Next-generation Sequencing Multigene Panel Research Articles

Mutation landscape in Chinese nodal diffuse large B-cell lymphoma by targeted next generation sequencing and their relationship with clinicopathological characteristics

BackgroundDiffuse large B-cell lymphoma (DLBCL), an aggressive and heterogenic malignant entity, is still a challenging clinical problem, since around one-third of patients are not cured with primary treatment. Next-generation sequencing (NGS) technologies have revealed common genetic mutations in DLBCL. We devised an NGS multi-gene panel to discover genetic features of Chinese nodal DLBCL patients and provide reference information for panel-based NGS detection in clinical laboratories.MethodsA panel of 116 DLBCL genes was designed based on the literature and related databases. We analyzed 96 Chinese nodal DLBCL biopsy specimens through targeted sequencing.ResultsThe most frequently mutated genes were KMT2D (30%), PIM1 (26%), SOCS1 (24%), MYD88 (21%), BTG1 (20%), HIST1H1E (18%), CD79B (18%), SPEN (17%), and KMT2C (16%). SPEN (17%) and DDX3X (6%) mutations were highly prevalent in our study than in Western studies. Thirty-three patients (34%) were assigned as genetic classification by the LymphGen algorithm, including 12 cases MCD, five BN2, seven EZB, seven ST2, and two EZB/ST2 complex. MYD88 L265P mutation, TP53 and BCL2 pathogenic mutations were unfavorable prognostic biomarkers in DLBCL.ConclusionsThis study presents the mutation landscape in Chinese nodal DLBCL, highlights the genetic heterogeneity of DLBCL and shows the role of panel-based NGS to prediction of prognosis and potential molecular targeted therapy in DLBCL. More precise genetic classification needs further investigations.

Molecular Characterization of Advanced-Stage Melanomas in Clinical Practice Using a Laboratory-Developed Next-Generation Sequencing Panel.

Cutaneous melanoma is one of the most lethal tumors among skin cancers, characterized by complex genetic and molecular alterations that result in uncontrolled cell proliferation and metastatic spread. Next-generation sequencing (NGS) enables the simultaneous examination of numerous genes, making this molecular technique essential for melanoma diagnosis, prognostic stratification, and therapy planning. Herein, we present the experience with our laboratory-designed NGS panel for the routine assessment of advanced-stage melanoma. A total of 260 specimens of advanced-stage melanomas were evaluated utilizing a laboratory-developed multi-gene NGS panel, which allowed the investigation of 229 amplicons in 25 oncogene/oncosuppressor genes. The NGS panel proved to be a reliable tool, failing to produce results in only 1.2% of the samples tested. BRAF and TERT were the two more commonly altered genes in 44.0% and 59.9% of samples, respectively. In 59.3% of the mutated cases, at least two concomitant variants were detected. In eight cases, both primary lesion and metastatic disease were analyzed by NGS. In all specimens (8/8, 100%), a perfect concordance in variants harbored by the primary and recurrence lesions was observed. Finally, this study described the validity of a laboratory-developed multi-gene NGS panel built specifically for advanced-stage melanomas in ordinary clinical practice.

Real-World Experiences of Next-Generation Sequencing in Oncology: From an Indian Multicenter Registry and Collaborative Centers

Abstract Background The integration of next-generation sequencing (NGS) in guiding personalized therapy for oncology faces the challenges, primarily, of cost and drug accessibility. Limited data from Indian academic centers accentuate the need for comprehensive insights into the real-world applications of NGS in oncology. Methods The Network of Oncology Clinical Trials in India (NOCI), accessible at www.noci-india.com, compiled data on patients who underwent NGS for solid organ cancers from January 1, 2018, to December 31, 2021. This study aimed to elucidate the testing indications, sample types analyzed, and the resultant impact on patient care. Results Analysis of data from six centers included 278 subjects, with 24 specimens (9%) excluded due to quality test failure. Tissue constituted 59.7% of specimens, blood 38.5%, and both 1.8%. Predominantly, NGS was employed for identifying BRCA1/2 mutations (56%) and for targeted therapy in non-small-cell lung cancer (NSCLC; 28%). Only 41 (16%) patients with other cancers underwent multigene NGS panels in pursuit of targetable mutations. Among them, 13 exhibited targetable mutations, and 3 received treatment based on NGS findings. Conclusion This study underscores that the majority of NGS applications focused on screening for BRCA1/2 mutations and identifying targetable mutations in NSCLC. However, among those undergoing NGS for advanced cancers, only a limited number received personalized therapy. The findings underscore the challenges of utilizing NGS in off-label indications within resource-constrained settings.

