Publications by authors named "Akhilesh Pandey"

437 Publications

Tyrosine Phosphoproteomics of Patient-Derived Xenografts Reveals Ephrin Type-B Receptor 4 Tyrosine Kinase as a Therapeutic Target in Pancreatic Cancer.

Cancers (Basel) 2021 Jul 7;13(14). Epub 2021 Jul 7.

Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN 55905, USA.

Pancreatic ductal adenocarcinoma is a recalcitrant tumor with minimal response to conventional chemotherapeutic approaches. Oncogenic signaling by activated tyrosine kinases has been implicated in cancers resulting in activation of diverse effector signaling pathways. Thus, the discovery of aberrantly activated tyrosine kinases is of great interest in developing novel therapeutic strategies in the treatment and management of pancreatic cancer. Patient-derived tumor xenografts (PDXs) in mice serve as potentially valuable preclinical models as they maintain the histological and molecular heterogeneity of the original human tumor. Here, we employed high-resolution mass spectrometry combined with immunoaffinity purification using anti-phosphotyrosine antibodies to profile tyrosine phosphoproteome across 13 pancreatic ductal adenocarcinoma PDX models. This analysis resulted in the identification of 1199 tyrosine-phosphorylated sites mapping to 704 proteins. The mass spectrometric analysis revealed widespread and heterogeneous activation of both receptor and non-receptor tyrosine kinases. Preclinical studies confirmed ephrin type-B receptor 4 (EphB4) as a potential therapeutic target based on the efficacy of human serum albumin-conjugated soluble EphB4 in mice bearing orthotopic xenografts. Immunohistochemistry-based validation using tissue microarrays from 346 patients with PDAC showed significant expression of EphB4 in >70% of patients. In summary, we present a comprehensive landscape of tyrosine phosphoproteome with EphB4 as a promising therapeutic target in pancreatic ductal adenocarcinoma.
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http://dx.doi.org/10.3390/cancers13143404DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8303779PMC
July 2021

DIA-Based Proteome Profiling of Nasopharyngeal Swabs from COVID-19 Patients.

J Proteome Res 2021 Jul 22. Epub 2021 Jul 22.

Department of Laboratory Medicine and Pathology, Mayo Clinic, 200 First Street Southwest, Rochester, Minnesota 55905, United States.

Since the recent outbreak of COVID-19, there have been intense efforts to understand viral pathogenesis and host immune response to combat SARS-CoV-2. It has become evident that different host alterations can be identified in SARS-CoV-2 infection based on whether infected cells, animal models or clinical samples are studied. Although nasopharyngeal swabs are routinely collected for SARS-CoV-2 detection by RT-PCR testing, host alterations in the nasopharynx at the proteomic level have not been systematically investigated. Thus, we sought to characterize the host response through global proteome profiling of nasopharyngeal swab specimens. A mass spectrometer combining trapped ion mobility spectrometry (TIMS) and high-resolution QTOF mass spectrometer with parallel accumulation-serial fragmentation (PASEF) was deployed for unbiased proteome profiling. First, deep proteome profiling of pooled nasopharyngeal swab samples was performed in the PASEF enabled DDA mode, which identified 7723 proteins that were then used to generate a spectral library. This approach provided peptide level evidence of five missing proteins for which MS/MS spectrum and mobilograms were validated with synthetic peptides. Subsequently, quantitative proteomic profiling was carried out for 90 individual nasopharyngeal swab samples (45 positive and 45 negative) in DIA combined with PASEF, termed as diaPASEF mode, which resulted in a total of 5023 protein identifications. Of these, 577 proteins were found to be upregulated in SARS-CoV-2 positive samples. Functional analysis of these upregulated proteins revealed alterations in several biological processes including innate immune response, viral protein assembly, and exocytosis. To the best of our knowledge, this study is the first to deploy diaPASEF for quantitative proteomic profiling of clinical samples and shows the feasibility of adopting such an approach to understand mechanisms and pathways altered in diseases.
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http://dx.doi.org/10.1021/acs.jproteome.1c00506DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8315246PMC
July 2021

The aftermath of COVID-19 pandemic on the diagnosis of TB at a tertiary care hospital in India.

J Infect Public Health 2021 Aug 8;14(8):1095-1098. Epub 2021 Jul 8.

Department of Microbiology, Kasturba Medical College, Manipal, Manipal Academy of Higher Education, Manipal, Karnataka 576104, India. Electronic address:

Background: The recent COVID-19 pandemic became a looming catastrophe over global public health and severely disrupted essential healthcare services like tuberculosis (TB). This study estimated the impact of the COVID-19 in the diagnosis of TB, a microbiology laboratory-based overview.

Method: This ambispective observational study was conducted at the Department of Microbiology in a tertiary care hospital in South Karnataka from January 2019 to December 2020. A standardized data collection sheet was prepared to collect the month-wise total number of suspected TB and confirmed TB samples. Data were analyzed using EZR 3.4.3 (R, open-source). Categorical variables were expressed in frequency and percentage. The Chi-square test was performed to test the difference in proportions and p < 0.05 indicated statistical significance.

Results: In this study, a significant drop was observed in suspected TB specimens in 2020 compared to 2019, i.e. 54.8% for microscopy, along with 34.2% and 49.7% for Xpert MTB/RIF and MGIT culture respectively. Also, a sharp decline in confirmed TB samples was noted in 2020 with 49%, 43.8%, and 59.7% reduction with microscopy, Xpert MTB/RIF, and MGIT culture respectively, compared to 2019. Another major finding from this study reveals the PTB: EPTB proportion changed from 2.7:1 in 2019 to 2.1:1 in 2020, divulging an overall increase in EPTB sample proportion in 2020 (p = 0.0385).

Conclusion: The COVID-19 pandemic adversely impacted the TB diagnostic services, resulting in a significant reduction of active TB case detection. It highlights an urgent need to revise the strategies to control and eliminate TB in this hour of the pandemic crisis.
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http://dx.doi.org/10.1016/j.jiph.2021.07.001DOI Listing
August 2021

High-resolution mass spectrometric analysis of cardiolipin profiles in Barth syndrome.

Mitochondrion 2021 Jul 15;60:27-32. Epub 2021 Jul 15.

Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN, United States; Center for Individualized Medicine, Mayo Clinic, Rochester, MN, United States. Electronic address:

Barth syndrome is an X-linked recessive disorder caused by pathogenic variants in TAZ, which leads to a reduction in cardiolipin with a concomitant elevation of monolysocardiolipins. There is a paucity of studies characterizing changes in individual species of monolysocardiolipins, dilysocardiolipins and cardiolipin in Barth syndrome using high resolution untargeted lipidomics that can accurately annotate and quantify diverse lipids. We confirmed the structural diversity monolysocardiolipins, dilysocardiolipins and cardiolipin and identified individual species that showed previously unreported alterations in BTHS. Development of mass spectrometry-based targeted assays for these lipid biomarkers should provide an important tool for clinical diagnosis of Barth syndrome.
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http://dx.doi.org/10.1016/j.mito.2021.07.003DOI Listing
July 2021

Analytical sensitivity and specificity of four point of care rapid antigen diagnostic tests for SARS-CoV-2 using real-time quantitative PCR, quantitative droplet digital PCR, and a mass spectrometric antigen assay as comparator methods.

Clin Chem 2021 Jul 8. Epub 2021 Jul 8.

Department of Laboratory Medicine and Pathology.

Background: We evaluated the analytical sensitivity and specificity of four rapid antigen diagnostic tests (Ag RDTs) for SARS-CoV-2, using reverse transcription quantitative PCR (RT-qPCR) as the reference method; and further characterizing samples using droplet digital quantitative PCR (ddPCR) and a mass spectrometric antigen test.

Methods: 350 (150 negative and 200 RT-qPCR positive) residual phosphate buffered saline (PBS) samples were tested for antigen using the BD Veritor lateral flow (LF), ACON LF, ACON fluorescence immunoassay (FIA), and LumiraDx FIA. ddPCR was performed on RT-qPCR positive samples to quantitate the viral load in copies/mL applied to each Ag RDT. Mass spectrometric antigen testing was performed on PBS samples to obtain a set of RT-qPCR positive, antigen positive samples for further analysis.

Results: All Ag RDTs had nearly 100% specificity compared to RT-qPCR. Overall analytical sensitivity varied from 66.5% to 88.3%. All methods detected antigen in samples with viral load >1,500,000 copies/mL RNA, and detected ≥75% of samples with viral load of 500,000 to 1,500,000 copies/mL. The BD Veritor LF detected only 25% of samples with viral load between 50,000-500,000 copies/mL, compared to 75% for the ACON LF device and >80% for LumiraDx and ACON FIA. The ACON FIA detected significantly more samples with viral load <50,000 copies/mL compared to the BD Veritor. Among samples with detectable antigen and viral load <50,000 copies/mL, sensitivity of the Ag RDT varied between 13.0% (BD Veritor) and 78.3% (ACON FIA).

Conclusions: Ag RDTs differ significantly in analytical sensitivity, particularly at viral load <500,000 copies/mL.
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http://dx.doi.org/10.1093/clinchem/hvab138DOI Listing
July 2021

A mass spectrometry-based targeted assay for detection of SARS-CoV-2 antigen from clinical specimens.

EBioMedicine 2021 Jul 3;69:103465. Epub 2021 Jul 3.

Department of Laboratory Medicine and Pathology, Division of Clinical Biochemistry and Immunology, Mayo Clinic, MN 55905, USA; Center for Individualized Medicine, Mayo Clinic, Rochester, MN 55905, USA; Center for Molecular Medicine, National Institute of Mental Health and Neurosciences, Hosur Road, Bangalore, Karnataka 560029, India. Electronic address:

Background: The COVID-19 pandemic caused by severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) has overwhelmed health systems worldwide and highlighted limitations of diagnostic testing. Several types of diagnostic tests including RT-PCR-based assays and antigen detection by lateral flow assays, each with their own strengths and weaknesses, have been developed and deployed in a short time.

Methods: Here, we describe an immunoaffinity purification approach followed a by high resolution mass spectrometry-based targeted qualitative assay capable of detecting SARS-CoV-2 viral antigen from nasopharyngeal swab samples. Based on our discovery experiments using purified virus, recombinant viral protein and nasopharyngeal swab samples from COVID-19 positive patients, nucleocapsid protein was selected as a target antigen. We then developed an automated antibody capture-based workflow coupled to targeted high-field asymmetric waveform ion mobility spectrometry (FAIMS) - parallel reaction monitoring (PRM) assay on an Orbitrap Exploris 480 mass spectrometer. An ensemble machine learning-based model for determining COVID-19 positive samples was developed using fragment ion intensities from the PRM data.

Findings: The optimized targeted assay, which was used to analyze 88 positive and 88 negative nasopharyngeal swab samples for validation, resulted in 98% (95% CI = 0.922-0.997) (86/88) sensitivity and 100% (95% CI = 0.958-1.000) (88/88) specificity using RT-PCR-based molecular testing as the reference method.

Interpretation: Our results demonstrate that direct detection of infectious agents from clinical samples by tandem mass spectrometry-based assays have potential to be deployed as diagnostic assays in clinical laboratories, which has hitherto been limited to analysis of pure microbial cultures.

Funding: This study was supported by DBT/Wellcome Trust India Alliance Margdarshi Fellowship grant IA/M/15/1/502023 awarded to AP and the generosity of Eric and Wendy Schmidt.
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http://dx.doi.org/10.1016/j.ebiom.2021.103465DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8253671PMC
July 2021

Cerebrospinal fluid lipidomics for biomarkers of Alzheimer's disease.

Mol Omics 2021 Jun;17(3):454-463

Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN, 55902, USA.

Alzheimer's disease (AD) is the most common cause of dementia and is associated with serious neurologic sequelae resulting from neurodegenerative changes. Identification of markers of early-stage AD could be important for designing strategies to arrest the progression of the disease. The brain is rich in lipids because they are crucial for signal transduction and anchoring of membrane proteins. Cerebrospinal fluid (CSF) is an excellent specimen for studying the metabolism of lipids in AD because it can reflect changes occurring in the brain. We aimed to identify CSF lipidomic alterations associated with AD, using untargeted lipidomics, carried out in positive and negative ion modes. We found CSF lipids that were significantly altered in AD cases. In addition, comparison of CSF lipid profiles between persons with mild cognitive impairment (MCI) and AD showed a strong positive correlation between the lipidomes of the MCI and AD groups. The novel lipid biomarkers identified in this study are excellent candidates for validation in a larger set of patient samples and as predictive biomarkers of AD through future longitudinal studies. Once validated, the lipid biomarkers could lead to early detection, disease monitoring and the ability to measure the efficacy of potential therapeutic interventions in AD.
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http://dx.doi.org/10.1039/d0mo00186dDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8210464PMC
June 2021

Mass Spectrometric Analysis of Urine from COVID-19 Patients for Detection of SARS-CoV-2 Viral Antigen and to Study Host Response.

