Next-generation Sequencing In Patients Research Articles

Germline targeted next-generation sequencing in patients with adrenal incidentalomas

Germline targeted next-generation sequencing in patients with adrenal incidentalomas

Mutational Profiling by Next-Generation Sequencing in Patients with Metastatic Non-Small Cell Lung Carcinoma: Our Experience

Introduction According to GLOBOCAN 2020, overall lung cancer is the second most common cancer in both sexes (11.4%) accounting for highest cancer-related mortality (18%).1 Adenocarcinoma subtype of non-small cell lung cancer (NSCLC) is the most common subtype and is divided into further molecular subtypes based on oncogenic driver mutations. Overall survival in these patients is poor with the use of conventional platinum-based double chemotherapy and various recent studies on targeted therapy studies have showed improved survival. Therefore, broad panel-based testing like next-generation sequencing (NGS) is strongly recommended to identify these targetable driver mutations. Aims and Objectives The aim of this study was to evaluate the mutational profile in patients with metastatic NSCLC (mNSCLC) by NGS method. Materials and Methods A hospital-based prospective observational study done on 88 patients under the Department of Medical Oncology, State Cancer Institute during a period of 1 year. All patients above 18 years of age diagnosed as mNSCLC having Eastern Cooperative Oncology Group performance status 0 to 2 and evaluated for mutational profiling by NGS method were included. Five gene panel tests including endothelial growth factor receptor (EGFR), echinoderm microtubule-associated protein-like 4 and anaplastic lymphoma kinase (EML4-ALK), BRAF, mesenchymal epithelial transition (MET), and ROS proto-oncogene 1 (ROS1) were used. Results and Observations Majority of mNSCLC cases were in the age group of 41 to 50 years (n = 30, 34.1%) with average age at presentation being 53.74 years. Male: female ratio was 1.14:1 and most patients were nonsmokers. Adenocarcinoma subtype of mNSCLC cases had the highest mutational burden (n = 55, 62.5%). EGFR (n = 32, 56.14%) was the most common mutation followed by EML4-ALK (n = 19, 33.33%). Most common EGFR mutation was in Exon 19. Other rare mutations were ROS1 (n = 4), BRAF V600E (n = 1), and MET (n = 1). Skeleton was the most common site of metastasis across all driver mutations. Conclusion EGFR and EML4-ALK were the commonest targetable mutations detected in the study. As there is very limited data from North Eastern region of India regarding mutational status in mNSCLC, this study opens up possibilities for further studies targeting multiple mutations to give us more comprehensive understanding of the mutational landscape of mNSCLC in this era of precision medicine.

Clinical practice recommendations for the use of next-generation sequencing in patients with solid cancer: a joint report from KSMO and KSP

Clinical practice recommendations for the use of next-generation sequencing in patients with solid cancer: a joint report from KSMO and KSP

Perspectives for next generation sequencing in patients with follicular lymphoma

Aim. To study the prognostic significance of gene mutations and intracellular signaling pathways involved in lymphomagenesis in patients with follicular lymphoma using next generation sequencing (NGS).Materials and methods. The prospective study included 26 patients with a median age of 51.5 years. Mutational screening was performed for cohort using custom NGS Panel of 118 genes. Gene set enrichment analysis (GSEA) was performed using Metascape. The data was analyzed in SPSS Statistics 26 and R 4.2.2.Results. The highest mutation frequency was noted in the genes: KMT2C – 50 %, KMT2D – 50 %, CREBBP – 31 %, NOTCH2 – 31 %, GNAS – 23 %. Missense mutations occurred with a frequency of 84.3 %. ARID1A gene mutation is an unfavorable prognostic factor according to progressive-free (p = 0.014) and event-free (p = 0.029) survival analysis. Tumor mutation burden (TMB) was defined as the number of mutations per megabase (Mb) of the coding sequence, the median TMB was 5.0 (3.3–8.3) mutations/Mb. The TMB threshold of 6 mutations/Mb divided patients into groups with high (44 %) and low (56 %) TMB. In the high TMB group, 2-year event-free survival was 27.3 % (95 % confidence interval 6.0–61.0), which was significantly lower than in low TMB group – 72.7 % (95 % confidence interval 41.9–91.6; p = 0.037). The most enriched cellular pathways according to GSEA results were regulation of cell activation (–log10(q-value) = 6.357), chromatin remodeling (–log10(q-value) = 5.707), histone modification (–log10(q-value) = 4.569). We have also demonstrated other possibilities of GSEA using follicular lymphoma as an example.Conclusion. TMB is a significant prognostic factor in patients with follicular lymphoma. We have shown that mutations in the MYC, CREBBP, EZH2, KMT2D genes lead to dysregulation in several intracellular processes, mediating complex molecular changes. The most enriched intracellular pathways in follicular lymphoma are those of chromatin remodeling, regulation of cell activation and histone modification.

