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BACKGROUND: Colorectal cancer is the third most diagnosed cancer in adults in the United States. Early detection could prevent more than 90% of colorectal cancer-related deaths, yet more than one third of the screening-eligible population is not up to date with screening despite multiple available tests. A blood-based test has the potential to improve screening adherence, detect colorectal cancer earlier, and reduce colorectal cancer-related mortality. METHODS: We assessed the performance characteristics of a cell-free DNA (cfDNA) blood-based test in a population eligible for colorectal cancer screening. The coprimary outcomes were sensitivity for colorectal cancer and specificity for advanced neoplasia (colorectal cancer or advanced precancerous lesions) relative to screening colonoscopy. The secondary outcome was sensitivity to detect advanced precancerous lesions. RESULTS: The clinical validation cohort included 10,258 persons, 7861 of whom met eligibility criteria and were evaluable. A total of 83.1% of the participants with colorectal cancer detected by colonoscopy had a positive cfDNA test and 16.9% had a negative test, which indicates a sensitivity of the cfDNA test for detection of colorectal cancer of 83.1% (95% confidence interval [CI], 72.2 to 90.3). Sensitivity for stage I, II, or III colorectal cancer was 87.5% (95% CI, 75.3 to 94.1), and sensitivity for advanced precancerous lesions was 13.2% (95% CI, 11.3 to 15.3). A total of 89.6% of the participants without any advanced colorectal neoplasia (colorectal cancer or advanced precancerous lesions) identified on colonoscopy had a negative cfDNA blood-based test, whereas 10.4% had a positive cfDNA blood-based test, which indicates a specificity for any advanced neoplasia of 89.6% (95% CI, 88.8 to 90.3). Specificity for negative colonoscopy (no colorectal cancer, advanced precancerous lesions, or nonadvanced precancerous lesions) was 89.9% (95% CI, 89.0 to 90.7). CONCLUSIONS: In an average-risk screening population, this cfDNA blood-based test had 83% sensitivity for colorectal cancer, 90% specificity for advanced neoplasia, and 13% sensitivity for advanced precancerous lesions. (Funded by Guardant Health; ECLIPSE ClinicalTrials.gov number, NCT04136002.).
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Ácidos Nucleicos Livres , Neoplasias Colorretais , Detecção Precoce de Câncer , Programas de Rastreamento , Lesões Pré-Cancerosas , Adulto , Humanos , Ácidos Nucleicos Livres/sangue , Colonoscopia , Neoplasias Colorretais/sangue , Neoplasias Colorretais/diagnóstico , Detecção Precoce de Câncer/métodos , Lesões Pré-Cancerosas/sangue , Lesões Pré-Cancerosas/diagnóstico , Programas de Rastreamento/métodos , Sensibilidade e EspecificidadeRESUMO
BACKGROUND: Circulating cell-free tumor DNA (ctDNA)-based mutation profiling, if sufficiently sensitive and comprehensive, can efficiently identify genomic targets in advanced lung adenocarcinoma. Therefore, the authors investigated the accuracy and clinical utility of a commercially available digital next-generation sequencing platform in a large series of patients with non-small cell lung cancer (NSCLC). METHODS: Plasma-based comprehensive genomic profiling results from 8388 consecutively tested patients with advanced NSCLC were analyzed. Driver and resistance mutations were examined with regard to their distribution, frequency, co-occurrence, and mutual exclusivity. RESULTS: Somatic alterations were detected in 86% of samples. The median variant allele fraction was 0.43% (range, 0.03%-97.62%). Activating alterations in actionable oncogenes were identified in 48% of patients, including EGFR (26.4%), MET (6.1%), and BRAF (2.8%) alterations and fusions (ALK, RET, and ROS1) in 2.3%. Treatment-induced resistance mutations were common in this cohort, including driver-dependent and driver-independent alterations. In the subset of patients who had progressive disease during EGFR therapy, 64% had known or putative resistance alterations detected in plasma. Subset analysis revealed that ctDNA increased the identification of driver mutations by 65% over standard-of-care, tissue-based testing at diagnosis. A pooled data analysis on this plasma-based assay demonstrated that targeted therapy response rates were equivalent to those reported from tissue analysis. CONCLUSIONS: Comprehensive ctDNA analysis detected the presence of therapeutically targetable driver and resistance mutations at the frequencies and distributions predicted for the study population. These findings add support for comprehensive ctDNA testing in patients who are incompletely tested at the time of diagnosis and as a primary option at the time of progression on targeted therapies.