Access, autonomy, and affordability: ethical and human rights issues surrounding multigene panel testing for cancer in Japan and Switzerland.

Introduction: Advancements in precision medicine and genomics have led to prospects in a wide range of clinical fields, including oncology. In particular, developments in next-generation sequencing multigene panel tests have led to the possibility of tailoring treatment to the specific genomic markers of a patient's cancer. However, findings from current literature suggest that the path to implementation and uptake of genomic medicine is not without uncertainties and challenges. Methods: To better understand the current challenges to the implementation of genomic medicine services, we investigated the current state of patient access to genomic medicine in Japan and Switzerland. In this investigation, we focused on equal access, patient autonomy, and healthcare affordability. Results: Results have shown that although multigene panel testing is in principle covered by health insurance in both countries, barriers exist in terms of where the tests are available, comprehensive information for patients, and the affordability of not only the test itself but the overall process from diagnosis to treatment. Discussion: These results suggest a need to continue examining a more diverse range of clinical landscapes for genomic medicine to reveal more nuanced understandings of barriers to implementation and thus better identify best practices for overcoming them.

Comprehensive Molecular Genetic Analysis in Glioma Patients by Next Generation Sequencing.

Glioma is caused by multiple genomic alterations. The evolving classification of gliomas emphasizes the significance of molecular testing. Next generation sequencing (NGS) offers the assessment of parallel combinations of multiple genetic alterations and identifying actionable mutations that guide treatment. This study comprehensively analyzed glioma patients using multi-gene NGS panels, providing powerful insights to inform diagnostic classification and targeted therapies. We conducted a targeted panel-based NGS analysis on formalin-fixed and paraffin-embedded nucleic acids extracted from a total of 147 glioma patients. These samples underwent amplicon capture-based library preparation and sequenced using the Oncomine Comprehensive Assay platform. The resulting sequencing data were then analyzed using the bioinformatics tools. A total of 301 mutations, were found in 132 out of 147 tumors (89.8%). These mutations were in 68 different genes. In 62 tumor samples (42.2%), copy number variations (CNVs) with gene amplifications occurred in 25 genes. Moreover, 25 tumor samples (17.0%) showed gene fusions in 6 genes and intragenic deletion in a gene. Our analysis identified actionable targets in several genes, including 11 with mutations, 8 with CNVs, and 3 with gene fusions and intragenic deletion. These findings could impact FDA-approved therapies, NCCN guideline-based treatments, and clinical trials. We analyzed precisely diagnosing the classification of gliomas, detailing the frequency and co-occurrence of genetic alterations and identifying genetic alterations with potential therapeutic targets by NGS-based molecular analysis. The high-throughput NGS analysis is an efficient and powerful tool to comprehensively support molecular testing in neurooncology.

ADCY5-related dyskinesia: a treatable inherited neurometabolic disorder

Case Report. A 40-year-old Brazilian man presented with long-standing loss of balance, dystonia, and choreoathetosis, with episodic paroxysmal worsening. Neuroimaging and cerebrospinal fluid analysis were both unremarkable. Quantitative urine organic acid analysis, plasma acylcarnitine quantitative profile, and quantitative analysis of plasma amino acids by high performance liquid chromatography were all within normal interval values. Multigene next-generation sequencing panel identified the heterozygous pathogenic variant c.1252C>T (p.Arg418Trp) in the ADCY5 gene, defining a diagnosis of ADCY5-related dyskinesia. Oral caffeine was associated to clonazepam and provided marked improvement of motor symptoms. Conclusion. This report adds evidence to the safety and efficacy of caffeine in the treatment of genetically confirmed cases of ADCY5-related dyskinesia in the context of gain-of-function variants.