Diagnostic Efficacy of Metagenomic Next-Generation Sequencing in Patients with Spinal Infections: A Retrospective Study

Diagnostic Efficacy of Metagenomic Next-Generation Sequencing in Patients with Spinal Infections: A Retrospective Study

Clinical Practice Recommendations for the Use of Next-Generation Sequencing in Patients with Solid Cancer: A Joint Report from KSMO and KSP.

In recent years, next-generation sequencing (NGS)-based genetic testing has become crucial in cancer care. While its primary objective is to identify actionable genetic alterations to guide treatment decisions, its scope has broadened to encompass aiding in pathological diagnosis and exploring resistance mechanisms. With the ongoing expansion in NGS application and reliance, a compelling necessity arises for expert consensus on its application in solid cancers. To address this demand, the forthcoming recommendations not only provide pragmatic guidance for the clinical use of NGS but also systematically classify actionable genes based on specific cancer types. Additionally, these recommendations will incorporate expert perspectives on crucial biomarkers, ensuring informed decisions regarding circulating tumor DNA panel testing.

First-Line Fixed-Duration Ibrutinib Plus Venetoclax (Ibr+Ven) Versus Chlorambucil Plus Obinutuzumab (Clb+O): 55-Month Follow-up from the Glow Study

Introduction : In the GLOW trial (NCT03462719), fixed-duration Ibr+Ven treatment has shown superiority to Clb+O in progression-free survival (PFS), better sustained undetectable minimal residual disease (uMRD) responses, and overall survival (OS) benefit in patients with previously untreated chronic lymphocytic leukemia (CLL) who are older and/or have comorbidities (CU Niemann, et al. Blood. 2022;140[suppl 1]:228-230). In this analysis, we report clinical outcomes, including PFS, OS, and MRD analysis, from the Ibr+Ven combination in GLOW with a median follow-up of 55 months, including subgroup analyses by IGHV and MRD. Methods : Patients aged ≥ 65 years or 18 to 64 years with a Cumulative Illness Rating Scale score > 6 or creatinine clearance < 70 mL/min were randomized 1:1 to Ibr+Ven (3 cycles of Ibr lead-in, followed by 12 cycles of Ibr+Ven; N = 106) or 6 cycles of Clb+O (N = 105). Each cycle was 28 days. Patients were excluded if they had del17p or known TP53 mutations at screening. End points included investigator-assessed PFS, uMRD rates, time to next treatment (TTNT), and OS. MRD was assessed sequentially over time in peripheral blood by next-generation sequencing for patients with partial response (PR) or better. Patients with < 1 CLL cell per 10,000 leukocytes (< 10 -4) were considered to have uMRD, whereas patients with ≥ 1 CLL cell per 10,000 leukocytes (≥ 10 -4) were considered to have detectable MRD (dMRD). All p values reported were nominal. Safety was not further assessed, as all patients were already past the treatment period in previous analyses. Results of updated analyses : With a median follow-up of 55 months, PFS remained superior for Ibr+Ven (hazard ratio [HR] 0.239 [95% confidence interval (CI), 0.159-0.359]; p < 0.0001); 54-month PFS rates were 65.8% for Ibr+Ven and 19.1% for Clb+O. In the Ibr+Ven arm, for patients with unmutated IGHV (uIGHV; n = 67) and mutated IGHV (mIGHV; n = 32), 54-month PFS rates were 58% and 90%, respectively. Also in the Ibr+Ven arm, PFS rates at 3 years post-treatment were 82% in patients who achieved uMRD at 3 months after end of treatment (EOT+3; n = 58) and 73% for patients with dMRD (n = 31). Among patients with uIGHV, PFS rates at 3 years post-treatment were 81% for those achieving uMRD at EOT+3 (n = 40) and 56% for those with dMRD (n = 16). In patients with mIGHV, PFS rates