MolDy: molecular dynamics simulation made easy.

MOTIVATION: Molecular dynamics (MD) is a computational experiment that is crucial for understanding the structure of biological macro and micro molecules, their folding, and the inter-molecular interactions. Accurate knowledge of these structural features is the cornerstone in drug development and elucidating macromolecules functions. The open-source GROMACS biomolecular MD simulation program is recognized as a reliable and frequently used simulation program for its precision. However, the user requires expertise, and scripting skills to carrying out MD simulations. RESULTS: We have developed an end-to-end interactive MD simulation application, MolDy for Gromacs. This front-end application provides a customizable user interface integrated with the Python and Perl-based logical backend connecting the Linux shell and Gromacs software. The tool performs analysis and provides the user with simulation trajectories and graphical representations of relevant biophysical parameters. The advantages of MolDy are (i) user-friendly, does not requiring the researcher to have prior knowledge of Linux; (ii) easy installation by a single command; (iii) freely available for academic research; (iv) can run with minimum configuration of operating systems; (v) has valid default prefilled parameters for beginners, and at the same time provides scope for modifications for expert users. AVAILABILITY AND IMPLEMENTATION: MolDy is available freely as compressed source code files with user manual for installation and operation on GitHub: https://github.com/AIBResearchMolDy/Moldyv01.git and on https://aibresearch.com/innovations.

Identification of potential biomarkers to predict organ morbidity in COVID-19: A repository based proteomics perspective.

SARS-CoV-2 causes substantial extrapulmonary manifestations in addition to pulmonary disease. Some of the major organs affected are cardiovascular, hematological and thrombotic, renal, neurological, and digestive systems. These types of muti-organ dysfunctions make it difficult and challenging for clinicians to manage and treat COVID-19 patients. The article focuses to identify potential protein biomarkers that can flag various organ systems affected in COVID-19. Publicly reposited high throughput proteomic data from human serum (HS), HEK293T/17 (HEK) and Vero E6 (VE) kidney cell culture were downloaded from ProteomeXchange consortium. The raw data was analyzed in Proteome Discoverer 2.4 to delineate the complete list of proteins in the three studies. These proteins were analyzed in Ingenuity Pathway Analysis (IPA) to associate them to various organ diseases. The shortlisted proteins were analyzed in MetaboAnalyst 5.0 to shortlist potential biomarker proteins. These were then assessed for disease-gene association in DisGeNET and validated by Protein-protein interactome (PPI) and functional enrichment studies (GO_BP, KEGG and Reactome pathways) in STRING. Protein profiling resulted in shortlisting 20 proteins in 7 organ systems. Of these 15 proteins showed at least 1.25-fold changes with a sensitivity and specificity of 70%. Association analysis further shortlisted 10 proteins with a potential association with 4 organ diseases. Validation studies established possible interacting networks and pathways affected, confirmingh the ability of 6 of these proteins to flag 4 different organ systems affected in COVID-19 disease. This study helps to establish a platform to seek protein signatures in different clinical phenotypes of COVID-19. The potential biomarker candidates that can flag organ systems involved are: (a) Vitamin K-dependent protein S and Antithrombin-III for hematological disorders; (b) Voltage-dependent anion-selective channel protein 1 for neurological disorders; (c) Filamin-A for cardiovascular disorder and, (d) Peptidyl-prolyl cis-trans isomerase A and Peptidyl-prolyl cis-trans isomerase FKBP1A for digestive disorders.

Structural Modelling of Platelet Activating Factor Acetyl Hydrolase in Leishmania donovani, Trypanosoma cruzi, and Trypanosoma brucei: Implications on Therapeutics for Leishmaniasis, Chagas Disease, and Sleeping Sickness.