Increasing targeted therapy options for mCRPC patients using multigene NGS panel.

e17056 Background: Prostate cancer (PCa) represents the second most common malignancy in males, characterized by a high level of clinical and molecular heterogeneity. Eventually, many patients develop metastatic Castration Resistant Prostate Cancer (mCRPC). mCRPC refers to prostate cancer that has spread beyond the prostate gland and is associated with increased morbidity and mortality. Next Generation Sequencing (NGS) has led to the identification of genomic alterations that may represent actionable targets with therapeutic potential in mCRPC. Such alterations can be identified on the somatic or hereditary level and implicate a variety of cell cycle mechanisms, like the DNA repair mechanism (Homologous Recombination Repair genes and Mismatch Repair Genes), as well as the AR, mTOR/AKT/PI3K, PTEN, ERK/MEK/Raf/Ras and WNT signaling pathways. Methods: In this study, 75 samples of patients with mCRPC were successfully processed. An NGS panel, targeting 513 genes associated with molecular and immuno-checkpoint inhibitor therapies, was used for the detection of gene alterations and fusions. In addition, genetic markers such as TMB, MSI and %gLOH were calculated for a subset of patients. Results: At least one mutation was detected in almost 90% of the cases tested, including SNVs, CNVs and gene fusions in a broad spectrum of the tested genes. In our cohort, the most commonly altered gene was CDK12 (9.3%), followed by ATM (6.7%) and PIK3CA (5.3%). Mutations were also found in FANCA (2.7%) and PTEN (2.7%). TMB was calculated in 52 patients, out of which 9.5% were characterized as TMB high ( > 10 muts/Mb). MSI status was high for 4.5% of patients and %gLOH was elevated in 37.5% of the tested cohort. A mutation in an HRR gene was found in 29% of patients, rendering them eligible for therapy with PARP inhibitors. In addition, 18.7% of the patients could benefit from off label treatment. The calculation of genetic biomarkers, like TMB, MSI, and LOH, led to an increase to the cohort of patients who could receive on label therapy, even in cases with no or non-targetable mutation on DNA or RNA level. Moreover, due to the continuous approval of new drug regimens, retrospective evaluation of tumor analysis results led to an increase of the percentage of patients that could benefit from targeted treatment by 20%. Conclusions: Our results indicate that tumor analysis is essential for the identification of actionable biomarkers in mCRPC, since more than half of the patients were positive for a biomarker related to either targeted therapy or immunotherapy. Hence, frequent re-evaluation of NGS results is essential due to continuous drug approvals.

Diagnostic utility of next generation sequencing for somatic variants in cancer: A seven year cumulative cohort.

e15060 Background: Next generation sequencing (NGS) makes it possible to identify the specific molecular biomarkers to define patient eligibility for targeted therapies via multigene panels. Since then, even though the interpretation of data is complex and not all tumors have genomic alterations within actionable genes, using somatic sequencing for cancer patients becomes a daily routine in clinical oncology. Thus, here we present the data derived from our experience in NGS testing of different cancer patients and biological samples over the course of 7 years in our genetic diagnosis center. Methods: The cohort was consisted of total 5511 cases. While 2590 (46.99%) of them were formalin fixed paraffin embedded (FFPE) tissues, 2906 (52.73%) were liquid biopsies and 15 (0.27%) were other types of biological samples. Total of 15 different cancers were classified (Table 1) and were sequenced using 3 different multigene NGS panels (additional Table 1, BRCAs’ testing for breast/ovarian cancer is not included). NGS was performed via GeneReader NGS System (Qiagen, Germany). Bioinformatics analyzes and variant classification were performed using QCI-Analyze and QCI-Interpret interfaces. Results: As a result, the cases of 93.36% (n = 5145) among all samples were successfully sequenced. The lung cancer group was the most frequent with 47.96% (n = 2643 patients), followed by 491 (%8.9) colorectal 288 (5.23%) and breast cancer patients respectively, while 781 cases (14.17%) grouped as other including many rare cancers. Overall positivity rate was 42.57% (n = 2346) and 366 samples (6.64%) were not sufficient enough to be sequenced. The distribution of positivity rates in relation to cancer type and biological samples used were listed in Table 1. Classification and diagnosis of all tested cases, specimens and Tier 1-2 variants included in this cohort are available on the Additional Table 2. Conclusions: This report is a comprehensive review of the aggregate data across the entire cohort of cancer patients referred to a genetics diagnosis center. While our findings highlight the importance of NGS testing in the somatic setting for cancer patients, also these data stress the importance of diagnosis, biological sample types tested and test results. [Table: see text]