J Proteome Res 2021 07 2;20(7):3404-3413. Epub 2021 Jun 2.

Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, Minnesota 55905, United States.

SARS-CoV-2 infection has become a major public health burden and affects many organs including lungs, kidneys, the liver, and the brain. Although the virus is readily detected and diagnosed using nasopharyngeal swabs by reverse transcriptase polymerase chain reaction (RT-PCR), detection of its presence in body fluids is fraught with difficulties. A number of published studies have failed to detect viral RNA by RT-PCR methods in urine. Although microbial identification in clinical microbiology using mass spectrometry is undertaken after culture, here we undertook a mass spectrometry-based approach that employed an enrichment step to capture and detect SARS-CoV-2 nucleocapsid protein directly from urine of COVID-19 patients without any culture. We detected SARS-CoV-2 nucleocapsid protein-derived peptides from 13 out of 39 urine samples. Further, a subset of COVID-19 positive and COVID-19 negative urine samples validated by mass spectrometry were used for the quantitative proteomics analysis. Proteins with increased abundance in urine of SARS-CoV-2 positive individuals were enriched in the acute phase response, regulation of complement system, and immune response. Notably, a number of renal proteins such as podocin (NPHS2), an amino acid transporter (SLC36A2), and sodium/glucose cotransporter 5 (SLC5A10), which are intimately involved in normal kidney function, were decreased in the urine of COVID-19 patients. Overall, the detection of viral antigens in urine using mass spectrometry and alterations of the urinary proteome could provide insights into understanding the pathogenesis of COVID-19.
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http://dx.doi.org/10.1021/acs.jproteome.1c00391DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8189038PMC
July 2021

Complement and Coagulation Cascades are Potentially Involved in Dopaminergic Neurodegeneration in α-Synuclein-Based Mouse Models of Parkinson's Disease.

J Proteome Res 2021 Jul 1;20(7):3428-3443. Epub 2021 Jun 1.

Neuroregeneration and Stem Cell Programs, Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore 21205-2105, Maryland, United States.

Parkinson's disease (PD) is the second most common neurodegenerative disorder that results in motor dysfunction and, eventually, cognitive impairment. α-Synuclein protein is known as a central protein to the pathophysiology of PD, but the underlying pathological mechanism still remains to be elucidated. In an effort to understand how α-synuclein underlies the pathology of PD, various PD mouse models with α-synuclein overexpression have been developed. However, systemic analysis of the brain proteome of those mouse models is lacking. In this study, we established two mouse models of PD by injecting α-synuclein preformed fibrils (PFF) or by inducing overexpression of human A53T α-synuclein to investigate common pathways in the two different types of the PD mouse models. For more accurate quantification of mouse brain proteome, the proteins were quantified using the method of stable isotope labeling with amino acids in mammals . We identified a total of 8355 proteins from the two mouse models; ∼6800 and ∼7200 proteins from α-synuclein PFF-injected mice and human A53T α-synuclein transgenic mice, respectively. Through pathway analysis of the differentially expressed proteins common to both PD mouse models, it was discovered that the complement and coagulation cascade pathways were enriched in the PD mice compared to control animals. Notably, a validation study demonstrated that complement component 3 (C3)-positive astrocytes were increased in the ventral midbrain of the intrastriatal α-synuclein PFF-injected mice and C3 secreted from astrocytes could induce the degeneration of dopaminergic neurons. This is the first study that highlights the significance of the complement and coagulation pathways in the pathogenesis of PD through proteome analyses with two sophisticated mouse models of PD.
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http://dx.doi.org/10.1021/acs.jproteome.0c01002DOI Listing
July 2021

Mitochondrial localization and moderated activity are key to murine erythroid enucleation.

Blood Adv 2021 05;5(10):2490-2504

Department of Cell, Developmental and Regenerative Biology.

Mammalian red blood cells (RBCs), which primarily contain hemoglobin, exemplify an elaborate maturation process, with the terminal steps of RBC generation involving extensive cellular remodeling. This encompasses alterations of cellular content through distinct stages of erythroblast maturation that result in the expulsion of the nucleus (enucleation) followed by the loss of mitochondria and all other organelles and a transition to anaerobic glycolysis. Whether there is any link between erythroid removal of the nucleus and the function of any other organelle, including mitochondria, remains unknown. Here we demonstrate that mitochondria are key to nuclear clearance. Using live and confocal microscopy and high-throughput single-cell imaging, we show that before nuclear polarization, mitochondria progressively move toward one side of maturing erythroblasts and aggregate near the nucleus as it extrudes from the cell, a prerequisite for enucleation to proceed. Although we found active mitochondrial respiration is required for nuclear expulsion, levels of mitochondrial activity identify distinct functional subpopulations, because terminally maturing erythroblasts with low relative to high mitochondrial membrane potential are at a later stage of maturation, contain greatly condensed nuclei with reduced open chromatin-associated acetylation histone marks, and exhibit higher enucleation rates. Lastly, to our surprise, we found that late-stage erythroblasts sustain mitochondrial metabolism and subsequent enucleation, primarily through pyruvate but independent of in situ glycolysis. These findings demonstrate the critical but unanticipated functions of mitochondria during the erythroblast enucleation process. They are also relevant to the in vitro production of RBCs as well as to disorders of the erythroid lineage.
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http://dx.doi.org/10.1182/bloodadvances.2021004259DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8152511PMC
May 2021

Galactose-Clicked Curcumin-Mediated Reversal of Meropenem Resistance among by Targeting Its Carbapenemases and the AcrAB-TolC Efflux System.

Antibiotics (Basel) 2021 Apr 4;10(4). Epub 2021 Apr 4.

Bacterial Biofilm and Drug Resistance Research Laboratory, Department of Microbiology, Institute of Medical Sciences, Banaras Hindu University, Varanasi 221005, India.

In over eighty years, despite successive antibiotics discoveries, the rapid advent of multidrug resistance among bacterial pathogens has jolted our misapprehension of success over them. Resistance is spreading faster than the discovery of new antibiotics/antimicrobials. Therefore, the search for better antimicrobials/additives becomes prudent. A water-soluble curcumin derivative (Cur) was synthesised, employing a Cu (I) catalysed 1, 3-cyclo addition reaction; it has been evaluated as a potential treatment for multidrug-resistant isolates and as an antibiotic adjuvant for meropenem against hypervirulent multidrug-resistant isolates. We also investigated its solubility and effect over carbapenemase activity. Additionally, we investigated its impact on the AcrAB-TolC system. We found that Cur inhibited bacterial growth at a minimal concentration of 16 µg/mL; at a 32 µg/mL concentration, it killed bacterial growth completely. Only nine (9.4%) isolates were sensitive to meropenem; however, after synergising with Cur (8 µg/mL), 85 (88.54%) hvKP isolates became sensitive to the drug. The Cur also inhibited the AcrAB-TolC efflux system at 1 µg/mL concentration by disrupting the membrane potential and causing depolarisation. The kinetic parameters obtained also indicated its promise as a carbapenemase inhibitor. These results suggest that Cur can be an excellent drug candidate as a broad-spectrum antibacterial and anti-efflux agent.
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http://dx.doi.org/10.3390/antibiotics10040388DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8066637PMC
April 2021

Developmental partitioning of SYK and ZAP70 prevents autoimmunity and cancer.