Purpose: Leishmaniasis, Chagas disease, and sleeping sickness are caused by protozoa Leishmania donovani, Trypanosoma cruzi, and Trypanosoma brucei, respectively. Platelet activating factor acetyl hydrolase (PAF-AH) is an inflammatory protein implicated in pathogenesis of these three infections, thereby making them attractive drug targets. Methods: PAF-AH sequences were retrieved from UniProt and aligned using Clustal Omega. Homologous models of parasitic proteins were built based on crystal structure of human PAF-AH and validated using PROCHECK server. Volumes of substrate-binding channel were calculated using the ProteinsPlus program. High throughput virtual screening using Glide program in Schrodinger was done with ZINC drug library against parasitic PAF-AH enzymes. Complexes with best hits were energy-minimized and subjected to 100 ns molecular dynamic simulation and analyzed. Results: PAF-AH enzyme sequences from protozoa Leishmania donovani, Trypanosoma cruzi, Trypanosoma brucei, and human have a minimum of 34% sequence similarity with each other. Corresponding structures show a globular conformation consisting of twisted ß-pleated sheets, flanked by α-helices on either side. Catalytic triad of serine-histidine-aspartate is conserved. Substrate-binding channel residues are conserved to an extent, with a lower channel volume in human as compared to target enzymes. Drug screening resulted in identification of three molecules that had better affinities than the substrate to the target enzymes. These molecules fulfill Lipinski's rules for drug likeness and also bind with less affinity to the human counterpart, thereby establishing a high selective index. Conclusion: Structures of PAF-AH from protozoan parasites and humans belong to the same family of enzymes and have a similar three-dimensional fold. However, they show subtle variations in residue composition, secondary structure composition, substrate-binding channel volume, and conformational stability. These differences result in certain specific molecules being potent inhibitors of the target enzymes while simultaneously having weaker binding to human homologue.

Label-Free Proteomics of Oral Mucosa Tissue to Identify Potential Biomarkers That Can Flag Predilection of Precancerous Lesions to Oral Cell Carcinoma: A Preliminary Study.

Oral squamous cell carcinomas are mostly preceded by precancerous lesions such as leukoplakia and erythroplakia. Our study is aimed at identifying potential biomarker proteins in precancerous lesions of leukoplakia and erythroplakia that can flag their transformation to oral cancer. Four biological replicate samples from clinical phenotypes of healthy control, leukoplakia, erythroplakia, and oral carcinoma were annotated based on clinical screening and histopathological evaluation of buccal mucosa tissue. Differentially expressed proteins were delineated using a label-free quantitative proteomic experiment done on an Orbitrap Fusion Tribrid mass spectrometer in three technical replicate sets of samples. Raw files were processed using MaxQuant version 2.0.1.0, and downstream analysis was done via Perseus version 1.6.15.0. Validation included functional annotation based on biological processes and pathways using the ClueGO plug-in of Cytoscape. Hierarchical clustering and principal component analysis were performed using the ClustVis tool. Across control, leukoplakia, and cancer, L-lactate dehydrogenase A chain, plectin, and WD repeat-containing protein 1 were upregulated, whereas thioredoxin 1 and spectrin alpha chain, nonerythrocytic 1 were downregulated. Across control, erythroplakia, and cancer, L-lactate dehydrogenase A chain was upregulated whereas aldehyde dehydrogenase 2, peroxiredoxin 1, heat shock 70 kDa protein 1B, and spectrin alpha chain, nonerythrocytic 1 were downregulated. We found that proteins involved in leukoplakia were associated with alteration in cytoskeletal disruption and glycolysis, while in erythroplakia, they were associated with alteration in response to oxidative stress and glycolysis across phenotypes. Hierarchical clustering subgrouped half of precancerous samples under the main branch of the control and the remaining half under carcinoma. Similarly, principal component analysis identified segregated clusters of control, precancerous lesions, and cancer, but erythroplakia phenotypes, in particular, overlapped more with the cancer cluster. Qualitative and quantitative protein signatures across control, precancer, and cancer phenotypes explain possible functional outcomes that dictate malignant transformation to oral carcinoma.

Development of sensitive magnetic nanoparticle assisted rapid sandwich assay(s-MARSA) to monitor Parkinson's disease and Schizophrenia pharmacotherapy.