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Adenocarcinoma de Pulmão/genética , Carcinoma Pulmonar de Células não Pequenas/genética , DNA Tumoral Circulante/genética , Neoplasias Pulmonares/genética , Mutação , Adenocarcinoma de Pulmão/sangue , Adenocarcinoma de Pulmão/tratamento farmacológico , Adenocarcinoma de Pulmão/patologia , Alelos , Biomarcadores Tumorais/sangue , Biomarcadores Tumorais/genética , Carcinoma Pulmonar de Células não Pequenas/sangue , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/patologia , DNA Tumoral Circulante/sangue , Estudos de Coortes , Progressão da Doença , Resistencia a Medicamentos Antineoplásicos/genética , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/genética , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Neoplasias Pulmonares/sangue , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Masculino , Oncogenes , Inibidores de Proteínas Quinases/uso terapêutico , Análise de Sequência de DNA , Transdução de Sinais/genética , Resultado do TratamentoRESUMO
BACKGROUND: Because cell-free DNA (cfDNA) analysis facilitates the noninvasive genomic profiling of metastatic castration-resistant prostate cancer (mCRPC), the authors evaluated the association between cfDNA alterations and outcomes and evolution with therapy. METHODS: Patients with mCRPC underwent cfDNA genomic profiling using Guardant360, which examines major cancer-associated genes. Clinical factors, therapy information, failure-free survival, and overall survival (OS) were obtained for select patients. The association between genomic alterations and outcomes was investigated. RESULTS: Of 514 men with mCRPC, 482 (94%) had ≥1 circulating tumor DNA (ctDNA) alteration. The most common recurrent somatic mutations were in TP53 (36%), androgen receptor (AR) (22%), adenomatous polyposis coli (APC) (10%), neurofibromin 1 (NF1) (9%), epidermal growth factor receptor (EGFR), catenin beta-1 (CTNNB1), and AT-rich interactive domain-containing protein 1A (ARID1A) (6% each); and BRCA1, BRCA2, and phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) (5% each) The most common genes with increased copy numbers were AR (30%), MYC (20%), and BRAF (18%). Clinical outcomes were available for 163 patients, 46 of whom (28.8%) were untreated for mCRPC. A higher number of ctDNA alterations, AR alterations, and amplifications of MYC and BRAF were associated with worse failure-free survival and/or OS. On multivariable analysis, MYC amplification remained significantly associated with OS. Prior therapy and serial profiling demonstrated the evolution of alterations in AR and other genes. CONCLUSIONS: ctDNA frequently was detected in this large cohort of "real-world" patients with mCRPC, and the alterations appeared to be similar to previously reported tumor tissue alterations. A higher number of alterations, and AR and MYC alterations, appear to compromise clinical outcomes, suggesting a role for immune checkpoint inhibitors and novel AR and BET inhibitors in selected patients.
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DNA Tumoral Circulante/genética , Mutação , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/análise , Biomarcadores Tumorais/sangue , Biomarcadores Tumorais/genética , DNA Tumoral Circulante/análise , DNA Tumoral Circulante/sangue , Variações do Número de Cópias de DNA , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , Prognóstico , Neoplasias de Próstata Resistentes à Castração/mortalidade , Neoplasias de Próstata Resistentes à Castração/terapia , Estudos Retrospectivos , Análise de SobrevidaRESUMO
BACKGROUND: Because imaging has a high sensitivity to diagnose hepatocellular carcinoma (HCC) and tissue biopsies carry risks such as bleeding, the latter are often not performed in HCC. Blood-derived circulating tumor DNA (ctDNA) analysis can identify somatic alterations, but its utility has not been characterized in HCC. MATERIALS AND METHODS: We evaluated 14 patients with advanced HCC (digital ctDNA sequencing [68 genes]). Mutant relative to wild-type allele fraction was calculated. RESULTS: All patients (100%) had somatic alterations (median = 3 alterations/patient [range, 1-8]); median mutant allele fraction, 0.29% (range, 0.1%-37.77%). Mutations were identified in several genes: TP53 (57% of patients), CTNNB1 (29%), PTEN (7%), CDKN2A (7%), ARID1A (7%), and MET (7%); amplifications, in CDK6 (14%), EGFR (14%), MYC (14%), BRAF (7%), RAF1 (7%), FGFR1 (7%), CCNE1 (7%), PIK3CA (7%), and ERBB2/HER2 (7%). Eleven patients (79%) had ≥1 theoretically actionable alteration. No two patients had identical genomic portfolios, suggesting the need for customized treatment. A patient with a CDKN2A-inactivating and a CTNNB1-activating mutation received matched treatment: palbociclib (CDK4/6 inhibitor) and celecoxib (COX-2/Wnt inhibitor); des-gamma-carboxy prothrombin level decreased by 84% at 2 months (1,410 to 242 ng/mL [normal: ≤7.4 ng/mL]; alpha fetoprotein [AFP] low at baseline). A patient with a PTEN-inactivating and a MET-activating mutation (an effect suggested by in silico molecular dynamic simulations) received sirolimus (mechanistic target of rapamycin inhibitor) and cabozantinib (MET inhibitor); AFP declined by 63% (8,320 to 3,045 ng/mL [normal: 0-15 ng/mL]). CONCLUSION: ctDNA derived from noninvasive blood tests can provide exploitable genomic profiles in patients with HCC. IMPLICATIONS FOR PRACTICE: This study reports that blood-derived circulating tumor DNA can provide therapeutically exploitable genomic profiles in hepatocellular cancer, a malignancy that is known to be difficult to biopsy.
