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Abstract Background Gastrointestinal cancers represent one of the most prevalent diseases worldwide. Strikingly, the incidence of Early Onset Gastrointestinal Cancer (EOGIC) has been rising during the last decades and changes in lifestyle and environmental exposure seem to play a role. EOGIC has been defined as a different entity compared to on-average gastrointestinal cancer, with distinct clinical and molecular characteristics. Inherent to the particularities of younger age, there is an unmet need for a tailored approach for the management of these patients. The TEOGIC proposes a comprehensive study to characterize EOGIC patients in the northern of Spain. Methods Patients with histologically confirmed new diagnosis of colorectal, gastroesophageal and pancreatic adenocarcinoma will be considered for two cohorts: EOGIC (≤ 50 years old) and non-EOGIC (60–75 years old), with a ratio of 1:2. Two hundred and forty patients will be recruited in 4 Public Hospitals from northern Spain. After receiving unified informed consent, demographic and clinical data of the patients will be collected in a REDCap database. Lifestyle related data will be obtained in questionnaires assessing diet, physical activity and the general quality of life of the patients before diagnosis. Biological samples prior to any onco-specific treatment will be obtained for the analyses of circulating inflammatory proteins, gut microbiota, and the proteome of the tumor microenvironment. Histologic characteristics and routine biomarkers will be also collected. Thereafter, data will be integrated and analyzed to assess tumor specific, pan-tumor and sex-associated differential characteristics of EOGIC. Discussion The underlying risk factors and differential characteristics of EOGIC remain poorly studied, particularly in our geographical area. Although limited by the exploratory nature and the small sample size estimated to be recruited, TEOGIC represents the first attempt to comprehensively characterize these young patients, and thus attend to their special needs. Findings derived from this study could contribute to raise awareness and preventive behaviors in the population. In parallel, molecular studies could lead to the identification of potential novel non-invasive biomarkers and therapeutic targets that would help in the development of the tailored clinical management of these patients, focusing on screening programs for early diagnosis and precision medicine.

Introduction: Gastric cancer (GC) burden is currently evolving with regional differences associated with complex behavioural, environmental, and genetic risk factors. The LEGACy study is a Horizon 2020-funded multi-institutional research project conducted prospectively to provide comprehensive data on the tumour biological characteristics of gastroesophageal cancer from European and LATAM countries., Material and Methods: Treatment-naïve advanced gastroesophageal adenocarcinoma patients were prospectively recruited in seven European and LATAM countries. Formalin-fixed paraffin-embedded primary tumour endoscopic biopsy samples were collected and submitted for central morphological and immunohistochemical characterization and TP53 molecular assessment and Helicobacter pylori infection., Results: A total of 259 patients were included in the study: 137 (53%) from LATAM and 122 (47%) from Europe. Significant biological differences were detected between European and LATAM patients. Low representation of chromosomal instability (CIN) and HER2 positive cases were found in LATAM. MUC6 and PD-L1 were more frequently overexpressed in European cases, showing a significant correlation across the entire study population, with this association being especially pronounced in MMRdeficient cases. Both TP53 mutation by next-generation sequencing and p53 immunohistochemical aberrant pattern were linked with features associated with chromosomal instability. No regional differences were observed in H. pylori prevalence or abundance, indicating that the afore mentioned variations cannot be attributed to this factor., Conclusion: Our findings underscore a need for region-specific approaches in gastroesophageal cancer diagnosis and treatment. MUC6 emerges as a putative immune regulator that needs further investigation. Research tailored to the unique biological profiles in different global regions is crucial to effectively address the observed disparities., Competing Interests: Declarations. Conflict of interest: Dr. Alsina discloses consultancy, advisory roles, honoraria from Amgen, AstraZeneca, Beigene, Drangofly Therapeutics, Jazz Pharmaceuticals, BMS, Lilly, Novartis and MSD. Institutional research funding from Merck. FL Dr. Lordick reports institutional grants from: Astra Zeneca, Beigene, BMS, Daiichi Sankyo and Gilead, and personal fees from: Amgen, ArtTempi. Astellas, Astra Zeneca, Bayer, Biontech, BMS, Boehringer Ingelheim, Daiichi