Parkinson's disease and Schizophrenia fall under low dopamine neurodegenerative and high dopamine psychiatric disorders respectively. Pharmacological interventions to correct mid-brain dopamine concentrations sometimes overshoots the physiological dopamine levels leading to psychosis in Parkinson's disease patients and, extra-pyramidal symptoms in schizophrenia patients. Currently no validated method is available to monitor side effects in such patients, Apolipoprotein E is one of the CSF biomarkers identified in the recent past that shows an inverse relation to mid-brain dopamine concentration. In this study, we have developed s-MARSA for the detection of Apolipoprotein E from ultra-small volume (2 µL) of CSF. s-MARSA exhibits a broad detection range (5 fg mL-1 to 4 µg mL-1) with a better detection limit and could be performed within an hour utilizing only a small volume of CSF sample. The values measured by s-MARSA strongly correlates with the values measured by ELISA. Our method has advantages over ELISA in having a lower detection limit, a broader linear detection range, shorter analysis time, and requiring a low volume of CSF samples. The developed s-MARSA method holds promise for the detection of Apolipoprotein E with clinical utility for monitoring pharmacotherapy of Parkinson's and Schizophrenia patients.

Mechanism of apoptosis activation by Curcumin rescued mutant p53Y220C in human pancreatic cancer.

The mutant p53Y220C (mutp53Y220C) is frequently observed in numerous tumors, including pancreatic cancer. The mutation creates a crevice in the DNA binding core domain and makes p53 a thermally unstable non-functional protein that assists tumor progression and confers resistance to chemotherapeutic drugs. Restoring mutp53 function to its wild type by selectively targeting this crevice with small molecules is a pivotal strategy to promote apoptosis. In this study, we have shown through different biophysical and cell-based studies that curcumin binds and rescues mutp53Y220C to an active wild-type conformation and restores its apoptotic transcription function in BxPC-3-pancreatic cancer cells. In addition, the curcumin-rescued-p53Y220C (CRp53) showed significant hyperphosphorylation at Ser15, Ser20, and acetylation at Lys382 with an 8-fold increase in transcription activity in the BxPC-3 cell lines. We also observed that the active CRp53 escapes Mdm2-mediated proteasomal degradation and the majority of the proteins were localized inside the nucleus with an increased half-life and transcription restoration compared to untreated BxPC-3 cells. By label-free proteomics analysis, we observed that upon curcumin treatment almost 227 proteins were dysregulated with the majority of them being transcriptional targets of p53. Based on our studies, it reflects that apoptosis in pancreatic cancer cells is mediated by curcumin-rescued mutant p53Y220C.

Comparative Structural Analysis of Human ACE2 Receptor with Spike Protein of SARS-CoV-2 Variants: Implications to Understand Infectivity of the Virus.

Purpose: Spike protein on SARS-CoV-2 virus plays an integral part during infection as cell entry depends on binding of this protein to human ACE2 receptor. Understanding of infectivity by these variants necessitates a comparative structural analysis of complexes of spike protein-receptor binding domain (RBD) of these variants to receptor. Methodology: Wild type SARS-CoV-2 spike protein sequence was retrieved from the UniProt database, and mutations of five variants at receptor binding domain were manually incorporated and aligned using Clustal Omega. Crystal structure complexes of human ACE2 receptor with spike protein RBD domain of SARS-CoV-2 variants of wild type, α, ß, and δ were extracted from the RCSB database. Wild type SARS-CoV-2 complex with receptor was used as template to generate model complexes of receptor with spike protein RBD of γ and omicron variants through WinCoot program. These were energy minimized and validated and molecular dynamic simulation was performed using Desmond simulation program. Results: Mutations are distributed across the entire length of RBD, but the maximum number of mutations are seen at 11 positions within binding interface motifs of six variant sequences. Interface of spike protein RBDs with human ACE2-receptor shows different mix of hydrogen bonded and ionic interactions. Alpha and ß variants have few interactions, while γ and δ variants have higher number of interactions compared to wild type variant. Omicron variant, with 10 polar interactions including two ionic bonds, has the highest binding energy. Conclusion: Different mutations on RBD of spike protein results in varying quantity and quality of interactions, thereby affecting potency of each variant. Variations in binding are due to interactions of mutant residues and induced conformational changes on loops of RBDs. Variants α and ß have a low potency, while, γ, δ, and omicron have a higher potency. These results correlate with viral infectivity and place clinical observations in the right perspective.

