RESUMO
Advancements in genomic technologies have shown remarkable promise for improving health trajectories. The Human Genome Project has catalyzed the integration of genomic tools into clinical practice, such as disease risk assessment, prenatal testing and reproductive genomics, cancer diagnostics and prognostication, and therapeutic decision making. Despite the promise of genomic technologies, their full potential remains untapped without including individuals of diverse ancestries and integrating social determinants of health (SDOHs). The NHGRI launched the 2020 Strategic Vision with ten bold predictions by 2030, including "individuals from ancestrally diverse backgrounds will benefit equitably from advances in human genomics." Meeting this goal requires a holistic approach that brings together genomic advancements with careful consideration to healthcare access as well as SDOHs to ensure that translation of genetics research is inclusive, affordable, and accessible and ultimately narrows rather than widens health disparities. With this prediction in mind, this review delves into the two paramount applications of genetic testing-reproductive genomics and precision oncology. When discussing these applications of genomic advancements, we evaluate current accessibility limitations, highlight challenges in achieving representativeness, and propose paths forward to realize the ultimate goal of their equitable applications.
Assuntos
Genômica , Medicina de Precisão , Humanos , Genômica/métodos , Medicina de Precisão/métodos , Genoma Humano , Testes Genéticos , Neoplasias/genética , Acessibilidade aos Serviços de SaúdeRESUMO
Liquid biopsies (LBs), particularly using circulating tumor DNA (ctDNA), are expected to revolutionize precision oncology and blood-based cancer screening. Recent technological improvements, in combination with the ever-growing understanding of cell-free DNA (cfDNA) biology, are enabling the detection of tumor-specific changes with extremely high resolution and new analysis concepts beyond genetic alterations, including methylomics, fragmentomics, and nucleosomics. The interrogation of a large number of markers and the high complexity of data render traditional correlation methods insufficient. In this regard, machine learning (ML) algorithms are increasingly being used to decipher disease- and tissue-specific signals from cfDNA. Here, we review recent insights into biological ctDNA features and how these are incorporated into sophisticated ML applications.
Assuntos
Ácidos Nucleicos Livres , DNA Tumoral Circulante , Neoplasias Hematológicas , Neoplasias , Humanos , Ácidos Nucleicos Livres/genética , Neoplasias/genética , Medicina de Precisão , DNA Tumoral Circulante/genética , DNA Tumoral Circulante/análise , Aprendizado de MáquinaRESUMO
The analysis of cell-free DNA (cfDNA) from plasma offers great promise for the earlier detection of cancer. At present, changes in DNA sequence, methylation, or copy number are the most sensitive ways to detect the presence of cancer. To further increase the sensitivity of such assays with limited amounts of sample, it would be useful to be able to evaluate the same template molecules for all these changes. Here, we report an approach, called MethylSaferSeqS, that achieves this goal, and can be applied to any standard library preparation method suitable for massively parallel sequencing. The innovative step was to copy both strands of each DNA-barcoded molecule with a primer that allows the subsequent separation of the original strands (retaining their 5-methylcytosine residues) from the copied strands (in which the 5-methylcytosine residues are replaced with unmodified cytosine residues). The epigenetic and genetic alterations present in the DNA molecules can then be obtained from the original and copied strands, respectively. We applied this approach to plasma from 265 individuals, including 198 with cancers of the pancreas, ovary, lung, and colon, and found the expected patterns of mutations, copy number alterations, and methylation. Furthermore, we could determine which original template DNA molecules were methylated and/or mutated. MethylSaferSeqS should be useful for addressing a variety of questions relating genetics and epigenetics.
Assuntos
Variações do Número de Cópias de DNA , Neoplasias , Feminino , Humanos , Metilação , 5-Metilcitosina , DNA/genética , Mutação , Neoplasias/genética , Metilação de DNARESUMO
In the TRIDENT-2 study, all pregnant women in the Netherlands are offered genome-wide non-invasive prenatal testing (GW-NIPT) with a choice of receiving either full screening or screening solely for common trisomies. Previous data showed that GW-NIPT can reliably detect common trisomies in the general obstetric population and that this test can also detect other chromosomal abnormalities (additional findings). However, evidence regarding the clinical impact of screening for additional findings is lacking. Therefore, we present follow-up results of the TRIDENT-2 study to determine this clinical impact based on the laboratory and perinatal outcomes of cases with additional findings. Between April 2017 and April 2019, additional findings were detected in 402/110,739 pregnancies (0.36%). For 358 cases, the origin was proven to be either fetal (n = 79; 22.1%), (assumed) confined placental mosaicism (CPM) (n = 189; 52.8%), or maternal (n = 90; 25.1%). For the remaining 44 (10.9%), the origin of the aberration could not be determined. Most fetal chromosomal aberrations were pathogenic and associated with severe clinical phenotypes (61/79; 77.2%). For CPM cases, occurrence of pre-eclampsia (8.5% [16/189] vs 0.5% [754/159,924]; RR 18.5), and birth weight <2.3rd percentile (13.6% [24/177] vs 2.5% [3,892/155,491]; RR 5.5) were significantly increased compared to the general obstetric population. Of the 90 maternal findings, 12 (13.3%) were malignancies and 32 (35.6%) (mosaic) pathogenic copy number variants, mostly associated with mild or no clinical phenotypes. Data from this large cohort study provide crucial information for deciding if and how to implement GW-NIPT in screening programs. Additionally, these data can inform the challenging interpretation, counseling, and follow-up of additional findings.
