Sequencing paired tumor DNA and white blood cells improves circulating tumor DNA tracking and detects pathogenic germline variants in localized colon cancer.

BACKGROUND: In the setting of localized colon cancer (CC), circulating tumor DNA (ctDNA) monitoring in plasma has shown potential for detecting minimal residual disease (MRD) and predicting a higher risk of recurrence. With the tumor-only sequencing approach, however, germline variants may be misidentified as somatic variations, precluding the possibility of tracking in up to 11% of patients due to a lack of known somatic mutations. In this study, we assess the potential value of adding white blood cells (WBCs) to tumor tissue sequencing to enhance the accuracy of sequencing results. PATIENTS AND METHODS: A total of 148 patients diagnosed with localized CC were prospectively recruited at the Hospital Clínico Universitario in Valencia (Spain). Employing a custom 29-gene panel, sequencing was conducted on tumor tissue, plasma and corresponding WBCs. Droplet digital PCR and amplicon-based NGS were performed on plasma samples post-surgery to track MRD. Oncogenic somatic variants were identified by annotating with COSMIC, OncoKB and an internal repository of pathogenic mutations database. A variant prioritization analysis, mainly characterized by the match of oncogenic mutations with the evidence levels defined in OncoKB, was carried out to select specific targeted therapies. RESULTS: Utilizing paired tumor and WBCs sequencing, we identified somatic mutations in all patients (100%) within our cohort, compared to 89% using only tumor tissue. Consequently, the top 10 most frequently mutated genes for plasma monitoring were altered. The sequencing of WBCs identified 9% of patients with pathogenic mutations in the germline, with APC and TP53 being the most frequently mutated genes. Additionally, mutations in genes related to clonal hematopoiesis of indeterminate potential were detected in 27% of the cohort, with TP53, KRAS, and KMT2C being the most frequently altered genes. There were no observed differences in the sensitivity of monitoring MRD using ddPCR or amplicon-based NGS (p = 1). Ultimately, 41% of the patients harbored potentially targetable alterations at diagnosis. CONCLUSION: The germline testing method not only enhanced sequencing results and raised the proportion of patients eligible for plasma monitoring, but also uncovered the existence of pathogenic germline variations, thereby aiding in the identification of patients at a higher risk of hereditary cancer syndromes.

ESMO recommendations on the use of circulating tumour DNA assays for patients with cancer: a report from the ESMO Precision Medicine Working Group.

Circulating tumour DNA (ctDNA) assays conducted on plasma are rapidly developing a strong evidence base for use in patients with cancer. The European Society for Medical Oncology convened an expert working group to review the analytical and clinical validity and utility of ctDNA assays. For patients with advanced cancer, validated and adequately sensitive ctDNA assays have utility in identifying actionable mutations to direct targeted therapy, and may be used in routine clinical practice, provided the limitations of the assays are taken into account. Tissue-based testing remains the preferred test for many cancer patients, due to limitations of ctDNA assays detecting fusion events and copy number changes, although ctDNA assays may be routinely used when faster results will be clinically important, or when tissue biopsies are not possible or inappropriate. Reflex tumour testing should be considered following a non-informative ctDNA result, due to false-negative results with ctDNA testing. In patients treated for early-stage cancers, detection of molecular residual disease or molecular relapse, has high evidence of clinical validity in anticipating future relapse in many cancers. Molecular residual disease/molecular relapse detection cannot be recommended in routine clinical practice, as currently there is no evidence for clinical utility in directing treatment. Additional potential applications of ctDNA assays, under research development and not recommended for routine practice, include identifying patients not responding to therapy with early dynamic changes in ctDNA levels, monitoring therapy for the development of resistance mutations before clinical progression, and in screening asymptomatic people for cancer. Recommendations for reporting of results, future development of ctDNA assays and future clinical research are made.

Identifying and preventing burnout in young oncologists, an overwhelming challenge in the COVID-19 era: a study of the Spanish Society of Medical Oncology (SEOM).

BACKGROUND: Young oncologists are at particular risk of professional burnout, and this could have a significant impact on their health and care of their patients. The coronavirus disease 2019 (COVID-19) pandemic has forced rapid changes in professionals' jobs and training, with the consequent physical and psychological effects. We aimed to characterize burnout levels and determinants in young oncologists, and the effects of the pandemic on their training and health. METHODS: Two online surveys were conducted among oncology residents and young oncology specialists in Spain. The first addressed professional burnout and its determinants before the COVID-19 pandemic, while the second analyzed the impact of the pandemic on health care organization, training, and physical and psychological health in the same population. RESULTS: In total, 243 respondents completed the first survey, and 263 the second; 25.1% reported significant levels of professional burnout. Burnout was more common among medical oncology residents (28.2%), mainly in their second year of training. It was significantly associated with a poor work-life balance, inadequate vacation time, and the burnout score. Nearly three-quarters of respondents (72%) were reassigned to COVID-19 care and 84.3% of residents missed part of their training rotations. Overall, 17.2% of this population reported that they had contracted COVID-19, 37.3% had scores indicating anxiety, and 30.4% moderate to severe depression. Almost a quarter of young oncologists (23.3%) had doubts about their medical vocation. CONCLUSIONS: Burnout affects a considerable number of young oncologists. The COVID-19 pandemic has had a profound impact on causes of burnout, making it even more necessary to periodically monitor it to define appropriate detection and prevention strategies.

