Clinical validation of a cell-free DNA fragmentome assay for augmentation of lung cancer early detection.

Lung cancer screening via annual low-dose computed tomography (LDCT) has poor adoption. We conducted a prospective case-control study among 958 individuals eligible for lung cancer screening to develop a blood-based lung cancer detection test that when positive is followed by an LDCT. Changes in genome-wide cell-free DNA (cfDNA) fragmentation profiles (fragmentomes) in peripheral blood reflected genomic and chromatin characteristics of lung cancer. We applied machine learning to fragmentome features to identify individuals who were more or less likely to have lung cancer. We trained the classifier using 576 cases and controls from study samples, and then validated it in a held-out group of 382 cases and controls. The validation demonstrated high sensitivity for lung cancer, and consistency across demographic groups and comorbid conditions. Applying test performance to the screening eligible population in a five-year model with modest utilization assumptions suggested the potential to prevent thousands of lung cancer deaths.

Genetic variations in immune mediators and prostate cancer risk: A field synopsis with Bayesian calculations.

OBJECTIVE: Our study aimed to revaluate the significant data from meta-analyses on genetic variations in immune mediators and the risk of prostate cancer (PCa) by Bayesian approaches. METHODS: We performed a search on the literature before September 5th, 2023, for meta-analytic studies on polymorphisms in immune mediator genes and the risk of PCa. Two Bayesian approaches were used to assess the level of noteworthiness in the meta-analytic data: the False-Positive Rate Probability (FPRP) and the Bayesian False Discovery Probability (BFDP) with a statistical power of 1.2 and 1.5 of Odds Ratio at a prior probability of 10-3 and 10-6. The quality evaluation of studies was performed with the Venice criteria. Gene-gene and protein-protein networks were designed for the genes and products enrolled in the results. RESULTS: As results, 18 meta-analyses on 17 polymorphisms in several immune mediator genes were included (IL1B rs16944/rs1143627, IL4 rs2243250/rs2227284/rs2070874, IL6 1800795/rs1800796/rs1800797, IL8 rs4073, IL10 rs1800896/rs1800871/rs1800872, IL18 rs1946518, COX2 rs2745557, TNFA rs361525 and PTGS2 rs20417/689470). The Bayesian calculations showed the rs1143627 and the rs1946518 polymorphisms in IL1B and IL18 genes, respectively, were noteworthy. The Venice criteria showed that only four studies received the highest level of evidence. The gene-gene and protein-protein networks reinforced the findings on IL1B and IL18 genes. CONCLUSION: In conclusion, this current Bayesian revaluation showed that the rs1143627 and the rs1946518 polymorphisms in the IL1B and IL18 genes, respectively, were noteworthy biomarker candidates for PCa risk.

Elucidating the Heterogeneity of Immunotherapy Response and Immune-Related Toxicities by Longitudinal ctDNA and Immune Cell Compartment Tracking in Lung Cancer.

PURPOSE: Although immunotherapy is the mainstay of therapy for advanced non-small cell lung cancer (NSCLC), robust biomarkers of clinical response are lacking. The heterogeneity of clinical responses together with the limited value of radiographic response assessments to timely and accurately predict therapeutic effect-especially in the setting of stable disease-calls for the development of molecularly informed real-time minimally invasive approaches. In addition to capturing tumor regression, liquid biopsies may be informative in capturing immune-related adverse events (irAE). EXPERIMENTAL DESIGN: We investigated longitudinal changes in circulating tumor DNA (ctDNA) in patients with metastatic NSCLC who received immunotherapy-based regimens. Using ctDNA targeted error-correction sequencing together with matched sequencing of white blood cells and tumor tissue, we tracked serial changes in cell-free tumor load (cfTL) and determined molecular response. Peripheral T-cell repertoire dynamics were serially assessed and evaluated together with plasma protein expression profiles. RESULTS: Molecular response, defined as complete clearance of cfTL, was significantly associated with progression-free (log-rank P = 0.0003) and overall survival (log-rank P = 0.01) and was particularly informative in capturing differential survival outcomes among patients with radiographically stable disease. For patients who developed irAEs, on-treatment peripheral blood T-cell repertoire reshaping, assessed by significant T-cell receptor (TCR) clonotypic expansions and regressions, was identified on average 5 months prior to clinical diagnosis of an irAE. CONCLUSIONS: Molecular responses assist with the interpretation of heterogeneous clinical responses, especially for patients with stable disease. Our complementary assessment of the peripheral tumor and immune compartments provides an approach for monitoring of clinical benefits and irAEs during immunotherapy.

