Anti-phosphatidyl-serine/prothrombin (aPS/PT) antibodies are superior predictors of LAC presence and APS diagnoses: A single center study.

BACKGROUND AND AIMS: Several non-criteria (NC) anti-phospholipid antibodies (APLA) have been proposed as candidates for antiphospholipid antibody syndrome (APS) diagnosis. The objectives of this study were 1) to determine the association of five different NC-APLA with positivity for Lupus anti-coagulant (LAC) and the criteria antibodies anti-cardiolipin (aCL) and anti-beta glycoprotein (aB2GPI), and 2) to assess the ability of NC-APLA to predict LAC presence and clinical APS diagnoses. MATERIAL AND METHODS: Results from 486 patients tested for LAC and APLA were retrieved. Patients were grouped according to LAC and serology positivity into three groups: Single-positives (SP) for LAC, aCL or aB2GPI; Double-positives for aCL and aB2GPI; Triple-positives (TP) for LAC, aCL and aB2GPI. NC-ALPA titers were compared between LAC-positive and negative and APS and non-APS patients. RESULTS: Forty-two of 486 patients were LAC-positive and 28 were diagnosed with APS. All criteria and NC-APLA titers were significantly higher in TP than SP patients. ROC analyses based on LAC status showed highest area under the curve (AUC, 95% CI) for aPS/PT IgG (0.75, 0.65-0.85) and aPS/PT IgM (0.73, 0.63-0.82). Based on APS diagnosis, aPS/PT IgM achieved highest AUC (0.87; 0.79-0.95). CONCLUSION: Anti-phosphatidyl-serine/prothrombin (aPS/PT) antibodies are superior predictors of LAC presence and APS diagnoses.

Performance of Anti-Carbamylated Protein Antibody Testing in the Routine Evaluation of Rheumatoid Arthritis from a Single Center.

BACKGROUND: Detection of anticyclic citrullinated peptide antibodies (anti-CCP) and rheumatoid factors (RF) in sera support the diagnosis of rheumatoid arthritis (RA); however, these markers are not detected in about 20% of RA patients. More recently, antibodies against carbamylated proteins (anti-CarP) have emerged with implications for preclinical RA diagnosis. The objective of this study was to assess the clinical performance of anti-CarP and correlate with disease severity in routine clinical practice. METHODS: Retrospective chart review of 331 subjects submitted for RA panel serology: 136 clinically defined RA-positive and 195 RA-negative patients. Fifty additional individuals were recruited for healthy controls. Patients' sera were tested for anti-CCP, anti-CarP, and RF antibodies. Clinical performance characteristics were evaluated for anti-CarP individually and in combination with anti-CCP and RF. Documented erosions and synovitis were correlated with anti-CarP positivity. RESULTS: Anti-CarP had a clinical sensitivity and specificity of 27% and 94%, respectively, for established RA. This sensitivity was lower than anti-CCP (79%) and RF (85%). The specificity of anti-CarP was similar to anti-CCP (93%) and higher than RF (69%). Anti-CarP in combination with anti-CCP and RF increased specificity (100%) but decreased sensitivity (21%). There was no correlation of anti-CarP positivity with presence of bone erosions; however, there was an increase in anti-CarP positivity among patients with synovitis. CONCLUSIONS: Anti-CarP demonstrates high specificity in diagnosis of established RA but lacks clinical sensitivity. In combination, anti-CarP does not improve clinical performance of anti-CCP and RF but may be useful in anti-CCP negative patients and in identifying patients with more active disease.

Performance Assessment of a Novel Multianalyte Methodology for Celiac Disease Biomarker Detection and Evaluation of the Serology-Alone Criteria for Biopsy-Free Diagnosis.

