Comparison of whole-exome, whole-transcriptome genomic profiling and targeted sequencing with 50-gene panels in advanced solid tumors

Genome Profiling of Actionable Cancer Targets (NYU LG-PACT) for Clinical Patient Molecular Diagnostics and Treatment

Transforming routine solid tumor profiling with automated next-generation sequencing: experience from a reference general hospital.

Screening tumor types for which a first-line, integrated DNA-RNA NGS strategy provides a valuable advantage for rapid therapeutic decisions demonstrates that integrated DNA-RNA high-throughput NGS enables timely, precise molecular profiling for personalized therapy in solid tumors.

Clinical utility of real-time comprehensive molecular testing in large B-cell lymphoma.

Due to the rapidly evolving landscape of targeted therapies, there is an unmet need for comprehensive molecular profiling to guide treatment decisions for patients with large B-cell lymphoma (LBCL). Therefore, we designed a pilot study to assess the feasibility and turnaround time (TAT) of a comprehensive whole exome sequencing (WES) and transcriptome sequencing (RNA-seq) assay in patients with LBCL (NCT05464823). Patients aged ≥18 years with LBCL were eligible. Formalin-fixed paraffin-embedded tissues at diagnosis or recurrence underwent WES, RNA-seq, and copy number analysis with concomitant germline DNA sequencing. Genomic and transcriptomic data were profiled to define various LBCL signatures such as cell-of-origin (COO), dark zone signature (DZsig), LymphGen and lymphoma microenvironment (LME) classification. Among 100 patients enrolled, samples from 71 patients (48 with newly diagnosed and 23 with relapsed/refractory disease) passed pathology quality control and underwent WES and RNA-seq analysis and reporting. The median TAT was 8 days for individual patient reports (range: 6-22 days), with 73% of reports delivered ≤10 days. Applying molecular risk classification, high risk event-free survival within 24 months (EFS24) signature was associated with DZsig and LME, but not with COO or LymphGen indicating the complex heterogeneity of the disease. We demonstrated the clinical utility and acceptable TAT of a comprehensive WES and RNA-seq assay for LBCL managed in routine clinical practice. These findings support the real-world feasibility of using integrated WES and RNA-seq to define molecular subgroups, guide clinical decision-making at the time of a new line of therapy, and enable biology based subtype-driven clinical trials.

Next-Generation Sequencing in Colorectal Cancer: Real-World Molecular Profiling and Clinical Correlations

Next-generation sequencing (NGS) has become a cornerstone of precision oncology in colorectal cancer (CRC), although its role in routine patient stratification remains incompletely defined. This retrospective single-centre study characterised the molecular landscape of clinically selected CRC patients undergoing routine NGS and explored associations between genomic alterations and clinicopathological features. 97 eligible patients who underwent targeted NGS using two validated sequencing platforms were selected. Demographic, clinicopathological, and molecular data were integrated, and associations were evaluated using descriptive statistics, exploratory association testing, principal component analysis, and multiple correspondence analysis. At least one reportable genetic alteration was identified in 91 patients (93.8%), comprising 198 alteration events across multiple cancer-related genes. The most frequently altered genes with pathogenic or likely pathogenic variants were TP53 (55.7%), KRAS (40.2%), PIK3CA (18.6%), and BRAF (11.5%), while microsatellite instability was detected in 19.1% of evaluable tumours. Exploratory analyses identified associations between selected molecular alterations and clinicopathological characteristics. However, these were generally modest, frequently limited by small subgroup sizes, and none survived Benjamini–Hochberg correction. Principal component analysis demonstrated substantial molecular heterogeneity without defining distinct clinicopathological subgroups. Routine targeted NGS provides detailed molecular characterisation of CRC and generates information relevant to biomarker-informed clinical decision-making in real-world practice. However, targeted panels alone were insufficient to establish robust molecular subgroups in this retrospective cohort. These findings highlight the biological complexity of CRC and support larger prospective studies incorporating broader molecular profiling to improve precision patient stratification and optimise personalised therapeutic strategies.

Beyond Targeted Gene Panels: Whole-Exome Sequencing as a Strategic Platform for Precision Therapeutics in Alzheimer’s Disease

Alzheimer’s disease (AD) continues to be one of the greatest challenges in public health due to its multifactorial and heterogeneous nature, involving multiple physiological axes that encompass a large number of genetic, metabolic, vascular, and inflammatory interactions. In current clinical practice, medical specialties, mainly neurology and psychiatry, still rely on targeted gene panels for genetic evaluation. Although these panels remain effective for certain predefined hypotheses, their restricted and predefined nature limits the detection of the broader spectrum of genetic variation that may contribute to the complex biological interactions underlying neurodegeneration. Whole-exome sequencing (WES) is, from our clinic-based perspective, one of the most comprehensive genomic approaches currently available, since it allows the analysis of the ~19,500 protein-coding regions, and depending on the library approximately 5500 additional clinically relevant genomic loci, including splice sites, untranslated regions, long non-coding RNAs (lncRNAs), pseudogenes, regulatory elements and mitochondrial DNA (mtDNA). It enables the identification of pathogenic variants and variants of uncertain significance (VUS) under the American College of Medical Genetics and Genomics and the Association for Molecular Pathology (ACMG/AMP) classification frameworks. It also expands biological interpretation to variants conventionally classified as benign, which, when interpreted collectively, may contribute to pathway-level contextualization within the hypothesis-generating theoretical framework proposed in this review without implying pathogenicity, causal inference, or immediate clinical actionability. Additionally, WES enables the identification of secondary and incidental findings that may provide clinically relevant information beyond the primary phenotype, thereby supporting preventive surveillance and clinical risk management. This review analyzes the use of WES as a strategic platform for personalized decision-making in contemporary practice given the multifactorial and heterogeneous complexity of AD. It also addresses the complexities and limitations of the ACMG/AMP recommendations for filtering and classification of variants, the lack of standardization between reports and platforms, and the need for physician training, which constitute a great challenge for the translation of data to therapeutic decision-making. While the clinical utility of whole-exome sequencing (WES) in genetic diagnosis and precision medicine is well established, this review additionally proposes a hypothesis-generating theoretical framework whereby variants conventionally classified as benign or of uncertain significance may contribute to pathway-level biological contextualization in Alzheimer’s disease.

Exome-based cancer driver gene comprehensive testing can provide a genetic diagnosis for individuals with triple-negative breast cancer

Triple-negative breast cancer (TNBC) is characterized by aggressive behaviour, high tumor heterogeneity, and an increased likelihood of recurrence and early metastasis. These factors hinder successful treatment. Genetic diagnosis enables personalized clinical recommendations and treatment options. The objective of this study was to validate whole-exome sequencing (WES) and variant prioritization in cancer susceptibility genes (CSG) associated with hereditary cancer (HC) predisposition in TNBC patients (n = 24). We present the development of a reproducible bioinformatic pipeline and its technical validation in a validation cohort (n = 25). This cohort comprised individuals with diverse primary tumors who had a previously confirmed molecular diagnosis of a hereditary cancer syndrome, serving as gold-standard cases to assess the pipeline’s analytical accuracy. We consolidated a comprehensive panel of cancer genes and determined all variants in the TNBC discovery cohort (12.5% of patients), identifying three pathogenic germline variants (gPV) in ATM, RAD51D, and BRCA1. These genes are involved in the molecular pathway of DNA repair by homologous recombination (HRD). Our results demonstrate that the developed bioinformatic pipeline provides reliable genetic diagnosis of cancer predisposition syndromes from exome data, applicable not only to TNBC patients but also to individuals with any cancer suspected of having a hereditary component.