MET increased gene copy number and primary resistance to gefitinib therapy in non-small-cell lung cancer patients

Background: MET amplification has been detected in ∼20% of non-small-cell lung cancer patients (NSCLC) with epidermal growth factor receptor (EGFR) mutations progressing after an initial response to tyrosine kinase inhibitor (TKI) therapy. Patients and methods: We analyzed MET gene copy number using FISH in two related NSCLC cell lines, one sensitive (HCC827) and one resistant (HCC827 GR6) to gefitinib therapy and in two different NSCLC patient populations: 24 never smokers or EGFR FISH-positive patients treated with gefitinib (ONCOBELL cohort) and 182 surgically resected NSCLC not exposed to anti-EGFR agents. Results: HCC827 GR6-resistant cell line displayed MET amplification, with a mean MET copy number >12, while sensitive HCC827 cell line had a mean MET copy number of 4. In the ONCOBELL cohort, no patient had gene amplification and MET gene copy number was not associated with outcome to gefitinib therapy. Among the surgically resected patients, MET was amplified in 12 cases (7.3%) and only four (2.4%) had a higher MET copy number than the resistant HCC827 GR6 cell line. Conclusions: MET gene amplification is a rare event in patients with advanced NSCLC. The development of anti-MET therapeutic strategies should be focused on patients with acquired EGFR-TKI resistanc

Improving the yield of circulating tumour cells facilitates molecular characterisation and recognition of discordant HER2 amplification in breast cancer

BACKGROUND: Circulating tumour cells (CTCs) offer a non-invasive approach to obtain and characterise metastatic tumour cells, but their usefulness has been limited by low CTC yields from conventional isolation methods. METHODS: To improve CTC yields and facilitate their molecular characterisation we compared the Food and Drug Administration-approved CellSearch Epithelial Kit (CEK) to a simplified CTC capture method, CellSearch Profile Kit (CPK), on paired blood samples from patients with metastatic breast (n=75) and lung (n=71) cancer. Molecular markers including Human Epidermal growth factor Receptor 2 (HER2) were evaluated on CTCs by fluorescence in situ hybridisation (FISH) and compared to patients' primary and metastatic cancer. RESULTS: The median cell count from patients with breast cancer using the CPK was 117 vs 4 for CEK (P<0.0001). Lung cancer samples were similar; CPK: 145 cells vs CEK:4 cells (P<0.0001). Recovered CTCs were relatively pure (60-70%) and were evaluable by FISH and immunofluorescence. A total of 10 of 30 (33%) breast cancer patients with HER2-negative primary and metastatic tissue had HER2-amplified CTCs. CONCLUSION: The CPK method provides a high yield of relatively pure CTCs, facilitating their molecular characterisation. Circulating tumour cells obtained using CPK technology demonstrate that significant discordance exists between HER2 amplification of a patient's CTCs and that of the primary and metastatic tumour

Primary resistance to cetuximab therapy in EGFR FISH-positive colorectal cancer patients

The impact of KRAS mutations on cetuximab sensitivity in epidermal growth factor receptor fluorescence in situ hybridisation-positive (EGFR FISH+) metastatic colorectal cancer patients (mCRC) has not been previously investigated. In the present study, we analysed KRAS, BRAF, PI3KCA, MET, and IGF1R in 85 mCRC treated with cetuximab-based therapy in whom EGFR status was known. KRAS mutations (52.5%) negatively affected response only in EGFR FISH+ patients. EGFR FISH+/KRAS mutated had a significantly lower response rate (P=0.04) than EGFR FISH+/KRAS wild type patients. Four EGFR FISH+ patients with KRAS mutations responded to cetuximab therapy. BRAF was mutated in 5.0% of patients and none responded to the therapy. PI3KCA mutations (17.7%) were not associated to cetuximab sensitivity. Patients overexpressing IGF1R (74.3%) had significantly longer survival than patients with low IGF1R expression (P=0.006), with no difference in response rate. IGF1R gene amplification was not detected, and only two (2.6%) patients, both responders, had MET gene amplification. In conclusion, KRAS mutations are associated with cetuximab failure in EGFR FISH+ mCRC, even if it does not preclude response. The rarity of MET and IGF1R gene amplification suggests a marginal role in primary resistance. The potential prognostic implication of IGF1R expression merits further evaluation

Mutations in the LKB1 tumour suppressor are frequently detected in tumours from Caucasian but not Asian lung cancer patients