Assuntos
Diagnóstico Pré-Natal , Trissomia , Estudos de Coortes , Feminino , Seguimentos , Humanos , Mosaicismo , Placenta , Gravidez , Diagnóstico Pré-Natal/métodosRESUMO
BACKGROUND: Delayed diagnosis and improper therapy for intraocular infections usually result in poor prognosis. Due to limitations of conventional culture and polymerase chain reaction methods, most causative pathogens cannot be identified from vitreous humor (VH) or aqueous humor (AH) samples with limited volume. METHODS: Patients with suspected intraocular infections were enrolled from January 2019 to August 2021. Metagenomic next-generation sequencing (mNGS) was used to detected causative pathogens. RESULTS: This multicenter prospective study enrolled 488 patients, from whom VH (152) and AH (336) samples were respectively collected and analyzed using mNGS of cell-free DNA (cfDNA). Taking final comprehensive clinical diagnosis as the gold standard, there were 39 patients with indefinite final diagnoses, whereas 288 and 161 patients were diagnosed as definite infectious and noninfectious diseases, respectively. Based on clinical adjudication, the sensitivity (92.2%) and total coincidence rate (81.3%) of mNGS using VH samples were slightly higher than those of mNGS using AH samples (85.4% and 75.4%, respectively). CONCLUSIONS: Using mNGS of cfDNA, an era with clinical experience for more rapid, independent, and impartial diagnosis of bacterial and other intraocular infections can be expected.
Assuntos
Ácidos Nucleicos Livres , Infecções Oculares , Humanos , Humor Aquoso , Ácidos Nucleicos Livres/genética , Estudos Prospectivos , Sequenciamento de Nucleotídeos em Larga Escala , Metagenômica , Sensibilidade e EspecificidadeRESUMO
Novel screening techniques for early detection of lung cancer are urgently needed. Profiling circulating tumor cell-free DNA (ctDNA) has emerged as a promising tool for biopsy-free tumor genotyping. However, both the scarcity and short half-life of ctDNA substantially limit the sensitivity and clinical utility of ctDNA detection methodologies. Our discovery that red blood cells (RBCs) sequester mitochondrial DNA opens a new avenue for detecting circulating nucleic acids, as RBCs represent an unrecognized reservoir of circulating nucleic acid. Here, we show that RBCs acquire tumor DNA following coculture with lung cancer cell lines harboring Kirsten rat sarcoma viral oncogene homolog (KRAS) and epidermal growth factor receptor (EGFR) mutations. RBC-bound tumor DNA is detectable in patients with early-stage non-small cell lung cancer (NSCLC) but not in healthy controls by qPCR. Our results collectively uncover a previously unrecognized yet easily accessible reservoir of tumor DNA, offering a promising foundation for future RBC-based tumor diagnostics.NEW & NOTEWORTHY We present a novel method for lung cancer detection by revealing RBCs as a reservoir for tumor DNA, overcoming the limitations of current circulating tumor ctDNA methodologies. By demonstrating that RBCs can capture tumor DNA, including critical mutations found in lung cancer, we provide a promising, biopsy-free avenue for early cancer diagnostics. This discovery opens up exciting possibilities for developing RBC-based diagnostic tools, significantly enhancing the sensitivity and clinical utility of noninvasive cancer detection.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , DNA Tumoral Circulante , Eritrócitos , Neoplasias Pulmonares , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/sangue , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/diagnóstico , Eritrócitos/metabolismo , DNA Tumoral Circulante/genética , DNA Tumoral Circulante/sangue , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/sangue , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Mutação , Linhagem Celular Tumoral , Receptores ErbB/genética , Receptores ErbB/metabolismo , DNA Mitocondrial/genética , DNA Mitocondrial/sangue , Proteínas Proto-Oncogênicas p21(ras)/genética , Masculino , Feminino , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/sangue , DNA de Neoplasias/sangue , DNA de Neoplasias/genéticaRESUMO
BACKGROUND: Gene expression profiles in breast tissue biopsies contain information related to chemotherapy efficacy. The promoter profiles in cell-free DNA (cfDNA) carrying gene expression information of the original tissues may be used to predict the response to neoadjuvant chemotherapy in breast cancer as a non-invasive biomarker. In this study, the feasibility of the promoter profiles in plasma cfDNA was evaluated as a novel clinical model for noninvasively predicting the efficacy of neoadjuvant chemotherapy in breast cancer. METHOD: First of all, global chromatin (5 Mb windows), sub-compartments and promoter profiles in plasma cfDNA samples from 94 patients with breast cancer before neoadjuvant chemotherapy (pCR = 31 vs. non-pCR = 63) were analyzed, and then classifiers were developed for predicting the efficacy of neoadjuvant chemotherapy in breast cancer. Further, the promoter profile