Mol Cell 2021 05 19;81(10):2094-2111.e9. Epub 2021 Apr 19.

Center of Molecular and Cellular Oncology, Yale Cancer Center, Yale School of Medicine, New Haven, CT, USA; Department of Immunobiology, Yale University School of Medicine, New Haven, CT, USA. Electronic address:

Even though SYK and ZAP70 kinases share high sequence homology and serve analogous functions, their expression in B and T cells is strictly segregated throughout evolution. Here, we identified aberrant ZAP70 expression as a common feature in a broad range of B cell malignancies. We validated SYK as the kinase that sets the thresholds for negative selection of autoreactive and premalignant clones. When aberrantly expressed in B cells, ZAP70 competes with SYK at the BCR signalosome and redirects SYK from negative selection to tonic PI3K signaling, thereby promoting B cell survival. In genetic mouse models for B-ALL and B-CLL, conditional expression of Zap70 accelerated disease onset, while genetic deletion impaired malignant transformation. Inducible activation of Zap70 during B cell development compromised negative selection of autoreactive B cells, resulting in pervasive autoantibody production. Strict segregation of the two kinases is critical for normal B cell selection and represents a central safeguard against the development of autoimmune disease and B cell malignancies.
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http://dx.doi.org/10.1016/j.molcel.2021.03.043DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8239336PMC
May 2021

Shukla-Vernon Syndrome: A Second Family with a Novel Variant in the Gene.

Genes (Basel) 2021 03 22;12(3). Epub 2021 Mar 22.

Manipal Academy of Higher Education, Manipal 576104, India.

Shukla-Vernon syndrome (SHUVER) is an extremely rare neurodevelopmental disorder characterized by global developmental delay, intellectual disability, behavioral anomalies, and dysmorphic features. Pathogenic variants in the gene have been identified as the molecular cause for this disorder. The gene encodes for BCL-6 corepressor-like protein 1, a transcriptional corepressor that is an integral component of protein complexes involved in transcription repression. In this study, we report an Indian family with two male siblings with features of Shukla-Vernon syndrome. The patients exhibited global developmental delay, intellectual disability, kyphosis, seizures, and dysmorphic features including bushy prominent eyebrows with synophrys, sharp beaked prominent nose, protuberant lower jaw, squint, and hypoplastic ears with fused ear lobes. No behavioral abnormalities were observed. Whole exome sequencing revealed a novel potentially pathogenic arginine to cysteine substitution (p.Arg1265Cys) in the BCORL1 protein. This is the second report of Shukla-Vernon syndrome with a novel missense variant in the gene. Our study confirms and expands the phenotypes and genotypes described previously for this syndrome and should aid in diagnosis and genetic counselling of patients and their families.
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http://dx.doi.org/10.3390/genes12030452DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8005212PMC
March 2021

Acute Kidney Injury Post-Percutaneous Nephrolithotomy (PNL): Prospective Outcomes from a University Teaching Hospital.

J Clin Med 2021 Mar 29;10(7). Epub 2021 Mar 29.

Department of Urology, Kasturba Medical College Hospital, Manipal Academy of Higher Education (MAHE), Manipal 576104, Karnataka, India.

Acute Kidney Injury (AKI) after percutaneous nephrolithotomy (PNL) is a significant complication, but evidence on its incidence is bereft in the literature. The objective of this prospective observational study was to analyze the incidence of post-PNL AKI and the potential risk factors and outcomes. Demographic data collected included age, gender, body mass index (BMI), comorbidities (hypertension, diabetes mellitus), and drug history-particularly angiotensin converting enzyme inhibitors (ACE inhibitors), angiotensin II receptor blockers and beta blockers. Laboratory data included serial serum creatinine measured pre- and postoperation (12, 24, and 48 h), hemoglobin (Hb), total leucocyte count (TLC), Prothrombin time (PT), serum uric acid and urine culture. Stone factors were assessed by noncontrast computerized tomography of kidneys, ureter and bladder (NCCT KUB) and included stone burden, location and Hounsfield values. Intraoperative factors assessed were puncture site, tract size, tract number, operative time, the need for blood transfusion and stone clearance. Postoperative complications were documented using the modified Clavien-Dindo grading system and patients with postoperative AKI were followed up with serial creatinine measurements up to 1 year. Among the 509 patients analyzed, 47 (9.23%) developed postoperative AKI. Older patients, with associated hypertension and diabetes mellitus, those receiving ACE inhibitors and with lower preoperative hemoglobin and higher serum uric acid, had higher incidence of AKI. Higher stone volume and density, staghorn stones, multiple punctures and longer operative time were significantly associated with postoperative AKI. Patients with AKI had an increased length of hospital stay and 17% patients progressed to chronic kidney disease (CKD). Cut-off values for patient age (39.5 years), serum uric acid (4.05 mg/dL) and stone volume (673.06 mm) were assessed by receiver operating characteristic (ROC) curve analysis. Highlighting the strong predictors of post-PNL AKI allows early identification, proper counseling and postoperative planning and management in an attempt to avoid further insult to the kidney.
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http://dx.doi.org/10.3390/jcm10071373DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8037383PMC
March 2021

Mapping the micro-proteome of the nuclear lamina and lamina-associated domains.

Life Sci Alliance 2021 05 23;4(5). Epub 2021 Mar 23.