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Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/genética , DNA Tumoral Circulante/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Adulto , Idoso , Carcinoma Hepatocelular/patologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos ProspectivosRESUMO
AIM: Insufficient adherence to colorectal cancer (CRC) screening impedes individual and population health benefits, with about one-third of individuals non-adherent to available screening options. The impact of poor adherence is inadequately considered in most health economics models, limiting the evaluation of real-world population-level screening outcomes. This study introduces the CAN-SCREEN (Colorectal cANcer SCReening Economics and adherENce) model, utilizing real-world adherence scenarios to assess the effectiveness of a blood-based test (BBT) compared to existing strategies. MATERIALS AND METHODS: The CAN-SCREEN model evaluates various CRC screening strategies per 1,000 screened individuals for ages 45-75. Adherence is modeled in two ways: (1) full adherence and (2) longitudinally declining adherence. BBT performance is based on recent pivotal trial data while existing strategies are informed using literature. The full adherence model is calibrated using previously published Cancer Intervention and Surveillance Modeling Network (CISNET) models. Outcomes, including life-years gained (LYG), CRC cases averted, CRC deaths averted, and colonoscopies, are compared to no screening. RESULTS: Longitudinal adherence modeling reveals differences in the relative ordering of health outcomes and resource utilization, as measured by the number of colonoscopies performed per 1,000, between screening modalities. BBT outperforms the fecal immunochemical test (FIT) and the multitarget stool DNA (mtsDNA) test with more CRC deaths averted (13) compared to FIT and mtsDNA (7, 11), more CRC cases averted (27 vs. 16, 22) and higher LYG (214 vs. 157, 199). BBT yields fewer CRC deaths averted compared to colonoscopy (13, 15) but requires fewer colonoscopies (1,053 vs. 1,928). LIMITATIONS: Due to limited data, the CAN-SCREEN model with longitudinal adherence leverages evidence-informed assumptions for the natural history and real-world longitudinal adherence to screening. CONCLUSIONS: The CAN-SCREEN model demonstrates that amongst non-invasive CRC screening strategies, those with higher adherence yield more favorable health outcomes as measured by CRC deaths averted, CRC cases averted, and LYG.
This study explored the impact of poor adherence to colorectal cancer (CRC) screening, where about one-third of people face barriers to screening. Common models don't consider real-world adherence, so we introduced the CAN-SCREEN model. It uses real-world data to determine how well a blood-based test (BBT) could work compared to existing tests. We studied people starting CRC screening at age 45. The model looked at two adherence scenarios: assuming everyone follows guidelines, and using real-world data about how people follow screening guidelines over time. The BBT's performance was based on a recent study, and we compared it to existing methods using data from the literature. Results per 1,000 simulated patients showed that the BBT outperforms two guideline-recommended stool-based tests, fecal immunochemical test (FIT) and the multitarget stool DNA (mtsDNA) test, with more CRC deaths averted (13) compared to FIT and mtsDNA (7, 11), more CRC cases averted (27 vs. 16, 22) and higher LYG (214 vs. 157, 199). BBT prevents less CRC deaths than colonoscopy (13 vs. 15), but it leads to fewer colonoscopies (1,053 compared to 1,928). Despite some limitations due to limited data, our model relies on informed assumptions for the natural history of CRC and real-world adherence. In conclusion, our CAN-SCREEN model shows that CRC screening strategies combining good test performance with high adherence give better health outcomes. Adding a blood test, which could be easier for people to use, could save lives and reduce the number of colonoscopies needed.
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Colonoscopia , Neoplasias Colorretais , Detecção Precoce de Câncer , Sangue Oculto , Cooperação do Paciente , Humanos , Neoplasias Colorretais/diagnóstico , Pessoa de Meia-Idade , Idoso , Detecção Precoce de Câncer/métodos , Masculino , Feminino , Análise Custo-BenefícioRESUMO
The enumeration of rare circulating epithelial cells (CEpCs) in the peripheral blood of metastatic cancer patients has shown promise for improved cancer prognosis. Moving beyond enumeration, molecular analysis of CEpCs may provide candidate surrogate endpoints to diagnose, treat, and monitor malignancy directly from the blood samples. Thorough molecular analysis of CEpCs requires the development of new sample preparation methods that yield easily accessible and purified CEpCs for downstream biochemical assays. Here, we describe a new immunomagnetic cell separator, the MagSweeper, which gently enriches target cells and eliminates cells that are not bound to magnetic particles. The isolated cells are easily accessible and can be extracted individually based on their physical characteristics to deplete any cells nonspecifically bound to beads. We have shown that our device can process 9 mL of blood per hour and captures >50% of CEpCs as measured in spiking experiments. We have shown that the separation process does not perturb the gene expression of rare cells. To determine the efficiency of our platform in isolating CEpCs from patients, we have isolated CEpCs from all 47 tubes of 9-mL blood samples collected from 17 women with metastatic breast cancer. In contrast, we could not find any circulating epithelial cells in samples from 5 healthy donors. The isolated CEpCs are all stored individually for further molecular analysis.