Comprehensive genomic profiling and therapeutic implications for patients with advanced cancers: The experience of an academic hospital.

e15162 Background: Next generation sequencing (NGS) can be used to detect tumor-specific genomic alterations, with the aim to identify actionable genomic alteration and provide additional treatment options for patients with advanced cancer. While smaller sequencing panels are being applied broadly, controversy exists around the routine implementation of comprehensive genomic profiling with large multigene NGS panels. Indeed, there is an ongoing discussion about the indications as well as timing of comprehensive genomic profiling in patients with advanced solid tumors. Therefore, there is a need for more data to increase understanding of the potential indications and added clinical benefit of routine implementation of broad comprehensive genomic profiling. However, available studies have often been reporting on a small patient population, which complicates the interpretation and translation to the clinical setting. This study aims to describe the feasibility of performing broad NGS panels in patients with advanced cancer in an academic hospital setting, as well as the targetable alterations detected. Methods: In this retrospective study, we analysed genomic alterations in solid tumor biopsies of 464 patients with locally advanced or metastatic cancer. Next generation sequencing was performed using the Foundation Medicine assay (FoundationOneCDx). The therapeutic implications of the genomic alterations were determined using the OncoKB classification. Results: The FoundationOne CDx was successfully applied in 464/521 patients (89%) with a range of cancer types. The most common altered genes were TP53 (61%), followed by KRAS (20%), CDKN2A (20%), TERT (16%), APC (16%), PTEN (14%), PIK3CA (14%) and CDKN2B (14%). Among 419 patients with successfully analyzed tumor mutational burden (TMB), the median TMB was 2.52 (range 0 – 67) mutations/Mb (mut/Mb). Forty-three patients presented with a high-TMB ( > = 10 mut/Mb) of which eight had microsatellite instability. Out of 126 patients with an actionable target (e.g. ERBB2, KRAS, PIK3CA, FGFR2), 40 patients received matched treatment (8.6%), of which 17 were included in clinical trials. Conclusions: This single-center retrospective study shows that broad application of NGS is feasible in an academic center, increases the detection of actionable genomic alterations, and opens opportunities to provide treatment with immune checkpoint inhibitors in case of high tumor mutational burden and microsatellite instability status regardless of tumor histology. The study brings to attention that strategies such as inclusion in clinical (basket) trials and development of new targeted drugs are necessary to improve matched treatment rates. Further research is needed to assess the possible clinical benefit of broad application of NGS and genotype matched treatment.

Multi-Gene Next-Generation Sequencing Panel for Analysis of BRCA1/BRCA2 and Homologous Recombination Repair Genes Alterations Metastatic Castration-Resistant Prostate Cancer.