CONTEXT.­: Serology plays a vital role in celiac disease (CD) diagnosis, and the latest European guidelines advocate for biopsy-free diagnoses in patients with ≥10× the upper limit of normal (ULN) of anti-tissue transglutaminase (tTG) immunoglobulin A (IgA) antibodies. OBJECTIVE.­: To assess performance characteristics of a novel automated particle-based multianalyte technology (Aptiva) for anti-tTG and anti-deamidated gliadin peptide (DGP) antibody detection as compared to the traditional enzyme-linked immunosorbent assay (QUANTA Lite). Performance characteristics of the ≥10× ULN anti-tTG IgA criteria for serologic diagnosis of CD were also evaluated. DESIGN.­: Sera samples from 703 patients were tested for anti-tTG IgA, anti-tTG immunoglobulin G (IgG), anti-DGP IgA, and anti-DGP IgG antibodies on both platforms. In total, 127 patients had medical information and were classified as CD-positive (n = 58) and CD-negative (n = 69) based on biopsy results. Clinical performance characteristics were evaluated. RESULTS.­: Anti-tTG IgA detection showed equal clinical sensitivity and specificity of 91% sensitivity and 99% specificity on both platforms. Anti-tTG IgG resulted in moderate sensitivity of 69% and 72%, but high specificity of 100% and 94% on Aptiva and QUANTA Lite, respectively. Anti-DGP IgG displayed comparable sensitivity of 90% and 81%, and a specificity of 94% and 99%, on Aptiva and QUANTA Lite, respectively. Anti-DGP IgA demonstrated greater sensitivity on QUANTA Lite (83%) than Aptiva (69%) and similar specificities of 97% and 98% on QUANTA Lite and Aptiva, respectively. At ≥10× ULN levels for anti-tTG IgA, Aptiva displayed a sensitivity of 72% and a specificity of 100%, and QUANTA Lite showed a sensitivity of 69% and a specificity of 100%. CONCLUSIONS.­: Aptiva is a reliable method to measure CD biomarkers with reduced hands-on necessity and high-throughput capabilities. This study supports the use of a ≥10× ULN anti-tTG IgA biopsy-free approach for serologic diagnosis of CD.

Presence of anti-gp210 or anti-sp100 antibodies in AMA-positive patients may help support a diagnosis of primary biliary cholangitis.

BACKGROUND: Anti-mitochondrial antibody (AMA) positivity is not always associated with primary biliary cholangitis (PBC). We aimed to determine the additional value of anti-sp100 or anti-gp210 antibody in AMA-positive patients for PBC. METHODS: Patients (n = 190) and healthy donors (n = 50) were evaluated for AMA, anti-gp210 and anti-sp100 antibodies by ELISA. Antibody frequencies in cohorts and performance characteristics in some patients categorized as 'definitive-', 'probable-', and 'no PBC' were determined following review of their charts. RESULTS: Of the patients (n = 190), 38.4% were AMA-positive (n = 73) and 61.6% AMA-negative (n = 117). Frequency of anti-sp100 or anti-gp210 antibody was 17.8%, 2.6%, and 0% in AMA-positive, AMA-negative and healthy controls, respectively. Clinical data was available for 63 of 73 AMA-positive patients with 28.6%, 22.2%, and 49.2% categorized as definite, probable, and no PBC, respectively. Patients with definite PBC had higher mean levels of AMA and frequencies of sp100 or gp210 antibody compared to other groups. Sensitivities were low (anti-sp100: 18.8% and anti-gp210: 16.7%) with specificities above 98.0% for both. CONCLUSION: AMA-positive patients positive for anti-sp100 or anti-gp210 antibody were more likely to have a diagnosis of definite or probable PBC than those with AMA alone. Use of all tests is likely to improve characterization of patients at-risk for PBC.

Evaluation of a Novel Multiplex Platform for Simultaneous Detection of IgG Antibodies Against the 4 Main SARS-CoV-2 Antigens.