at 3 years post-treatment were ≥ 92% regardless of MRD status at EOT+3. Overall, at 38 months after end of treatment (EOT+38), 32.1% of patients had uMRD in the Ibr+Ven arm. Of the patients who achieved uMRD at EOT+3 (n = 58), 53.4% sustained uMRD status at EOT+38 in the Ibr+Ven arm. In addition, TTNT was prolonged for patients receiving Ibr+Ven versus Clb+O. The risk of needing second-line therapy was significantly reduced by 83% with first-line Ibr+Ven versus Clb+O (HR 0.174 [95% CI, 0.088-0.342]; p < 0.0001), and by 85% among patients with uIGHV (HR 0.146 [95% CI, 0.069-0.310]) compared with a 29% risk reduction in those with mIGHV (HR 0.708 [95% CI, 0.118-4.256]). In the Ibr+Ven arm, 4 patients received single-agent ibrutinib as subsequent therapy as part of the study; while 1 patient had not yet had a disease assessment, the best response for the other 3 patients were complete response (n = 1) and PR (n = 2). Finally, Ibr+Ven continues to demonstrate improved OS versus Clb+O, reducing the risk of death by 58% (HR 0.421 [95% CI, 0.237-0.747]; p = 0.0023). Estimated 54-month OS rates were 84.5% for the Ibr+Ven arm and 63.1% for the Clb+O arm. Conclusion : With prolonged follow-up of 55 months in the GLOW study, all-oral, once-daily, fixed-duration Ibr+Ven continues to show superior PFS versus Clb+O. Among patients treated with Ibr+Ven, benefit in PFS was particularly observed in patients with uIGHV who achieved uMRD at EOT+3 and in patients with mIGHV regardless of MRD status at EOT+3. Moreover, Ibr+Ven fixed-duration combination treatment continues to demonstrate an OS advantage versus chemoimmunotherapy in patients with previously untreated CLL.

T(11;14) status is stable between diagnosis and relapse, and concordant between detection methodologies based on fluorescence in situ hybridization and next-generation sequencing in patients with multiple myeloma.

Multiple myeloma (MM) is associated with a wide variety of recurrent genomic alterations. The most common translocation in MM is t(11;14). In this retrospective, single-center, non-interventional study, patient bone marrow samples were examined at diagnosis and at relapse(s) following treatment with anti-myeloma regimens to determine whether t(11;14) status was stable over time. This Stability Cohort consisted of 272 patients, of whom 118 were t(11;14)-positive at diagnosis and 154 were negative. All patients in the Stability Cohort retained the same t(11;14) status at relapse that they had at diagnosis of MM. Sixteen patients who had t(11;14)-positive MM at diagnosis had multiple longitudinal FISH assessments at relapse events, which remained t(11;14)-positive across all time points. Patients who had t(11;14)-positive disease at diagnosis of monoclonal gammopathy of unknown significance (MGUS) or smoldering multiple myeloma (SMM) also retained t(11;14) positivity through MM diagnosis and relapse. The t(11;14) fusion patterns also remained constant for 90% of patients. For detection of t(11;14), results from fluorescence in situ hybridization (FISH) and next generation sequencing (NGS) were compared to determine the rate of concordance between these 2 methods. This Concordance Cohort contained 130 patients, of whom 66 had t(11;14)-positive disease and 64 were t(11;14)-negative. In this sample set, the concordance between FISH and NGS-based detection of t(11;14) was 100%. These results strongly suggest that the t(11;14) rearrangement remains stable during the full disease course in patients with multiple myeloma and can be detected by FISH- and NGS-based methodologies.

Detecting circulating microbial cell-free DNA by next-generation sequencing in patients with Mycobacterium avium complex-lung disease: A pilot study.