Sankyo, Eli Lilly, Gilead, Elsevier, Incyte, Medscape, MedUpdate, Merck Serono, MSD, PAGE, Roche, Servier, StreamedUp!, VJ Oncology, all outside the submitted work. Dr. Fleitas discloses advisory roles honoraria from Amgen, AstraZeneca, Beigene, BMS and MSD. Institutional research funding from Gilead. Speaker honoraria from Amgen, Servier, BMS, MSD, Lilly, Roche, Bayer. The other authors has no conflict of interests to declare. Ethics approval and consent to participate: The study has been conducted according to the principles of the Declaration of Helsinki (Fortaleza, Brazil, October 2013), following the Medical Research Involving Human Subjects Act and Good Clinical Practice standards. The study can be found under ClinicalTrials.gov Identifier: NCT04015466, July 11, 2019. The study protocol has been approved by the ethics committee of all patient-recruiting centers: the ethics committee of University Clinical Hospital of Valencia, Spain (reference number 2018/205), the institutional review board of VU University Medical Center Amsterdam (reference number 2019.355. NL 69480.02919), the ethics committee of Instituto de Previsión Social, Asuncion-Paraguay (reference number CA N°11–020/19), the ethical research committee of Instituto Alexander Fleming, Buenos Aires Argentina (Resolution July 25th, 2019, for LEGACy study 1 and 2 and October 3rd, 2019 for LEGACy study 3); the ethical committee of Instituto Nacional de Cancerología (INCAN, México (reference number INCAN/CEI/0486/19). The ethics committee of the University Center of Sao Joao and Medicine Faculty of Porto University, Portugal (reference 100/019), the scientific ethical Committee Pontificia University of Chili, reference 180806007, and the Drug research ethics committee of Valld’Hebron University Hospital, Barcelona, Spain with references PR (AG)387/2019 approved on October 29th, 2019 for LEGACy study 1, PR (AG)388/2019 approved in December 13th 2019 for LEGACy study 2 and PR (AG)419/2019 approved in January 30th, 2020 for Legacy study 3 respectively. All participants provided written informed consent before study enrolment. Each data-contributing partner has undergone online ethical and data training before the beginning of data collection and has managed access to the data of their center through this security system. Inside this system, a patient ID generator has generated a unique code for each participating patient to maintain data privacy., (© 2025. The Author(s).)

Background: Gastrointestinal cancers represent one of the most prevalent diseases worldwide. Strikingly, the incidence of Early Onset Gastrointestinal Cancer (EOGIC) has been rising during the last decades and changes in lifestyle and environmental exposure seem to play a role. EOGIC has been defined as a different entity compared to on-average gastrointestinal cancer, with distinct clinical and molecular characteristics. Inherent to the particularities of younger age, there is an unmet need for a tailored approach for the management of these patients. The TEOGIC proposes a comprehensive study to characterize EOGIC patients in the northern of Spain., Methods: Patients with histologically confirmed new diagnosis of colorectal, gastroesophageal and pancreatic adenocarcinoma will be considered for two cohorts: EOGIC (≤ 50 years old) and non-EOGIC (60-75 years old), with a ratio of 1:2. Two hundred and forty patients will be recruited in 4 Public Hospitals from northern Spain. After receiving unified informed consent, demographic and clinical data of the patients will be collected in a REDCap database. Lifestyle related data will be obtained in questionnaires assessing diet, physical activity and the general quality of life of the patients before diagnosis. Biological samples prior to any onco-specific treatment will be obtained for the analyses of circulating inflammatory proteins, gut microbiota, and the proteome of the tumor microenvironment. Histologic characteristics and routine biomarkers will be also collected. Thereafter, data will be integrated and analyzed to assess tumor specific, pan-tumor and sex-associated differential characteristics of EOGIC., Discussion: The underlying risk factors and differential characteristics of EOGIC remain poorly studied, particularly in our geographical area. Although limited by the exploratory nature and the small sample size estimated to be recruited, TEOGIC represents the first attempt to comprehensively characterize these young patients, and thus attend to their special needs. Findings derived from this study could contribute to raise awareness and preventive behaviors in the population. In parallel, molecular studies could lead to the identification of potential novel non-invasive biomarkers and therapeutic targets that would help in the development of the tailored clinical management of these patients, focusing on screening programs for early diagnosis and precision medicine., (© 2024. The Author(s).)

