Intratumoral heterogeneity in gastric cancer: a new challenge to face

Abstract

Gastric cancer (GC) is the fifth most common malignancy and the third leading cause of cancer-death worldwide [1.Ferlay J. Bray F. Pisani P. Parkin M. Globocan 2002. IARC CancerBase No. 5.Cancer Incidence, Mortality and Prevalence Worldwide. IARC, Lyon2004Google Scholar], with an overall survival for advanced disease of ∼1 year. Although classically considered as a single disease, GC is highly heterogeneous from the morphological and molecular standpoints. Noteworthy, GC heterogeneity encompasses not only inter-patient variability (intertumoral heterogeneity) but also variations within the same tumor (intratumoral heterogeneity). The latter includes spatial heterogeneity in different tumor areas, and temporal heterogeneity, along progression from primary to recurrent and/or metastatic disease [2.I Gullo , FCarneiro, COliveira, GAlmeida . Heterogeneity in gastric cancer: from pure morphology to molecular classifications. Pathobiology; in press.Google Scholar]. Despite this feature, for therapeutic purposes, GC patients have been treated with identical chemo and/or radiotherapy regimens and targeted therapies [3.Wilke H. Muro K. Van Cutsem E. et al.Ramucirumab plus paclitaxel versus placebo plus paclitaxel in patients with previously treated advanced gastric or gastro-oesophageal junction adenocarcinoma (RAINBOW): a double-blind, randomised phase 3 trial.Lancet Oncol. 2014; 15: 1224-1235Abstract Full Text Full Text PDF PubMed Scopus (1641) Google Scholar, 4.Cunningham D. Starling N. Rao S. et al.Capecitabine and oxaliplatin for advanced esophagogastric cancer.N Engl J Med. 2008; 358: 36-46Crossref PubMed Scopus (1847) Google Scholar], except for the HER2 positive population [5.Bang Y.-J. Van Cutsem E. Feyereislova A. et al.Trastuzumab in combination with chemotherapy versus chemotherapy alone for treatment of HER2-positive advanced gastric or gastro-oesophageal junction cancer (ToGA): a phase 3, open-label, randomised controlled trial.Lancet. 2010; 376: 687-697Abstract Full Text Full Text PDF PubMed Scopus (5210) Google Scholar]. Intratumoral heterogeneity has been described as an inherent characteristic in most human cancers [6.Burrell R.A. McGranahan N. Bartek J. Swanton C. The causes and consequences of genetic heterogeneity in cancer evolution.Nature. 2013; 501: 338-345Crossref PubMed Scopus (1480) Google Scholar] and it is a major obstacle for an efficient diagnosis and successful molecularly driven treatment. Along with intertumoral heterogeneity, intratumoral heterogeneity has definitely affected the negative results of most targeted therapies tested in phase III trials. 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Nebozhyn M. et al.Molecular analysis of gastric cancer identifies subtypes associated with distinct clinical outcomes.Nat Med. 2015; 21: 449-456Crossref PubMed Scopus (1209) Google Scholar] highlight this heterogeneity. The landmark study of GC molecular-based stratification (based on genomic alterations) was carried out by The Cancer Genome Atlas (TCGA) research network [19.Lei Z. Tan I.B. Das K. et al.Identification of molecular subtypes of gastric cancer with different responses to PI3-kinase inhibitors and 5-fluorouracil.Gastroenterology. 2013; 145: 554-565Abstract Full Text Full Text PDF PubMed Scopus (319) Google Scholar]. It proposed a four-tiered molecular classification encompassing GCs with: (i) Epstein–Barr infection (EBV+), (ii) microsatellite instability (MSI-high), (iii) genomic stability (GS) and (iv) chromosomal instability (CIN). EBV + GC (EBVaGC) displays a specific profile identified in different studies and confirmed in Western and Asian populations [22.Setia N. Agoston A.T. Han H.S. et al.A protein and mRNA expression-based classification of gastric cancer.Mod Pathol. 2016; 29: 772-784Crossref PubMed Scopus (103) Google Scholar, 23.Ahn S. Lee S.J. Kim Y. et al.High-throughput protein and mRNA expression-based classification of gastric cancers can identify clinically distinct subtypes, concordant with recent molecular classifications.Am J Surg Pathol. 