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Células Sanguíneas/citologia , Separação Celular/instrumentação , Células Epiteliais/citologia , Magnetismo/instrumentação , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Simulação por Computador , Feminino , Regulação da Expressão Gênica , Antígeno HLA-A2/imunologia , Humanos , Modelos ImunológicosRESUMO
PURPOSE: Detection of persistent circulating tumor DNA (ctDNA) after curative-intent surgery can identify patients with minimal residual disease (MRD) who will ultimately recur. Most ctDNA MRD assays require tumor sequencing to identify tumor-derived mutations to facilitate ctDNA detection, requiring tumor and blood. We evaluated a plasma-only ctDNA assay integrating genomic and epigenomic cancer signatures to enable tumor-uninformed MRD detection. EXPERIMENTAL DESIGN: A total of 252 prospective serial plasma specimens from 103 patients with colorectal cancer undergoing curative-intent surgery were analyzed and correlated with recurrence. RESULTS: Of 103 patients, 84 [stage I (9.5%), II (23.8%), III (47.6%), IV (19%)] had evaluable plasma drawn after completion of definitive therapy, defined as surgery only (n = 39) or completion of adjuvant therapy (n = 45). In "landmark" plasma drawn 1-month (median, 31.5 days) after definitive therapy and >1 year follow-up, 15 patients had detectable ctDNA, and all 15 recurred [positive predictive value (PPV), 100%; HR, 11.28 (P < 0.0001)]. Of 49 patients without detectable ctDNA at the landmark timepoint, 12 (24.5%) recurred. Landmark recurrence sensitivity and specificity were 55.6% and 100%. Incorporating serial longitudinal and surveillance (drawn within 4 months of recurrence) samples, sensitivity improved to 69% and 91%. Integrating epigenomic signatures increased sensitivity by 25%-36% versus genomic alterations alone. Notably, standard serum carcinoembryonic antigen levels did not predict recurrence [HR, 1.84 (P = 0.18); PPV = 53.9%]. CONCLUSIONS: Plasma-only MRD detection demonstrated favorable sensitivity and specificity for recurrence, comparable with tumor-informed approaches. Integrating analysis of epigenomic and genomic alterations enhanced sensitivity. These findings support the potential clinical utility of plasma-only ctDNA MRD detection.See related commentary by Bent and Kopetz, p. 5449.
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DNA Tumoral Circulante/sangue , Neoplasias Colorretais/sangue , Neoplasias Colorretais/cirurgia , Neoplasia Residual/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias Colorretais/patologia , Feminino , Testes Hematológicos , Humanos , Masculino , Pessoa de Meia-Idade , Estudos ProspectivosRESUMO
Current methods used for analyzing biomarkers involve expensive and time consuming techniques like the Sandwich ELISA which require lengthy incubation times, high reagent costs, and bulky optical equipment. We have developed a technique involving the use of a micro-channel with integrated electrodes, functionalized with receptors specific to target biomarkers. We have applied our biochip to the rapid electrical detection and quantification of target protein biomarkers using protein functionalized micro-channels. We successfully demonstrate detection of anti-hCG antibody, at a concentration of 1 ng ml(-1) and a dynamic range of three orders of magnitude, in less than one hour. We envision the use of this technique in a handheld device for multiplex high throughput analysis using an array of micro-channels for probing various protein biomarkers in clinically relevant samples such as human serum for cancer detection.
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Biomarcadores/análise , Técnicas Eletroquímicas , Técnicas Analíticas Microfluídicas , Proteínas/análise , Gonadotropina Coriônica , Diagnóstico Precoce , Impedância Elétrica , Técnicas Eletroquímicas/instrumentação , Técnicas Eletroquímicas/métodos , Eletrodos , Eletrólitos , Desenho de Equipamento , Técnicas Analíticas Microfluídicas/instrumentação , Técnicas Analíticas Microfluídicas/métodos , MicroesferasRESUMO
Traditionally, expensive and time consuming techniques such as mass spectrometry and Western Blotting have been used for characterization of protein-protein interactions. In this paper, we describe the design, fabrication, and testing of a rapid and inexpensive sensor, involving the use of microelectrodes in a microchannel, which can be used for real-time electrical detection of specific interactions between proteins. We have successfully demonstrated detection of target glycoprotein-glycoprotein interactions, antigen-antibody interactions, and glycoprotein-antigen interactions. We have also demonstrated the ability of this technique to distinguish between strong and weak interactions. Using this approach, it may be possible to multiplex an array of these sensors onto a chip and probe a complex mixture for various types of interactions involving protein molecules.