The search for immunotherapy biomarkers in Microsatellite Instability High/Deficient Mismatch Repair system (MSI-H/dMMR) metastatic colorectal cancer (mCRC) is an unmet need. Sixteen patients with mCRC and MSI-H/dMMR (determined by either immunohistochemistry or polymerase chain reaction) treated with PD-1/PD-L1 inhibitors at our institution were included. According to whether the progression-free survival with PD-1/PD-L1 inhibitors was longer than 6 months or shorter, patients were clustered into the IT-responder group ( n : 9 patients) or IT-resistant group ( n : 7 patients), respectively. In order to evaluate determinants of benefit with PD-1/PD-L1 inhibitors, we performed multimodal analysis including genomics (through NGS panel tumour-only with 431 genes) and the immune microenvironment (using CD3, CD8, FOXP3 and PD-L1 antibodies). The following mutations were more frequent in IT-resistant compared with IT-responder groups: B2M (4/7 versus 2/9), CTNNB1 (2/7 versus 0/9), and biallelic PTEN (3/7 versus 1/9). Biallelic ARID1A mutations were found exclusively in the IT-responder group (4/9 patients). Tumour mutational burden did not correlate with immunotherapy benefit, neither the rate of indels in homopolymeric regions. Of note, biallelic ARID1A mutated tumours had the highest immune infiltration and PD-L1 scores, contrary to tumours with CTNNB1 mutation. Immune microenvironment analysis showed higher densities of different T cell subpopulations and PD-L1 expression in IT-responders. Misdiagnosis of MSI-H/dMMR inferred by discordances between immunohistochemistry and polymerase chain reaction was only found in the IT-resistant population (3/7 patients). Biallelic ARID1A mutations and Wnt signalling activation through CTNNB1 mutation were associated with high and low T cell immune infiltrates, respectively, and deserve special attention as determinants of response to PD-1/PD-L1 inhibitors. The non-MSI-H phenotype in dMMR is associated with poor benefit to immunotherapy. Our results suggest that mechanisms of resistance to immunotherapy are multi-factorial.

Background: Eribulin is a microtubule-targeting agent approved for the treatment of advanced or metastatic breast cancer (BC) previously treated with anthracycline- and taxane-based regimens. PIK3CA mutation is associated with worse response to chemotherapy in oestrogen receptor-positive (ER+)/human epidermal growth factor receptor 2-negative (HER2-) metastatic BC. We aimed to evaluate the role of phosphoinositide 3-kinase (PI3K)/AKT pathway mutations in eribulin resistance., Methods: Resistance to eribulin was evaluated in HER2- BC cell lines and patient-derived tumour xenografts, and correlated with a mutation in the PI3K/AKT pathway., Results: Eleven out of 23 HER2- BC xenografts treated with eribulin exhibited disease progression. No correlation with ER status was detected. Among the resistant models, 64% carried mutations in PIK3CA, PIK3R1 or AKT1, but only 17% among the sensitive xenografts (P = 0.036). We observed that eribulin treatment induced AKT phosphorylation in vitro and in patient tumours. In agreement, the addition of PI3K inhibitors reversed primary and acquired resistance to eribulin in xenograft models, regardless of the genetic alterations in PI3K/AKT pathway or ER status. Mechanistically, PI3K blockade reduced p21 levels likely enabling apoptosis, thus sensitising to eribulin treatment., Conclusions: PI3K pathway activation induces primary resistance or early adaptation to eribulin, supporting the combination of PI3K inhibitors and eribulin for the treatment of HER2- BC patients.