Despite significant therapeutic advances, metastatic CRPC (mCRPC) remains a lethal disease. Mutations in homologous recombination repair (HRR) genes are frequent in mCRPC, and tumors harboring these mutations are known to be sensitive to PARP inhibitors. The aim of this study was to verify the technical effectiveness of this panel in the analysis of mCRPC, the frequency and type of mutations in the BRCA1/BRCA2 genes, as well as in the homologous recombination repair (HRR) genes. A total of 50 mCRPC cases were analyzed using a multi-gene next-generation sequencing panel evaluating a total of 1360 amplicons in 24 HRR genes. Of the 50 cases, 23 specimens (46.0%) had an mCRPC harboring a pathogenic variant or a variant of uncertain significance (VUS), whereas in 27 mCRPCs (54.0%), no mutations were detected (wild-type tumors). BRCA2 was the most commonly mutated gene (14.0% of samples), followed by ATM (12.0%), and BRCA1 (6.0%). In conclusion, we have set up an NGS multi-gene panel that is capable of analyzing BRCA1/BRCA2 and HRR alterations in mCRPC. Moreover, our clinical algorithm is currently being used in clinical practice for the management of patients with mCRPC.

Pompe disease ascertained through The Lantern Project, 2018-2021: Next-generation sequencing and enzymatic testing to overcome obstacles to diagnosis.

The Lantern Project is an ongoing complimentary diagnostic program for patients in the United States sponsored by Sanofi and implemented by PerkinElmer Genomics. It combines specific enzymatic, biomarker, and genetic testing to facilitate rapid, accurate laboratory diagnosis of Pompe disease and several other lysosomal storage diseases, and a multigene next-generation sequencing panel including Pompe disease, LGMD, and other neuromuscular disorders. This article reports data for Pompe disease collected from October 2018 through December 2021, including acid α-glucosidase (GAA) enzyme assay and GAA sequencing (standard or expedited for positive newborn screening [NBS] to rule out infantile-onset Pompe disease [IOPD]) and the Focused Neuromuscular Panel, which includes GAA. One hundred forty patients (12 received only GAA enzyme testing, 128 had GAA sequencing alone or in addition to enzyme assay) have been confirmed with Pompe disease in this project. Eight of the 140 had a variant of unknown significance, but GAA activity ≤2.10μmol/L/h, thus were confirmed with Pompe disease. Three diagnosed patients 0-2years old had cross-reactive immunologic material (CRIM)-negative GAA variants and thus IOPD. One additional infant with presumptive IOPD had a homozygous frameshift c.1846del, likely CRIM-negative; symptoms were not provided. Among the 128 patients with molecular results, the c.-32-13T>G splice variant was homozygous in 11, compound-heterozygous in 98, and absent in 19. Proximal muscle weakness (58 patients) was the most common sign reported at testing; elevated creatine kinase (29 patients) was the most common laboratory result. The most common symptom categories were muscular (73 patients), musculoskeletal (13 patients), and respiratory (23 patients). Clinical information was not available for 42 samples, and 17 infants had only "abnormal NBS" or "low GAA" reported. Cardiac symptoms in 7 included potentially age-related conditions in five c.-32-13T>G-compound-heterozygous adults (myocardial infarction, heart murmur/palpitations, congestive heart failure: 1 each; 2 with atrial fibrillation) and hypertrophic cardiomyopathy in 2 children (1 and 2years old) with presumptive IOPD. One novel GAA variant was observed in a patient with enzyme activity 0.31μmol/L/h: c.1853_1854ins49, a frameshift pathogenic variant. The Lantern Project demonstrates the combinatorial utility of enzyme assay, targeted single-gene testing, and a focused neuromuscular next-generation sequencing panel in diagnosing Pompe disease.

Commercial Gene Panels for Congenital Anterior Segment Anomalies: Are They All the Same?

