Intratumoral heterogeneity of oncogenic drivers in mixed histology lung adenocarcinomas: How tissue selection impacts molecular testing?

Majority of the lung adenocarcinomas show a mixture of different histological patterns. The possibility of histologically heterogeneous areas of the adenocarcinoma showing genetic heterogeneity and harboring different driver mutations, with potentially significant clinical impact, has not been adequately addressed. Currently, there are no guidelines to suggest how to submit tumor tissue in adenocarcinomas with mixed histological features for molecular testing. The objective of this study is to assess intra-tumoral heterogeneity in prominent driver mutations among different morphological patterns of lung adenocarcinoma, its implications on the future of molecular testing as well as its potential impact on patient management. Twenty-three cases of mixed histology lung adenocarcinoma resected between 2018 and 2023 were retrieved from the archives. H&E slides were reviewed to identify the predominant and second most predominant histological patterns. The morphologically different tumor areas were manually macro-dissected for DNA extraction. Next-Generation Sequencing with Ion AmpliSeq™ Cancer Hotspot Panel v2 (Thermo Fisher Scientific, USA). Thirteen cases showed the same pathological variant in both histological components tested. Three cases (13 %) exhibited disparities in the variants detected across the different histological patterns tested (p=0.025). The discrepant findings had a direct therapeutic impact in 4.3 % cases. Seven cases showed no pathogenic variants detected on either of the histological components tested. This study elucidates the presence of infrequent yet significant intra-tumoral heterogeneity in the molecular profiles of mixed histology adenocarcinomas, highlighting the need for guidelines directing tissue selection for molecular testing to avoid missed therapeutic opportunities and mitigate disease relapse.

Cancer-related gene mutations and intratumoral genetic heterogeneity in human epidermal growth factor receptor 2 heterogeneous gastric cancer.

Human epidermal growth factor receptor 2 (HER2) protein overexpression is associated with HER2 gene amplification, a critical driver oncogenetic change in gastric cancer. HER2 heterogeneity in advanced gastric cancer is associated with a poor prognosis and affects the clinical efficacy of trastuzumab. However, the mechanisms of HER2 heterogeneity are not fully understood. Here, we examined whether HER2 heterogeneous gastric cancer exhibited intratumoral genetic heterogeneity in other cancer-related genes. Two cases of advanced gastric cancer with HER2 heterogeneity were selected, and samples of HER2-positive and HER2-negative areas in each case were analyzed using a cancer-associated multiple gene panel. In both cases, TP53 mutations were observed in both HER2-positive and HER2-negative areas, whereas many of the potential driver and passenger mutations differed between HER2-positive and HER2-negative areas. Overall, our findings demonstrated that HER2 heterogeneous gastric cancer exhibited intratumoral genetic heterogeneity in other cancer-related genes and that the molecular mechanisms could differ between HER2-positive and -negative areas.

Rare genetic heterogeneity within single tumor discovered for the first time in colorectal liver metastases after liver resection.

Effective individualized treatment of patients with colorectal liver metastases (CLM) requires tumor genotyping, usually based on the analysis of one single sample per patient. Therapy failure may partially be explained by sampling errors and/or intratumoral genetic heterogeneity. We aimed to demonstrate intratumoral genetic heterogeneity in CLM and enable pathologists to select tumor tissue for genotyping. All the tumors of 86 patients who underwent liver resection for a single CLM were reviewed. Of the 86 patients, 66 patients received chemotherapy and 20 patients did not receive chemotherapy before liver resection. All the tumor areas sampled were analyzed for KRAS, BRAF, PIK3CA, and NRAS mutations. The mutational status was tested in 74 cases, 7 cases had no tumoral cells due to complete responses and 5 blocks were unavailable. Of the 59/74 CLM with > 1 sample, 56 showed the same mutational status between the samples. The remaining 3 cases (5% of all cases) showed genetic heterogeneity for KRAS in 2 and BRAF in 1 patient. Genetic heterogeneity correlated with lower rate of viable tumor cells (p=0.009) and higher rate of mucin pools (p=0.013). We demonstrate for the first time the existence of genetic intratumoral heterogeneity in 5% of CLM. In routine practice, this low incidence does not require the genotyping of additional tumor samples. The correlation between the genetic heterogeneity and some histological components of the CLM should be verified by further in situ mutation assay.

Frequent somatic TERT promoter mutations and CTNNB1 mutations in hepatocellular carcinoma.

Genetic alterations of TERT and CTNNB1 have been documented in hepatocellular carcinoma. TERT promoter mutations are the earliest genetic events in the multistep process of hepatocarcinogenesis related to cirrhosis. However, analyses of TERT promoter and CTNNB1 mutations in hepatocellular carcinoma tumor samples have not been performed in the Korean population, where hepatitis B virus-related hepatocellular carcinoma is prevalent. In order to identify the role of TERT promoter and CTNNB1 mutations in the hepatocarcinogenesis and pathogenesis of recurrent hepatocellular carcinoma, we performed the sequence analyses in 140 hepatocellular nodules (including 107 hepatocellular carcinomas), and 8 pairs of matched primary and relapsed hepatocellular carcinomas. TERT promoter and CTNNB1 mutations were only observed in hepatocellular carcinomas but not in precursor lesions. Of 109 patients with hepatocellular carcinoma, 41 (39.0%) and 15 (14.6%) harbored TERT and CTNNB1 mutations, respectively. TERT promotermutations were significantly more frequent in hepatocellular carcinomas related to hepatitis C virus infection (5/6; 83.3%) compared to tumors of other etiologies (P = 0.001). In two cases, discordance in TERT promoter mutation status was observed between the primary and the corresponding recurrent hepatocellular carcinoma. The two patients with discordant cases had early relapses. In conclusion, we identified TERT promoter and CTNNB1 mutations as the most frequent somatic genetic alterations observed in hepatocellular carcinoma, indicating its pivotal role in hepatocarcinogenesis. Furthermore, we suggest the possibility of intratumoral genetic heterogeneity of TERT promoter mutations in hepatocellular carcinoma as indicated by the discordance in TERT promoter mutations between primary and corresponding recurrent hepatocellular carcinoma.

Diagnostic implications of intrapatient genetic tumor heterogeneity.

The complex genetic composition of neuroblastoma emphasizes the importance of conscientious and meticulous diagnosis. Clones with amplification or segmental chromosomal aberrations sometimes remain hidden. Several determinations should be performed when sufficient tumor material is available to establish the final diagnosis by combining the results of different techniques on tumor fragments or liquid biopsies.