changes in sequential cfDNA samples from 30 patients (pCR = 8 vs. non-pCR = 22) during neoadjuvant chemotherapy were analyzed to explore the potential benefits of cfDNA promoter profile changes as a novel potential biomarker for predicting the treatment efficacy. RESULTS: The results showed significantly distinct promoter profile in plasma cfDNA of pCR patients compared with non-pCR patients before neoadjuvant chemotherapy. The classifier based on promoter profiles in a Random Forest model produced the largest area under the curve of 0.980 (95% CI: 0.978-0.983). After neoadjuvant chemotherapy, 332 genes with significantly differential promoter profile changes in sequential cfDNA samples of pCR patients was observed, compared with non-pCR patients, and their functions were closely related to treatment response. CONCLUSION: These results suggest that promoter profiles in plasma cfDNA may be a powerful, non-invasive tool for predicting the efficacy of neoadjuvant chemotherapy breast cancer patients before treatment, and the on-treatment cfDNA promoter profiles have potential benefits for predicting the treatment efficacy.
Assuntos
Biomarcadores Tumorais , Neoplasias da Mama , Ácidos Nucleicos Livres , Terapia Neoadjuvante , Regiões Promotoras Genéticas , Humanos , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Neoplasias da Mama/sangue , Neoplasias da Mama/patologia , Feminino , Biomarcadores Tumorais/sangue , Biomarcadores Tumorais/genética , Pessoa de Meia-Idade , Ácidos Nucleicos Livres/sangue , Ácidos Nucleicos Livres/genética , Adulto , Prognóstico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Idoso , Resultado do Tratamento , Perfilação da Expressão GênicaRESUMO
T-cell lymphoma is a highly invasive tumor with significant heterogeneity. Invasive tissue biopsy is the gold standard for acquiring molecular data and categorizing lymphoma patients into genetic subtypes. However, surgical intervention is unfeasible for patients who are critically ill, have unresectable tumors, or demonstrate low compliance, making tissue biopsies inaccessible to these patients. A critical need for a minimally invasive approach in T-cell lymphoma is evident, particularly in the areas of early diagnosis, prognostic monitoring, treatment response, and drug resistance. Therefore, the clinical application of liquid biopsy techniques has gained significant attention in T-cell lymphoma. Moreover, liquid biopsy requires fewer samples, exhibits good reproducibility, and enables real-time monitoring at molecular levels, thereby facilitating personalized health care. In this review, we provide a comprehensive overview of the current liquid biopsy biomarkers used for T-cell lymphoma, focusing on circulating cell-free DNA (cfDNA), circulating tumor DNA (ctDNA), circulating tumor cells (CTCs), Epstein-Barr virus (EBV) DNA, antibodies, and cytokines. Additionally, we discuss their clinical application, detection methodologies, ongoing clinical trials, and the challenges faced in the field of liquid biopsy.
Assuntos
Infecções por Vírus Epstein-Barr , Linfoma de Células T , Humanos , Reprodutibilidade dos Testes , Biomarcadores Tumorais/genética , Herpesvirus Humano 4 , Biópsia Líquida/métodos , Linfoma de Células T/diagnóstico , Linfoma de Células T/genéticaRESUMO
BACKGROUND: Current diagnostics for the detection of pancreato-biliary cancers (PBCs) need to be optimized. We therefore propose that methylated cell-free DNA (cfDNA) derived from non-invasive liquid biopsies serves as a novel biomarker with the ability to discriminate pancreato-biliary cancers from non-cancer pancreatitis patients. METHODS: Differentially methylated regions (DMRs) from plasma cfDNA between PBCs, pancreatitis and clinical control samples conditions were identified by next-generation sequencing after enrichment using methyl-binding domains and database searches to generate a discriminatory panel for a hybridization and capture assay with subsequent targeted high throughput sequencing. RESULTS: The hybridization and capture panel, covering around 74 kb in total, was applied to sequence a cohort of 25 PBCs, 25 pancreatitis patients, 25 clinical controls, and seven cases of Intraductal Papillary Mucinous Neoplasia (IPMN). An unbiased machine learning approach identified the 50 most discriminatory methylation markers for the discrimination of PBC from pancreatitis and controls resulting in an AUROC of 0.85 and 0.88 for a training (n = 45) and a validation (n = 37) data set, respectively. The panel was also able to distinguish high grade from low grade IPMN samples. CONCLUSIONS: We present a proof of concept for a methylation biomarker panel with better performance and improved discriminatory power than the current clinical marker CA19-9 for the discrimination of pancreato-biliary cancers from non-cancerous pancreatitis patients and clinical controls. This workflow might be used in future diagnostics for the detection of precancerous lesions, e.g. the identification of high grade IPMNs vs. low grade IPMNs.