2D-DIGE based urinary proteomics and functional enrichment studies to reveal novel Mycobacterium tuberculosis and human protein biomarker candidates for pulmonary tuberculosis.

In the absence of a sensitive and specific diagnostic modality capable of detecting all forms of tuberculosis (TB), proteomics may identify specific Mycobacterium tuberculosis (M.tb) proteins in urine, with a potential as biomarkers. To identify candidate biomarkers for TB, proteome profile of urine from pulmonary TB patients was compared with non-disease controls (NDC) and disease controls (DC, Streptococcus pneumonia infected patients) using a combination of two-dimensional difference gel electrophoresis (2D-DIGE) and liquid chromatography tandem mass spectrometry (LCMS/MS). Eleven differentially expressed host proteins and Eighteen high abundant M.tb proteins were identified. Protein-protein interactome (PPI) and functional enrichment analyses like Gene Ontologies, Reactome pathway etc. demonstrated that the human proteins mainly belong to extracellular space and show physiological pathways for immune response and hematological disorders. Whereas, M.tb proteins belong to the cell periphery, plasma membrane and cell wall, and demonstrated catalytic, nucleotide binding and ATPase activities along with other functional processes. The study findings provide valuable inputs about the biomarkers of TB and shed light on the probable disease consequences as an outcome of the bacterial pathogenicity.

iTRAQ proteomics of sentinel lymph nodes for identification of extracellular matrix proteins to flag metastasis in early breast cancer.

Patients with early breast cancer are affected by metastasis to axillary lymph nodes. Metastasis to these nodes is crucial for staging and quality of surgery. Sentinel Lymph Node Biopsy that is currently used to assess lymph node metastasis is not effective. This necessitates identification of biomarkers that can flag metastasis. Early stage breast cancer patients were recruited. Surgical resection of breast was followed by identification of sentinel lymph nodes. Fresh frozen section biopsy was used to assign metastatic and non-metastatic sentinel lymph nodes. Discovery phase included iTRAQ proteomics coupled with mass spectrometric analysis to identify differentially expressed proteins. Data is available via ProteomeXchange with identifier PXD027668. Validation was done by bioinformatic analysis and ELISA. There were 2398 unique protein groups and 109 differentially expressed proteins comparing metastatic and non-metastatic lymph nodes. Forty nine proteins were up-regulated, and sixty proteins that were down regulated in metastatic group. Bioinformatic analysis showed ECM-receptor interaction pathways to be implicated in lymph node metastasis. ELISA confirmed up-regulation of ECM proteins in metastatic lymph nodes. ECM proteins have requisite parameters to be developed as a diagnostic tool to assess status of sentinel lymph nodes to guide surgical intervention in early breast cancer.

Structural modeling of Omicron spike protein and its complex with human ACE-2 receptor: Molecular basis for high transmissibility of the virus.

Omicron is a new variant of SARS-CoV-2, which is currently infecting people around the world. Spike glycoprotein, an important molecule in pathogenesis of infection has been modeled and the interaction of its Receptor Binding Domain with human ACE-receptor has been analysed by simulation studies. Structural analysis of Omicron spike glycoprotein shows the 30 mutations to be distributed over all domains of the trimeric protein, wherein the mutant residues are seen to be participating in higher number of intra-molecular interactions including two salt bridges emanating from mutant residues thereby stabilizing their conformation, as compared to wild type. Complex of Receptor Binding Domain (RBD) with human ACE-2 receptor shows seven mutations at interacting interface comprising of two ionic interactions, eight hydrogen bonds and seven Van der Waals interactions. The number and quality of these interactions along with other binding biophysical parameters suggests more potency of RBD domain to the receptor as compared to the wild type counterpart. Results of this study explains the high transmissibility of Omicron variant of SARS-CoV-2 that is currently observed across the world.

Structural Modeling of Wild and Mutant Forms of Human Plasma Platelet Activating Factor-Acetyl Hydrolase Enzyme.