BACKGROUND: Numerous serology assays are available for detection of SARS-CoV-2 antibodies but are limited in that only 1 or 2 target antigen(s) can be tested at a time. Here, we describe a novel multiplex assay that simultaneously detects and quantifies IgG antibodies to SARS-CoV-2 antigens, spike (S), nucleocapsid (N), receptor-binding domain (RBD), and N-terminal domain (NTD) in a single well. METHODS: Sensitivity was determined using samples (n = 124) from confirmed SARS-CoV-2 RT-PCR positive individuals. Prepandemic (n = 100) and non-COVID respiratory infection positive samples (n = 100) were used to evaluate specificity. Samples were analyzed using COVID-19 IgG multiplex serology assay from Meso Scale Discovery (MSD) and using commercial platforms from Abbott, EUROIMMUN, and Siemens. RESULTS: At >14 days post-PCR, MSD assay displayed >98.0% sensitivity [S 100% (95% CI 98.0%-100.0%); N 98.0% (95% CI 97.2%-98.9%); RBD 94.1% (95% CI 92.6%-95.6%); NTD 98.0% (95% CI, 97.2%-98.9%)] and 99% specificity (95% CI 99.3%-99.7%) for antibodies to all 4 antigens. Parallel assessment of antibodies to more than 1 antigen improved the sensitivity to 100% (95% CI 98.0%-100.0%) while maintaining 98% (95% CI 97.6%-98.4%) specificity regardless of the combinations used. When AU/mL concentrations of IgG antibodies from the MSD assay were compared against the corresponding IgG signals acquired from the single target commercial assays, the following correlations were observed: Abbott (vs MSD N, R2 = 0.73), Siemens (vs MSD RBD, R2 = 0.92), and EUROIMMUN (vs MSD S, R2 = 0.82). CONCLUSION: MSD assay offers an accurate and a comprehensive assessment of SARS-CoV-2 antibodies with higher sensitivity and equivalent specificity compared to the commercial IgG serology assays.

Effect of pH on the Quantification of Common Chemistry Analytes in Body Fluid Specimens Using the Roche cobas Analyzer for Clinical Diagnostic Testing.

OBJECTIVES: To determine the influence of pH on recovery of analytes in body fluids (BFs), investigate the mechanism of pH interference, measure the frequency of abnormal-pH BFs received, and compare pH measured by meter and paper. METHODS: We performed pH titration in residual BFs. A low-pH BF was spiked and neutralized to investigate pH interference. We measured analytes on a Roche cobas c501 analyzer (Roche Diagnostics) and calculated the percent recovery. Measurement of pH using a meter and paper was conducted on 122 BF samples received in the laboratory. RESULTS: Enzyme activity in BFs was unaffected when pH = 7.4-8.5 lactate dehydrogenase, pH = 7.3-10.2 amylase, pH = 6.0-9.9 lipase, and pH = 1.3-11.7 all other analytes. BFs had mean (range) pH of 8.0 (5.1-8.9), with a mean (range) difference (paper ‒ meter) of ‒0.4 (‒0.6 to 1.1). CONCLUSIONS: Irreversible loss of enzyme activity occurs in BFs at low pH. Few clinical BFs have pH < 7.0, but laboratories should incorporate pH measurement in BF workflows.

Evaluation of a Surrogate Enzyme-Linked Immunosorbent Assay-Based Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) cPass Neutralization Antibody Detection Assay and Correlation With Immunoglobulin G Commercial Serology Assays.

CONTEXT.­: Emerging evidence shows correlation between the presence of neutralization antibodies (nAbs) and protective immunity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Currently available commercial serology assays lack the ability to specifically identify nAbs. An enzyme-linked immunosorbent assay-based nAb assay (GenScript cPass neutralization antibody assay) has recently received emergency use authorization from the Food and Drug Administration. OBJECTIVE.­: To evaluate the performance characteristics of this assay and compare and correlate it with the commercial assays that detect SARS-CoV-2-specific immunoglobulin G (IgG). DESIGN.­: Specimens from SARS-COV-2 infected patients (n = 124), healthy donors obtained prepandemic (n = 100), and patients with non-coronavirus disease 2019 (COVID-19) respiratory infections (n = 92) were analyzed using this assay. Samples with residual volume were also tested on 3 commercial serology platforms (Abbott, Euroimmun, Siemens). Twenty-eight randomly selected specimens from patients with COVID-19 and 10 healthy controls were subjected to a plaque reduction neutralization test. RESULTS.­: The cPass assay exhibited 96.1% (95% CI, 94.9%-97.3%) sensitivity (at >14 days post-positive PCR), 100% (95% CI, 98.0%-100.0%) specificity, and zero cross-reactivity for the presence of non-COVID-19 respiratory infections. When compared with the plaque reduction assay, 97.4% (95% CI, 96.2%-98.5%) qualitative agreement and a positive correlation (R2 = 0.76) was observed. Comparison of IgG signals from each of the commercial assays with the nAb results from plaque reduction neutralization test/cPass assays displayed greater than 94.7% qualitative agreement and correlations with R2 = 0.43/0.68 (Abbott), R2 = 0.57/0.85 (Euroimmun), and R2 = 0.39/0.63 (Siemens), respectively. CONCLUSIONS.­: The combined data support the use of cPass assay for accurate detection of the nAb response. Positive IgG results from commercial assays associated reasonably with nAbs presence and can serve as a substitute.