Department of Biological Chemistry, Johns Hopkins University of Medicine, Baltimore, MD, USA

The nuclear lamina is a proteinaceous network of filaments that provide both structural and gene regulatory functions by tethering proteins and large domains of DNA, the so-called lamina-associated domains (LADs), to the periphery of the nucleus. LADs are a large fraction of the mammalian genome that are repressed, in part, by their association to the nuclear periphery. The genesis and maintenance of LADs is poorly understood as are the proteins that participate in these functions. In an effort to identify proteins that reside at the nuclear periphery and potentially interact with LADs, we have taken a two-pronged approach. First, we have undertaken an interactome analysis of the inner nuclear membrane bound LAP2β to further characterize the nuclear lamina proteome. To accomplish this, we have leveraged the BioID system, which previously has been successfully used to characterize the nuclear lamina proteome. Second, we have established a system to identify proteins that bind to LADs by developing a chromatin-directed BioID system. We combined the BioID system with the m6A-tracer system which binds to LADs in live cells to identify both LAD proximal and nuclear lamina proteins. In combining these datasets, we have further characterized the protein network at the nuclear lamina, identified putative LAD proximal proteins and found several proteins that appear to interface with both micro-proteomes. Importantly, several proteins essential for LAD function, including heterochromatin regulating proteins related to H3K9 methylation, were identified in this study.
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http://dx.doi.org/10.26508/lsa.202000774DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8008952PMC
May 2021

The mitochondrial carrier SFXN1 is critical for complex III integrity and cellular metabolism.

Cell Rep 2021 Mar;34(11):108869

Department of Physiology, The Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA. Electronic address:

Mitochondrial carriers (MCs) mediate the passage of small molecules across the inner mitochondrial membrane (IMM), enabling regulated crosstalk between compartmentalized reactions. Despite MCs representing the largest family of solute carriers in mammals, most have not been subjected to a comprehensive investigation, limiting our understanding of their metabolic contributions. Here, we functionally characterize SFXN1, a member of the non-canonical, sideroflexin family. We find that SFXN1, an integral IMM protein with an uneven number of transmembrane domains, is a TIM22 complex substrate. SFXN1 deficiency leads to mitochondrial respiratory chain impairments, most detrimental to complex III (CIII) biogenesis, activity, and assembly, compromising coenzyme Q levels. The CIII dysfunction is independent of one-carbon metabolism, the known primary role for SFXN1 as a mitochondrial serine transporter. Instead, SFXN1 supports CIII function by participating in heme and α-ketoglutarate metabolism. Our findings highlight the multiple ways that SFXN1-based amino acid transport impacts mitochondrial and cellular metabolic efficiency.
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http://dx.doi.org/10.1016/j.celrep.2021.108869DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8048093PMC
March 2021

Quantitative Proteomics Reveals that the OGT Interactome Is Remodeled in Response to Oxidative Stress.

Mol Cell Proteomics 2021 Mar 12;20:100069. Epub 2021 Mar 12.

Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland, United States; Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, Maryland, United States. Electronic address:

The dynamic modification of specific serine and threonine residues of intracellular proteins by O-linked N-acetyl-β-D-glucosamine (O-GlcNAc) mitigates injury and promotes cytoprotection in a variety of stress models. The O-GlcNAc transferase (OGT) and the O-GlcNAcase are the sole enzymes that add and remove O-GlcNAc, respectively, from thousands of substrates. It remains unclear how just two enzymes can be specifically controlled to affect glycosylation of target proteins and signaling pathways both basally and in response to stress. Several lines of evidence suggest that protein interactors regulate these responses by affecting OGT and O-GlcNAcase activity, localization, and substrate specificity. To provide insight into the mechanisms by which OGT function is controlled, we have used quantitative proteomics to define OGT's basal and stress-induced interactomes. OGT and its interaction partners were immunoprecipitated from OGT WT, null, and hydrogen peroxide-treated cell lysates that had been isotopically labeled with light, medium, and heavy lysine and arginine (stable isotopic labeling of amino acids in cell culture). In total, more than 130 proteins were found to interact with OGT, many of which change their association upon hydrogen peroxide stress. These proteins include the major OGT cleavage and glycosylation substrate, host cell factor 1, which demonstrated a time-dependent dissociation after stress. To validate less well-characterized interactors, such as glyceraldehyde 3-phosphate dehydrogenase and histone deacetylase 1, we turned to parallel reaction monitoring, which recapitulated our discovery-based stable isotopic labeling of amino acids in cell culture approach. Although the majority of proteins identified are novel OGT interactors, 64% of them are previously characterized glycosylation targets that contain varied domain architecture and function. Together these data demonstrate that OGT interacts with unique and specific interactors in a stress-responsive manner.
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http://dx.doi.org/10.1016/j.mcpro.2021.100069DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8079276PMC
March 2021

Integrated Proteomic and Phosphoproteomics Analysis of DKK3 Signaling Reveals Activated Kinase in the Most Aggressive Gallbladder Cancer.

Cells 2021 Feb 28;10(3). Epub 2021 Feb 28.

Institute of Bioinformatics, International Tech Park, Bangalore 560066, India.

DKK3 is a secreted protein, which belongs to a family of Wnt antagonists and acts as a potential tumor suppressor in gallbladder cancer. To further understand its tumor suppressor functions, we overexpressed DKK3 in 3 GBC cell lines. We have employed high-resolution mass spectrometry and tandem mass tag (TMT) multiplexing technology along with immobilized metal affinity chromatography to enrich phosphopeptides to check the downstream regulators. In this study, we reported for the first time the alteration in the phosphorylation of 14 kinases upon DKK3 overexpression. In addition, we observed DKK3 induced hyper phosphorylation of 2 phosphatases: PPP1R12A and PTPRA, which have not been reported previously. Canonical pathway analysis of altered molecules indicated differential enrichment of signaling cascades upon DKK3 overexpression in all the 3 cell lines. Protein kinase A signaling, Sirtuin signaling pathway, and Cell Cycle Control of Chromosomal Replication were observed to be differentially activated in the GBC cell lines. Our study revealed, DKK3 overexpression has differential effect based on the aggressive behavior of the cell lines. This study expands the understanding of DKK3-mediated signaling events and can be used as a primary factor for understanding the complex nature of this molecule.
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http://dx.doi.org/10.3390/cells10030511DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7997438PMC
February 2021

Ethylmalonic encephalopathy ETHE1 p. D165H mutation alters the mitochondrial function in human skeletal muscle proteome.

Mitochondrion 2021 05 24;58:64-71. Epub 2021 Feb 24.