PURPOSE: To investigate the structural features of wild and mutant forms of the pPAF-AH enzyme that are responsible for coronary artery disease. METHODS: Mutant variants of human pPAF-AH having either V279F, Q281R, or both were modelled and evaluated for stereo chemical and structural correctness. The 3D coordinates of substrate PAF were retrieved from the PubChem database was solvated and minimized on Discovery Studio, and docked to the wild and mutant enzyme models. The top docked pose complex was refined by MD simulation. RESULTS: pPAF-AH model comprises of 420 amino acids in a α/ß-hydrolase fold that contains a substrate-binding hydrophobic channel with an active site pocket having a catalytic triad of Ser273, Asp296 and His351. Mutations at positions 279 and 281 are opposite one another on the middle of 12 residues long H5 helix that forms the hydrophobic core of the enzyme. V279F causes a tilt on the axis of the mutation bearing helix to avoid steric clashes with the hydrophobic residues on the ß-sheets adjacent to it, inducing subtle conformational changes on the H5-ß8 loop, ß8 sheet, and the loop bearing Asp296. A cascade of conformational changes induces a change in the orientation of His351 resulting in loss of hydrogen bonded interaction with catalytic Ser273. Q281R causes a shortening of H5 and ß8, which induces conformational changes of the loops bearing Ser273 and Asp296, respectively. Simultaneous conformational changes of secondary structural elements result in the flipping of His351 causing a break in the catalytic triad. Also, there is a compromise in the substrate-binding area and volume in the mutants resulting in loss of binding to its substrate. CONCLUSION: Mutant enzymes show changes at the site of the mutation, secondary motif conformations and global structural conformations that adversely affect the active site, decrease substrate channel volume and decrease stability, thereby affecting enzymatic function.

Human Secretary Phospholipase A2 Mutations and Their Clinical Implications.

Phospholipases A2 (PLA2s) belong to a superfamily of enzymes responsible for hydrolysis of the sn-2 fatty acids of membrane phospholipids to release arachidonic acid. PLA2s are the rate limiting enzyme for the downstream synthesis of prostaglandins and leukotrienes that are the main mediators of inflammation. The extracellular forms of this enzyme are also called the secretary phospholipase A2 (sPLA2) and are distributed extensively in most of the tissues in the human body. Their integral role in inflammatory pathways has been the primary reason for the extensive research on this molecule. The catalytic mechanism of sPLA2 is initiated by a histidine/aspartic acid/calcium complex within the active site. Though they are known to have certain housekeeping functions, certain mutations of sPLA2 are known to be implicated in causation of certain pathologies leading to diseases such as atherosclerosis, cardiovascular diseases, benign fleck retina, neurodegeneration, and asthma. We present an overview of human sPLA2 and a comprehensive compilation of the mutations that result in various disease phenotypes. The study not only helps to have a holistic understanding of human sPLA2 mutations and their clinical implications, but is also a useful platform to initiate research pertaining to structure-function relationship of the mutations to develop effective therapies for management of these diseases.

Proteomics of Sentinel Lymph Nodes in Early Breast Cancer for Identification of Thymidylate Synthase as a Potential Biomarker to Flag Metastasis: A Preliminary Study.

INTRODUCTION: Breast cancer is the second most common cancer in women across the world. Some of the patients who present in the early stage of disease are affected by metastasis to the axillary group of lymph nodes. The first among this group that is affected is called as sentinel lymph node, and its diagnosis is crucial for the staging of cancer thereby dictating the type of surgical therapy. Therefore, the sentinel lymph node status provides the most relevant information to the surgeon and patient prognosis. The expanded utilization of breast conservation surgery has declined the morbidity associated with mastectomy and axillary lymph node surgery. Recent interest is, therefore, centered on techniques that allow accurate assessment of the sentinel lymph node metastasis. A current procedure such as sentinel lymph node biopsy (SLNB) that is used to assess axillary lymph node metastasis is neither specific nor sensitive, and besides, it is time-consuming. OBJECTIVE: To compare the protein profiles between metastatic and non-metastatic lymph nodes to identify a biomarker that can flag lymph node metastasis. MATERIALS AND METHODS: Women with early breast cancer were screened using mammography imaging and recruited to the study. Surgical resection was done to remove the breast tissue, and sentinel lymph node was identified using fluorescein and methylene blue tracer. Lymph node was sliced, and one set was sent for histopathology, which was considered the gold standard to assess the metastatic status of the lymph node. One set of slices was taken for proteomic experiments. Proteins were labelled with fluorescent cyanine tags and were subjected to difference gel electrophoresis experiment. Differentially expressed spots that had at least a twofold relative ratio and consistent pattern across three sets of biological replicate experiments were marked. Gel spots were trypsin digested and identified on mass spectrometry machine. Validation study was done by Western blot experiment on the same set of samples. RESULTS: Thymidylate synthase has a twofold higher expression in the metastatic sentinel lymph nodes as compared to non-metastatic lymph nodes in early breast cancer patients. CONCLUSION: Differential in gel expression proteomics is an ideal platform for the identification of potential protein biomarker candidates that can differentiate metastatic from non-metastatic lymph nodes in early breast cancer. The identification of thymidylate synthase offers a scope to develop an on-table diagnostic kit to assess the status of sentinel lymph nodes during mastectomy procedure to guide surgical management of axillary lymph nodes in early breast cancer.