Background: Accumulating evidence has identified Fusobacterium as an important pathogenic gut bacterium associated with colorectal cancer. Nevertheless, only limited data exist about the role of this bacterium in locally advanced rectal cancer (LARC). In this study, we quantified Fusobacterium nucleatum in untreated and post-neoadjuvant chemoradiotherapy (nCRT) samples from LARC patients and investigated its association with therapy response and survival., Patients and Methods: A total of 254 samples from 143 patients with rectal adenocarcinomas were analyzed for the presence and abundance of F. nucleatum using RNA in situ hybridization and digital image analysis. Assay accuracy was determined using infected cell lines and tumor samples with available quantitative PCR data. We studied the impact of F. nucleatum load on pathologic complete response and relapse-free survival. Treatment-induced changes were evaluated in paired pre- and post-nCRT samples (n = 71). Finally, tumor microenvironment changes during nCRT were assessed in paired samples (n = 45) by immune contexture analysis., Results: F. nucleatum tissue levels by RNA in situ hybridization strongly correlated with quantitative PCR (r = 0.804, P < 0.001). F. nucleatum abundance was higher in untreated [median, 7.4; 95% confidence interval (3.7-16.2)] compared with treated [median, 1.6; 95% confidence interval (1.3-2.4)] tumors (P <0.001) with 58% (73/126) and 26% (22/85) positive tumors, respectively (P < 0.001). Baseline F. nucleatum levels were not associated with pathologic complete response. F. nucleatum positivity after nCRT, but not baseline status, significantly increased risk of relapse [hazard ratio = 7.5, 95% confidence interval (3.0-19.0); P < 0.001]. Tumors that turned F. nucleatum-negative after nCRT had a strong increase in CD8+ T cells post-nCRT (P < 0.001), while those that persisted F. nucleatum-positive after nCRT lacked CD8+ T cells induction in post-nCRT samples compared with baseline (P = 0.69)., Conclusion: F. nucleatum persistence post-nCRT is associated with high relapse rates in LARC, potentially linked to suppression of immune cytotoxicity., Competing Interests: Disclosure JT reports personal financial interest in the form of scientific consultancy role for Array Biopharma, AstraZeneca, Bayer, BeiGene, Boehringer Ingelheim, Chugai, Genentech, Inc., Genmab A/S, Halozyme, Imugene Limited, Inflection Biosciences Limited, Ipsen, Kura Oncology, Lilly, Merck Sharp & Dohme, Menarini, Merck Serono, Merrimack, Merus, Molecular Partners, Novartis, Peptomyc, Pfizer, Pharmacyclics, ProteoDesign SL, Rafael Pharmaceuticals, F. Hoffmann-La Roche Ltd, Sanofi, SeaGen, Seattle Genetics, Servier, Symphogen, Taiho, VCN Biosciences, Biocartis, Foundation Medicine, HalioDX SAS, and Roche Diagnostics. JC reports scientific consultancy role (speaker and advisory roles) from Novartis, Pfizer, Ipsen, Exelixis, Bayer, Eisai, Advanced Accelerator Applications, Amgen, Sanofi, ITM, Sirlex and Merck Serono. Research grants from Novartis, Pfizer, AstraZeneca, Advanced Accelerator Applications, Eisai, and Bayer. RD has an advisory role at Roche and Boehringer-Ingelheim, has received a speaker’s fee from Roche, Ipsen, Amgen, Sanofi, Servier, Merck Sharp & Dohme and further received direct research funding from Merck and Pierre Fabre. PN has consulted for Bayer, Novartis, Merck Sharp & Dohme, and Targos, and received compensation. The other authors have declared no conflicts of interest., (Copyright © 2020 