Evaluation of Phi Clinical Performance for the Detection of Prostate Cancer in Routine Clinical Practice.

Prostate Health Index (phi) and percent free PSA (%fPSA) are used in patients with a PSA concentration between 4-10 µg/L as an aid to distinguish prostate cancer (PCa) from benign conditions, and to assist in the decision of whether to proceed to biopsy. This study assesses the clinical performance and diagnostic accuracy of phi versus %fPSA in a cohort of Mayo Clinic's patients. Of 4065 phi orders received from May 2017-August 2018, concordance between phi and %fPSA results was evaluated on 2845 results with a total PSA within 4-10 µg/L. Retrospective chart review was performed on 201 Mayo Clinic patients, and %fPSA and phi results were compared with both the decision to biopsy and presence of PCa at biopsy. Receiver operating characteristic (ROC) curve analysis to evaluate the diagnostic accuracy of PSA, %fPSA and phi was performed. In this study 2.5% of the 2845 orders exhibited discordant PCa risk classifications between %fPSA and phi results. In the phi high risk category, 41.7% (versus 26.1% by %fPSA) of patients had biopsy and 100% (versus 66.6% by %fPSA) were positive for PCa. Phi exhibited the highest specificity and ROC area under the curve compared to %fPSA and PSA. phi was a better predictor than %fPSA for finding PCa at biopsy. These findings support continued utilization of phi in the evaluation of patients with a PSA in the 4-10 µg/L range.

A critical role for CARD9 in pneumocystis pneumonia host defence.

Caspase recruitment domains-containing protein 9 (CARD9) is an adaptor molecule critical for key signalling pathways initiated through C-type lectin receptors (CLRs). Previous studies demonstrated that Pneumocystis organisms are recognised through a variety of CLRs. However, the role of the downstream CARD9 adaptor signalling protein in host defence against Pneumocystis infection remains to be elucidated. Herein, we analysed the role of CARD9 in host defence against Pneumocystis both in CD4-depleted CARD9-/- and immunocompetent hosts. Card9 gene-disrupted (CARD9-/- ) mice were more susceptible to Pneumocystis, as evidenced by reduced fungal clearance in infected lungs compared to wild-type (WT) infected mice. Our data suggests that this defect was due to impaired proinflammatory responses. Furthermore, CARD9-/- macrophages were severely compromised in their ability to differentiate and express M1 and M2 macrophage polarisation markers, to enhanced mRNA expression for Dectin-1 and Mincle, and most importantly, to kill Pneumocystis in vitro. Remarkably, compared to WT mice, and despite markedly increased organism burdens, CARD9-/- animals did not exhibit worsened survival during pneumocystis pneumonia (PCP), perhaps related to decreased lung injury due to altered influx of inflammatory cells and decreased levels of proinflammatory cytokines in response to the organism. Finally, although innate phase cytokines were impaired in the CARD9-/- animals during PCP, T-helper cell cytokines were normal in immunocompetent CARD9-/- animals infected with Pneumocystis. Taken together, our data demonstrate that CARD9 has a critical function in innate immune responses against Pneumocystis.