Insights into SARS-CoV-2 genome, structure, evolution, pathogenesis and therapies: Structural genomics approach.

The sudden emergence of severe respiratory disease, caused by a novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has recently become a public health emergency. Genome sequence analysis of SARS-CoV-2 revealed its close resemblance to the earlier reported SARS-CoV and Middle East respiratory syndrome coronavirus (MERS-CoV). However, initial testing of the drugs used against SARS-CoV and MERS-CoV has been ineffective in controlling SARS-CoV-2. The present study highlights the genomic, proteomic, pathogenesis, and therapeutic strategies in SARS-CoV-2 infection. We have carried out sequence analysis of potential drug target proteins in SARS-CoV-2 and, compared them with SARS-CoV and MERS viruses. Analysis of mutations in the coding and non-coding regions, genetic diversity, and pathogenicity of SARS-CoV-2 has also been done. A detailed structural analysis of drug target proteins has been performed to gain insights into the mechanism of pathogenesis, structure-function relationships, and the development of structure-guided therapeutic approaches. The cytokine profiling and inflammatory signalling are different in the case of SARS-CoV-2 infection. We also highlighted possible therapies and their mechanism of action followed by clinical manifestation. Our analysis suggests a minimal variation in the genome sequence of SARS-CoV-2, may be responsible for a drastic change in the structures of target proteins, which makes available drugs ineffective.

Cerebrospinal Fluid Proteomics For Identification Of α2-Macroglobulin As A Potential Biomarker To Monitor Pharmacological Therapeutic Efficacy In Dopamine Dictated Disease States Of Parkinson's Disease And Schizophrenia.

AIM: Parkinson's disease and schizophrenia are clinical end points of dopaminergic deficit and excess, respectively, in the mid-brain. In accordance, current pharmacological interventions aim to restore normal dopamine levels, the overshooting of which culminates in adverse effects which results in psychotic symptoms in Parkinson's disease and extra-pyramidal symptoms in schizophrenia. Currently, there are no laboratory assays to assist treatment decisions or help foresee these drug side-effect outcomes. Therefore, the aim was to discover a protein biomarker that had a varying linear expression across the clinical dopaminergic spectrum. MATERIALS AND METHODS: iTRAQ-based proteomic experiments along with mass spectrometric analysis was used for comparative proteomics using cerebrospinal fluid (CSF). CSF fluid was collected from 36 patients with Parkinson's disease, 15 patients with urological diseases that served as neurological controls, and seven schizophrenic patients with hallucinations. Validation included ELISA and pathway analysis to highlight the varying expression and provide plausible molecular pathways for differentially expressed proteins in the three clinical phenotypes. RESULTS: Protein profiles were delineated in CSF from Parkinson's disease patients, neurological control and schizophrenia, respectively. Ten of the proteins that were identified had a linear relationship across the dopaminergic spectrum. α-2-Macroglobulin showed to be having high statistical significance on inter-group comparison on validation studies using ELISA. CONCLUSIONS: Non-gel-based proteomic experiments are an ideal platform to discover potential biomarkers that can be used to monitor pharmaco-therapeutic efficacy in dopamine-dictated clinical scenarios. α-2 Macroglobulin is a potential biomarker to monitor pharmacological therapy in Parkinson's disease and schizophrenia.