Elucidating the heterogeneity of immunotherapy response and immune-related toxicities by longitudinal ctDNA and immune cell compartment tracking in lung cancer.

Purpose: Although immunotherapy is the mainstay of therapy for advanced non-small cell lung cancer (NSCLC), robust biomarkers of clinical response are lacking. The heterogeneity of clinical responses together with the limited value of radiographic response assessments to timely and accurately predict therapeutic effect -especially in the setting of stable disease-call for the development of molecularly-informed real-time minimally invasive predictive biomarkers. In addition to capturing tumor regression, liquid biopsies may be informative in evaluating immune-related adverse events (irAEs). Experimental design: We investigated longitudinal changes in circulating tumor DNA (ctDNA) in patients with metastatic NSCLC who received immunotherapy-based regimens. Using ctDNA targeted error-correction sequencing together with matched sequencing of white blood cells and tumor tissue, we tracked serial changes in cell-free tumor load (cfTL) and determined molecular response for each patient. Peripheral T-cell repertoire dynamics were serially assessed and evaluated together with plasma protein expression profiles. Results: Molecular response, defined as complete clearance of cfTL, was significantly associated with progression-free (log-rank p=0.0003) and overall survival (log-rank p=0.01) and was particularly informative in capturing differential survival outcomes among patients with radiographically stable disease. For patients who developed irAEs, peripheral blood T-cell repertoire reshaping, assessed by significant TCR clonotypic expansions and regressions were noted on-treatment. Conclusions: Molecular responses assist with interpretation of heterogeneous clinical responses especially for patients with stable disease. Our complementary assessment of the tumor and immune compartments by liquid biopsies provides an approach for monitoring of clinical benefit and immune-related toxicities for patients with NSCLC receiving immunotherapy. Statement of translational relevance: Longitudinal dynamic changes in cell-free tumor load and reshaping of the peripheral T-cell repertoire capture clinical outcomes and immune-related toxicities during immunotherapy for patients with non-small cell lung cancer.

What We Know about Euterpe Genus and Neuroprotection: A Scoping Review.

The Euterpe genus (mainly Euterpe oleracea Martius, Euterpe precatoria Martius, and Euterpe edulis Martius) has recently gained commercial and scientific notoriety due to the high nutritional value of its fruits, which are rich in polyphenols (phenolic acids and anthocyanins) and have potent antioxidant activity. These characteristics have contributed to the increased number of neuropharmacological evaluations of the three species over the last 10 years, especially açaí of the species Euterpe oleracea Martius. The fruits of the three species exert neuroprotective effects through the modulation of inflammatory and oxidative pathways and other mechanisms, including the inhibition of the mTOR pathway and protection of the blood-brain barrier, all of them intimately involved in several neuropathologies. Thus, a better understanding of the neuropharmacological properties of these three species may open new paths for the development of therapeutic tools aimed at preventing and treating a variety of neurological conditions.

Cell-Free DNA Fragmentomes in the Diagnostic Evaluation of Patients With Symptoms Suggestive of Lung Cancer.