Biases in study design, implementation, and data analysis that distort the appraisal of clinical benefit and ESMO-Magnitude of Clinical Benefit Scale (ESMO-MCBS) scoring.

BACKGROUND: The European Society for Medical Oncology-Magnitude of Clinical Benefit Scale (ESMO-MCBS) is a validated, widely used tool developed to score the clinical benefit from cancer medicines reported in clinical trials. ESMO-MCBS scores assume valid research methodologies and quality trial implementation. Studies incorporating flawed design, implementation, or data analysis may generate outcomes that exaggerate true benefit and are not generalisable. Failure to either indicate or penalise studies with bias undermines the intention and diminishes the integrity of ESMO-MCBS scores. This review aimed to evaluate the adequacy of the ESMO-MCBS to address bias generated by flawed design, implementation, or data analysis and identify shortcomings in need of amendment. METHODS: As part of a refinement of the ESMO-MCBS, we reviewed trial design, implementation, and data analysis issues that could bias the results. For each issue of concern, we reviewed the ESMO-MCBS v1.1 approach against standards derived from Helsinki guidelines for ethical human research and guidelines from the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use, the Food and Drugs Administration, the European Medicines Agency, and European Network for Health Technology Assessment. RESULTS: Six design, two implementation, and two data analysis and interpretation issues were evaluated and in three, the ESMO-MCBS provided adequate protections. Seven shortcomings in the ability of the ESMO-MCBS to identify and address bias were identified. These related to (i) evaluation of the control arm, (ii) crossover issues, (iii) criteria for non-inferiority, (iv) substandard post-progression treatment, (v) post hoc subgroup findings based on biomarkers, (vi) informative censoring, and (vii) publication bias against quality-of-life data. CONCLUSION: Interpretation of the ESMO-MCBS scores requires critical appraisal of trials to understand caveats in trial design, implementation, and data analysis that may have biased results and conclusions. These will be addressed in future iterations of the ESMO-MCBS.

The role of tumor-associated macrophages in gastric cancer development and their potential as a therapeutic target.

Gastric cancer (GC) represents the fifth cause of cancer-related death worldwide. Molecular biology has become a central area of research in GC and there are currently at least three major classifications available to elucidate the mechanisms that drive GC oncogenesis. Further, tumor microenvironment seems to play a crucial role, and tumor-associated macrophages (TAMs) are emerging as key players in GC development. TAMs are cells derived from circulating chemokine- receptor-type 2 (CCR2) inflammatory monocytes in blood and can be divided into two main types, M1 and M2 TAMs. M2 TAMs play an important role in tumor progression, promoting a pro-angiogenic and immunosuppressive signal in the tumor. The diffuse GC subtype, in particular, seems to be strongly characterized by an immuno-suppressive and pro-angiogenic phenotype. No molecular targets in this subgroup have yet been identified. There is an urgent need to understand the molecular pathways and tumor microenvironment features in the GC molecular subtypes. The role of anti-angiogenics and checkpoint inhibitors has recently been clinically validated in GC. Both ramucirumab, a fully humanized IgG1 monoclonal anti-vascular endothelial growth factor receptor 2 (VEGFR2) antibody, and checkpoint inhibitors in Epstein Bar Virus (EBV) and Microsatellite Instable (MSI) subtypes, have proved beneficial in advanced GC. Nevertheless, there is a need to identify predictive markers of response to anti-angiogenics and immunotherapy in clinical practice for a personalized treatment approach. The importance of M2 TAMs in development of solid tumors is currently gaining increasing interest. In this literature review we analyze immune microenvironment composition and signaling related to M1 and M2 TAMs in GC as well as its potential role as a therapeutic target.

EPDR1 up-regulation in human colorectal cancer is related to staging and favours cell proliferation and invasiveness.

