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Background: circulating tumor DNA (ctDNA) is a source of tumor genetic material for EGFR testing in NSCLC. Real-word data about liquid biopsy (LB) clinical practice are lacking. The aim of the study was to describe the LB practice for EGFR detection in North Eastern Italy. Methods: we conducted a multi-regional survey on ctDNA testing practices in lung cancer patients. Results: Median time from blood collection to plasma separation was 50 min (20&ndash, 120 min), median time from plasma extraction to ctDNA analysis was 24 h (30 min&ndash, 5 days) and median turnaround time was 24 h (6 h&ndash, 5 days). Four hundred and seventy five patients and 654 samples were tested. One hundred and ninety-two patients were tested at diagnosis, with 16% EGFR mutation rate. Among the 283 patients tested at disease progression, 35% were T790M+. Main differences in LB results between 2017 and 2018 were the number of LBs performed for each patient at disease progression (2.88 vs. 1.2, respectively) and the percentage of T790M+ patients (61% vs. 26%).

Aside from the CTNNB1 and adenomatous polyposis coli (APC) mutations, the genetic profile of pediatric aggressive fibromatosis (AF) has remained poorly characterized. The aim of this study was to shed more light on the mutational spectrum of pediatric AF, comparing it with its adult counterpart, with a view to identifying biomarkers for use as prognostic factors or new potential therapeutic targets. CTNNB1,APC,AKT1,BRAF TP53, and RET Sanger sequencing and next‐generation sequencing (NGS) with the 50‐gene Ion AmpliSeq Cancer Hotspot Panel v2 were performed on formalin‐fixed samples from 28 pediatric and 33 adult AFs. The prognostic value of CTNNB1,AKT1, and BRAF mutations in pediatric AF patients was investigated. Recurrence‐free survival (RFS) curves were estimated with the Kaplan–Meier method and statistical comparisons were drawn using the log‐rank test. In addition to the CTNNB1 mutation (64%), pediatric AF showed AKT1 (31%), BRAF (19%), and TP53 (9%) mutations, whereas only the CTNNB1 mutation was found in adult AF. The polymorphism Q472H VEGFR was identified in both pediatric (56%) and adult (40%) AF. Our results indicate that the mutational spectrum of pediatric AF is more complex than that of adult AF, with multiple gene mutations involving not only CTNNB1 but also AKT1 and BRAF. This intriguing finding may have clinical implications and warrants further investigations.

IntroductionLittle is known about genotypic and phenotypic correlations in undifferentiated large-cell carcinoma (LCC) of the lung.MethodsThirty LCC were dissected by unsupervised targeted next generation sequencing analysis for 50 cancer-associated oncogenes and tumor suppressor genes. Cell differentiation lineages were unveiled by using thyroid transcription factor-1 (TTF1) for adenocarcinoma (ADC) and p40 for squamous cell carcinoma (SQC), dichotomizing immunohistochemistry (IHC) results for TTF1 as negative or positive (whatever its extent) and for p40 as negative, positive, or focal (if

We present a very interesting case of lung adenocarcinoma carrying an uncommon EGFR exon 19 insertion with concomitant PIK3CA mutation showing dramatic and persisting improvement upon erlotinib therapy, after progression during PI3K inhibitor.

Background There is lack of evidence about systemic treatment of pseudomyxoma peritonei (PMP) relapsing after cytoreductive surgery and hyperthermic intraperitoneal chemotherapy. There is also lack of biomarkers able to predict outcomes beyond known clinical and pathological prognostic features. Methods Fifteen patients with relapsed PMP and progressive disease within the last 6 months were included and received metronomic capecitabine (625 mg/mq/day b.i.d.) and bevacizumab (7.5 mg/Kg three-weekly) until progressive disease/unacceptable toxicity. The primary endpoint was progression-free survival (PFS). Ion Torrent® next generation sequencing technology (Hot-spot Cancer Panel) was used to characterize molecular features. Results At a median follow up of 12 months, median PFS was 8.2 months and 1-year overall survival was 91 %. Partial responses were observed in 20 % of cases, but a significant reduction of tumor markers in up to 79 %. Treatment was very well tolerated without no new safety signals. All tumor samples except one had KRAS mutations. Patients with GNAS mutations had a significantly shorter median PFS as compared to GNAS wild-type ones (5.3 months vs. not reached; p

A new somatic mutation in the coding region of Kirsten rat sarcoma viral oncogene homolog gene ( KRAS ), G48A, has been identified in a patient with non–small cell lung cancer (NSCLC). No other mutations were found by screening several genes known to be mutated in NSCLC. The patient responded to first-line therapy and is still under maintenance treatment 18 months after diagnosis. Normal and cancer cells were engineered to express the KRAS(G48A) mutation. KRAS(G48A) overexpression did not change the growth or the response to treatment compared with KRAS(wild type)-expressing cells. Analysis of the structure of the KRAS(G48A) mutant predicted altered interactions with other proteins. Analysis of KRAS binding to B-Raf proto-oncogene, serine/threonine kinase showed that the KRAS(G48A) mutant behaves more like a wild-type than a classical KRAS(G12) mutant. In conclusion, this new mutation in the coding region of KRAS , found in NSCLC, does not induce phenotypic changes similar to those induced by G12 mutants but presumably affects KRAS binding to proteins other than B-Raf proto-oncogene, serine/threonine kinase.