We compared next generation sequencing multigene panels (NGS-MGP) from 5 commercial laboratories to inform ophthalmologists' decision making in diagnostic genetic testing for congenital anterior segment anomalies (CASAs). Comparison of commercial genetic testing panels. This observational study gathered publicly available information on NGS-MGP from 5 commercial laboratories for the following: cataracts, glaucoma, anterior segment dysgenesis (ASD), microphthalmia-anophthalmia-coloboma (MAC), corneal dystrophies, and Axenfeld-Rieger syndrome (ARS). We compared gene panel composition, consensus rate (genes covered by all the panels per condition, "concurrent"), dissensus rate (genes covered by only 1 panel per condition, "standalone"), and intronic variant coverage. For individual genes, we compared publication history and association with systemic conditions. Altogether, cataract, glaucoma, corneal dystrophies, MAC, ASD, and ARS panels tested 239, 60, 36, 292, and 10 discrete genes, respectively. The consensus rate varied between 16% and 50%, and the dissensus rate varied between 14% and 74%. After pooling concurrent genes from all conditions, 20% of these genes were concurrent in 2 or more conditions. For both cataract and glaucoma, concurrent genes had significantly stronger correlation with the condition than standalone genes. The genetic testing of CASAs using NGS-MGPs is complicated, owing to their number, variety, and phenotypic and genetic overlap. Although the inclusion of additional genes, such as the standalone ones, might increase diagnostic yield, these genes are also less well studied, indicating uncertainty over their role in CASA pathogenesis. Rigorous prospective diagnostic yield studies of NGS-MGPs will aid in making decisions of panel selection for the diagnosis of CASAs.

Clinical usefulness of NGS multi-gene panel testing in hereditary cancer analysis.

Introduction: A considerable number of families with pedigrees suggestive of a Mendelian form of Breast Cancer (BC), Ovarian Cancer (OC), or Pancreatic Cancer (PC) do not show detectable BRCA1/2 mutations after genetic testing. The use of multi-gene hereditary cancer panels increases the possibility to identify individuals with cancer predisposing gene variants. Our study was aimed to evaluate the increase in the detection rate of pathogenic mutations in BC, OC, and PC patients when using a multi-gene panel. Methods: 546 patients affected by BC (423), PC (64), or OC (59) entered the study from January 2020 to December 2021. For BC patients, inclusion criteria were i) positive cancer family background, ii) early onset, and iii) triple negative BC. PC patients were enrolled when affected by metastatic cancer, while OC patients were all submitted to genetic testing without selection. The patients were tested using a Next-Generation Sequencing (NGS) panel containing 25 genes in addition to BRCA1/2. Results: Forty-four out of 546 patients (8%) carried germline pathogenic/likely pathogenic variants (PV/LPV) on BRCA1/2 genes, and 46 (8%) presented PV or LPV in other susceptibility genes. Discussion: Our findings demonstrate the utility of expanded panel testing in patients with suspected hereditary cancer syndromes, since this approach increased the mutation detection rate of 15% in PC, 8% in BC and 5% in OC cases. In absence of multi-gene panel analysis, a considerable percentage of mutations would have been lost.

Molecular profiling of male breast cancer by multigene panel testing: Implications for precision oncology.

Compared with breast cancer (BC) in women, BC in men is a rare disease with genetic and molecular peculiarities. Therapeutic approaches for male BC (MBC) are currently extrapolated from the clinical management of female BC, although the disease does not exactly overlap in males and females. Data on specific molecular biomarkers in MBC are lacking, cutting out male patients from more appropriate therapeutic strategies. Growing evidence indicates that Next Generation Sequencing (NGS) multigene panel testing can be used for the detection of predictive molecular biomarkers, including Tumor Mutational Burden (TMB) and Microsatellite Instability (MSI). In this study, NGS multigene gene panel sequencing, targeting 1.94 Mb of the genome at 523 cancer-relevant genes (TruSight Oncology 500, Illumina), was used to identify and characterize somatic variants, Copy Number Variations (CNVs), TMB and MSI, in 15 Formalin-Fixed Paraffin-Embedded (FFPE) male breast cancer samples. A total of 40 pathogenic variants were detected in 24 genes. All MBC cases harbored at least one pathogenic variant. PIK3CA was the most frequently mutated gene, with six (40.0%) MBCs harboring targetable PIK3CA alterations. CNVs analysis showed copy number gains in 22 genes. No copy number losses were found. Specifically, 13 (86.7%) MBCs showed gene copy number gains. MYC was the most frequently amplified gene with eight (53.3%) MBCs showing a median fold-changes value of 1.9 (range 1.8-3.8). A median TMB value of 4.3 (range 0.8-12.3) mut/Mb was observed, with two (13%) MBCs showing high-TMB. The median percentage of MSI was 2.4% (range 0-17.6%), with two (13%) MBCs showing high-MSI. Overall, these results indicate that NGS multigene panel sequencing can provide a comprehensive molecular tumor profiling in MBC. The identification of targetable molecular alterations in more than 70% of MBCs suggests that the NGS approach may allow for the selection of MBC patients eligible for precision/targeted therapy.