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Most academic precision oncology programs have been designed to facilitate enrollment of patients in early clinical trials with matched targeted agents. Over the last decade, major changes were seen both in the targetable molecular alteration landscape and in drug development trends. In this article, we describe how the Vall d'Hebron Institute of Oncology molecular prescreening program adapted to a dynamic model of biomarker-drug codevelopment. We started with a tumor-agnostic hotspot mutation panel plus in situ hybridization and immunohistochemistry of selected markers and subsequently transitioned to tumor-specific amplicon-based next-generation sequencing (NGS) tests together with custom copy number, fusion, and outlier gene expression panels. All assays are optimized for archived formalin-fixed paraffin-embedded tumor tissues without matched germline sequencing. In parallel, biomarker-matched trials evolved from a scenario of few targets and large populations (such as PI3K inhibitors in PIK3CA mutants) to a complex situation with many targets and small populations (such as multiple targetable fusion events). Recruitment rates in clinical trials with mandatory biomarkers decreased over the last 3 years. Molecular tumor board meetings proved critical to guide oncologists on emerging biomarkers for clinical testing and interpretation of NGS results. The substantial increase of immunotherapy trials had a major impact in target prioritization and guided clinical implementation of new markers, such as tumor mutational burden, with larger exon-based NGS assays and gene expression signatures to capture microenvironment infiltration patterns. This new multiomics era of precision oncology is expected to increase the opportunities for early clinical trial matching., Competing Interests: The following represents disclosure information provided by authors of this manuscript. All relationships are considered compensated unless otherwise noted. Relationships are self-held unless noted. I = Immediate Family Member, Inst = My Institution. Relationships may not relate to the subject matter of this manuscript. For more information about ASCO's conflict of interest policy, please refer to www.asco.org/rwc or ascopubs.org/po/author-center. Open Payments is a public database containing information reported by companies about payments made to US-licensed physicians (Open Payments). Rodrigo DienstmannConsulting or Advisory Role: Roche Speakers' Bureau: Roche, Ipsen, Sanofi, MSD Oncology, SERVIER, Amgen Research Funding: MerckElena GarraldaConsulting or Advisory Role: Roche, Ellipses Pharma, Neomed Therapeutics, Janssen, Boehringer Ingelheim, Seattle Genetics, TFS, Alkermes, Thermo Fisher Scientific, Bristol-Myers Squibb Speakers' Bureau: MSD, Roche, Thermo Fisher Scientific Research Funding: Novartis, Roche (Inst), Thermo Fisher Scientific (Inst) Travel, Accommodations, Expenses: Bristol-Myers Squibb, Menarini, Glycotope, Merck Sharp & DohmeAna OakninConsulting or Advisory Role: Roche, AstraZeneca, PharmaMar, Clovis Oncology, Tesaro, Immunogen, Genmab Research Funding: AbbVie Deutschland (Inst), Ability Pharmaceuticals (Inst), Advaxis (Inst), Aeterna Zentaris (Inst), Amgen (Inst), Aprea Therapeutics (Inst), Clovis Oncology (Inst), Eisai (Inst), F. Hoffmann–La Roche (Inst), Regeneron Pharmaceuticals (Inst) Travel, Accommodations, Expenses: AstraZeneca, Clovis Oncology, PharmaMar, RocheCristina SauraConsulting or Advisory Role: Puma