Assuntos
Carcinoma Ductal Pancreático , Ácidos Nucleicos Livres , Neoplasias Intraductais Pancreáticas , Neoplasias Pancreáticas , Pancreatite , Humanos , Biomarcadores Tumorais/genética , Neoplasias Pancreáticas/diagnóstico , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Pancreatite/diagnóstico , Pancreatite/genética , Biópsia Líquida , Carcinoma Ductal Pancreático/patologiaRESUMO
BACKGROUND: Molecular characterization has significantly improved the management of advanced endometrial cancer (EC). It distinguishes four molecular subclasses associated with prognosis and personalized therapeutic strategies. This study assesses the clinical utility of cell-free DNA (cfDNA) profiling in EC to identify targetable alterations. METHODS: Women with metastatic or recurrent EC were prospectively recruited within the framework of the STING trial (NCT04932525), during which cfDNA was analyzed. Genomic alterations were identified with the FoundationOne CDx assay. Each molecular report underwent review by a molecular tumor board. Alterations were categorized via the European Society of Medical Oncology Scale for Clinical Actionability of Molecular Targets (ESCAT). RESULTS: A total of 61 patients were enrolled. The median age was 66.9 years, with 43% presenting frontline metastatic disease. All histologic subgroups were represented. Notably, 89% of patients yielded informative cfDNA analysis. Six tumors were classified with deficient mismatch repair/microsatellite instability (11%) and 37 as TP53 gene mutant (67%), and 12 had nonspecific molecular profiles (22%). Molecular classification based on liquid biopsy showed 87.5% accuracy in correlating with tissue results. Moreover, 65% of cases exhibited ≥1 actionable alteration, including 25% ESCAT I alterations and 13% ESCAT II alterations. Consequently, 16% of patients received a molecularly matched therapy, and presented with a 56% response rate and median progression-free survival of 7.7 months. CONCLUSIONS: cfDNA sequencing in EC is a feasible approach that produces informative results in 89% of cases and accurately categorizes patients into the main molecular subclasses. It also reveals multiple actionable alterations, which offers the potential for personalized therapeutic strategies.
RESUMO
The diagnosis and management of hepatocellular carcinoma (HCC) have improved significantly in recent years. With the introduction of immunotherapy-based combination therapy, there has been a notable expansion in treatment options for patients with unresectable HCC. Simultaneously, innovative molecular tests for early detection and management of HCC are emerging. This progress prompts a key question: as liquid biopsy techniques rise in prominence, will they replace traditional tissue biopsies, or will both techniques remain relevant? Given the ongoing challenges of early HCC detection, including issues with ultrasound sensitivity, accessibility, and patient adherence to surveillance, the evolution of diagnostic techniques is more relevant than ever. Furthermore, the accurate stratification of HCC is limited by the absence of reliable biomarkers which can predict response to therapies. While the advantages of molecular diagnostics are evident, their potential has not yet been fully harnessed, largely because tissue biopsies are not routinely performed for HCC. Liquid biopsies, analysing components such as circulating tumour cells, DNA, and extracellular vesicles, provide a promising alternative, though they are still associated with challenges related to sensitivity, cost, and accessibility. The early results from multi-analyte liquid biopsy panels are promising and suggest they could play a transformative role in HCC detection and management; however, comprehensive clinical validation is still ongoing. In this review, we explore the challenges and potential of both tissue and liquid biopsy, highlighting that these diagnostic methods, while distinct in their approaches, are set to jointly reshape the future of HCC management.