2017; 41: 106-115Crossref PubMed Scopus (63) Google Scholar]. The major features of this subtype are the male predominance, the proximal location, the good prognosis and special molecular features, such as extreme CpG island methylator phenotype and CDKN2A (p16) promoter hypermethylation, PIK3CA, ARID1A and BCOR mutations, PD-L1, PD-L2 and JAK 2 amplification [20.TCGA Comprehensive molecular characterization of gastric adenocarcinoma.Nature. 2014; 513: 202-209Crossref PubMed Scopus (3953) Google Scholar]. In this issue of Annals of Oncology, Böger et al. [24.Böger C. Krüger S. Behrens H.M. et al.Epstein–Barr virus-associated gastric cancer reveals intratumoral heterogeneity of PIK3CA mutations.Ann Oncol. 2017; 28: 1005-1024Abstract Full Text Full Text PDF PubMed Scopus (52) Google Scholar] report the results of a large scale, meticulous study, encompassing 484 GCs, in which the authors searched for EBV infection by EBER in situ hybridization and PIK3CA mutations by pyrosequencing, further extended by Sanger sequencing and next generation sequencing. Twenty-two (22/484, 5%) GCs were EBER-positive (EBVaGCs), all being of latency type I. Intratumoral heterogeneity of EBER-positivity was found in 18% of EBVaGCs. Twenty-three (23/477; 5%) GCs had PIK3CA mutations in hot spot regions of exon 9 or 20, being significantly more common in EBVaGCs than in EBV negative GCs (P<0.001). Subsequent extended sequencing of PIK3CA also demonstrated heterogeneity of PIK3CA mutations in EBVaGCs (3/9, 33%); in some cases, three to five different PIK3CA-genotypes (including wild-type) were observed in the same primary tumor, albeit in histologically and spatially distinct tumor areas. Furthermore, intratumoral heterogeneity of PIK3CA was also present in the corresponding lymph node metastases. This is the first report of intratumoral heterogeneity of PIK3CA mutations in GC, and the findings show that PIK3CA mutant and wild-type tumor sub-clones are skilled to metastasize independently to different regional lymph nodes. This study validated the enrichment of PIK3CA mutations in EBVaGCs in a large Central European patient cohort. However, the study highlighted the occurrence of intratumoral heterogeneity that, ultimately, can deprive GC patients of a suitable targeted therapy. The EBV infection and PIK3CA mutations are considered potential surrogates for target therapies, immunotherapy [25.Jin Z. Yoon HH. The promise of PD-1 inhibitors in gastro-esophageal cancers: microsatellite instability vs. PD-L1.J Gastrointest Oncol. 2016; 7: 771-788Crossref PubMed Scopus (71) Google Scholar] and mTOR-inhibitors [26.Das K. Chan X.B. Epstein D. et al.NanoString expression profiling identifies candidate biomarkers of RAD001 response in metastatic gastric cancer.ESMO Open. 2016; 1: e000009Abstract Full Text Full Text PDF PubMed Scopus (16) Google Scholar], respectively, and new diagnostic approaches should take into consideration this heterogeneous pattern of expression. This aspect is particularly relevant in GC, because a significant number of patients are diagnosed at an advanced, inoperable stage where only biopsies are available for diagnosis and subsequent biomarker testing. As emphasized by Böger et al., clinicians and pathologists need to be aware that the analysis of biopsy samples carries the risk of a non-representative, i.e. false-negative EBER–ISH, test result, which might hamper correct classification of the GC subtype and, in turn, might influence the selection of suitable patients for targeted therapy (e.g. immune therapy directed to PD-L1/PD1 checkpoint). For the progression to ‘Precision’ beyond ‘Personalized’ medicine, omics data should be validated in clinical grounds, for the development of new strategies for diagnosis (using small samples, such as liquid and endoscopic biopsies), as well as for prognostic evaluation and targeted therapy. Tumor heterogeneity introduces an additional level of complexity in this context that has to be taken into consideration in clinical applications. Prospective randomized studies in GC should be tailored according to the molecular characterization of the tumors and, therefore, pathological analyses should be expanded to identify tumor heterogeneity, also including the tumoral niche (tumor micro-environment heterogeneity), due to its putative role in cancer development and progression [27.Junttila M.R. de Sauvage FJ. Influence of tumour micro-environment heterogeneity on therapeutic response.Nature. 2013; 501: 346-354Crossref PubMed Scopus (1705) Google Scholar].