BACKGROUND: The diagnostic workup of individuals suspected of having lung cancer can be complex and protracted because conventional symptoms of lung cancer have low specificity and sensitivity. RESEARCH QUESTION: Among individuals with symptoms of lung cancer, can a blood-based approach to analyze cell-free DNA (cfDNA) fragmentation (the DNA evaluation of fragments for early interception [DELFI] score) enhance evaluation for the possible presence of lung cancer? STUDY DESIGN AND METHODS: Adults were referred to Bispebjerg Hospital (Copenhagen, Denmark) for diagnostic evaluation of initial imaging anomalies and symptoms consistent with lung cancer. Numbers and types of symptoms were extracted from medical records. cfDNA from plasma samples obtained at the prediagnostic visit was isolated, sequenced, and analyzed for genome-wide cfDNA fragmentation patterns. The relationships among clinical presentation, cancer status, and DELFI score were examined. RESULTS: A total of 296 individuals were analyzed. Median DELFI scores were higher for those with lung cancer (n = 98) than those without cancer (n = 198; 0.94 vs 0.19; P < .001). In a multivariate model adjusted for age, smoking history, and presenting symptoms, the addition of the DELFI score improved the prediction of lung cancer for those who demonstrated symptoms (area under the receiver operating characteristic curve, 0.74-0.94). INTERPRETATION: The DELFI score distinguishes individuals with lung cancer from those without cancer better than suspicious symptoms do. These results represent proof-of-concept support that fragmentation-based biomarker approaches may facilitate diagnostic resolution for patients with concerning symptoms of lung cancer.

Gene variations related to the hepatocellular carcinoma: Results from a field synopsis and Bayesian revaluation.

Hepatocellular carcinoma (HCC) is considered as the second cause of cancer-related deaths worldwide. Genetic variations are associated with HCC risk, an issue that has been the subject of several meta-analyses. However, meta-analyses have an important limitation on the likelihood of false positive data. Henceforth, this study aimed to assess the level of noteworthiness in the meta-analyses by means of a Bayesian approach. A systematic search was performed for meta-analyses with associations between gene polymorphisms and HCC. The calculations for the False-Positive Rate Probability (FPRP) and the Bayesian False Discovery Probability (BFDP) were performed to assess the noteworthiness with a statistical power of 1.2 and 1.5 of Odds Ratio at a prior probability of 10-3 and 10-5. The quality of studies was evaluated by the Venice criteria. As additional analyses, the gene-gene and protein-protein networks were designed for these genes and products. As results, we found 33 meta-analytic studies on 45 polymorphisms occurring in 35 genes. A total of 1,280 values for FPRP and BFDP were obtained. Seventy-five for FPRP (5.86%) and 95 for BFDP (14.79%) were noteworthy. In conclusion, the polymorphisms in CCND1, CTLA4, EGF, IL6, IL12A, KIF1B, MDM2, MICA, miR-499, MTHFR, PNPLA3, STAT4, TM6SF2, and XPD genes were considered as noteworthy biomarkers for HCC risk.

Metastatic Colorectal Cancer Treatment Response Evaluation by Ultra-Deep Sequencing of Cell-Free DNA and Matched White Blood Cells.

PURPOSE: Circulating tumor DNA (ctDNA) has the potential to guide therapy selection and monitor treatment response in patients with metastatic cancer. However, germline and clonal hematopoiesis-associated alterations can confound identification of tumor-specific mutations in cell-free DNA (cfDNA), often requiring additional sequencing of tumor tissue. The current study assessed whether ctDNA-based treatment response monitoring could be performed in a tumor tissue-independent manner by combining ultra-deep targeted sequencing analyses of cfDNA with patient-matched white blood cell (WBC)-derived DNA. EXPERIMENTAL DESIGN: In total, 183 cfDNA and 49 WBC samples, along with 28 tissue samples, from 52 patients with metastatic colorectal cancer participating in the prospective phase III CAIRO5 clinical trial were analyzed using an ultra-deep targeted sequencing liquid biopsy assay. RESULTS: The combined cfDNA and WBC analysis prevented false-positives due to germline or hematopoietic variants in 40% of patients. Patient-matched tumor tissue sequencing did not provide additional information. Longitudinal analyses of ctDNA were more predictive of overall survival than standard-of-care radiological response evaluation. ctDNA mutations related to primary or acquired resistance to panitumumab were identified in 42% of patients. CONCLUSIONS: Accurate calling of ctDNA mutations for treatment response monitoring is feasible in a tumor tissue-independent manner by combined cfDNA and patient-matched WBC genomic DNA analysis. This tissue biopsy-independent approach simplifies sample logistics and facilitates the application of liquid biopsy ctDNA testing for evaluation of emerging therapy resistance, opening new avenues for early adaptation of treatment regimens.

