumor in China.For targeted therapy,genomic data of 4,901 patients covering 63 subtypes of rare tumor from c Bio Portal were used as the western cohort.The Chinese cohort was comprised of next generation sequencing(NGS)data of 1,312 patients from across China covering 67 subtypes.Forty-one subtypes were common between two cohorts.The accumulative prevalence of TGAs was 20.40%(1000/4901)in c Bio Portal cohort,and 53.43%(701/1312)in Chinese cohort(p < 0.001).Among those 41 overlapping subtypes,the prevalence was still significantly higher in Chinese cohort compared with c Bio Portal cohort(54.1%% vs.26.1%,p < 0.001).Generally,actionable mutations in BRAF,BRCA2,CDKN2 A,EGFR,ERBB2,KIT,MET,NF1,ROS1 were ?3 times more frequent in Chinese cohort compared with that of the c Bio Portal cohort.A total of 701 target-related mutations were detected in 1312 specimens.The top 10 targeted mutation genes were EGFR,KIT,CDKN2 A,PIK3CA,PTEN,NF1,ERBB2,BRAF,BRCA2,and FGFR1/2/3.Further analysis of the genetic variant typing of 4rare tumors(gallbladder cancer,astrocytoma,gastrointestinal stromal tumor and unidentified malignant solid tumor)with equal sample sizes in the two cohorts revealed that rare tumors in China cohort,the overall incidence of target mutations is higher.For gallbladder cancer,ERBB2 and BRCA2 mutations are more frequent in the Chinese rare tumor cohort,while ATM mutations are more frequent in the c Bio Portal cohort.For astrocytoma,BRAF,ATM,CDKN2 A and EGFR mutation/amplification are highly enriched in the cohort of rare tumors in China.For gastrointestinal stromal tumors,the incidence of KIT mutations was similar between the two groups,but the incidence of CDKN2 A and NF1 in the Chinese rare tumor cohort was significantly higher.For CUP,EGFR mutations and ALK fusions are highly enriched in rare Chinese tumors.At the same time,it was found that the incidence of TGA in rare tumors in the world is higher(26.1%),and even higher in the rare tumor population in my country(54.1%).For immunotherapy,the prevalence of PD-L1 positive,TMB-H,MSI-,and HLA-I-heterozygous were 47.8%,15.5%,7.4%,and 78.9%,respectively.The consistency ratio of TMB with PD-L1 expression,TMB with MSI status,and HLA-I type with PD-L1 expression were 54.8%(78/135),87.3%(598/685),and 47.4%(54/114),respectively.The prevalence of the four indicators varied widely across tumors subtypes.The probability that neuroendocrine tumors(NETs)and biliary tumors may benefit from immunotherapy is high,since the proportions of TMB-H are as high as 50% and 25.4% respectively.The rates of PDL1 positive,TMB-H and MSI-H patients in CUP were relatively high,while the rates of TMBH and MSI-H patients in soft tissue tumors were both relatively low.Based on the above research on molecular markers of targeted therapy and immunotherapy for rare tumors,we designed a multi-arm and multi-stage phase II platform(PLATFORM)study(NCT04423185),in order to carry out large-scale clinical research through the real-world PCT model to change the rare Current status of tumor diagnosis and treatment and improvement of prognosis.For the OTSCC and ACC,in addition to the well-characterized alterations previously reported in Caucasian patients,such as TP53 mutations(11/13,84.6%)in OTSCC and MYB-NFIB fusion in ACC(11/30,36.7%),we also observed several novel genetic changes,including FOXP1-TEX261 fusion(1/13,7.7%)and MYB-FMO5 fusion(1/30,3.3%)in OTSCC and ACC respectively,which might play a role in facilitating tumor progression and were delineated to exert functions in other tumor types.The data from this study exhibited promising translational value as numerous actionable genes were identified with mutations.Among the cohort,76.9%OTSCC and 20% salivary ACC harbored at least one alteration in actionable genes,mutations in CDKN2A(4/13,30.8%),NTRK3(3/13,23.1%)in OTSCC and FGFR2(2/30,6.7%),BRAF(2/30,6.7%)in salivary gland ACC for instance,suggesting targeted immunotherapy might be an effective intervention for these two rare malignancies.Of particular interest,instead of MYB-independent NFIB fusions previously reported in ACC,only NFIB-independent MYB fusions were found in our cohort,indicating the previous conclusion that alterations in NFIB rather than MYB might be essential to ACC tumorigenesis might be flawed.According to this study,challenges are faced in rare tumor treatment and drug development.Clinical trials are underway with innovative trial design to overcome the problem of sample exhaustion,hoping to open a new gate for rare tumor treatment.All of the above indicated a bright future for targeted therapies as well as PD-1/PD-L1 blockades in treating rare tumors.