Institute of Bioinformatics, International Tech Park, Bangalore, India; Center for Molecular Medicine, National Institute of Mental Health and Neurosciences (NIMHANS), Bangalore, India; Manipal Academy of Higher Education, Manipal, India. Electronic address:

Ethylmalonic encephalopathy (EE) is a rare autosomal recessive inborn error of metabolism. To study the molecular effects of ETHE1 p. D165H mutation, we employed mass spectrometry-based mitochondrial proteome and phosphoproteome profiling in the human skeletal muscle. Eighty-six differentially altered proteins were identified, of which thirty-seven mitochondrial proteins were differentially expressed, and most of the proteins (37%) were down-regulated in the OXPHOS complex-IV. Also, nine phosphopeptides that correspond to eight mitochondrial proteins were significantly affected in EE patient. These altered proteins recognized are involved in several pathways and molecular functions, predominantly in oxidoreductase activity. This is the first study that has integrated proteome and phosphoproteome of skeletal muscle and identified multiple proteins associated in the pathogenesis of EE.
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http://dx.doi.org/10.1016/j.mito.2021.02.011DOI Listing
May 2021

Proteomics-based approach for differentiation of age-related macular degeneration sub-types.

Indian J Ophthalmol 2021 03;69(3):647-654

L&T Opthalmic Pathology, Vision Research Foundation, Sankara Nethralaya, Chennai, India.

Purpose: Age-related macular degeneration (AMD) is one of the leading causes of irreversible central vision loss in the elderly population. The current study aims to find non-invasive prognostic biomarkers in the urine specimens of the AMD patients.

Methods: Peripheral blood and urine samples were collected from 23 controls and 61 AMD patients. Genomic DNA was extracted from the buffy coat of peripheral blood. Allele specific PCR was used to assay SNPs in complement factor H (CFH), complement component 3 (C3). Comparative proteomic analysis of urine samples from early AMD, choroidal neovascular membrane (CNVM), geographic atrophy (GA), and healthy controls was performed using isobaric labelling followed by mass spectrometry. Validation was performed using enzyme-linked immunosorbent assay (ELISA).

Results: Comparative proteomic analysis of urine samples identified 751 proteins, of which 383 proteins were found to be differentially expressed in various groups of AMD patients. Gene ontology classification of differentially expressed proteins revealed the majority of them were involved in catalytic functions and binding activities. Pathway analysis showed cell adhesion molecule pathways (CAMs), Complement and coagulation cascades, to be significantly deregulated in AMD. Upon validation by ELISA, SERPINA-1 (Alpha1 antitrypsin), TIMP-1 (Tissue inhibitor of matrix metaloprotease-1), APOA-1 (Apolipoprotein A-1) were significantly over-expressed in AMD (n = 61) patients compared to controls (n = 23). A logistic model of APOA-1 in combination with CFH and C3 polymorphisms predicted the risk of developing AMD with 82% accuracy.

Conclusion: This study gives us a preliminary data on non-invasive predictive biomarkers for AMD, which can be further validated in a large cohort and translated for diagnostic use.
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http://dx.doi.org/10.4103/ijo.IJO_470_20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7942106PMC
March 2021

Persistently Elevated mTOR Complex 1-S6 Kinase 1 Disrupts DARPP-32-Dependent D Dopamine Receptor Signaling and Behaviors.

Biol Psychiatry 2021 06 27;89(11):1058-1072. Epub 2020 Oct 27.

Solomon Snyder Department of Neuroscience, School of Medicine, Johns Hopkins University, Baltimore, Maryland; Department of Neurology, School of Medicine, Johns Hopkins University, Baltimore, Maryland. Electronic address:

Background: The serine-threonine kinase mTORC1 (mechanistic target of rapamycin complex 1) is essential for normal cell function but is aberrantly activated in the brain in both genetic-developmental and sporadic diseases and is associated with a spectrum of neuropsychiatric symptoms. The underlying molecular mechanisms of cognitive and neuropsychiatric symptoms remain controversial.

Methods: The present study examines behaviors in transgenic models that express Rheb, the most proximal known activator of mTORC1, and profiles striatal phosphoproteomics in a model with persistently elevated mTORC1 signaling. Biochemistry, immunohistochemistry, electrophysiology, and behavior approaches are used to examine the impact of persistently elevated mTORC1 on D dopamine receptor (D1R) signaling. The effect of persistently elevated mTORC1 was confirmed using D1-Cre to elevate mTORC1 activity in D1R neurons.

Results: We report that persistently elevated mTORC1 signaling blocks canonical D1R signaling that is dependent on DARPP-32 (dopamine- and cAMP-regulated neuronal phosphoprotein). The immediate downstream effector of mTORC1, ribosomal S6 kinase 1 (S6K1), phosphorylates and activates DARPP-32. Persistent elevation of mTORC1-S6K1 occludes dynamic D1R signaling downstream of DARPP-32 and blocks multiple D1R responses, including dynamic gene expression, D1R-dependent corticostriatal plasticity, and D1R behavioral responses including sociability. Candidate biomarkers of mTORC1-DARPP-32 occlusion are increased in the brain of human disease subjects in association with elevated mTORC1-S6K1, supporting a role for this mechanism in cognitive disease.

Conclusions: The mTORC1-S6K1 intersection with D1R signaling provides a molecular framework to understand the effects of pathological mTORC1 activation on behavioral symptoms in neuropsychiatric disease.
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http://dx.doi.org/10.1016/j.biopsych.2020.10.012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8076344PMC
June 2021

Is the Proteome of Bronchoalveolar Lavage Extracellular Vesicles a Marker of Advanced Lung Cancer?

Cancers (Basel) 2020 Nov 20;12(11). Epub 2020 Nov 20.

Computational and Experimental Biology Group, Chronic Diseases Research Centre, NOVA Medical School, Faculdade de Ciencias Medicas, Universidade NOVA de Lisboa, Campo dos Martires da Patria, 130, 1169-056 Lisboa, Portugal.

Acellular bronchoalveolar lavage (BAL) proteomics can partially separate lung cancer from non-lung cancer patients based on principal component analysis and multivariate analysis. Furthermore, the variance in the proteomics data sets is correlated mainly with lung cancer status and, to a lesser extent, smoking status and gender. Despite these advances BAL small and large extracellular vehicles (EVs) proteomes reveal aberrant protein expression in paracrine signaling mechanisms in cancer initiation and progression. We consequently present a case-control study of 24 bronchoalveolar lavage extracellular vesicle samples which were analyzed by state-of-the-art liquid chromatography-mass spectrometry (LC-MS). We obtained evidence that BAL EVs proteome complexity correlated with lung cancer stage 4 and mortality within two years´ follow-up ( value = 0.006). The potential therapeutic target DNMT3B complex is significantly up-regulated in tumor tissue and BAL EVs. The computational analysis of the immune and fibroblast cell markers in EVs suggests that patients who deceased within the follow-up period display higher marker expression indicative of innate immune and fibroblast cells (four out of five cases). This study provides insights into the proteome content of BAL EVs and their correlation to clinical outcomes.
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http://dx.doi.org/10.3390/cancers12113450DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7699733PMC
November 2020

Determinants of Metabolic Syndrome and 5-Year Cardiovascular Risk Estimates among HIV-Positive Individuals from an Indian Tertiary Care Hospital.