The finding of novel molecular markers for prediction or prognosis of invasiveness in colorectal cancer (CRC) constitutes an appealing challenge. Here we show the up-regulation of EPDR1 in a prospective cohort of 101 CRC patients, in a cDNA array of 43 patients and in in silico analyses. EPDR1 encodes a protein related to ependymins, a family of glycoproteins involved in intercellular contacts. A thorough statistical model allowed us to conclude that the gene is significantly up-regulated in tumour tissues when compared with normal mucosa. These results agree with those obtained by the analysis of three publicly available databases. EPDR1 up-regulation correlates with the TNM staging parameters, especially T and M. Studies with CRC cell lines revealed that the methylation of a CpG island controls EPDR1 expression. siRNA knocking-down and overexpression of the gene following transient plasmid transfection, showed that EPDR1 favours cell proliferation, migration, invasiveness and adhesion to type I collagen fibres, suggesting a role in epithelial to mesenchymal transition. Both statistical and functional analysis correlated EPDR1 overexpression with invasiveness and dissemination of tumour cells, supporting the inclusion of EPDR1 in panels of genes used to improve molecular subtyping of CRC. Eventually, EPDR1 may be an actionable target.

Targeted next-generation sequencing of circulating-tumor DNA for tracking minimal residual disease in localized colon cancer.

BACKGROUND: A high percentage of patients diagnosed with localized colon cancer (CC) will relapse after curative treatment. Although pathological staging currently guides our treatment decisions, there are no biomarkers determining minimal residual disease (MRD) and patients are at risk of being undertreated or even overtreated with chemotherapy in this setting. Circulating-tumor DNA (ctDNA) can to be a useful tool to better detect risk of relapse. PATIENTS AND METHODS: One hundred and fifty patients diagnosed with localized CC were prospectively enrolled in our study. Tumor tissue from those patients was sequenced by a custom-targeted next-generation sequencing (NGS) panel to characterize somatic mutations. A minimum variant allele frequency (VAF) of 5% was applied for variant filtering. Orthogonal droplet digital PCR (ddPCR) validation was carried out. We selected known variants with higher VAF to track ctDNA in the plasma samples by ddPCR. RESULTS: NGS found known pathological mutations in 132 (88%) primary tumors. ddPCR showed high concordance with NGS (r = 0.77) for VAF in primary tumors. Detection of ctDNA after surgery and in serial plasma samples during follow-up were associated with poorer disease-free survival (DFS) [hazard ratio (HR), 17.56; log-rank P = 0.0014 and HR, 11.33; log-rank P = 0.0001, respectively]. Tracking at least two variants in plasma increased the ability to identify MRD to 87.5%. ctDNA was the only significantly independent predictor of DFS in multivariable analysis. In patients treated with adjuvant chemotherapy, presence of ctDNA after therapy was associated with early relapse (HR 10.02; log-rank P < 0.0001). Detection of ctDNA at follow-up preceded radiological recurrence with a median lead time of 11.5 months. CONCLUSIONS: Plasma postoperative ctDNA detected MRD and identified patients at high risk of relapse in localized CC. Mutation tracking with more than one variant in serial plasma samples improved our accuracy in predicting MRD.

Towards precision oncology for HER2 blockade in gastroesophageal adenocarcinoma.

Gastroesophageal adenocarcinoma (GEA) represents a very heterogeneous disease and patients in advanced stages have a very poor prognosis. Although several molecular classifications have been proposed, precision medicine for HER2-amplified GEA patients still represents a challenge. Despite improvement in clinical outcomes obtained by adding trastuzumab to first-line platinum-based chemotherapy, no other anti-HER2 agents used first-line or beyond progression have demonstrated any benefit. Several factors contribute to this failure. Among them, variable HER2 amplification assessment, tumour heterogeneity, molecular mechanisms of resistance and microenvironmental factors could limit the effectiveness of anti-HER2 blockade. Identifying the factors responsible for both primary and acquired resistance is a priority for providing an improved, personalised approach. In this review, we examine current treatments for HER2-amplified GEA, their potential mechanisms of resistance and the ways to overcome them, investigating the most relevant translational studies with anti-HER2 agents in GEA, as well as novel agents under development in this field.

Carbon roadmap from syngas to polyhydroxyalkanoates in Rhodospirillum rubrum.

The gasification of organic waste materials to synthesis gas (syngas), followed by microbial fermentation, provides a significant resource for generating bioproducts such as polyhydroxyalkanoates (PHA). The anaerobic photosynthetic bacterium, Rhodospirillum rubrum, is an organism particularly attractive for the bioconversion of syngas into PHAs. In this study, a quantitative physiological analysis of R. rubrum was carried out by implementing GC-MS and HPLC techniques to unravel the metabolic pathway operating during syngas fermentation that leads to PHA production. Further, detailed investigations of the central carbon metabolites using (13) C-labelled substrate showed significant CO2 assimilation (of 40%) into cell material and PHA from syngas carbon fraction. By a combination of quantitative gene expression and enzyme activity analyses, the main role of carboxylases from the central carbon metabolism in CO2 assimilation was shown, where the Calvin-Benson-Bassham cycle (CBB) played a minor role. This knowledge sheds light about the biochemical pathways that contribute to synthesis of PHA during syngas fermentation being valuable information to further optimize the fermentation process.