Natural Language Processing Approaches for Retrieval of Clinically Relevant Genomic Information in Cancer.

The accelerating impact of genomic data in clinical decision-making has generated a paradigm shift from treatment based on the anatomic origin of the tumor to the incorporation of key genomic features to guide therapy. Assessing the clinical validity and utility of the genomic background of a patient's cancer represents one of the emerging challenges in oncology practice, demanding the development of automated platforms for extracting clinically relevant genomic information from medical texts. We developed PubMiner, a natural language processing tool to extract and interpret cancer type, therapy, and genomic information from biomedical abstracts. Our initial focus has been the retrieval of gene names, variants, and negations, where PubMiner performed highly in terms of total recall (91.7%) with a precision of 79.7%. Our next steps include developing a web-based interface to promote personalized treatment based on each tumor's unique genomic fingerprints.

KRAS A146 Mutations Are Associated With Distinct Clinical Behavior in Patients With Colorectal Liver Metastases.

Somatic KRAS mutations occur in approximately half of the patients with metastatic colorectal cancer (mCRC). Biologic tumor characteristics differ on the basis of the KRAS mutation variant. KRAS mutations are known to influence patient prognosis and are used as predictive biomarker for treatment decisions. This study examined clinical features of patients with mCRC with a somatic mutation in KRAS G12, G13, Q61, K117, or A146. METHODS: A total of 419 patients with colorectal cancer with initially unresectable liver-limited metastases, who participated in a multicenter prospective trial, were evaluated for tumor tissue KRAS mutation status. For the subgroup of patients who carried a KRAS mutation and were treated with bevacizumab and doublet or triplet chemotherapy (N = 156), pretreatment circulating tumor DNA levels were analyzed, and total tumor volume (TTV) was quantified on the pretreatment computed tomography images. RESULTS: Most patients carried a KRAS G12 mutation (N = 112), followed by mutations in G13 (N = 15), A146 (N = 12), Q61 (N = 9), and K117 (N = 5). High plasma circulating tumor DNA levels were observed for patients carrying a KRAS A146 mutation versus those with a KRAS G12 mutation, with median mutant allele frequencies of 48% versus 19%, respectively. Radiologic TTV revealed this difference to be associated with a higher tumor load in patients harboring a KRAS A146 mutation (median TTV 672 cm3 [A146] v 74 cm3 [G12], P = .036). Moreover, KRAS A146 mutation carriers showed inferior overall survival compared with patients with mutations in KRAS G12 (median 10.7 v 26.4 months; hazard ratio = 2.5; P = .003). CONCLUSION: Patients with mCRC with a KRAS A146 mutation represent a distinct molecular subgroup of patients with higher tumor burden and worse clinical outcomes, who might benefit from more intensive treatments. These results highlight the importance of testing colorectal cancer for all KRAS mutations in routine clinical care.

Detection and characterization of lung cancer using cell-free DNA fragmentomes.

Non-invasive approaches for cell-free DNA (cfDNA) assessment provide an opportunity for cancer detection and intervention. Here, we use a machine learning model for detecting tumor-derived cfDNA through genome-wide analyses of cfDNA fragmentation in a prospective study of 365 individuals at risk for lung cancer. We validate the cancer detection model using an independent cohort of 385 non-cancer individuals and 46 lung cancer patients. Combining fragmentation features, clinical risk factors, and CEA levels, followed by CT imaging, detected 94% of patients with cancer across stages and subtypes, including 91% of stage I/II and 96% of stage III/IV, at 80% specificity. Genome-wide fragmentation profiles across ~13,000 ASCL1 transcription factor binding sites distinguished individuals with small cell lung cancer from those with non-small cell lung cancer with high accuracy (AUC = 0.98). A higher fragmentation score represented an independent prognostic indicator of survival. This approach provides a facile avenue for non-invasive detection of lung cancer.