Determining a diagnosis for non-Tuberculous mycobacterium (NTM)-lung disease (LD) remains difficult. The value of circulating cell-free DNA (cfDNA) secreted from microbes has been established in the detection of pathogens in septic patients. However, it is unknown whether NTM-derived cfDNA is detectable in plasma from patients with NTM-LD and whether this is associated with the disease status of NTM-LD, especially in patients with Mycobacterium avium complex (MAC)-LD. In this pilot study, from 2018 to 2019, we enrolled adult patients with MAC-LD at Taipei Veterans General Hospital in Taiwan for the detection of circulating cfDNA. We performed cfDNA extraction from plasma, next-generation sequencing (NGS) for nonhuman cfDNA, and sequence matching to a microbial database and then assessed the association between pathogen cfDNA and MAC-LD. Two (40%) plasma samples from MAC-LD patients had detectable MAC-specific cfDNA, namely one instance of DNA polymerase III alpha subunit and one instance of ATP-binding cassette transporters permease. The plasma samples from the three other MAC-LD cases and the one tuberculosis control were negative for either NTM-derived cfDNA or tuberculosis-related cfDNA. In addition to MAC-specific cfDNA, Ralstonia solanacearum, Staphylococcus aureus, and Pasteurella multocida were the most observed bacteria in our patients. The two patients with MAC-cfDNA positivity yielded higher radiographic scores (P = 0.076) and presented a higher number of nonhuman reads than those without MAC-cfDNA positivity (P = 0.083). Using NGS method, we demonstrated MAC-cfDNA was detectable in patients with MAC-LD. Further large-scale research is warranted to assess the clinical value of detecting MAC-specific cfDNA in MAC-LD patients.

Identification of Potential Genomic Alterations Using Pan-Cancer Cell-Free DNA Next-Generation Sequencing in Patients With Gastric Cancer.

Molecular cancer profiling may lead to appropriate trials for molecularly targeted therapies. Cell-free DNA (cfDNA) is a promising diagnostic and/or prognostic biomarker in gastric cancer (GC). We characterized somatic genomic alterations in cfDNA of patients with GC. Medical records and cfDNA data of 81 patients diagnosed as having GC were reviewed. Forty-nine and 32 patients were tested using the Oncomine Pan-Cancer Cell-Free Assay on the Ion Torrent platform and AlphaLiquid 100 kit on the Illumina platform, respectively. Tier I or II alterations were detected in 64.2% (52/81) of patients. Biomarkers for potential targeted therapy were detected in 55.6% of patients (45/81), and clinical trials are underway. ERBB2 amplification is actionable and was detected in 4.9% of patients (4/81). Among biomarkers showing potential for possible targeted therapy, TP53 mutation (38.3%, 35 variants in 31 patients, 31/81) and FGFR2 amplification (6.2%, 5/81) were detected the most. Next-generation sequencing of cfDNA is a promising technique for the molecular profiling of GC. Evidence suggests that cfDNA analysis can provide accurate and reliable information on somatic genomic alterations in patients with GC, potentially replacing tissue biopsy as a diagnostic and prognostic tool. Through cfDNA analysis for molecular profiling, it may be possible to translate the molecular classification into therapeutic targets and predictive biomarkers, leading to personalized treatment options for patients with GC in the future.

Efficacy of olaparib in advanced cancers with germline or somatic mutations in BRCA1, BRCA2, CHEK2 and ATM, a Belgian Precision tumor-agnostic phase II study