Implementation and assessment of a fast-track programme to improve communication between primary and specialized care in patients with suspected cancer: how to shorten time between initial symptoms of cancer, diagnosis and initiation of treatment.

PURPOSE: This study aims to asses a cancer fast-track programme (CFP) to shorten the time since a patient with suspicion of cancer is referred by the primary care (PC) physician to the specialized medical team. METHODS: Guidelines for main suspected tumours were designed to help PC physicians to detect and rapidly refer cases to the CFP oncology coordinator, who sent them to the appropriate department to accelerate diagnosis, staging and therapy. All patients analysed in this report were referred from June 2009 to July 2012. RESULTS: A total of 897 suspected cancer cases were submitted and finally 705 were studied. In 205 (29 %) a cancer diagnosis was confirmed within 23 days (median). Therapy was initiated within 46 days after referral (median). Early diagnoses with a potential curative approach were made in 166 (82 %). CONCLUSIONS: This CFP decreased the waiting time for cancer diagnosis, by improving communication between PC physician and specialized care teams. Most patients included in this program could get therapy with curative intent.

Current questions for the treatment of advanced gastric cancer.

BACKGROUND: Gastric cancer remains a major health problem worldwide. Treatment of advanced gastric cancer is controversial and there is no standard regimen for first- or second-line chemotherapy (CT). This review aims to give an overview of the hot topics concerning treatment, prognostic factors and new strategies in advanced gastric cancer. MATERIAL AND METHODS: Seven questions of special clinical interest have been formulated previously to the literature review. With the aim of answering each of these questions, a specific search of the relevant trials and meta-analyses published or communicated from 1990 to date was performed. RESULTS: Patients treated with CT have a survival benefit over those treated with only best supportive care (BSC). Such active cytotoxic drugs as cisplatin or docetaxel and targeted agents as trastuzumab showed superiority in randomized trials. Other agents such as oxaliplatin, oral fluoropyrimidines and irinotecan showed non-inferiority or less toxic results, positioning them as valuable alternatives to classical schedules. Combination regimens seem to be an improvement over single agent therapy. However, increased toxicity of some regimens makes their general use difficult. Second-line CT is of value for selected patients with good performance status. Trastuzumab is the only targeted agent showing better survival when added to chemotherapy in HER2-driven tumors. CONCLUSIONS: With the introduction of new agents, management of advanced gastric cancer has experienced important changes. First and second-line CT improve survival in patients with good performance status. Future trials should address how to better select patients for new, targeted agents, based upon validated predictive biomarkers.

Percutaneous endocardial transfer and expression of genes to the myocardium utilizing fluoroscopic guidance.

Experimental studies indicate that administration of angiogenic proteins or genes by the epicardial or intracoronary route can stimulate development of new collateral vessels and improve myocardial perfusion. An endocardial catheter-based approach to this therapy would obviate the need for surgery, while preserving the effectiveness of direct intramyocardial administration. Fluoroscopic guidance and prototype, preformed, coaxial catheters were used to examine the feasibility of percutaneous catheter-based adenovirus (Ad)-mediated gene transfer and expression in normal swine myocardium. The feasibility of intramyocardial administration (100 microl/injection) of a radiocontrast agent and black tissue dye to all regions of the left ventricle (septum, anterior, lateral, and inferior wall) was confirmed fluoroscopically and on postmortem examination. Injections of replication-deficient adenovirus (10 injections of 10(11) particle units/100 microl each) coding for beta-galactosidase (Adbetagal) or vascular endothelial growth factor (Ad(GV)VEGF121.10) were administered to the left ventricular free wall to examine endocardial based gene transfer and expression. beta-Galactosidase activity was detected by histochemical staining and quantitative assay in targeted regions of the myocardium. Regional VEGF expression was found to be significantly greater in targeted regions (1.3 +/- 0.4 ng/mg protein) as compared with non-targeted regions (0.3 +/- 0.1 ng/mg protein) or regions injected with control (Adbetagal) virus (0.2 +/- 0.03 ng/mg protein, P < 0.001). Catheter-based Ad mediated endocardial gene transfer and expression is feasible using percutaneous, fluoroscopically guided, preformed, coaxial catheters. This approach should be clinically useful to administer angiogenic genes to the ischemic myocardium.