Real-world applications of tumor mutation burden (TMB) analysis using ctDNA and FFPE samples in various cancer types of Turkish population

Tumor mutation burden (TMB) has become one of the most popular approaches in the last decade as a molecular genetic testing strategy for cancer therapeutics that represents the somatic variations per Mbase in coding regions of the genome and which can be performed via comprehensive genomic profiling (CGP) by next generation sequencing (NGS). TMB is most commonly used to stratify the patients for immunotherapy as well as the actionable variant detection for possible other therapeutics. In this context, within this study, we share our results of the TMB score distribution of cancer patients together with distinctive diagnoses and specimens. The study was conducted from a total of 278 samples. One hundred seventy six (176) of them were formalin-fixed paraffin-embedded (FFPE) tissue samples and 102 liquid biopsy samples. Samples were sequenced using a multi-gene NGS panel consisting of 486 cancer-related genes (Illumina-NextSeq500/550). Bioinformatics analyzes were performed using an optimized in-house bioinformatics pipeline. As a result, the studies of 91.7% (n=255) among all samples were successfully performed in which total of 21 different cancer types were included. The lung cancer group was the most frequent (n=43 patients), followed by 31 colorectal cancer and 22 ovarian cancer patients. The classification of TMB scoring was very high (>50), high (20-50), moderate (5-20) and low (<5). The shared data of this study represents a cancer genome atlas-like data set for TMBs of Turkish cancer patients in relation to various cancer types and specimens in comparison with The Cancer Genome Atlas (TCGA) data.

Clinical Impact of Next-Generation Sequencing Multi-Gene Panel Highlighting the Landscape of Germline Alterations in Ovarian Cancer Patients.

BRCA1 and BRCA2 are the most frequently mutated genes in ovarian cancer (OC) crucial both for the identification of cancer predisposition and therapeutic choices. However, germline variants in other genes could be involved in OC susceptibility. We characterized OC patients to detect mutations in genes other than BRCA1/2 that could be associated with a high risk of developing OC and permit patients to enter the most appropriate treatment and surveillance program. Next-generation sequencing analysis with a 94-gene panel was performed on germline DNA of 219 OC patients. We identified 34 pathogenic/likely pathogenic variants in BRCA1/2 and 38 in other 21 genes. The patients with pathogenic/likely pathogenic variants in the non-BRCA1/2 genes mainly developed OC alone compared to the other groups that also developed breast cancer or other tumors (p = 0.001). Clinical correlation analysis showed that the low-risk patients were significantly associated with platinum sensitivity (p < 0.001). Regarding PARP inhibitors (PARPi) response, the patients with pathogenic mutations in the non-BRCA1/2 genes had worse PFS and OS. Moreover, a statistically significantly worse PFS was found for every increase of one thousand platelets before PARPi treatment. To conclude, knowledge about molecular alterations in genes beyond BRCA1/2 in OC could allow for more personalized diagnostic, predictive, prognostic, and therapeutic strategies for OC patients.

Challenges in integrating molecular profiles into clinical cancer care

SummaryProfiling of malignancies with next-generation sequencing (NGS) is now routine in clinical practice. While many cases of approved targeted therapies are straightforward based on well-characterized alterations, applying large NGS multigene panels to therapeutic use is frequently challenging. In this article, variant interpretation, therapy matching, and final treatment selection challenges are discussed.

Frequency of Parkinson's Disease Genes and Role of PARK2 in Amyotrophic Lateral Sclerosis: An NGS Study.