The Belgian Precision initiative aims to maximize the implementation of tumor-agnostic next-generation sequencing in patients with advanced cancer and enhance access to molecularly guided treatment options. Academic tumor-agnostic basket phase II studies are part of this initiative. The current investigator-driven trial aimed to investigate the efficacy of olaparib in advanced cancers with a (likely) pathogenic mutation (germline or somatic) in a gene that plays a role in homologous recombination (HR). This open-label, multi-cohort, phase II study examines the efficacy of olaparib in patients with an HR gene mutation in their tumor and disease progression on standard of care. Patients with a somatic or germline mutation in the same gene define a cohort. For each cohort, a Simon minimax two-stage design was used. If a response was observed in the first 13 patients, 14 additional patients were included. Here, we report the results on four completed cohorts: patients with a BRCA1, BRCA2, CHEK2 or ATM mutation. The overall objective response rate across different tumor types was 11% in the BRCA1-mutated (n= 27) and 21% in the BRCA2-mutated (n= 27) cohorts. Partial responses were seen in pancreatic cancer, gallbladder cancer, endocrine carcinoma of the pancreas and parathyroid cancer. One patient with a BRCA2 germline-mutated colon cancer has an ongoing complete response with 19+ months on treatment. Median progression-free survival in responding patients was 14+ months (5-34+ months). The clinical benefit rate was 63% in the BRCA1-mutated and 46% in the BRCA2-mutated cohorts. No clinical activity was observed in the ATM (n= 13) and CHEK2 (n= 14) cohorts. Olaparib showed efficacy in different cancer types harboring somatic or germline mutations in the BRCA1/2 genes but not in ATM and CHEK2. Patients with any cancer type harboring BRCA1/2 mutations should have access to olaparib.

Diagnostic value of plasma and blood cells metagenomic next-generation sequencing in patients with sepsis

BackgroundSeveral studies have investigated the detection of plasma cell-free DNA (cfDNA) using metagenomic next-generation sequencing (mNGS). However, to our knowledge, no study has evaluated the diagnostic value of mNGS detection using blood cells. In this study, we aimed to evaluate the performance of a whole blood mNGS assay which includes the results of plasma and blood cells mNGS detection. MethodsWe selected a panel of seven microorganisms to validate both the plasma and blood cells assay for their limits of detection (LoD), linearity, precision, and robustness to interference. In a multicentered prospective study conducted from January 2021 to April 2022, we tested 253 septic patients with plasma and blood cells mNGS and compared it with blood cultures (BCs). The performance of pathogen detection was compared between mNGS and BCs. ResultsThe LoD for plasma and blood cells mNGS was 8.3–140 genome equivalents (GE)/mL and 26 to 534 colony-forming units (CFU) or copies/mL, respectively. The inter- and intra-assay reproducibility of both plasma and blood cells mNGS was 100%. Compared to plasma mNGS alone, the sensitivity of whole blood mNGS was increased by 18.04% when using BCs as the standard (67.21% vs 85.25%). Furthermore, the sensitivity of whole blood mNGS in diagnosing bloodstream infections (BSIs) was 85.21%, which was significantly higher than that of BCs (36.09%, P＜0.0001) and plasma mNGS (69.82%; P = 0.0007). Additional analysis showed that blood cells mNGS was able to detect bacteria missed by plasma mNGS, while plasma mNGS was effective at detecting viruses. ConclusionsOur findings indicate that whole blood mNGS shows great potential as a promising diagnostic technique for BSIs owing to its ability to identify pathogens with higher sensitivity.

Mutation Profile via Next-Generation Sequencing in Patients with Colorectal Adenocarcinoma and Its Clinicopathological Correlation.

Recent studies that reveal the molecular profiles of colorectal carcinomas have demonstrated tumor heterogeneity. Characterization of colorectal carcinoma-specific genomic alterations is essential for developing more successful and targeted treat- ment protocols. Moreover, it is vital in elucidating the pathogenesis and mechanisms of resistance against treatment and predicting prognosis. The study included 73 cases diagnosed with colorectal carcinomas and subjected to molecular analysis by the next-generation sequencing. The association between the clinicopathologic parameters and pathogenic mutations detected in 32 genes was evaluated. Pathogenic mutations were determined in a total of 24 genes. The Cell Division Cycle 27 (CDC27), Kirsten rat sarcoma viral proto-oncogene (KRAS), serine/threonine protein kinase B-raf (BRAF), phosphatase and tensin homolog, breast cancer 2 (BRCA2), and phosphotidylinositol-4,5-biphosphate 3-kinase (PIK3CA) mutations were determined at higher rates, with the adenomatous polypo- sis coli mutation determined at a lower rate than in the literature. There were significant positive correlations between CDC27 and phosphatase and tensin homolog (PTEN), PTEN and BRCA2, and PTEN and adenomatous polyposis coli (APC) concomitant muta- tions, whereas negative correlations were present between BRAF and KRAS. Statistically significant relationships were present between KRAS exon 2 and mucinous morphology, PIK3CA and absence of perineural invasion, BRAF and tumor differentiation/localization, MutS homolog 3 (MSH3) and tumor diameter, and BRCA2 and absence of lymph node metastasis. It is necessary to have a comprehensive database of genomic alterations of colorectal carcinomas to interpret mutations more accurately clinically. There are no studies on the frequency of mutations in colorectal carcinomas in the Turkish population; thus, follow-up and treatment protocols are organized following the European and American databases and guidelines. A comprehensive study of the colorectal carcinoma patients' mutation profile in the Turkish patient cohort by the next-generation sequencing method will help to provide significant therapeutic, prognostic, and predictive data and design more successful treatment and follow-up strategies.