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PURPOSE: Genomic alterations in blood-derived circulating tumor DNA (ctDNA) from patients with colorectal cancers were correlated with clinical outcomes. PATIENTS AND METHODS: Next-generation sequencing of ctDNA (54- to 73-gene panel) was performed in 94 patients with colorectal cancer. RESULTS: Most patients (96%) had metastatic or recurrent disease at the time of blood draw. The median number of nonsynonymous alterations per patient was three (range, zero to 30). The most frequently aberrant genes were TP53 (52.1% of patients), KRAS (34%), and APC (28.7%). Concordance between tissue and blood next-generation sequencing ranged from 63.2% (APC) to 85.5% (BRAF). Altogether, 74 patients (79%) had one or more nonsynonymous alterations, 69 (73%) had one or more potentially actionable alterations, and 61 (65%) had an alteration actionable by a drug approved by the US Food and Drug Administration (on or off label). Lung metastases correlated with improved survival from diagnosis in univariable analysis. ctDNA of 5% or more from blood tests as well as EGFR and ERBB2 (HER2) nonsynonymous alterations correlated with worse survival (but only ERBB2 remained significant in multivariable analysis). No two patients had identical molecular portfolios. Overall, 65% versus 31% of patients treated with matched (n = 17) versus unmatched therapy (n = 18) after ctDNA testing achieved stable disease for 6 months or more, partial response, or complete response (P = .045); progression-free survival, 6.1 versus 2.3 months (P = .08); and survival not reached versus 9.4 months (P = .146; all by multivariable analysis). CONCLUSION: Patients with colorectal cancer have heterogeneous ctDNA profiles, and most harbor potentially actionable ctDNA alterations. Matched therapy yielded higher rates of stable disease for 6 months or more, partial response, or complete response. ctDNA assessment may have clinical utility and merits further investigation.
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BACKGROUND: Major guidelines do not recommend routine molecular profiling of lung squamous-cell carcinoma (LUSC) because the prevalence of actionable alterations is thought to be low. Increased utilization of next-generation sequencing (NGS), particularly with cell-free circulating tumor DNA, facilitates reevaluation of this premise. PATIENTS AND METHODS: We retrospectively evaluated the prevalence of actionable alterations in 2 distinct LUSC cohorts totaling 492 patients. A total of 410 consecutive patients with stage 3B or 4 LUSC were tested with a targeted cell-free circulating DNA NGS assay, and 82 patients with LUSC of any stage were tested with a tissue NGS cancer panel. RESULTS: In the overall cohort, 467 patients (94.9%) had a diagnosis of LUSC, and 25 patients (5.1%) had mixed histology with a squamous component. A total of 10.5% of the LUSC subgroup had somatic alterations with therapeutic relevance, including in EGFR (2.8%), ALK/ROS1 (1.3%), BRAF (1.5%), and MET amplification or exon 14 skipping (5.1%). Sixteen percent of patients with mixed histology had an actionable alteration. In the LUSC subgroup, 3 evaluable patients were treated with targeted therapy for an actionable alteration; all of them experienced partial response. CONCLUSION: In this large, real-world LUSC cohort, we observed a clinically significant prevalence of actionable alterations. Accurate local histopathologic assessment in advanced-stage LUSC can be challenging. Further evaluation of the genomic landscape in this setting is warranted to potentially identify underappreciated treatment options.
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Quinase do Linfoma Anaplásico/genética , Carcinoma de Células Escamosas/genética , DNA Tumoral Circulante/genética , Neoplasias Pulmonares/genética , Mutação/genética , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas c-met/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Coortes , Feminino , Genes erbB-1/genética , Marcadores Genéticos/genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Pessoa de Meia-Idade , Estudos RetrospectivosRESUMO
PURPOSE: Molecular profiling has been used to select patients for targeted therapy and determine prognosis. Noninvasive strategies are critical to hepatocellular carcinoma (HCC) given the challenge of obtaining liver tissue biopsies. EXPERIMENTAL DESIGN: We analyzed blood samples from 206 patients with HCC using comprehensive genomic testing (Guardant Health) of circulating tumor DNA (ctDNA). RESULTS: A total of 153/206 (74.3%) were men; median age, 62 years (range, 18-91 years). A total of 181/206 patients had ≥1 alteration. The total number of alterations was 680 (nonunique); median number of alterations/patient was three (range, 1-13); median mutant allele frequency (% cfDNA), 0.49% (range, 0.06%-55.03%). TP53 was the common altered gene [>120 alterations (non-unique)] followed by EGFR, MET, ARID1A, MYC, NF1, BRAF, and ERBB2 [20-38 alterations (nonunique)/gene]. Of the patients with alterations, 56.9% (103/181) had ≥1 actionable alterations, most commonly in MYC, EGFR, ERBB2, BRAF, CCNE1, MET, PIK3CA, ARID1A, CDK6, and KRAS. In these genes, amplifications occurred more frequently than mutations. Hepatitis B (HBV)-positive patients were more likely to have ERBB2 alterations, 35.7% (5/14) versus 8.8% HBV-negative (P = 0.04). CONCLUSIONS: This study represents the first large-scale analysis of blood-derived ctDNA in HCC in United States. The genomic distinction based on HCC risk factors and the high percentage of potentially actionable genomic alterations suggests potential clinical utility for this technology.