Amyotrophic lateral sclerosis (ALS) and Alzheimer’s disease (AD) patients show a higher prevalence of Lewy body disease than the general population. Additionally, parkinsonian features were found in about 30% of ALS patients. We aimed to explore the frequency of Parkinson’s disease (PD)-causative genes in ALS patients, compared to AD and healthy controls (HCs). We used next-generation sequencing multigene panels by analyzing SNCA, LRRK2, PINK1, PARK2, PARK7, SYNJ1, CHCHD2, PLA2G6, GCH1, ATP13A2, DNAJC6 and FBXO genes. GBA gene, a risk factor for PD, was also analyzed. In total, 130 ALS and 100 AD patients were investigated. PD-related genes were found to be altered in 26.2% of ALS, 20% of AD patients and 19.2% of HCs. Autosomal recessive genes were significantly more involved in ALS as compared to AD and HCs (p = 0.021). PARK2 variants were more frequent in ALS than in AD and HCs, although not significantly. However, the p.Arg402Cys variant was increased in ALS than in HCs (p = 0.025). This finding is consistent with current literature, as parkin levels were found to be decreased in ALS animal models and patients. Our results confirm the possible role of PD-related genes as risk modifier in ALS pathogenesis.

A hypothesis-generating analysis on the role of TERT promoter mutation in advanced urothelial carcinoma treated with immunotherapy.

BackgroundThe therapeutic scenario of urothelial carcinoma is constantly expanding with the widening of the knowledge on molecular characteristics, thus claiming for the need of prognostic and predictive factors to guide treatment strategy. TERT promoter mutation is one of the most frequent genomic alterations in urothelial carcinoma and could present several implications, from diagnostic to prognostic or potentially even predictive. MethodsWe performed a single-center retrospective analysis on patients with advanced urothelial carcinoma treated with an immune checkpoint inhibitor as second line of therapy to assess the status of the TERT promoter and the potential implication of its mutation on survival outcomes. ResultsWe analyzed tissue samples from 11 patients with a next-generation sequencing multi-gene panel. The most frequently altered genes were TP53 (54.5%, n = 6) and TERT promoter (36.3%, n = 4). Other mutations found were BRAF, SMAD4, PIK3CA / PDGRFA. The only type of detected TERT promoter mutation was the c 0.124 C>T (n = 4/4, 100%). Of the 4 TERT mutated patients, 2 presented a co-mutation of TP53. Patients with TERT promoter mutation treated with immunotherapy presented a low median overall survival (16.5 months) and progression-free survival (3.8 months). ConclusionsOur hypothesis-generating analysis suggests that the presence of TERT promoter mutation could have a negative prognostic value and should be further evaluated in wider cohorts.

Germline MUTYH mutations and high-grade gliomas: Novel evidence for a potential association.

There is growing body of evidence supporting the role of germline mutations in the pathogenesis of pediatric central nervous system (CNS) tumors, and the widespread use of next‐generation sequencing (NGS) panels facilitates their detection. Variants of the MUTYH gene are increasingly recognized as suspected germline background of various extraintestinal malignancies, besides their well‐characterized role in the polyposis syndrome associated with biallelic mutations. Using a multigene NGS panel (Illumina TruSight Oncology 500), we detected one H3 G34V‐ and one H3 K27M‐mutant pediatric high‐grade diffuse glioma, in association with c.1178G>A (p.G393D) and c.916C>T (p.R306C) MUTYH variants, respectively. Both MUTYH mutations were germline, heterozygous and inherited, according to the subsequent genetic testing of the patients and their first‐degree relatives. In the H3 K27M‐mutant glioma, amplifications affecting the 4q12 region were also detected, in association with KDR‐PDGFRA, KIT‐PDGFRA, and KDR‐CHIC2 fusions, previously unreported in this entity. Among 47 other CNS tumors of various histological types tested with the same NGS panel in our institution, only one adult glioblastoma harbored MUTYH mutation. Together with a single previous report, our data raises the possibility of an association between germline MUTYH mutations and CNS malignancies, particularly in pediatric histone H3‐mutant gliomas.