Assuntos
Carcinoma Hepatocelular , Biópsia Líquida , Neoplasias Hepáticas , Humanos , Biomarcadores Tumorais , Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/terapia , Carcinoma Hepatocelular/genética , Biópsia Líquida/métodos , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/terapia , Neoplasias Hepáticas/genética , Células Neoplásicas CirculantesRESUMO
PURPOSE: Although changes in circulating tumour DNA (ctDNA) in breast cancer are well described, the kinetics of their fluctuations has not been described over short timescales. We investigated ctDNA dynamics during alternating cycles of chemotherapy and hormonal treatment in pre-treated patients with oestrogen receptor-positive metastatic breast cancer. METHODS: Patients received alternating, 9-week cycles of eribulin and aromatase inhibitors (AIs). The clinical primary endpoint, progression-free survival (PFS), was monitored at 3, 6 and 9 months; secondary endpoints, clinical benefit rate (CBR), safety and tolerability profiles, were also assessed. Importantly, ctDNA fluctuations were monitored using the Oncomine™ Breast cfDNA assay to test whether biomarkers may change rapidly between chemotherapy and aromatase inhibitor (AI) treatment in the setting of advanced breast cancer, potentially reflecting disease dynamics. RESULTS: The median PFS was 202 days (95% CI: 135-undefined) and 235 days (95% CI: 235-undefined) at 6 and 9 months, respectively, with a 50% CBR at both 6 and 9 months. Dynamic changes in ctDNA were observed in short timescales between chemotherapy and AI treatment and support the clinical benefit (CB) seen in individual patients and, critically, appear informative of acquired resistance in real time. CONCLUSION: Changes in ctDNA can occur rapidly and reflect changes in patients' clinical tumour responses (NCT02681523).
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica , Inibidores da Aromatase , Neoplasias da Mama , DNA Tumoral Circulante , Adulto , Idoso , Feminino , Humanos , Pessoa de Meia-Idade , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Inibidores da Aromatase/uso terapêutico , Inibidores da Aromatase/administração & dosagem , Biomarcadores Tumorais/sangue , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Neoplasias da Mama/sangue , Neoplasias da Mama/mortalidade , Neoplasias da Mama/genética , DNA Tumoral Circulante/sangue , DNA Tumoral Circulante/genética , Furanos/uso terapêutico , Furanos/administração & dosagem , Cetonas , Metástase Neoplásica , Policetídeos de Poliéter , Resultado do TratamentoRESUMO
BACKGROUND: Uterine corpus endometrial carcinoma (UCEC) is a prevalent gynecologic malignancy with a favorable prognosis if detected early. However, there is a lack of accurate and reliable early detection tests for UCEC. This study aims to develop a precise and non-invasive diagnostic method for UCEC using circulating cell-free DNA (cfDNA) fragmentomics. METHODS: Peripheral blood samples were collected from all participants, and cfDNA was extracted for analysis. Low-coverage whole-genome sequencing was performed to obtain cfDNA fragmentomics data. A robust machine learning model was developed using these features to differentiate between UCEC and healthy conditions. RESULTS: The cfDNA fragmentomics-based model showed high predictive power for UCEC detection in training (n = 133; AUC 0.991) and validation cohorts (n = 89; AUC 0.994). The model manifested a specificity of 95.5% and a sensitivity of 98.5% in the training cohort, and a specificity of 95.5% and a sensitivity of 97.8% in the validation cohort. Physiological variables and preanalytical procedures had no significant impact on the classifier's outcomes. In terms of clinical benefit, our model would identify 99% of Chinese UCEC patients at stage I, compared to 21% under standard care, potentially raising the 5-year survival rate from 84 to 95%. CONCLUSION: This study presents a novel approach for the early detection of UCEC using cfDNA fragmentomics and machine learning showing promising sensitivity and specificity. Using this model in clinical practice could significantly improve UCEC management and control, enabling early intervention and better patient outcomes. Further optimization and validation of this approach are warranted to establish its clinical utility.