AIDS Res Treat 2020 28;2020:5019025. Epub 2020 Oct 28.

Department of Infectious Diseases, Kasturba Medical College, Manipal Academy of Higher Education, Manipal, Karnataka 576104, India.

Longer survival due to use of antiretroviral therapy (ART) has made human immunodeficiency virus- (HIV-) infected individuals prone to chronic diseases such as diabetes, hypertension, and cardiovascular diseases (CVD). Metabolic syndrome (MS), a constellation of risk factors which increase chances of the cardiovascular disease and diabetes, can increase the morbidity and mortality among this population. Hence, the present study was conducted with the objectives of estimating the prevalence and determinants of MS among ART naïve and ART-treated patients and assess their 5-year CVD risk using the reduced version of Data Collection on Adverse Effects of Anti-HIV Drugs (D : A : D) risk prediction model (D : A : D(R)). This hospital-based cross-sectional study included 182 adults aged ≥ 18 years. MS was defined using the National Cholesterol Education Program-Adult Treatment Panel-3 (NCEP ATP-3) criteria. Univariate and multivariate logistic regressions were done to identify the factors associated with MS. Prevalence of MS was 40.1% (95% confidence interval (CI) = 33.0%-47.2%). About 24.7% of the participants had at least a single criterion for MS. Age >45 years (adjusted odds ratio (AOR) = 2.3; 95% CI = 1.1-4.9, < 0.018) and body mass index (BMI) > 23 kg/m (AOR = 6.4; 95% CI = 3.1-13.1, < 0.001) were positively associated with MS, whereas daily consumption of high sugar items was inversely associated (AOR = 0.2; 95% CI = 0.1-0.5, < 0.001). More than 50% of the participants were found to have moderate or high 5-year CVD risk. Observed prevalence of MS among HIV patients was higher than other studies done in India. Considering a sizeable number of participants to be having moderate to high CVD risk, culturally appropriate lifestyle interventions need to be planned.
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http://dx.doi.org/10.1155/2020/5019025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7641704PMC
October 2020

Expanding the clinical and metabolic phenotype of DPM2 deficient congenital disorders of glycosylation.

Mol Genet Metab 2021 01 17;132(1):27-37. Epub 2020 Oct 17.

Mayo Clinic, Department of Clinical Genomics, Rochester, MN, USA; Mayo Clinic, Department of Laboratory of Medical Pathology, Rochester, MN, USA.

Pathogenic alterations in the DPM2 gene have been previously described in patients with hypotonia, progressive muscle weakness, absent psychomotor development, intractable seizures, and early death. We identified biallelic DPM2 variants in a 23-year-old male with truncal hypotonia, hypertonicity, congenital heart defects, intellectual disability, and generalized muscle wasting. His clinical presentation was much less severe than that of the three previously described patients. This is the second report on this ultra-rare disorder. Here we review the characteristics of previously reported individuals with a defect in the DPM complex while expanding the clinical phenotype of DPM2-Congenital Disorders of Glycosylation. In addition, we offer further insights into the pathomechanism of DPM2-CDG disorder by introducing glycomics and lipidomics analysis.
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http://dx.doi.org/10.1016/j.ymgme.2020.10.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7855207PMC
January 2021

PASS-DIA: A Data-Independent Acquisition Approach for Discovery Studies.

Anal Chem 2020 11 20;92(21):14466-14475. Epub 2020 Oct 20.

Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, Minnesota 55905, United States.

A data-independent acquisition (DIA) approach is being increasingly adopted as a promising strategy for identification and quantitation of proteomes. As most DIA data sets are acquired with wide isolation windows, highly complex MS/MS spectra are generated, which negatively impacts obtaining peptide information through classical protein database searches. Therefore, the analysis of DIA data mainly relies on the evidence of the existence of peptides from prebuilt spectral libraries. Consequently, one major weakness of this method is that it does not account for peptides that are not included in the spectral library, precluding the use of DIA for discovery studies. Here, we present a strategy termed Precursor ion And Small Slice-DIA (PASS-DIA) in which MS/MS spectra are acquired with small isolation windows (slices) and MS/MS spectra are interpreted with accurately determined precursor ion masses. This method enables the direct application of conventional spectrum-centric analysis pipelines for peptide identification and precursor ion-based quantitation. The performance of PASS-DIA was observed to be superior to both data-dependent acquisition (DDA) and conventional DIA experiments with 69 and 48% additional protein identifications, respectively. Application of PASS-DIA for the analysis of post-translationally modified peptides again highlighted its superior performance in characterizing phosphopeptides (77% more), N-terminal acetylated peptides (56% more), and N-glycopeptides (83% more) as compared to DDA alone. Finally, the use of PASS-DIA to characterize a rare proteome of human fallopian tube organoids enabled 34% additional protein identifications than DDA alone and revealed biologically relevant pathways including low abundance proteins. Overall, PASS-DIA is a novel DIA approach for use as a discovery tool that outperforms both conventional DDA and DIA experiments to provide additional protein information. We believe that the PASS-DIA method is an important strategy for discovery-type studies when deeper proteome characterization is required.
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http://dx.doi.org/10.1021/acs.analchem.0c02513DOI Listing
November 2020

High-quality nuclear genome for Sarcoptes scabiei-A critical resource for a neglected parasite.

PLoS Negl Trop Dis 2020 10 1;14(10):e0008720. Epub 2020 Oct 1.

Cell and Molecular Biology Department, Infectious Diseases Program, QIMR Berghofer Medical Research Institute, Brisbane, Queensland, Australia.

The parasitic mite Sarcoptes scabiei is an economically highly significant parasite of the skin of humans and animals worldwide. In humans, this mite causes a neglected tropical disease (NTD), called scabies. This disease results in major morbidity, disability, stigma and poverty globally and is often associated with secondary bacterial infections. Currently, anti-scabies treatments are not sufficiently effective, resistance to them is emerging and no vaccine is available. Here, we report the first high-quality genome and transcriptomic data for S. scabiei. The genome is 56.6 Mb in size, has a a repeat content of 10.6% and codes for 9,174 proteins. We explored key molecules involved in development, reproduction, host-parasite interactions, immunity and disease. The enhanced 'omic data sets for S. scabiei represent comprehensive and critical resources for genetic, functional genomic, metabolomic, phylogenetic, ecological and/or epidemiological investigations, and will underpin the design and development of new treatments, vaccines and/or diagnostic tests.
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http://dx.doi.org/10.1371/journal.pntd.0008720DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7591027PMC
October 2020

Mutation-Specific and Common Phosphotyrosine Signatures of G12D and G13D Alleles.