Evaluation of plasma ACTH stability using the Roche Elecsys immunoassay.

BACKGROUND: Adrenocorticotropic hormone (ACTH) has been reported to be labile in blood due to proteolytic degradation and stringent procedures are followed to prevent in vitro degradation after sample collection. OBJECTIVE: The purpose of this study was to examine the effect of time and temperature before and after separation of plasma from cells in the quantitation of plasma ACTH. METHODS: Our current protocol includes sample collection in a pre-chilled tube, transport on ice and immediate centrifugation at 4 °C. These reference conditions were compared against sample processing in tubes and centrifuge set at room-temperature; using delayed centrifugation at 4 °C. ACTH stability was evaluated at ambient and refrigerated temperatures after collection and plasma separation using the reference protocol. Plasma samples were analyzed using the Roche Elecsys ACTH immunoassay. RESULTS: Quantification of ACTH was not impacted by the use of non-chilled tubes and centrifuge and up to a 4 h delay in separation of plasma from cells. Average percent differences in plasma ACTH concentration from time 0 was <10% up to 12 h at ambient temperature. Refrigeration of plasma did not preserve ACTH stability at 12 h and longer storage resulted in significant ACTH degradation at both ambient and refrigerated temperatures. CONCLUSIONS: As supported by these data, previously recommended strict specimen collection and processing requirements are not necessary for measuring ACTH with the Roche Elecsys immunoassay.

The Interaction of Pneumocystis with the C-Type Lectin Receptor Mincle Exerts a Significant Role in Host Defense against Infection.

Pneumocystis pneumonia (PCP) remains a major cause of morbidity and mortality within immunocompromised patients. In this study, we examined the potential role of macrophage-inducible C-type lectin (Mincle) for host defense against Pneumocystis Binding assays implementing soluble Mincle carbohydrate recognition domain fusion proteins demonstrated binding to intact Pneumocystis carinii as well as to organism homogenates, and they purified major surface glycoprotein/glycoprotein A derived from the organism. Additional experiments showed that rats with PCP expressed increased Mincle mRNA levels. Mouse macrophages overexpressing Mincle displayed increased binding to P. carinii life forms and enhanced protein tyrosine phosphorylation. The binding of P. carinii to Mincle resulted in activation of FcRγ-mediated cell signaling. RNA silencing of Mincle in mouse macrophages resulted in decreased activation of Syk kinase after P. carinii challenge, critical in downstream inflammatory signaling. Mincle-deficient CD4-depleted (Mincle-/-) mice showed a significant defect in organism clearance from the lungs with higher organism burdens and altered lung cytokine responses during Pneumocystis murina pneumonia. Interestingly, Mincle-/- mice did not demonstrate worsened survival during PCP compared with wild-type mice, despite the markedly increased organism burdens. This may be related to increased expression of anti-inflammatory factors such as IL-1Ra during infection in the Mincle-/- mice. Of note, the P. murina-infected Mincle-/- mice demonstrated increased expression of known C-type lectin receptors Dectin-1, Dectin-2, and MCL compared with infected wild-type mice. Taken together, these data support a significant role for Mincle in Pneumocystis modulating host defense during infection.

Differential Macrophage Polarization from Pneumocystis in Immunocompetent and Immunosuppressed Hosts: Potential Adjunctive Therapy during Pneumonia.

We explored differential polarization of macrophages during infection using a rat model of Pneumocystis pneumonia. We observed enhanced pulmonary M1 macrophage polarization in immunosuppressed (IS) hosts, but an M2 predominant response in immunocompetent (IC) hosts following Pneumocystis carinii challenge. Increased inflammation and inducible nitric oxide synthase (iNOS) levels characterized the M1 response. However, macrophage ability to produce nitric oxide was defective. In contrast, the lungs of IC animals revealed a prominent M2 gene signature, and these macrophages effectively elicited an oxidative burst associated with clearance of Pneumocystis In addition, during P. carinii infection the expression of Dectin-1, a critical receptor for recognition and clearance of P. carinii, was upregulated in macrophages of IC animals but suppressed in IS animals. In the absence of an appropriate cytokine milieu for M2 differentiation, Pneumocystis induced an M1 response both in vitro and in vivo The M1 response induced by P. carinii was plastic in nature and reversible with appropriate cytokine stimuli. Finally, we tested whether macrophage polarization can be modulated in vivo and used to help manage the pathogenesis of Pneumocystis pneumonia by adoptive transfer. Treatment with both M1 and M2 cells significantly improved survival of P. carinii-infected IS hosts. However, M2 treatment provided the best outcomes with efficient clearance of P. carinii and reduced inflammation.