Assuntos
Ácidos Nucleicos Livres , Detecção Precoce de Câncer , Neoplasias do Endométrio , Humanos , Feminino , Neoplasias do Endométrio/diagnóstico , Neoplasias do Endométrio/sangue , Neoplasias do Endométrio/genética , Pessoa de Meia-Idade , Ácidos Nucleicos Livres/sangue , Detecção Precoce de Câncer/métodos , Idoso , Aprendizado de Máquina , Adulto , Biomarcadores Tumorais/sangue , Biomarcadores Tumorais/genética , Sensibilidade e EspecificidadeRESUMO
Aspergillus species and Mucorales agents are the primary etiologies of invasive fungal disease (IFD). Biomarkers that predict outcomes are needed to improve care. Patients diagnosed with invasive aspergillosis and mucormycosis using plasma cell-free DNA (cfDNA) PCR were retested weekly for 4 weeks. The primary outcome included all-cause mortality at 6 weeks and 6 months based on baseline cycle threshold (CT) values and results of follow-up cfDNA PCR testing. Forty-five patients with Aspergillus and 30 with invasive Mucorales infection were retested weekly for a total of 197 tests. Using the European Organization for Research and Treatment of Cancer and the Mycoses Study Group Education and Research Consortium (EORTC/MSG) criteria, 30.7% (23/75), 25.3% (19/75), and 38.7% (29/75) had proven, probable, and possible IFD, respectively. In addition, 97.3% (73/75) were immunocompromised. Baseline CT increased significantly starting at week 1 for Mucorales and week 2 for Aspergillus. Aspergillosis and mucormycosis patients with higher baseline CT (CT >40 and >35, respectively) had a nonsignificantly higher survival rate at 6 weeks, compared with patients with lower baseline CT. Mucormycosis patients with higher baseline CT had a significantly higher survival rate at 6 months. Mucormycosis, but not aspergillosis patients, with repeat positive cfDNA PCR results had a nonsignificantly lower survival rate at 6 weeks and 6 months compared with patients who reverted to negative. Aspergillosis patients with baseline serum Aspergillus galactomannan index <0.5 and <1.0 had significantly higher survival rates at 6 weeks when compared with those with index ≥0.5 and ≥1.0, respectively. Baseline plasma cfDNA PCR CT can potentially be used to prognosticate survival in patients with invasive Aspergillus and Mucorales infections. IMPORTANCE: We show that Aspergillus and Mucorales plasma cell-free DNA PCR can be used not only to noninvasively diagnose patients with invasive fungal disease but also to correlate the baseline cycle threshold with survival outcomes, thus potentially allowing the identification of patients at risk for poor outcomes, who may benefit from more targeted therapies.
Assuntos
Ácidos Nucleicos Livres , DNA Fúngico , Infecções Fúngicas Invasivas , Mucormicose , Reação em Cadeia da Polimerase , Humanos , Mucormicose/diagnóstico , Mucormicose/mortalidade , Mucormicose/sangue , Mucormicose/microbiologia , Masculino , Feminino , Pessoa de Meia-Idade , Prognóstico , Idoso , Ácidos Nucleicos Livres/sangue , Reação em Cadeia da Polimerase/métodos , Adulto , DNA Fúngico/genética , DNA Fúngico/sangue , Infecções Fúngicas Invasivas/diagnóstico , Infecções Fúngicas Invasivas/mortalidade , Infecções Fúngicas Invasivas/microbiologia , Aspergillus/genética , Aspergillus/isolamento & purificação , Aspergilose/diagnóstico , Aspergilose/mortalidade , Aspergilose/microbiologia , Mucorales/genética , Mucorales/isolamento & purificação , Biomarcadores/sangue , Idoso de 80 Anos ou mais , Estudos ProspectivosRESUMO
BACKGROUND: The current precision medicine relies on biomarkers, which are mainly obtained through next-generation sequencing (NGS). However, this model failed to find effective drugs for most cancer patients. This study tried to combine liquid biopsy with functional drug tests using organoid models to find potential drugs for cancer patients. METHODS: Colorectal cancer (CRC) patients were prospectively enrolled and blood samples were collected from patients before the start of treatment. Targeted deep sequencing of cfDNA samples was performed using a 14-gene panel. Gastrointestinal (GI) cancer organoids were established and PI3K and mTOR inhibitors were evaluated on organoid models. RESULTS: A total of 195 mutations were detected across 58 cfDNA samples. The most frequently mutated genes were KRAS, TP53, PIK3CA, and BRAF, all of which exhibited higher mutation rates than tissue biopsy. Although 81% of variants had an allele frequency of less than 1%, certain mutations in KRAS, TP53, and SMAD4 had high allele frequencies exceeding 10%. Notably, among the seven patients with high allele frequency mutations, six had metastatic tumors, indicating that a high allele frequency of ctDNA could potentially serve as a biomarker of later-stage cancer. A high rate of PIK3CA mutation (31 out of 67, or 46.3%) was discovered in CRC patients, suggesting possible tumor progression mechanisms and targeted therapy opportunities. To evaluate the value of anti PI3K strategy in GI cancer, different lines of GI cancer organoids were established. The organoids recapitulated the morphologies of the original tumors. Organoids were generally insensitive to PI3K inhibitors. However, CRC-3 and GC-4 showed response to mTOR inhibitor Everolimus, and GC-3 was sensitive to PI3Kδ inhibitor Idelalisib. The CRC organoid with a PIK3CA mutation showed greater sensitivity to the PI3K inhibitor Alpelisib than wildtype organoids, suggesting potential treatment options for the corresponding patients. CONCLUSION: Liquid biopsy holds significant promise for improving precision treatment and tumor prognosis in colorectal cancer patients. The combination of biomarker-based drug prediction with organoid-based functional drug sensitivity assay may lead to more effective cancer treatment.