Comparison of microbial detection rates in microbial culture methods versus next-generation sequencing in patients with prosthetic joint infection: a systematic review and meta-analysis

BackgroundAccurate diagnosis of prosthetic joint infection (PJI) enables early and effective treatment. However, there is currently no gold standard test for microbial detection of PJI and traditional synovial fluid culture is relatively insensitive. Recently, it has been reported that sonicating fluid culture and next-generation sequencing (NGS) improve microbial detection rates. Hence, we performed a systematic review and meta-analysis to compare microbial detection rates in microbial culture methods with and without sonication versus NGS.MethodsWe systematically searched EMBASE, PubMed, Scopus, CINAHL, and Ichushi databases and other sources (previous reviews) until August 2022. We evaluated the detection rates of pathogens in NGS and microbial cultures using samples of synovial or sonicated fluid.ResultsOf the 170 citations identified for screening, nine studies were included. Pooled analysis indicated that NGS had the highest detection rate among the microbial detection methods (NGS vs. sonicated, odds ratios [OR] 5.09, 95% confidential interval [CI] 1.67–15.50; NGS vs. synovial, OR 4.52, 95% CI 2.86–7.16). Sonicated fluid culture showed a higher detection rate than synovial fluid culture (OR 2.11, 95% CI 1.23–3.62).ConclusionNGS might be useful as a screening tool for culture-negative patients. In clinical settings, sonicated fluid culture is a practical method for diagnosing PJI.

Clinical Value of Sampling Time of Metagenomic Next-Generation Sequencing in Patients with Severe Pneumonia.

Severe pneumonia is a common infectious disease with high morbidity and mortality. Early etiological diagnosis is crucial for improving the prognosis. The aim of this study is to evaluate the clinical value ofsamplingtime of mNGS in patients with severe pneumonia. This retrospective study enrolled 105 patients with severe pneumonia. mNGS was performed on bronchoalveolar lavage fluid (BALF). Patients were divided into the sampling time ≤ 72h vs sampling time >72h groups and survivors vs non-survivors groups according to their sampling time and prognosis. Clinical characteristics, the adjustment of antibiotics and clinical prognostic value were evaluated. Our study showed that, early sampling of mNGS can significantly shorten the mechanical ventilation time (p = 0.007) and hospitalization time (p = 0.004). In the non-survivors group, CURB-65, SOFA, and APACHE II scores were higher. Age (OR: 1.051, 95% CI: 1.004-1.100, p = 0.034), chronic respiratory diseases (OR: 4.639, 95% CI: 1.260-17.082, p = 0.021), immunosuppression (OR: 5.008, 95% CI: 1.617-15.510, p = 0.005) and SOFA score on the day of mNGS sampling (OR: 1.492, 95% CI: 1.212-1.837, p < 0.001) were independent risk factors of in-hospital mortality. The most common pathogens were Klebsiella pneumoniae and Human gammaherpesvirus 4. The proportion of appropriate and targeted antibiotics adjusted was significantly higher than that in the sampling time > 72h group, and the proportion of antifungal and antiviral agents adjusted was lower. In the early sampling group, it was significantly decreased in the CRP, PCT level and NEU% at discharge. This study demonstrated that early sampling of mNGS could shorten the time of mechanical ventilation and hospitalization of patients with severe pneumonia. Patients with higher SOFA score on the day of sampling had a poorer prognosis. It emphasizes that early sampling of mNGS has a positive value.