Somatic mutations of LKB1 tumour suppressor gene have been detected in human cancers including non-small cell lung cancer (NSCLC). The relationship between LKB1 mutations and clinicopathological characteristics and other common oncogene mutations in NSCLC is inadequately described. In this study we evaluated tumour specimens from 310 patients with NSCLC including those with adenocarcinoma, adenosquamous carcinoma, and squamous cell carcinoma histologies. Tumours were obtained from patients of US (n=143) and Korean (n=167) origin and screened for LKB1, KRAS, BRAF, and EGFR mutations using RT—PCR-based SURVEYOR-WAVE method followed by Sanger sequencing. We detected mutations in the LKB1 gene in 34 tumours (11%). LKB1 mutation frequency was higher in NSCLC tumours of US origin (17%) compared with 5% in NSCLCs of Korean origin (P=0.001). They tended to occur more commonly in adenocarcinomas (13%) than in squamous cell carcinomas (5%) (P=0.066). LKB1 mutations associated with smoking history (P=0.007) and KRAS mutations (P=0.042) were almost mutually exclusive with EGFR mutations (P=0.002). The outcome of stages I and II NSCLC patients treated with surgery alone did not significantly differ based on LKB1 mutation status. Our study provides clinical and molecular characteristics of NSCLC, which harbour LKB1 mutations

Acquired Resistance to KRAS (G12C) Inhibition in Cancer

BACKGROUND: Clinical trials of the KRAS inhibitors adagrasib and sotorasib have shown promising activity in cancers harboring KRAS glycine-to-cysteine amino acid substitutions at codon 12 (KRAS(G12C)). The mechanisms of acquired resistance to these therapies are currently unknown. METHODS: Among patients with KRAS(G12C) -mutant cancers treated with adagrasib monotherapy, we performed genomic and histologic analyses that compared pretreatment samples with those obtained after the development of resistance. Cell-based experiments were conducted to study mutations that confer resistance to KRAS(G12C) inhibitors. RESULTS: A total of 38 patients were included in this study: 27 with non-small-cell lung cancer, 10 with colorectal cancer, and 1 with appendiceal cancer. Putative mechanisms of resistance to adagrasib were detected in 17 patients (45% of the cohort), of whom 7 (18% of the cohort) had multiple coincident mechanisms. Acquired KRAS alterations included G12D/R/V/W, G13D, Q61H, R68S, H95D/Q/R, Y96C, and high-level amplification of the KRAS(G12C) allele. Acquired bypass mechanisms of resistance included MET amplification; activating mutations in NRAS, BRAF, MAP2K1, and RET; oncogenic fusions involving ALK, RET, BRAF, RAF1, and FGFR3; and loss-of-function mutations in NF1 and PTEN. In two of nine patients with lung adenocarcinoma for whom paired tissue-biopsy samples were available, histologic transformation to squamous-cell carcinoma was observed without identification of any other resistance mechanisms. Using an in vitro deep mutational scanning screen, we systematically defined the landscape of KRAS mutations that confer resistance to KRAS(G12C) inhibitors. CONCLUSIONS: Diverse genomic and histologic mechanisms impart resistance to covalent KRAS(G12C) inhibitors, and new therapeutic strategies are required to delay and overcome this drug resistance in patients with cancer. (Funded by Mirati Therapeutics and others; ClinicalTrials.gov number, NCT03785249.)

Lack of EGFR-activating mutations in European patients with triple-negative breast cancer could emphasise geographic and ethnic variations in breast cancer mutation profiles

INTRODUCTION: Triple-negative breast cancers (TNBCs) are characterised by lack of expression of hormone receptors and epidermal growth factor receptor 2 (HER-2). As they frequently express epidermal growth factor receptors (EGFRs), anti-EGFR therapies are currently assessed for this breast cancer subtype as an alternative to treatments that target HER-2 or hormone receptors. Recently, EGFR-activating mutations have been reported in TNBC specimens in an East Asian population. Because variations in the frequency of EGFR-activating mutations in East Asians and other patients with lung cancer have been described, we evaluated the EGFR mutational profile in tumour samples from European patients with TNBC. METHODS: We selected from a DNA tumour bank 229 DNA samples isolated from frozen, histologically proven and macrodissected invasive TNBC specimens from European patients. PCR and high-resolution melting (HRM) analyses were used to detect mutations in exons 19 and 21 of EGFR. The results were then confirmed by bidirectional sequencing of all samples. RESULTS: HRM analysis allowed the detection of three EGFR exon 21 mutations, but no exon 19 mutations. There was 100% concordance between the HRM and sequencing results. The three patients with EGFR exon 21 abnormal HRM profiles harboured the rare R836R SNP, but no EGFR-activating mutation was identified. CONCLUSIONS: This study highlights variations in the prevalence of EGFR mutations in TNBC. These variations have crucial implications for the design of clinical trials involving anti-EGFR treatments in TNBC and for identifying the potential target population