Impact of COVID-19 pandemic on care of oncological patients: experience of a cancer center in a Latin American pandemic epicenter / Impacto da COVID-19 sobre o atendimento de pacientes oncológicos: experiência de um centro oncológico localizado em um epicentro Latino-Americano da pandemia

ABSTRACT Objective Since the rising of coronavirus disease 2019 (COVID-19) pandemic, there is uncertainty regarding the impact of transmission to cancer patients. Evidence on increased severity for patients undergoing antineoplastic treatment is posed against deferring oncologic treatment. We aimed to evaluate the impact of COVID-19 pandemic on patient volumes in a cancer center in an epicenter of the pandemic. Methods Outpatient and inpatient volumes were extracted from electronic health record database. Two intervals were compared: pre-COVID-19 (March to May 2019) and COVID-19 pandemic (March to May 2020) periods. Results The total number of medical appointments declined by 45% in the COVID-19 period, including a 56.2% decrease in new visits. There was a 27.5% reduction in the number of patients undergoing intravenous systemic treatment and a 57.4% decline in initiation of new treatments. Conversely, there was an increase by 309% in new patients undergoing oral chemotherapy regimens and a 5.9% rise in new patients submitted to radiation therapy in the COVID-19 period. There was a 51.2% decline in length of stay and a 60% reduction in the volume of surgical cases during COVID-19. In the stem cell transplant unit, we observed a reduction by 36.5% in length of stay and a 62.5% drop in stem cell transplants. Conclusion A significant decrease in the number of patients undergoing cancer treatment was observed after COVID-19 pandemic. Although this may be partially overcome by alternative therapeutic options, avoiding timely health care due to fear of getting COVID-19 infection might impact on clinical outcomes. Our findings may help support immediate actions to mitigate this hypothesis.

RESUMO Objetivo Desde o surgimento da pandemia da doença pelo coronavírus 2019 (COVID-19), há incerteza quanto ao impacto da transmissão para pacientes com câncer. As evidências sobre o aumento da gravidade para pacientes submetidos a tratamento antineoplásico são contra o adiamento do tratamento oncológico. Nosso objetivo foi avaliar o impacto da pandemia de COVID-19 em volumes de pacientes em um centro oncológico, em um epicentro da pandemia. Métodos Os volumes de pacientes ambulatoriais e de internação foram extraídos do banco de dados de prontuários eletrônicos. Dois intervalos foram comparados: períodos pré-COVID-19 (março a maio de 2019) e pandemia COVID-19 (março a maio de 2020). Resultados O número total de consultas médicas diminuiu 45% no período pandemia COVID-19, inclusive com redução de 56,2% nas novas consultas. Houve redução de 27,5% no número de pacientes em tratamento sistêmico intravenoso e de 57,4% no início de novos tratamentos. Por outro lado, ocorreram aumento de 309% em novos pacientes submetidos a regimes de quimioterapia oral e elevação de 5,9% em novos pacientes submetidos à radioterapia no período pandemia COVID-19. Observaram-se queda de 51,2% nos dias de internação e redução de 60% no volume de casos cirúrgicos durante a COVID-19. Na unidade de transplante de células-tronco, a redução foi de 36,5% nos dias de internação e de 62,5% nos transplantes de células-tronco. Conclusão Foi observado declínio significativo no número de pacientes em tratamento de câncer após a pandemia de COVID-19. Embora isso possa ser parcialmente superado por opções terapêuticas alternativas, evitar cuidados de saúde oportunos devido ao medo de contrair COVID-19 pode impactar nos resultados clínicos. Nossos resultados podem ajudar a apoiar ações imediatas para mitigar essa hipótese.