Biotechnology, Roche, Pfizer, AstraZeneca, Celgene, Daiichi Sankyo, Eisai, Genomic Health, Novartis, Pierre Fabre, Synthon, Piqur Therapeutics, BMS, Philips Healthwork, MSD, Sanofi Research Funding: Genentech (Inst), AstraZeneca (Inst), Roche (Inst), Macrogenics (Inst), Novartis (Inst), Pfizer (Inst), Puma Biotechnology (Inst), Synthon (Inst), Piqur Therapeutics (Inst) Travel, Accommodations, Expenses: Pfizer, Novartis, Roche, AstraZeneca, Genomic Health, Puma BiotechnologyMafalda OliveiraHonoraria: Roche, Novartis, Seattle Genetics Consulting or Advisory Role: Roche/Genentech, GlaxoSmithKline, Puma Biotechnology, AstraZeneca, Seattle Genetics Research Funding: Philips Healthcare (Inst), Roche/Genentech (Inst), Novartis (Inst), AstraZeneca (Inst), Immunomedics (Inst), Seattle Genetics (Inst), Boehringer Ingelheim (Inst), GlaxoSmithKline (Inst), Cascadian Therapeutics (Inst), Sanofi (Inst), Celldex (Inst), Bayer (Inst), Piqur (Inst), Puma Biotechnology (Inst), Zenith Epigenetics (Inst) Travel, Accommodations, Expenses: Roche, Novartis, Grünenthal Group, Pierre Fabre, GP Pharm, EisaiJudith BalmañaConsulting or Advisory Role: AstraZeneca, Pfizer Research Funding: AstraZeneca (Inst), PharmaMar (Inst) Patents, Royalties, Other Intellectual Property: European patent request submitted (EP17382884.9) not related to this work Travel, Accommodations, Expenses: AstraZeneca, PharmaMarJoan CarlesConsulting or Advisory Role: Bayer, Johnson & Johnson, Bristol-Myers Squibb, Astellas Pharma, Pfizer, Sanofi, MSD Oncology, Roche, AstraZeneca Speakers' Bureau: Bayer, Johnson & Johnson, Asofarma, Astellas Pharma Research Funding: AB Science (Inst), Aragon Pharmaceuticals (Inst), Arog Pharmaceuticals (Inst), Astellas Pharma (Inst), Aveo Pharmaceuticals (Inst), Bayer AG (Inst), Blueprint Medicines (Inst), BN Immunotherapeutics (Inst), Boehringer Ingelheim España, (Inst), Bristol-Myers Squibb International (BMS) (Inst), Clovis Oncology (Inst), Cougar Biotechnology (Inst), Deciphera Pharmaceuticals (Inst), Exelixis (Inst), F. Hoffmann–La Roche (Inst), Genentech (Inst), GlaxoSmithKline (Inst), Incyte (Inst), Janssen-Cilag International (Inst), Karyopharm Therapeutics (Inst), Medimmune (Inst), Millennium Pharmaceuticals (Inst), Nanobiotix (Inst), Novartis Farmacéutica (Inst), Pfizer (Inst), Puma Biotechnology (Inst), Sanofi-AventisA (Inst), SFJ Pharma (Inst), Teva Pharma (Inst), Mediolanum Laboratoires Leurquin (Inst), Eli Lilly (Inst), AstraZeneca (Inst) Travel, Accommodations, Expenses: BMS (Inst), Ipsen (Inst), Roche (Inst), AstraZéneca (Inst)Teresa MacarullaConsulting or Advisory Role: Sanofi/Aventis, Shire, Celgene, Roche, Baxalta, QED Therapeutics, Baxter, Incyte, SERVIER, Eli Lilly, Ipsen Research Funding: Celgene (Inst), Agios Pharmaceuticals (Inst), Aslan Pharmaceuticals (Inst), Bayer (Inst), Roche (Inst), Genentech (Inst), AstraZeneca (Inst), Halozyme (Inst), Immunomedics (Inst), Eli Lilly (Inst), Merrimack (Inst), Millennium (Inst), Novartis Farmacéutica (Inst), Novocure (Inst), Pfizer (Inst), Pharmacyclics (Inst) Travel, Accommodations, Expenses: Merck, H3 Biomedicine, Sanofi, Celgene, SERVIERElena ElezConsulting or Advisory Role: Amgen, Roche, Merck Serono, Sanofi, SERVIER, Bayer, Pierre Fabre Research Funding: Merck Serono, Sanofi/Aventis (Inst) Travel, Accommodations, Expenses: Roche, Merck Serono, Sanofi, AmgenMaria AlsinaConsulting or Advisory Role: SERVIER, Bristol-Myers Squibb, Eli Lilly, MSD Oncology Speakers' Bureau: Eli Lilly, Bristol-Myers Squibb, MSD, Roche/Genentech, Amgen, SERVIER