Assuntos
Ácidos Nucleicos Livres , Neoplasias Colorretais , Humanos , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Neoplasias Colorretais/diagnóstico , Fosfatidilinositol 3-Quinases/genética , Avaliação Pré-Clínica de Medicamentos , Proteínas Proto-Oncogênicas p21(ras)/genética , Detecção Precoce de Câncer , Biópsia Líquida , Inibidores de Fosfoinositídeo-3 Quinase , Biomarcadores , Classe I de Fosfatidilinositol 3-Quinases/genética , Mutação/genéticaRESUMO
BACKGROUND: Cell free DNA (cfDNA)-based assays hold great potential in detecting early cancer signals yet determining the tissue-of-origin (TOO) for cancer signals remains a challenging task. Here, we investigated the contribution of a methylation atlas to TOO detection in low depth cfDNA samples. METHODS: We constructed a tumor-specific methylation atlas (TSMA) using whole-genome bisulfite sequencing (WGBS) data from five types of tumor tissues (breast, colorectal, gastric, liver and lung cancer) and paired white blood cells (WBC). TSMA was used with a non-negative least square matrix factorization (NNLS) deconvolution algorithm to identify the abundance of tumor tissue types in a WGBS sample. We showed that TSMA worked well with tumor tissue but struggled with cfDNA samples due to the overwhelming amount of WBC-derived DNA. To construct a model for TOO, we adopted the multi-modal strategy and used as inputs the combination of deconvolution scores from TSMA with other features of cfDNA. RESULTS: Our final model comprised of a graph convolutional neural network using deconvolution scores and genome-wide methylation density features, which achieved an accuracy of 69% in a held-out validation dataset of 239 low-depth cfDNA samples. CONCLUSIONS: In conclusion, we have demonstrated that our TSMA in combination with other cfDNA features can improve TOO detection in low-depth cfDNA samples.
Assuntos
Metilação de DNA , Genoma Humano , Neoplasias , Redes Neurais de Computação , Humanos , Metilação de DNA/genética , Neoplasias/genética , Neoplasias/sangue , Neoplasias/diagnóstico , Ácidos Nucleicos Livres/sangue , Ácidos Nucleicos Livres/genética , Especificidade de Órgãos/genética , AlgoritmosRESUMO
The cumulative pool of cell-free DNA (cfDNA) molecules within bodily fluids represents a highly dense and multidimensional information repository. This "biological mirror" provides real-time insights into the composition, function, and dynamics of the diverse genomes within the body, enabling significant advancements in personalized molecular medicine. However, effective use of this information necessitates meticulous classification of distinct cfDNA subtypes with exceptional precision. While cfDNA molecules originating from different sources exhibit numerous genetic, epigenetic, and physico-chemical variations, they also share common features that complicate analyses. Considerable progress has been achieved in mapping the landscape of cfDNA features, their clinical correlations, and optimizing extraction procedures, analytical approaches, bioinformatics pipelines, and machine learning algorithms. Nevertheless, preanalytical workflows, despite their profound impact on cfDNA measurements, have not progressed at a corresponding pace. In this perspective article, we emphasize the pivotal role of robust preanalytical procedures in the development and clinical integration of cfDNA assays, highlighting persistent obstacles and emerging challenges.
Assuntos
Ácidos Nucleicos Livres , Neoplasias , Humanos , Ácidos Nucleicos Livres/genética , Neoplasias/genética , Biomarcadores Tumorais/genética , Biologia Computacional , Medicina de PrecisãoRESUMO
BACKGROUND: Circulating cell-free DNA (cfDNA, liquid biopsy) is a powerful tool to detect molecular alterations. However, depending on tumor characteristics, biology and anatomic localization, cfDNA detection and analysis may be challenging. Gliomas are enclosed into an anatomic sanctuary, which obstacles the release of cfDNA into the peripheral blood. Therefore, the advantages of using liquid biopsy for brain tumors is still to be confirmed. The present study evaluates the ability of liquid biopsy to detect IDH1 mutations and its correlation with survival and clinical characteristics of glioma patients. METHODS: Blood samples obtained from glioma patients were collected after surgery prior to the adjuvant therapy. cfDNA was extracted from plasma and IDH1 p.R132H mutation analysis was performed on a digital droplet PCR. χ2-test and Cohen k were used to assess the correlation between plasma and tissue IDH1 status, while Kaplan Meier curve and Cox regression analysis were applied to survival analysis. Statistical calculations were performed by MedCalc and GraphPad Prism software. RESULTS: A total of 67 samples were collected. A concordance between IDH1 status in tissue and in plasma was found (p = 0.0024), and the presence of the IDH1 mutation both in tissue (138.8 months vs 24.4, p < 0.0001) and cfDNA (116.3 months vs 35.8, p = 0.016) was associated with longer median OS. A significant association between IDH1 mutation both in tissue and cfDNA, age, tumor grade and OS was demonstrated by univariate Cox regression analysis. No statistically significant association between IDH1 mutation and tumor grade was found (p = 0.10). CONCLUSIONS: The present study demonstrates that liquid biopsy may be used in brain tumors to detect IDH1 mutation which represents an important prognostic biomarker in patients with different types of gliomas, being associated to OS.