Impact of COVID-19 pandemic on care of oncological patients: experience of a cancer center in a Latin American pandemic epicenter.

OBJECTIVE: Since the rising of coronavirus disease 2019 (COVID-19) pandemic, there is uncertainty regarding the impact of transmission to cancer patients. Evidence on increased severity for patients undergoing antineoplastic treatment is posed against deferring oncologic treatment. We aimed to evaluate the impact of COVID-19 pandemic on patient volumes in a cancer center in an epicenter of the pandemic. METHODS: Outpatient and inpatient volumes were extracted from electronic health record database. Two intervals were compared: pre-COVID-19 (March to May 2019) and COVID-19 pandemic (March to May 2020) periods. RESULTS: The total number of medical appointments declined by 45% in the COVID-19 period, including a 56.2% decrease in new visits. There was a 27.5% reduction in the number of patients undergoing intravenous systemic treatment and a 57.4% decline in initiation of new treatments. Conversely, there was an increase by 309% in new patients undergoing oral chemotherapy regimens and a 5.9% rise in new patients submitted to radiation therapy in the COVID-19 period. There was a 51.2% decline in length of stay and a 60% reduction in the volume of surgical cases during COVID-19. In the stem cell transplant unit, we observed a reduction by 36.5% in length of stay and a 62.5% drop in stem cell transplants. CONCLUSION: A significant decrease in the number of patients undergoing cancer treatment was observed after COVID-19 pandemic. Although this may be partially overcome by alternative therapeutic options, avoiding timely health care due to fear of getting COVID-19 infection might impact on clinical outcomes. Our findings may help support immediate actions to mitigate this hypothesis.

Diagnostic Strategies toward Clinical Implementation of Liquid Biopsy RAS/BRAF Circulating Tumor DNA Analyses in Patients with Metastatic Colorectal Cancer.

Detection of KRAS, NRAS, and BRAF mutations in tumor tissue is currently used to predict resistance to treatment with anti-epidermal growth factor receptor (EGFR) antibodies in patients with metastatic colorectal cancer (mCRC). Liquid biopsies are minimally invasive, and cell-free circulating tumor DNA (ctDNA) mutation analyses may better represent tumor heterogeneity. This study examined the incorporation of liquid biopsy RAS/BRAF ctDNA analyses into diagnostic strategies to determine mCRC patient eligibility for anti-EGFR therapy. Tumor tissue and liquid biopsies were collected from 100 mCRC patients with liver-only metastases in a multicenter prospective clinical trial. Three diagnostic strategies incorporating droplet digital PCR ctDNA analyses were compared with routine tumor tissue RAS/BRAF mutation profiling using decision tree analyses. Tissue DNA mutations in KRAS, NRAS, and BRAF were present in 54%, 0%, and 3% of mCRC patients, respectively. A 93% concordance was observed between tissue DNA and liquid biopsy ctDNA mutations. The proportion of patients with RAS/BRAF alterations increased from 57% to 60% for diagnostic strategies that combined tissue and liquid biopsy mutation analyses. Consecutive RAS/BRAF ctDNA analysis followed by tissue DNA analysis in case of a liquid biopsy-negative result appeared to be the most optimal diagnostic strategy to comprehensively determine eligibility for anti-EGFR therapy in a cost-saving manner. These results highlight the potential clinical utility of liquid biopsies for detecting primary resistance to anti-EGFR-targeted therapies.

Applications of cell-free circulating tumor DNA detection in EGFR mutant lung cancer.