Evaluation of α-synuclein and apolipoprotein E as potential biomarkers in cerebrospinal fluid to monitor pharmacotherapeutic efficacy in dopamine dictated disease states of Parkinson's disease and schizophrenia.

BACKGROUND AND OBJECTIVE: Dopamine plays an important role in the disease pathology of Parkinson's disease and schizophrenia. These two neuropsychiatric disorders represent disease end points of the dopaminergic spectrum where Parkinson's disease represents dopamine deficit and schizophrenia represents dopamine hyperactivity in the mid-brain. Therefore, current treatment strategies aim to restore normal dopamine levels. However, during treatment patients develop adverse effects due to overshooting of physiological levels of dopamine leading to psychosis in Parkinson's disease, and extrapyramidal symptoms in schizophrenia. Absence of any laboratory tests hampers modulation of pharmacotherapy. Apolipoprotein E and α-synuclein have an important role in the neuropathology of these two diseases. The objective of this study was to evaluate cerebrospinal fluid (CSF) concentrations of apolipoprotein E and α-synuclein in patients with these two diseases so that they may serve as biomarkers to monitor therapy in Parkinson's disease and schizophrenia. METHODS: Drug-naïve Parkinson's disease patients and Parkinson's disease patients treated with dopaminergic therapy, neurological controls, schizophrenic patients treated with antidopaminergic therapy, and drug-naïve schizophrenic patients were recruited for the study and CSF was collected. Enzyme-linked immunosorbent assays were carried out to estimate the concentrations of apolipoprotein E and α-synuclein. Pathway analysis was done to establish a possible role of these two proteins in various pathways in these two dopamine dictated diseases. RESULTS: Apolipoprotein E and α-synuclein CSF concentrations have an inverse correlation along the entire dopaminergic clinical spectrum. Pathway analysis convincingly establishes a plausible hypothesis for their co-regulation in the pathogenesis of Parkinson's disease and schizophrenia. Each protein by itself or as a combination has encouraging sensitivity and specificity values of more than 55%. CONCLUSION: The dynamic variation of these two proteins along the spectrum is ideal for them to be pursued as pharmacotherapeutic biomarkers in CSF to monitor pharmacological efficacy in Parkinson's disease and schizophrenia.

Chemotherapy Resistance in Advanced Ovarian Cancer Patients.

Ovarian cancer is the seventh most common gynaecologic malignancy seen in women. Majority of the patients with ovarian cancer are diagnosed at the advanced stage making prognosis poor. The standard management of advanced ovarian cancer includes tumour debulking surgery followed by chemotherapy. Various types of chemotherapeutic regimens have been used to treat advanced ovarian cancer, but the most promising and the currently used standard first-line treatment is carboplatin and paclitaxel. Despite improved clinical response and survival to this combination of chemotherapy, numerous patients either undergo relapse or succumb to the disease as a result of chemotherapy resistance. To understand this phenomenon at a cellular level, various macromolecules such as DNA, messenger RNA and proteins have been developed as biomarkers for chemotherapy response. This review comprehensively summarizes the problem that pertains to chemotherapy resistance in advanced ovarian cancer and provides a good overview of the various biomarkers that have been developed in this field.

2D-DIGE as a strategy to identify serum protein biomarkers to monitor pharmacological efficacy in dopamine-dictated states of Parkinson's disease and schizophrenia.