Comparative analysis of genomic profiles between tissue-based and plasma-based next-generation sequencing in patients with non-small cell lung cancer

ObjectivesGenotype-guided personalized therapy has become an essential part of routine clinical care in non-small cell lung cancer (NSCLC) patients. However, small tissue specimens often yield inadequate molecular testing material. Plasma ctDNA-based liquid biopsy is an increasingly common non-invasive alternative to tissue biopsy. This study examined the similarities and differences in the molecular profiling of tissue and plasma samples to provide insight into sample selection in clinical practice. Materials and MethodsSequencing data from 190 NSCLC patients who underwent concurrent tissue-based next-generation sequencing (tissue-NGS) and plasma-based NGS (plasma-NGS) using a 168-gene panel were analyzed. ResultsTissue-NGS identified genomic alterations in 97.4% (185/190) of the enrolled patients and plasma-NGS identified genomic alterations in 72.1% (137/190) of the enrolled patients. Considering all NSCLC guideline-recommended biomarkers in the entire cohort of 190 cases, 81 patients had positive concordant mutations detected in both tissue and plasma samples, while 69 patients had no predefined alterations detected in either tissue or plasma samples. Additional mutations were found in the tissues of 34 patients and the plasma of six patients. The overall concordance rate between tissue and plasma samples was 78.9% (150/190). The tissue-NGS and plasma-NGS sensitivities were 95.0% and 71.9%, respectively. In the 137 patients with detectable ctDNA in plasma samples, the concordance rate between tissue and plasma samples reached 91.2%, and the sensitivity of plasma-NGS reached 93.5%. ConclusionOur findings indicate that plasma-NGS is less capable of detecting genetic alterations than tissue-NGS, especially for copy number variations and gene fusions. Tissue-NGS remains the preferred method for evaluating the molecular profile of NSCLC patients when tumor tissue is available. We suggest that the concurrent use of liquid biopsy and tissue biopsy is the optimal approach in clinical practice; alternatively, plasma can be used as substitute material when tissue is unavailable.

The microbiological diagnostic performance of metagenomic next-generation sequencing in patients with infectious diseases.

Metagenomic next-generation sequencing (mNGS) can be used to detect pathogens in clinical infectious diseases through the sequencing analysis of microbial and host nucleic acids in clinical samples. This study aimed to assess the diagnostic performance of mNGS in patients with infections. In this study, 641 patients with infectious diseases were enrolled. These patients simultaneously underwent pathogen detection by both mNGS and microbial culture. Through statistical analysis, we judged the diagnostic performance of mNGS and microbial culture on different pathogens. Among 641 patients, 276 cases of bacteria and 95 cases of fungi were detected by mNGS, whereas 108 cases of bacteria and 41 cases of fungi were detected by traditional cultures. Among all mixed infections, combined bacterial and viral infections were the highest (51%, 87/169), followed by combined bacterial with fungal infections (16.57%, 28/169) and mixed bacterial, fungal, and viral infections (13.61%, 23/169). Among all sample types, bronchoalveolar lavage fluid (BALF) samples had the highest positive rate (87.8%, 144/164), followed by sputum (85.4%, 76/89) and blood samples (61.2%, 158/258). For the culture method, sputum samples had the highest positive rate (47.2%, 42/89), followed by BALF (37.2%, 61/164). The positive rate of mNGS was 69.89% (448/641), which was significantly higher than that of traditional cultures (22.31% [143/641]) (P < .05). Our results show that mNGS is an effective tool for the rapid diagnosis of infectious diseases. Compared with traditional detection methods, mNGS also showed obvious advantages in mixed infections and infections with uncommon pathogens.

Assessing the Diagnostic Accuracy of Next-Generation Sequencing in Patients With Antibiotic Spacers Before Reimplantation.