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Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/genética , DNA Tumoral Circulante/genética , Testes Genéticos/métodos , Neoplasias Hepáticas/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/sangue , Carcinoma Hepatocelular/sangue , Carcinoma Hepatocelular/terapia , DNA Tumoral Circulante/sangue , Tomada de Decisão Clínica/métodos , Estudos de Coortes , Análise Mutacional de DNA , Feminino , Frequência do Gene , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Neoplasias Hepáticas/sangue , Neoplasias Hepáticas/terapia , Masculino , Pessoa de Meia-Idade , Mutação , Seleção de Pacientes , Prognóstico , Estados Unidos , Adulto JovemRESUMO
PURPOSE: To analytically and clinically validate microsatellite instability (MSI) detection using cell-free DNA (cfDNA) sequencing. EXPERIMENTAL DESIGN: Pan-cancer MSI detection using Guardant360 was analytically validated according to established guidelines and clinically validated using 1,145 cfDNA samples for which tissue MSI status based on standard-of-care tissue testing was available. The landscape of cfDNA-based MSI across solid tumor types was investigated in a cohort of 28,459 clinical plasma samples. Clinical outcomes for 16 patients with cfDNA MSI-H gastric cancer treated with immunotherapy were evaluated. RESULTS: cfDNA MSI evaluation was shown to have high specificity, precision, and sensitivity, with a limit of detection of 0.1% tumor content. In evaluable patients, cfDNA testing accurately detected 87% (71/82) of tissue MSI-H and 99.5% of tissue microsatellite stable (863/867) for an overall accuracy of 98.4% (934/949) and a positive predictive value of 95% (71/75). Concordance of cfDNA MSI with tissue PCR and next-generation sequencing was significantly higher than IHC. Prevalence of cfDNA MSI for major cancer types was consistent with those reported for tissue. Finally, robust clinical activity of immunotherapy treatment was seen in patients with advanced gastric cancer positive for MSI by cfDNA, with 63% (10/16) of patients achieving complete or partial remission with sustained clinical benefit. CONCLUSIONS: cfDNA-based MSI detection using Guardant360 is highly concordant with tissue-based testing, enabling highly accurate detection of MSI status concurrent with comprehensive genomic profiling and expanding access to immunotherapy for patients with advanced cancer for whom current testing practices are inadequate.See related commentary by Wang and Ajani, p. 6887.
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Biomarcadores Tumorais/genética , DNA Tumoral Circulante/sangue , DNA Tumoral Circulante/genética , Instabilidade de Microssatélites , Neoplasias/genética , Biomarcadores Tumorais/sangue , Estudos de Casos e Controles , Seguimentos , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Neoplasias/sangue , Neoplasias/patologia , PrognósticoRESUMO
The VIKTORY (targeted agent eValuation In gastric cancer basket KORea) trial was designed to classify patients with metastatic gastric cancer based on clinical sequencing and focused on eight different biomarker groups (RAS aberration, TP53 mutation, PIK3CA mutation/amplification, MET amplification, MET overexpression, all negative, TSC2 deficient, or RICTOR amplification) to assign patients to one of the 10 associated clinical trials in second-line (2L) treatment. Capivasertib (AKT inhibitor), savolitinib (MET inhibitor), selumetinib (MEK inhibitor), adavosertib (WEE1 inhibitor), and vistusertib (TORC inhibitor) were tested with or without chemotherapy. Seven hundred seventy-two patients with gastric cancer were enrolled, and sequencing was successfully achieved in 715 patients (92.6%). When molecular screening was linked to seamless immediate access to parallel matched trials, 14.7% of patients received biomarker-assigned drug treatment. The biomarker-assigned treatment cohort had encouraging response rates and survival when compared with conventional 2L chemotherapy. Circulating tumor (ctDNA) analysis demonstrated good correlation between high MET copy number by ctDNA and response to savolitinib. SIGNIFICANCE: Prospective clinical sequencing revealed that baseline heterogeneity between tumor samples from different patients affected response to biomarker-selected therapies. VIKTORY is the first and largest platform study in gastric cancer and supports both the feasibility of tumor profiling and its clinical utility.This article is highlighted in the In This Issue feature, p. 1325.