OBJECTIVES: Parkinson's disease and schizophrenia are clinical scenarios that occur due to dopaminergic deficit and hyperactivity in the midbrain, respectively. Current pharmacological interventions for these two diseases therefore aim to restore normal dopamine levels in the midbrain. But during therapy, there is a overshooting of dopamine concentrations that result in hallucinations in Parkinson's disease patients and extra-pyramidal symptoms in schizophrenic patients. This causes a lot of inconvenience to the patents and the clinicians. There are no tests currently available to monitor drug efficacy in these two neuropsychiatric diseases. MATERIALS AND METHODS: Parkinson's disease and schizophrenic naïve patients were recruited. Serum proteins isolated from these two clinical phenotypes were labeled with fluorescent cyanine dyes and analyzed by two-dimensional difference in gel electrophoresis proteomic experiment. Differentially expressed spots that had consistent expression pattern across five sets of biological replicate gels were trypsin digested and subjected to mass spectrometric analysis for protein identification. Validation experiments were done for the identified proteins using antibody-based assay on a patient cohort that included naïve, treated, and those who had side effects. RESULTS: Serum α- and ß-globin chains were identified as differentially expressed proteins having threefold higher expressions in Parkinson's patients as compared to schizophrenia. Interestingly, concentrations of these two proteins had an inverse correlation across clinical phenotypes in the dopaminergic spectrum. RBC contamination as a source for these proteins was ruled out. CONCLUSION: There is a clear association of free serum globin with dopaminergic clinical states. This lays a platform for protein biomarker-based monitoring of pharmacological efficacy in Parkinson's disease and schizophrenia.

Evaluation of Serum Apolipoprotein E as a Potential Biomarker for Pharmacological Therapeutic Efficacy Monitoring in Dopamine Dictated Disease Spectrum of Schizophrenia and Parkinson's disease: A Preliminary Study.

AIM OF THE STUDY: Parkinson's disease and schizophrenia are disease end points of dopaminergic deficit and hyperactivity, respectively, in the mid brain. Accordingly, current medications aim to restore normal dopamine levels, overshooting of which results in adverse effects of psychosis and extra-pyramidal symptoms, respectively. There are currently no available laboratory tests to guide treatment decisions or help predict adverse side effects of the drugs. The aim was to therefore explore the possibility of using apolipoprotein E as a biomarker to monitor pharmacological intervention in dopamine dictated states of Parkinson's disease and schizophrenia for optimum therapy. METHODS: Naïve and treated, Parkinson's disease and schizophrenic patients were recruited from neurology and psychiatry clinics. Serum of healthy volunteers was collected as controls. Serum concentrations of apolipoprotein E was estimated by enzyme-linked immunosorbent assay (ELISA). Pathway analysis was carried out to delineate the interactions of apolipoprotein E in Parkinson's disease and schizophrenia. RESULTS: Apolipoprotein E levels are higher in Parkinson's disease patients as compared with schizophrenic samples (P < .05). Also, post-treatment apolipoprotein E levels in both disease states were at par with levels seen in healthy controls. The interactions of apolipoprotein E validate the results and place the differential expression of the protein in Parkinson's disease and schizophrenia in the right perspective. CONCLUSION: Apolipoprotein E concentration across the dopaminergic spectrum suggests that it can be pursued not only as a potential biomarker in schizophrenia and Parkinson's disease, but can also be an effective tool for clinicians to determine efficacy of drug-based therapy.

Is there an association between enhanced choline and ß-catenin pathway in breast cancer? A pilot study by MR Spectroscopy and ELISA.

Total choline (tCho) was documented as a biomarker for breast cancer diagnosis by in vivo MRS. To understand the molecular mechanisms behind elevated tCho in breast cancer, an association of tCho with ß-catenin and cyclin D1 was evaluated. Hundred fractions from 20 malignant, 10 benign and 20 non-involved breast tissues were isolated. Cytosolic and nuclear expressions of ß-catenin and cyclin D1 were estimated using ELISA. Higher tCho was seen in malignant compared to benign tissues. Malignant tissues showed higher cytosolic and nuclear ß-catenin expressions than benign and non-involved tissues. Within malignant tissues, ß-catenin and cyclin D1 expressions were higher in the nucleus than cytosol. Cyclin D1 expression was higher in the cytosolic fractions of benign and non-involved than malignant tissues. Furthermore, in malignant tissues, tCho showed a positive correlation with the cytosolic and nuclear expression of ß-catenin and cyclin D1 and also a correlation between nuclear expressions of both these proteins was seen. Higher cytosolic ß-catenin expression was seen in progesterone receptor negative than positive patients. Results provide an evidence of correlation between non-invasive biomarker, tCho and the Wnt/ß-catenin pathway. The findings explain the molecular mechanism of tCho elevation which may facilitate exploration of additional therapeutic targets for breast cancer.