Background: Acquired resistance to anti-EGFR MoAbs (cetuximab and panitumumab) represents a challenge in the treatment of mCRC, but its molecular mechanisms are not completely understood. Even in presence of RAS-BRAF-PI3KCA ”quadruple wt” and HER-2/MET negative status for protein expression and gene amplification, pts who primarily respond to anti-EGFR MoAbs will eventually develop secondary resistance. Prior retrospective and small series uniquely investigated a single biomarker, showing in some cases the emergence of KRAS mutations,HER-2 or MET amplifications. Our study aimed at comprehensively describing all known molecular alterations possibly associated with acquired resistance. Methods: Pts with mCRC were prospectively treated with cetuximab- or panitumumab-based therapy until progressive disease. All archival tumors were defined as RAS-BRAF-PI3KCA”quadruple wt” by Sanger sequencing, as well as HER-2/MET negative by both immunohistochemistry (IHC) and in-situ hybridization (ISH). At the time of disease progression, tumor re-biopsy was performed on the most accessible site of metastasis, as institutional procedure for a wide phase 1 screening program. On both archival tissue and re-biopsy, next generation sequencing of 50 genes' hotspot regions included in the Hotspot Cancer Panel v2 (Life Technologies) was performed by using the Ion Torrent Personal Genome Machine platform (Life Technologies). Moreover,HER-2/MET status were repeated on tumor re-biopsy by IHC and ISH. Results: Seventeen pts were recruited. All had prior objective response to anti-EGFRs. Next-generation sequencing confirmed RAS-BRAF-PI3KCA wt status on archival tumors. The results of our analyses on tumor re-biopsies are shown in the table: IDNGS: Acquired mutations (% mutant alleles)cellularity% mutant alleles normalized for cellularityHER2 ISHMET ISH#1KRAS Q61H (37%)80%46%--#2-Amplified-#3BRAF V600E (13%)90%14%--#4NRAS Q61R (13%)40%32%--#5--Amplified#6KRAS Q61K (4%)70%6% Acquired RAS or BRAF mutations were found in 4 (23%) and 1 (6%) cases, respectively. Acquired HER-2 or MET amplification were found in 4 (23%) and 2 (12%) cases, respectively. As shown for patient #6, some degree of intra-tumor heterogeneity may exist due to concomitant presence of low represented RAS-mutated and HER-2 amplified sub-clones. In some cases (35%), a detectable acquired mechanism of resistance remains unknown. Conclusions: Based on our results, currently known molecular alterations associated with acquired resistance were mainly mutually exclusive. In a relevant subset of cases additional molecular profiling is warranted. Citation Format: Filippo Pietrantonio, Rosa Berenato, Federica Perrone, Annunziata Gloghini, Elena Tamborini, Benedetta Picciani, Adele Busico, Giulio Settanni, Chiara Costanza Volpi, Ambra Vittoria Gualeni, Alessio Pellegrinelli, Massimo Milione, Marta Caporale, Monica Niger, Maria Di Bartolomeo, Filippo de Braud. Comprehensive molecular characterization of acquired resistance to anti-EGFR monoclonal antibodies (MoAbs) in patients with metastatic colorectal cancer (mCRC). [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B57.

Background Outcome of pseudomyxoma peritonei (PMP) after cytoreductive surgery (CRS) and hypertermic intraperitoneal chemotherapy (HIPEC) is heterogeneous even after adjusting for clinico-pathological prognostic variables. The identification of additional prognostic or even predictive biomarkers is an unmet clinical need. Patients and methods Forty patients with mucinous appendiceal tumors and PMP were clinically eligible and had evaluable tumor samples obtained after CRS and HIPEC. We carried out next-generations sequencing (NGS) of 50 gene's hotspot regions contained in the Hotspot Cancer Panel v2 using the Ion Torrent Personal Genome Machine platform (Life Technologies). Results KRAS and GNAS mutations were found in 72% and 52%, and their allelic frequency was below 10% in 55% and 43% of samples, respectively. KRAS and GNAS mutations were associated with worse progression-free survival (PFS) at univariate analysis (P = 0.006 and 0.011, respectively). At multivariate analysis, only KRAS mutations were independently associated with PFS (P = 0.012); GNAS mutations were not—being significantly associated with other poor prognostic features such as incomplete cytoreduction or KRAS mutations. Validation of results was carried out in an independent bi-institutional cohort of 25 patients and the prognostic effect of KRAS mutations was again confirmed in the multivariate model (P = 0.029). NGS approach allowed the discovery of other potentially druggable mutations such as those in PI3K, AKT, LKB1, FGFR3 and PDGFRA. Conclusions Given the homogeneity of this series and the sensitivity of NGS in this low-cellularity tumor, we demonstrated for the first time a poor prognostic role of KRAS mutations.