Assuntos
Neoplasias Encefálicas , Ácidos Nucleicos Livres , Glioma , Humanos , Glioma/patologia , Mutação , Neoplasias Encefálicas/patologia , Reação em Cadeia da Polimerase , Ácidos Nucleicos Livres/genética , Isocitrato Desidrogenase/genéticaRESUMO
BACKGROUND: Detection of cancer and identification of tumor origin at an early stage improve the survival and prognosis of patients. Herein, we proposed a plasma cfDNA-based approach called TOTEM to detect and trace the cancer signal origin (CSO) through methylation markers. METHODS: We performed enzymatic conversion-based targeted methylation sequencing on plasma cfDNA samples collected from a clinical cohort of 500 healthy controls and 733 cancer patients with seven types of cancer (breast, colorectum, esophagus, stomach, liver, lung, and pancreas) and randomly divided these samples into a training cohort and a testing cohort. An independent validation cohort of 143 healthy controls, 79 liver cancer patients and 100 stomach cancer patients were recruited to validate the generalizability of our approach. RESULTS: A total of 57 multi-cancer diagnostic markers and 873 CSO markers were selected for model development. The binary diagnostic model achieved an area under the curve (AUC) of 0.907, 0.908 and 0.868 in the training, testing and independent validation cohorts, respectively. With a training specificity of 98%, the specificities in the testing and independent validation cohorts were 100% and 98.6%, respectively. Overall sensitivity across all cancer stages was 65.5%, 67.3% and 55.9% in the training, testing and independent validation cohorts, respectively. Early-stage (I and II) sensitivity was 50.3% and 45.7% in the training and testing cohorts, respectively. For cancer patients correctly identified by the binary classifier, the top 1 and top 2 CSO accuracies were 77.7% and 86.5% in the testing cohort (n = 148) and 76.0% and 84.0% in the independent validation cohort (n = 100). Notably, performance was maintained with only 21 diagnostic and 214 CSO markers, achieving a training AUC of 0.865, a testing AUC of 0.866, and an integrated top 2 accuracy of 83.1% in the testing cohort. CONCLUSIONS: TOTEM demonstrates promising potential for accurate multi-cancer detection and localization by profiling plasma methylation markers. The real-world clinical performance of our approach needs to be investigated in a much larger prospective cohort.
Assuntos
Biomarcadores Tumorais , DNA Tumoral Circulante , Metilação de DNA , Neoplasias , Humanos , Biomarcadores Tumorais/sangue , Biomarcadores Tumorais/genética , Neoplasias/genética , Neoplasias/sangue , Neoplasias/diagnóstico , Feminino , Masculino , DNA Tumoral Circulante/sangue , DNA Tumoral Circulante/genética , Pessoa de Meia-Idade , Idoso , Detecção Precoce de Câncer/métodos , Estudos de Casos e Controles , Sensibilidade e Especificidade , Adulto , PrognósticoRESUMO
PURPOSE: Genomic alterations are fundamental for molecular-guided therapy in patients with breast and lung cancer. However, the turn-around time of standard next-generation sequencing assays is a limiting factor in the timely delivery of genomic information for clinical decision-making. METHODS: In this study, we evaluated genomic alterations in 54 cerebrospinal fluid samples from 33 patients with metastatic lung cancer and metastatic breast cancer to the brain using the Oncomine Precision Assay on the Genexus sequencer. There were nine patients with samples collected at multiple time points. RESULTS: Cell-free total nucleic acids (cfTNA) were extracted from CSF (0.1-11.2 ng/µl). Median base coverage was 31,963× with cfDNA input ranging from 2 to 20 ng. Mutations were detected in 30/54 CSF samples. Nineteen (19/24) samples with no mutations detected had suboptimal DNA input (< 20 ng). The EGFR exon-19 deletion and PIK3CA mutations were detected in two patients with increasing mutant allele fraction over time, highlighting the potential of CSF-cfTNA analysis for monitoring patients. Moreover, the EGFR T790M mutation was detected in one patient with prior EGFR inhibitor treatment. Additionally, ESR1 D538G and ESR1::CCDC170 alterations, associated with endocrine therapy resistance, were detected in 2 mBC patients. The average TAT from cfTNA-to-results was < 24 h. CONCLUSION: In summary, our results indicate that CSF-cfTNA analysis with the Genexus-OPA can provide clinically relevant information in patients with brain metastases with short TAT.