Identification of Genetic Mutations in Human Lung Cancer by Targeted Sequencing.

Lung cancer remains the most prevalent malignancy and the primary cause of cancer-related deaths worldwide. Unique mutations patterns can be found in lung cancer subtypes, in individual cancers, or within a single tumor, and drugs that target these genetic mutations and signal transduction pathways are often beneficial to patients. In this study, we used the Ion Torrent AmpliSeq Cancer Panel to sequence 737 loci from 45 cancer-related genes and oncogenes to identify genetic mutations in 48 formalin-fixed, paraffin-embedded (FFPE) human lung cancer samples from Chinese patients. We found frequent mutations in EGFR, KRAS, PIK3CA, and TP53 genes. Moreover, we observed that a portion of the lung cancer samples harbored two or more mutations in these key genes. This study demonstrates the feasibility of using the Ion Torrent sequencing to efficiently identify genetic mutations in individual tumors for targeted lung cancer therapy.

Epigenetic control of dendritic cell development and fate determination of common myeloid progenitor by Mysm1.

The mechanisms controlling the development of dendritic cells (DCs) remain incompletely understood. Using an Mysm1 knockout (Mysm1(-/-)) mouse model, we identified the histone H2A deubiquitinase Mysm1, as a critical regulator in DC differentiation. Mysm1(-/-) mice showed a global reduction of DCs in lymphoid organs, whereas development of granulocytes and macrophages were not severely affected. Hematopoietic progenitors and DC precursors were significantly decreased in Mysm1(-/-) mice and defective in Fms-like tyrosine kinase-3(Flt3) ligand-induced, but not in granulocyte macrophage-colony-stimulating factor (GM-CSF)-induced DC differentiation in vitro. Molecular studies demonstrated that the developmental defect of DCs from common myeloid progenitor (CMP) in Mysm1(-/-) mice is associated with decreased Flt3 expression and that Mysm1 derepresses transcription of the Flt3 gene by directing histone modifications at the Flt3 promoter region. Two molecular mechanisms were found to be responsible for the selective role of Mysm1 in lineage determination of DCs from CMPs: the selective expression of Mysm1 in a subset of CMPs and the different requirement of Mysm1 for PU.1 recruitment to the Flt3 locus vs GM-CSF-α and macrophage-colony-stimulating factor receptor loci. In conclusion, this study reveals an essential role of Mysm1 in epigenetic regulation of Flt3 transcription and DC development, and it provides a novel mechanism for lineage determination from CMP.

Frequent KIT mutations in human gastrointestinal stromal tumors.

Identifying gene mutations in individual tumors is critical to improve the efficacy of cancer therapy by matching targeted drugs to specific mutations. Gastrointestinal stromal tumors (GIST) are stromal or mesenchymal subepithelial neoplasms affecting the gastrointestinal tract and frequently contain activating gene mutations in either KIT or platelet-derived growth factor A (PDGFRA). Although GIST is highly responsive to several selective tyrosine kinase inhibitors, combined use of inhibitors targeting other mutations is needed to further prolong survival in patients with GIST. In this study, we aim to screen and identify genetic mutations in GIST for targeted therapy using the new Ion Torrent next-generation sequencing platform. Utilizing the Ion Ampliseq Cancer Panel, we sequenced 737 loci from 45 cancer-related genes using DNA extracted from formalin-fixed and paraffin-embedded (FFPE) samples of 121 human gastrointestinal stromal tumors, set up stringent parameters for reliable variant calling by filtering out potential raw base calling errors, and identified frequent mutations in the KIT gene. This study demonstrates the utility of using Ion Torrent sequencing to efficiently identify human cancer mutations. This may provide a molecular basis for clinically developing new drugs targeting these gene mutations for GIST therapy.