J Proteome Res 2021 01 27;20(1):670-683. Epub 2020 Oct 27.

Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, United States.

is one of the most frequently mutated genes across all cancer subtypes. Two of the most frequent oncogenic mutations observed in patients result in glycine to aspartic acid substitution at either codon 12 (G12D) or 13 (G13D). Although the biochemical differences between these two predominant mutations are not fully understood, distinct clinical features of the resulting tumors suggest involvement of disparate signaling mechanisms. When we compared the global phosphotyrosine proteomic profiles of isogenic colorectal cancer cell lines bearing either G12D or G13D mutation, we observed both shared as well as unique signaling events induced by the two mutations. Remarkably, while the G12D mutation led to an increase in membrane proximal and adherens junction signaling, the G13D mutation led to activation of signaling molecules such as nonreceptor tyrosine kinases, MAPK kinases, and regulators of metabolic processes. The importance of one of the cell surface molecules, MPZL1, which was found to be hyperphosphorylated in G12D cells, was confirmed by cellular assays as its knockdown led to a decrease in proliferation of G12D but not G13D expressing cells. Overall, our study reveals important signaling differences across two common mutations and highlights the utility of our approach to systematically dissect subtle differences between related oncogenic mutants and potentially lead to individualized treatments.
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http://dx.doi.org/10.1021/acs.jproteome.0c00587DOI Listing
January 2021

Mutational Landscape of Esophageal Squamous Cell Carcinoma in an Indian Cohort.

Front Oncol 2020 20;10:1457. Epub 2020 Aug 20.

Institute of Bioinformatics, International Technology Park, Bangalore, India.

Esophageal squamous cell carcinoma (ESCC) is the most common histological subtype of esophageal cancer in India. Cigarette smoking and chewing tobacco are known risk factors associated with ESCC. However, genomic alterations associated with ESCC in India are not well-characterized. In this study, we carried out exome sequencing to characterize the mutational landscape of ESCC tumors from subjects with a varied history of tobacco usage. Whole exome sequence analysis of ESCC from an Indian cohort revealed several genes that were mutated or had copy number changes. ESCC from tobacco chewers had a higher frequency of C:G > A:T transversions and 2-fold enrichment for mutation signature 4 compared to smokers and non-users of tobacco. Genes, such as , and were found to be frequently mutated in Indian cohort. Mutually exclusive mutation patterns were observed in ---, and - gene pairs. Recurrent amplifications were observed in 3q22-3q29, 11q13.3-q13.4, 7q22.1-q31.1, and 8q24 regions. Approximately 53% of tumors had genomic alterations in making this pathway a promising candidate for targeted therapy. In conclusion, we observe enrichment of mutation signature 4 in ESCC tumors from patients with a history of tobacco chewing. This is likely due to direct exposure of esophagus to tobacco carcinogens when it is chewed and swallowed. Genomic alterations were frequently observed in PIK3CA-AKT pathway members independent of the history of tobacco usage. PIK3CA pathway can be potentially targeted in ESCC which currently has no effective targeted therapeutic options.
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http://dx.doi.org/10.3389/fonc.2020.01457DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7469928PMC
August 2020

Urinary glycoproteomic profiling of non-muscle invasive and muscle invasive bladder carcinoma patients reveals distinct N-glycosylation pattern of CD44, MGAM, and GINM1.

Oncotarget 2020 Aug 25;11(34):3244-3255. Epub 2020 Aug 25.

Institute of Bioinformatics, International Technology Park, Bangalore 560066, India.

Clinical management of bladder carcinomas (BC) remains a major challenge and demands comprehensive multi-omics analysis for better stratification of the disease. Identification of patients on risk requires identification of signatures predicting prognosis risk of the patients. Understanding the molecular alterations associated with the disease onset and progression could improve the routinely used diagnostic and therapy procedures. In this study, we investigated the aberrant changes in N-glycosylation pattern of proteins associated with tumorigenesis as well as disease progression in bladder cancer. We integrated and compared global N-glycoproteomic and proteomic profile of urine samples from bladder cancer patients at different clinicopathological stages (non-muscle invasive and muscle-invasive patients [ = 5 and 4 in each cohort]) with healthy subjects ( = 5) using SPEG method. We identified 635 N-glycopeptides corresponding to 381 proteins and 543 N-glycopeptides corresponding to 326 proteins in NMIBC and MIBC patients respectively. Moreover, we identified altered glycosylation in 41 NMIBC and 21 MIBC proteins without any significant change in protein abundance levels. In concordance with the previously published bladder cancer cell line N-glycoproteomic data, we also observed dysregulated glycosylation in ECM related proteins. Further, we identified distinct N-glycosylation pattern of CD44, MGAM, and GINM1 between NMIBC and MIBC patients, which may be associated with disease progression in bladder cancer. These aberrant protein glycosylation events would provide a novel approach for bladder carcinoma diagnosis and further define novel mechanisms of tumor initiation and progression.
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http://dx.doi.org/10.18632/oncotarget.27696DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7456616PMC
August 2020

Integrated genomic analysis reveals mutated ELF3 as a potential gallbladder cancer vaccine candidate.

Nat Commun 2020 08 24;11(1):4225. Epub 2020 Aug 24.

Department of Surgery, Division of Hepatobiliary and Pancreatic Surgery, Seoul National University Hospital, Seoul National University College of Medicine, Seoul, 08826, South Korea.

Gallbladder cancer (GBC) is an aggressive gastrointestinal malignancy with no approved targeted therapy. Here, we analyze exomes (n = 160), transcriptomes (n = 115), and low pass whole genomes (n = 146) from 167 gallbladder cancers (GBCs) from patients in Korea, India and Chile. In addition, we also sequence samples from 39 GBC high-risk patients and detect evidence of early cancer-related genomic lesions. Among the several significantly mutated genes not previously linked to GBC are ETS domain genes ELF3 and EHF, CTNNB1, APC, NSD1, KAT8, STK11 and NFE2L2. A majority of ELF3 alterations are frame-shift mutations that result in several cancer-specific neoantigens that activate T-cells indicating that they are cancer vaccine candidates. In addition, we identify recurrent alterations in KEAP1/NFE2L2 and WNT pathway in GBC. Taken together, these define multiple targetable therapeutic interventions opportunities for GBC treatment and management.
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http://dx.doi.org/10.1038/s41467-020-17880-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7445288PMC
August 2020
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