Comparison of CT and integrated PET-CT based radiation therapy planning in patients with malignant pleural mesothelioma

<p>Abstract</p> <p>Background</p> <p>When combined with adequate tumoricidal doses, accurate target volume delineation remains to be the one of the most important predictive factors for radiotherapy (RT) success in locally advanced or medically inoperable malignant pleural mesothelioma (MPM) patients. Recently, 18-fluorodeoxyglucose positron emission tomography (PET) has demonstrated significant improvements in diagnosis and accurate staging of MPM. However, role of additional PET data has not been studied in RT planning (RTP) of patients with inoperable MPM or in those who refuse surgery. Therefore, we planned to compare CT with co-registered PET-CT as the basis for delineating target volumes in these patients group.</p> <p>Methods</p> <p>Retrospectively, the CT and co-registered PET-CT data of 13 patients with histologically proven MPM were utilized to delineate target volumes separately. For each patient, target volumes (gross tumor volume [GTV], clinical target volume [CTV], and planning target volume [PTV]) were defined using the CT and PET-CT fusion data sets. The PTV was measured in two ways: PTV1 was CTV plus a 1-cm margin, and PTV2 was GTV plus a 1-cm margin. We analyzed differences in target volumes.</p> <p>Results</p> <p>In 12 of 13 patients, compared to CT-based delineation, PET-CT-based delineation resulted in a statistically significant decrease in the mean GTV, CTV, PTV1, and PTV2. In these 12 patients, mean GTV decreased by 47.1% ± 28.4%, mean CTV decreased by 38.7% ± 24.7%, mean PTV1 decreased by 31.1% ± 23.1%, and mean PTV2 decreased by 40.0% ± 24.0%. In 4 of 13 patients, hilar lymph nodes were identified by PET-CT that was not identified by CT alone, changing the nodal status of tumor staging in those patients.</p> <p>Conclusion</p> <p>This study demonstrated the usefulness of PET-CT-based target volume delineation in patients with MPM. Co-registration of PET and CT information reduces the likelihood of geographic misses, and additionally, significant reductions observed in target volumes may potentially allow escalation of RT dose beyond conventional limits potential clinical benefits in tumor control rates, which needs to be tested in future studies.</p

Assessing the clinical utility of cancer genomic and proteomic data across tumor types

Molecular profiling of tumors promises to advance the clinical management of cancer, but the benefits of integrating molecular data with traditional clinical variables have not been systematically studied. Here we retrospectively predict patient survival using diverse molecular data (somatic copy-number alteration, DNA methylation and mRNA, miRNA and protein expression) from 953 samples of four cancer types from The Cancer Genome Atlas project. We found that incorporating molecular data with clinical variables yielded statistically significantly improved predictions (FDR < 0.05) for three cancers but those quantitative gains were limited (2.2–23.9%). Additional analyses revealed little predictive power across tumor types except for one case. In clinically relevant genes, we identified 10,281 somatic alterations across 12 cancer types in 2,928 of 3,277 patients (89.4%), many of which would not be revealed in single-tumor analyses. Our study provides a starting point and resources, including an open-access model evaluation platform, for building reliable prognostic and therapeutic strategies that incorporate molecular data

Genomic and biological study of fusion genes as resistance mechanisms to EGFR inhibitors

The clinical significance of gene fusions detected by DNA-based next generation sequencing remains unclear as resistance mechanisms to EGFR tyrosine kinase inhibitors in EGFR mutant non-small cell lung cancer. By studying EGFR inhibitor-resistant patients treated with a combination of an EGFR inhibitor and a drug targeting the putative resistance-causing fusion oncogene, we identify patients who benefit and those who do not from this treatment approach. Through evaluation including RNA-seq of potential drug resistance-imparting fusion oncogenes in 504 patients with EGFR mutant lung cancer, we identify only a minority of them as functional, potentially capable of imparting EGFR inhibitor resistance. We further functionally validate fusion oncogenes in vitro using CRISPR-based editing of EGFR mutant cell lines and use these models to identify known and unknown drug resistance mechanisms to combination therapies. Collectively, our results partially reveal the complex nature of fusion oncogenes as potential drug resistance mechanisms and highlight approaches that can be undertaken to determine their functional significance.</p