Genetic mutation analysis of human gastric adenocarcinomas using ion torrent sequencing platform.

Gastric cancer is the one of the major causes of cancer-related death, especially in Asia. Gastric adenocarcinoma, the most common type of gastric cancer, is heterogeneous and its incidence and cause varies widely with geographical regions, gender, ethnicity, and diet. Since unique mutations have been observed in individual human cancer samples, identification and characterization of the molecular alterations underlying individual gastric adenocarcinomas is a critical step for developing more effective, personalized therapies. Until recently, identifying genetic mutations on an individual basis by DNA sequencing remained a daunting task. Recent advances in new next-generation DNA sequencing technologies, such as the semiconductor-based Ion Torrent sequencing platform, makes DNA sequencing cheaper, faster, and more reliable. In this study, we aim to identify genetic mutations in the genes which are targeted by drugs in clinical use or are under development in individual human gastric adenocarcinoma samples using Ion Torrent sequencing. We sequenced 737 loci from 45 cancer-related genes in 238 human gastric adenocarcinoma samples using the Ion Torrent Ampliseq Cancer Panel. The sequencing analysis revealed a high occurrence of mutations along the TP53 locus (9.7%) in our sample set. Thus, this study indicates the utility of a cost and time efficient tool such as Ion Torrent sequencing to screen cancer mutations for the development of personalized cancer therapy.

PIK3CA and TP53 gene mutations in human breast cancer tumors frequently detected by ion torrent DNA sequencing.

Breast cancer is the most common malignancy and the leading cause of cancer deaths in women worldwide. While specific genetic mutations have been linked to 5-10% of breast cancer cases, other environmental and epigenetic factors influence the development and progression of the cancer. Since unique mutations patterns have been observed in individual cancer samples, identification and characterization of the distinctive breast cancer molecular profile is needed to develop more effective target therapies. Until recently, identifying genetic cancer mutations via personalized DNA sequencing was impractical and expensive. The recent technological advancements in next-generation DNA sequencing, such as the semiconductor-based Ion Torrent sequencing platform, has made DNA sequencing cost and time effective with more reliable results. Using the Ion Torrent Ampliseq Cancer Panel, we sequenced 737 loci from 45 cancer-related genes to identify genetic mutations in 105 human breast cancer samples. The sequencing analysis revealed missense mutations in PIK3CA, and TP53 genes in the breast cancer samples of various histologic types. Thus, this study demonstrates the necessity of sequencing individual human cancers in order to develop personalized drugs or combination therapies to effectively target individual, breast cancer-specific mutations.

Frequent mutations in EGFR, KRAS and TP53 genes in human lung cancer tumors detected by ion torrent DNA sequencing.

Lung cancer is the most common malignancy and the leading cause of cancer deaths worldwide. While smoking is by far the leading cause of lung cancer, other environmental and genetic factors influence the development and progression of the cancer. Since unique mutations patterns have been observed in individual cancer samples, identification and characterization of the distinctive lung cancer molecular profile is essential for developing more effective, tailored therapies. Until recently, personalized DNA sequencing to identify genetic mutations in cancer was impractical and expensive. The recent technological advancements in next-generation DNA sequencing, such as the semiconductor-based Ion Torrent sequencing platform, has made DNA sequencing cost and time effective with more reliable results. Using the Ion Torrent Ampliseq Cancer Panel, we sequenced 737 loci from 45 cancer-related genes to identify genetic mutations in 76 human lung cancer samples. The sequencing analysis revealed missense mutations in KRAS, EGFR, and TP53 genes in the breast cancer samples of various histologic types. Thus, this study demonstrates the necessity of sequencing individual human cancers in order to develop personalized drugs or combination therapies to effectively target individual, breast cancer-specific mutations.