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Biomarcadores Tumorais , Genômica , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Biópsia , DNA Tumoral Circulante , Tomada de Decisão Clínica , Biologia Computacional/métodos , Gerenciamento Clínico , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Genômica/métodos , Humanos , Terapia de Alvo Molecular , Metástase Neoplásica , Estadiamento de Neoplasias , Prognóstico , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/mortalidade , Resultado do TratamentoRESUMO
The present study reports on the retention of conformational flexibility of a model allosteric protein upon immobilization on self-assembled monolayers (SAMs) on gold. Organothiolated SAMs of different compositions were utilized for adsorptive and covalent attachment of bovine liver glutamate dehydrogenase (GDH), a well-characterized allosteric enzyme. Sensitive fluorimetric assays were developed to determine immobilization capacity, specific activity, and allosteric properties of the immobilized preparations as well as the potential for repeated use and continuous catalytic transformations. The allosteric response of the free and immobilized forms towards ADP, L-leucine and high concentrations of NAD(+), some of the well-known activators for this enzyme, were determined and compared. The enzyme immobilized by adsorption or chemical binding responded similarly to the activators with a greater degree of activation, as compared to the free form. Also loss of activity involving the two immobilization procedures were similar, suggesting that residues essential for catalytic activity or allosteric properties of GDH remained unchanged in the course of chemical modification. A recently established method was used to predict GDH orientation upon immobilization, which was found to explain some of the experimental results presented. The general significance of these observations in connection with retention of native properties of protein structures upon immobilization on SAMs is discussed.
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Materiais Revestidos Biocompatíveis/química , Glutamato Desidrogenase/química , Glutamato Desidrogenase/ultraestrutura , Ouro/química , Modelos Químicos , Modelos Moleculares , Adsorção , Animais , Sítios de Ligação , Bovinos , Simulação por Computador , Ativação Enzimática , Estabilidade Enzimática , Enzimas Imobilizadas/química , Ligação Proteica , Conformação ProteicaRESUMO
PURPOSE: Hotspot blood cell-free DNA (cfDNA) biomarker assays have limited utility in profiling tumor heterogeneity and burden and in capturing regional metastasis with low disease burden in patients with melanoma. We investigated the utility of a sensitive 54-cancer gene digital next-generation sequencing approach targeting blood cfDNA single nucleotide variants (SNVs) and copy number amplification for monitoring disease in patients with melanoma with regional or distant organ metastasis (DOM). PATIENTS AND METHODS: A total of 142 blood samples were evaluated by digital next-generation sequencing across two patient cohorts. Cohort 1 contained 44 patients with stage II, III, or IV disease with matched tumor DNA at the time of surgery or DOM. Cohort 2 consisted of 12 overlapping patients who were longitudinally monitored after complete lymph node dissection to DOM. RESULTS: In cohort 1, cfDNA SNVs were detected in 75% of patients. Tumor-cfDNA somatic SNV concordance was 85% at a variant allele fraction of ≥ 0.5%. An SNV load (number of unique SNVs detected) of greater than two SNVs and an SNV burden (total cumulative SNV VAF) of > 0.5% were significantly associated with worse overall survival (P < .05) in stage IV patients. In cohort 2, 98 longitudinal blood samples along with matched regional and distant metastases from 12 stage III patients were analyzed before complete lymph node dissection and throughout disease progression. cfDNA SNV levels correlated with tumor burden (P = .019), enabled earlier detection of recurrence compared with radiologic imaging (P < .01), captured tumor heterogeneity, and identified increasing SNVs levels before recurrence. CONCLUSION: This study demonstrates significant utility for cfDNA profiling in patients with melanoma with regional and/or distant metastasis for earlier detection of recurrence and progression and in capturing tumor evolution and heterogeneity, thus impacting how patients with melanoma are monitored.
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"Liquid biopsy" approaches analyzing cell-free DNA (cfDNA) from the blood of patients with cancer are increasingly utilized in clinical practice. However, it is not yet known whether cfDNA sequencing from large cohorts of patients with cancer can detect genomic alterations at frequencies similar to those observed by direct tumor sequencing, and whether this approach can generate novel insights. Here, we report next-generation sequencing data from cfDNA of 1,397 patients with colorectal cancer. Overall, frequencies of genomic alterations detected in cfDNA were comparable to those observed in three independent tissue-based colorectal cancer sequencing compendia. Our analysis also identified a novel cluster of extracellular domain (ECD) mutations in EGFR, mediating resistance by blocking binding of anti-EGFR antibodies. Patients with EGFR ECD mutations displayed striking tumor heterogeneity, with 91% harboring multiple distinct resistance alterations (range, 1-13; median, 4). These results suggest that cfDNA profiling can effectively define the genomic landscape of cancer and yield important biological insights.Significance: This study provides one of the first examples of how large-scale genomic profiling of cfDNA from patients with colorectal cancer can detect genomic alterations at frequencies comparable to those observed by direct tumor sequencing. Sequencing of cfDNA also generated insights into tumor heterogeneity and therapeutic resistance and identified novel EGFR ectodomain mutations. Cancer Discov; 8(2); 164-73. ©2017 AACR.This article is highlighted in the In This Issue feature, p. 127.