Analyses of cell-free tumor DNA (ctDNA) have provided a non-invasive strategy for cancer diagnosis, the identification of molecular aberrations for treatment identification, and evaluation of tumor response. Sensitive and specific ctDNA sequencing strategies have allowed for implementation into clinical practice for the initial genotyping of patients and resistance monitoring. The specific need for EGFR mutation detection for the management of lung cancer patients has been an early imperative and has set the stage for non-invasive molecular profiling across other oncogenic drivers. Ongoing efforts are demonstrating the utility of ctDNA analyses in the initial genotyping of patients, the monitoring resistance clones, and the initial evaluation of response.

White blood cell and cell-free DNA analyses for detection of residual disease in gastric cancer.

Liquid biopsies are providing new opportunities for detection of residual disease in cell-free DNA (cfDNA) after surgery but may be confounded through identification of alterations arising from clonal hematopoiesis. Here, we identify circulating tumor-derived DNA (ctDNA) alterations through ultrasensitive targeted sequencing analyses of matched cfDNA and white blood cells from the same patient. We apply this approach to analyze samples from patients in the CRITICS trial, a phase III randomized controlled study of perioperative treatment in patients with operable gastric cancer. After filtering alterations from matched white blood cells, the presence of ctDNA predicts recurrence when analyzed within nine weeks after preoperative treatment and after surgery in patients eligible for multimodal treatment. These analyses provide a facile method for distinguishing ctDNA from other cfDNA alterations and highlight the utility of ctDNA as a predictive biomarker of patient outcome to perioperative cancer therapy and surgical resection in patients with gastric cancer.

Tissue and Cell-Free DNA-Based Epigenomic Approaches for Cancer Detection.

BACKGROUND: Over 9 million people die of cancer each year worldwide, reflecting the unmet need for effective biomarkers for both cancer diagnosis and prognosis. Cancer diagnosis is complex because the majority of malignant tumors present with long periods of latency and lack of clinical presentation at early stages. During carcinogenesis, premalignant cells experience changes in their epigenetic landscapes, such as differential DNA methylation, histone modifications, nucleosome positioning, and higher orders of chromatin changes that confer growth advantage and contribute to determining the biologic phenotype of human cancers. CONTENT: Recent progress in microarray platforms and next-generation sequencing approaches has allowed the characterization of abnormal epigenetic patterns genome wide in a large number of cancer cases. The sizable amount of processed data also comes with challenges regarding data management and assessment for effective biomarker exploration to be further applied in prospective clinical trials. Epigenetics-based single or panel tests of genes are being explored for clinical management to fulfill unmet needs in oncology. The advance of these tests to the clinical routine will depend on rigorous, extensive, and independent validation in well-annotated cohort of patients and commercial development of clinical routine-friendly and adequate procedures. SUMMARY: In this review we discuss the analytic validation of tissue and cell-free DNA-based epigenomic approaches for early cancer detection, diagnosis, and treatment monitoring and the clinical utility of candidate epigenetic alterations applied to colorectal, glioblastoma, breast, prostate, bladder, and lung cancer management.

Phase I Study of Rapid Alternation of Sunitinib and Regorafenib for the Treatment of Tyrosine Kinase Inhibitor Refractory Gastrointestinal Stromal Tumors.

PURPOSE: Polyclonal emergence of KIT secondary mutations is a main mechanism of imatinib progression in gastrointestinal stromal tumor (GIST). Approved KIT inhibitors sunitinib and regorafenib have complementary activity against KIT resistance mutations. Preclinical evidence suggests that rapid alternation of sunitinib and regorafenib broadens the spectrum of imatinib-resistant subclones targeted. PATIENTS AND METHODS: Phase Ib study investigating continuous treatment with cycles of sunitinib (3 days) followed by regorafenib (4 days) in patients with tyrosine kinase inhibitor (TKI)-refractory GIST. A 3+3 dosing schema was utilized to determine the recommended phase II dose (RP2D). Plasma samples were analyzed for pharmacokinetics and circulating tumor DNA (ctDNA) studies using targeted error correction sequencing (TEC-seq) and droplet digital PCR (ddPCR). RESULTS: Of the 14 patients enrolled, 2 experienced dose-limiting toxicities at dose level 2 (asymptomatic grade 3 hypophosphatemia). Sunitinib 37.5 mg/day and regorafenib 120 mg/day was the RP2D. Treatment was well-tolerated and no unexpected toxicities resulted from the combination. Stable disease was the best response in 4 patients, and median progression-free survival was 1.9 months. Combined assessment of ctDNA with TEC-seq and ddPCR detected plasma mutations in 11 of 12 patients (92%). ctDNA studies showed that KIT secondary mutations remain the main mechanism of resistance in TKI-refractory GIST, revealing effective suppression of KIT-mutant subpopulations in patients benefiting from the combination. CONCLUSIONS: Sunitinib and regorafenib combination is feasible and tolerable. Rapid alternation of TKIs with complementary activity might be effective when combining drugs with favorable pharmacokinetics, potentially allowing active doses while minimizing adverse events. Serial monitoring with ctDNA may guide treatment in patients with GIST.