Travel, Accommodations, Expenses: Roche/Genentech, Amgen, Eli LillyIrene BrañaConsulting or Advisory Role: Bristol-Myers Squibb Speakers' Bureau: Bristol-Myers Squibb, AstraZeneca Spain, Merck Serono Research Funding: AstraZeneca, Bristol-Myers Squibb, Celgene, Gliknik, GlaxoSmithKline, Janssen Oncology, Kura, Merck Sharp & Dohme, Novartis, Orion Pharma, Pfizer, Roche Travel, Accommodations, Expenses: AstraZeneca Spain, Merck SeronoEnriqueta FelipConsulting or Advisory Role: Pfizer, Roche, Boehringer Ingelheim, AstraZeneca, Bristol-Myers Squibb, Guardant Health, Novartis, Takeda, AbbVie, Blue Print Medicines, Eli Lilly, Merck, Merck Sharp & Dohme, Janssen, Samsung Speakers' Bureau: Pfizer, Roche, AstraZeneca, Bristol-Myers Squibb, Novartis, Takeda, Eli Lilly, Merck Sharp & Dohme, Medscape, Prime Oncology, Touchtime Research Funding: Fundación Merck Salud, Grant for Oncology Innovation Other Relationship: GrífolsJosep TaberneroConsulting or Advisory Role: Bayer, Boehringer Ingelheim, Eli Lilly, MSD, Merck Serono, Novartis, Sanofi, Taiho Pharmaceutical, Merrimack, Peptomyc, Rafael Pharmaceuticals, Symphogen, Chugai Pharma, Ipsen, Merus, Pfizer, Seattle Genetics, Array BioPharma, AstraZeneca, BeiGene, Genentech, Genmab, Halozyme, Imugene Limited, Inflection Biosciences Limited, Kura, Menarini, Molecular Partners, Pharmacyclics, ProteoDesign, Roche, Seattle Genetics, Servier, VCN Biosciences, Biocartis, Foundation Medicine, HalioDX, Roche DiagnosticsJordi RodonConsulting or Advisory Role: Peptomyc, Kelun Pharmaceuticals, Merck Sharp & Dohme, Spectrum Pharmaceuticals, Pfizer, Roche/Genentech, Ellipses Pharma, Certera, Bayer, Ionctura Research Funding: Novartis, BayerPaolo NuciforoHonoraria: Bayer, Novartis, MSD Oncology, MSD Oncology Consulting or Advisory Role: Bayer, MSD Oncology Travel, Accommodations, Expenses: NovartisAna VivancosConsulting or Advisory Role: Guardant Health, Novartis, Bayer Health No other potential conflicts of interest were reported., (© 2020 by American Society of Clinical Oncology.)

Protein biomarkers are widely used in cancer diagnosis, prognosis, and prediction of treatment response. Here we introduce the use of targeted multiplex proteomics (TMP) as a tool to simultaneously measure a panel of 54 proteins involved in oncogenic, tumour suppression, drug metabolism and resistance, in patients with metastatic colorectal cancer (mCRC). TMP provided valuable diagnostic information by unmasking an occult neuroendocrine differentiation and identifying a misclassified case based on abnormal proteins phenotype. No significant differences in protein levels between unpaired primary and metastatic samples were observed. Four proteins were found differentially expressed in KRAS-mutant as compared to wild-type tumours (overexpressed in mutant: KRAS, EGFR; overexpressed in wild-type: TOPO1, TOP2A). Survival analyses revealed the association between mesothelin expression and poor overall survival, whereas lack of PTEN protein expression associated with lower progression-free survival with anti-EGFR-based therapy in the first-line setting for patients with RAS wild-type tumour. Finally, outlier analysis identified putative targetable proteins in 65% of patients lacking a targetable genomic alteration. Our data show that TMP constitutes a promising, novel molecular prescreening tool in mCRC to identify protein expression alterations that may impact on patient outcomes and more precisely guide patient eligibility to clinical trials with novel targeted experimental therapies.