Use of molecular sequencing modalities in periprosthetic joint infection diagnosis and organism identification has gained popularity recently. To date, there is no diagnostic test that reliably predicts infection eradication in patients with antibiotic spacers. The purpose of this study was to compare the diagnostic accuracy of next-generation sequencing (NGS), culture, the Musculoskeletal Infection Society (MSIS) criteria, and the criteria by Parvizi et al in patients with antibiotic spacers. In this retrospective study, aspirate or tissue samples were collected from 38 knee and 19 hip antibiotic spacers for routine diagnostic workup for the presence of persistent infection and sent to the laboratory for NGS. The kappa statistic along with statistical differences between diagnostic studies were calculated using the chi-square test for categorical data. The kappa coefficient for agreement between NGS and culture was 0.27 (fair agreement). The percentages of positive and negative agreement were 22.8% and 42.1%, respectively, with a total concordance of 64.9%. There were 12 samples that were culture positive and NGS negative. Eight samples were NGS positive but culture negative. The kappa coefficient was 0.42 (moderate agreement) when comparing NGS with MSIS criteria. In our series, NGS did not provide sufficient agreement compared with culture or MSIS criteria in the setting of an antibiotic spacer. A reliable diagnostic indicator for reimplantation has yet to be identified. [Orthopedics. 2024;47(1);46-51.].

The role of the genomic mutation signature and tumor mutation burden on relapse risk prediction in head and neck squamous cell carcinoma after concurrent chemoradiotherapy

Personalized genetic profiling has focused on improving treatment efficacy and predicting risk stratification by identifying mutated genes and selecting targeted agents according to genetic testing. Therefore, we evaluated the role of genetic profiling and tumor mutation burden (TMB) using next-generation sequencing in patients with head and neck squamous cell carcinoma (HNSC). The relapse mutation signature (RMS) and chromatin remodeling mutation signature (CRMS) were explored to predict the risk of relapse in patients with HNSC treated with concurrent chemoradiotherapy (CCRT) with platinum-based chemotherapy. Patients in the high RMS and CRMS groups showed significantly shorter relapse-free survival than those in the low RMS and CRMS groups, respectively (p < 0.001 and p = 0.006). Multivariate Cox regression analysis showed that extranodal extension, CCRT response, and three somatic mutation profiles (TMB, RMS, and CRMS) were independent risk predictors for HNSC relapse. The predictive nomogram showed satisfactory performance in predicting relapse-free survival in patients with HNSC treated with CCRT.

Clinical values of metagenomic next-generation sequencing in patients with severe pneumonia: a systematic review and meta-analysis.

Clinical values of metagenomic next-generation sequencing (mNGS) in patients with severe pneumonia remain controversial. Therefore, we conduct this meta-analysis to evaluate the diagnostic performance of mNGS for pathogen detection and its role in the prognosis of severe pneumonia. We systematically searched the literature published in PubMed, Embase, Cochrane Library, Web of Science, Clinical Trials.gov, CNKI, Wanfang Data, and CBM from the inception to the 28th September 2022. Relevant trials comparing mNGS with conventional methods applied to patients with severe pneumonia were included. The primary outcomes of this study were the pathogen-positive rate, the 28-day mortality, and the 90-day mortality; secondary outcomes included the duration of mechanical ventilation, the length of hospital stay, and the length of stay in the ICU. Totally, 24 publications with 3220 patients met the inclusion criteria and were enrolled in this study. Compared with conventional methods (45.78%, 705/1540), mNGS (80.48%, 1233/1532) significantly increased the positive rate of pathogen detection [OR = 6.81, 95% CI (4.59, 10.11, P < 0.001]. The pooled 28-day and 90-day mortality in mNGS group were 15.08% (38/252) and 22.36% (36/161), respectively, which were significantly lower than those in conventional methods group 33.05% (117/354) [OR = 0.35, 95% CI (0.23, 0.55), P < 0.001, I2 = 0%] and 43.43%(109/251) [OR = 0.34, 95% CI (0.21, 0.54), P < 0.001]. Meanwhile, adjusted treatment based on the results of mNGS shortened the length of hospital stay [MD = -2.76, 95% CI (- 3.56, - 1.96), P < 0.001] and the length of stay in ICU [MD = -4.11, 95% CI (- 5.35, - 2.87), P < 0.001]. The pathogen detection positive rate of mNGS was much higher than that of conventional methods. Adjusted treatment based on mNGS results can reduce the 28-day and 90-day mortality of patients with severe pneumonia, and shorten the length of hospital and ICU stay. Therefore, mNGS advised to be applied to severe pneumonia patients as early as possible in addition to conventional methods to improve the prognosis and reduce the length of hospital stay.