Genome-wide cell-free DNA fragmentation in patients with cancer.

Cell-free DNA in the blood provides a non-invasive diagnostic avenue for patients with cancer1. However, characteristics of the origins and molecular features of cell-free DNA are poorly understood. Here we developed an approach to evaluate fragmentation patterns of cell-free DNA across the genome, and found that profiles of healthy individuals reflected nucleosomal patterns of white blood cells, whereas patients with cancer had altered fragmentation profiles. We used this method to analyse the fragmentation profiles of 236 patients with breast, colorectal, lung, ovarian, pancreatic, gastric or bile duct cancer and 245 healthy individuals. A machine learning model that incorporated genome-wide fragmentation features had sensitivities of detection ranging from 57% to more than 99% among the seven cancer types at 98% specificity, with an overall area under the curve value of 0.94. Fragmentation profiles could be used to identify the tissue of origin of the cancers to a limited number of sites in 75% of cases. Combining our approach with mutation-based cell-free DNA analyses detected 91% of patients with cancer. The results of these analyses highlight important properties of cell-free DNA and provide a proof-of-principle approach for the screening, early detection and monitoring of human cancer.

Dynamics of Tumor and Immune Responses during Immune Checkpoint Blockade in Non-Small Cell Lung Cancer.

Despite the initial successes of immunotherapy, there is an urgent clinical need for molecular assays that identify patients more likely to respond. Here, we report that ultrasensitive measures of circulating tumor DNA (ctDNA) and T-cell expansion can be used to assess responses to immune checkpoint blockade in metastatic lung cancer patients (N = 24). Patients with clinical response to therapy had a complete reduction in ctDNA levels after initiation of therapy, whereas nonresponders had no significant changes or an increase in ctDNA levels. Patients with initial response followed by acquired resistance to therapy had an initial drop followed by recrudescence in ctDNA levels. Patients without a molecular response had shorter progression-free and overall survival compared with molecular responders [5.2 vs. 14.5 and 8.4 vs. 18.7 months; HR 5.36; 95% confidence interval (CI), 1.57-18.35; P = 0.007 and HR 6.91; 95% CI, 1.37-34.97; P = 0.02, respectively], which was detected on average 8.7 weeks earlier and was more predictive of clinical benefit than CT imaging. Expansion of T cells, measured through increases of T-cell receptor productive frequencies, mirrored ctDNA reduction in response to therapy. We validated this approach in an independent cohort of patients with early-stage non-small cell lung cancer (N = 14), where the therapeutic effect was measured by pathologic assessment of residual tumor after anti-PD1 therapy. Consistent with our initial findings, early ctDNA dynamics predicted pathologic response to immune checkpoint blockade. These analyses provide an approach for rapid determination of therapeutic outcomes for patients treated with immune checkpoint inhibitors and have important implications for the development of personalized immune targeted strategies.Significance: Rapid and sensitive detection of circulating tumor DNA dynamic changes and T-cell expansion can be used to guide immune targeted therapy for patients with lung cancer.See related commentary by Zou and Meyerson, p. 1038.