Objectives: Aberrant PI3K/AKT/mTOR activation is common in gynaecological malignancies. However, predictive biomarkers of response to PI3K pathway inhibitors (PAMi) have yet to be identified., Methods: We analysed the outcomes of patients with advanced gynaecological cancer with available genomic data, treated with PAMi as single agents or in combination in phase I clinical trials. Clinical relevance of the PIK3CA mutant allele fraction (MAF) was investigated. MAF of each variant was normalised for tumour purity in the sample (adjMAFs) to infer clonality of PIK3CA mutations, defined as clonal (≥0.4) or subclonal (<0.4)., Results: A total of 50 patients with gynaecological cancer (24 ovarian; 15 endometrial; 11 cervical) with available targeted mutation profiling were selected. PAMi therapy was matched to PIK3CA/PTEN mutation in 30 patients (60%). The overall response rate, median time to progression (mTTP) and clinical benefit rate (CBR) of the entire population were 10% (N=5), 3.57 months (2.57-4.4) and 40% (N=18), respectively. Genotype-matched therapy did not lead to a favourable CBR (OR 0.91, p=1 (0.2-3.7)) or mTTP (3.57 months (2.6-4.4) vs 3.73 months (1.9-13.2); HR 1.41; p=0.29). We did not detect differences in mTTP according to therapy or PIK3CA codon mutation (HR 1.71, p=0.24). Overall, 41% of patients had a TTP ratio (TTP PAMi/TTP on immediately prior or subsequent palliative chemotherapy) ≥1.3, without statistically significant differences according to tumour type (p=0.39), molecular alteration status (p=0.13) or therapy (p=0.54). In univariate analysis, genotype-matched therapy in patients with PIK3CA clonal events was associated with improved mTTP (HR 3.6; p=0.03)., Conclusions: Our study demonstrates that patients with advanced gynaecological cancer, refractory to standard therapies, achieved meaningful clinical benefit from PAMi. The impact of PI3KCA clonality on response to selected PAMi in patients with gynaecological cancer deserves further investigation., Competing Interests: Competing interests: AO is a consultant/advisory board member of AstraZeneca, Clovis, Tesaro, Roche and PharmaMar. RD reports personal fees from Roche, outside the submitted work.

Sequencing of tumors is now routine and guides personalized cancer therapy. Mutant allele fractions (MAFs, or the 'mutation dose') of a driver gene may reveal the genomic structure of tumors and influence response to targeted therapies. We performed a comprehensive analysis of MAFs of driver alterations in unpaired primary and metastatic colorectal cancer (CRC) at our institution from 2010 to 2015 and studied their potential clinical relevance. Of 763 CRC samples, 622 had detailed annotation on overall survival in the metastatic setting (OSmet) and 89 received targeted agents matched to KRAS (MEK inhibitors), BRAF (BRAF inhibitors), or PIK3CA mutations (PI3K pathway inhibitors). MAFs of each variant were normalized for tumor purity in the sample (adjMAFs). We found lower adjMAFs for BRAF V 600E and PIK3CA than for KRAS, NRAS, and BRAF non-V600 variants. TP53 and BRAF V 600E adjMAFs were higher in metastases as compared to primary tumors, and high KRAS adjMAFs were found in CRC metastases of patients with KRAS wild-type primary tumors previously exposed to EGFR antibodies. Patients with RAS- or BRAF V 600E -mutated tumors, irrespective of adjMAFs, had worse OSmet. There was no significant association between adjMAFs and time to progression on targeted therapies matched to KRAS, BRAF, or PIK3CA mutations, potentially related to the limited antitumor activity of the employed drugs (overall response rate of 4.5%). In conclusion, the lower BRAF V 600E and PIK3CA adjMAFs in subsets of primary CRC tumors indicate subclonality of these driver genes. Differences in adjMAFs between metastases and primary tumors suggest that approved therapies may result in selection of BRAF V 600E - and KRAS-resistant clones and an increase in genomic heterogeneity with acquired TP53 alterations. Despite significant differences in prognosis according to mutations in driver oncogenes, adjMAFs levels did not impact on survival and did not help predict benefit with matched targeted agents in the metastatic setting., (© 2017 The Authors. Published by FEBS Press and John Wiley & Sons Ltd.)