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Biomarcadores Tumorais , Ácidos Nucleicos Livres , Neoplasias Colorretais/genética , DNA de Neoplasias , Estudo de Associação Genômica Ampla , Genômica , Evolução Clonal/genética , Estudos de Coortes , Neoplasias Colorretais/sangue , Receptores ErbB/genética , Genômica/métodos , Humanos , MutaçãoRESUMO
Purpose: To analytically and clinically validate a circulating cell-free tumor DNA sequencing test for comprehensive tumor genotyping and demonstrate its clinical feasibility.Experimental Design: Analytic validation was conducted according to established principles and guidelines. Blood-to-blood clinical validation comprised blinded external comparison with clinical droplet digital PCR across 222 consecutive biomarker-positive clinical samples. Blood-to-tissue clinical validation comprised comparison of digital sequencing calls to those documented in the medical record of 543 consecutive lung cancer patients. Clinical experience was reported from 10,593 consecutive clinical samples.Results: Digital sequencing technology enabled variant detection down to 0.02% to 0.04% allelic fraction/2.12 copies with ≤0.3%/2.24-2.76 copies 95% limits of detection while maintaining high specificity [prevalence-adjusted positive predictive values (PPV) >98%]. Clinical validation using orthogonal plasma- and tissue-based clinical genotyping across >750 patients demonstrated high accuracy and specificity [positive percent agreement (PPAs) and negative percent agreement (NPAs) >99% and PPVs 92%-100%]. Clinical use in 10,593 advanced adult solid tumor patients demonstrated high feasibility (>99.6% technical success rate) and clinical sensitivity (85.9%), with high potential actionability (16.7% with FDA-approved on-label treatment options; 72.0% with treatment or trial recommendations), particularly in non-small cell lung cancer, where 34.5% of patient samples comprised a directly targetable standard-of-care biomarker.Conclusions: High concordance with orthogonal clinical plasma- and tissue-based genotyping methods supports the clinical accuracy of digital sequencing across all four types of targetable genomic alterations. Digital sequencing's clinical applicability is further supported by high rates of technical success and biomarker target discovery. Clin Cancer Res; 24(15); 3539-49. ©2018 AACR.
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Ácidos Nucleicos Livres/genética , DNA Tumoral Circulante/genética , Genômica , Neoplasias/genética , Biomarcadores Tumorais , Ácidos Nucleicos Livres/sangue , DNA Tumoral Circulante/sangue , Feminino , Genótipo , Técnicas de Genotipagem , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Mutação , Neoplasias/sangue , Neoplasias/patologiaRESUMO
Purpose: Cell-free DNA (cfDNA) sequencing provides a noninvasive method for obtaining actionable genomic information to guide personalized cancer treatment, but the presence of multiple alterations in circulation related to treatment and tumor heterogeneity complicate the interpretation of the observed variants.Experimental Design: We describe the somatic mutation landscape of 70 cancer genes from cfDNA deep-sequencing analysis of 21,807 patients with treated, late-stage cancers across >50 cancer types. To facilitate interpretation of the genomic complexity of circulating tumor DNA in advanced, treated cancer patients, we developed methods to identify cfDNA copy-number driver alterations and cfDNA clonality.Results: Patterns and prevalence of cfDNA alterations in major driver genes for non-small cell lung, breast, and colorectal cancer largely recapitulated those from tumor tissue sequencing compendia (The Cancer Genome Atlas and COSMIC; r = 0.90-0.99), with the principal differences in alteration prevalence being due to patient treatment. This highly sensitive cfDNA sequencing assay revealed numerous subclonal tumor-derived alterations, expected as a result of clonal evolution, but leading to an apparent departure from mutual exclusivity in treatment-naïve tumors. Upon applying novel cfDNA clonality and copy-number driver identification methods, robust mutual exclusivity was observed among predicted truncal driver cfDNA alterations (FDR = 5 × 10-7 for EGFR and ERBB2), in effect distinguishing tumor-initiating alterations from secondary alterations. Treatment-associated resistance, including both novel alterations and parallel evolution, was common in the cfDNA cohort and was enriched in patients with targetable driver alterations (>18.6% patients).Conclusions: Together, these retrospective analyses of a large cfDNA sequencing data set reveal subclonal structures and emerging resistance in advanced solid tumors. Clin Cancer Res; 24(15); 3528-38. ©2018 AACR.