RESUMO
We report an autosomal recessive, multi-organ tumor predisposition syndrome, caused by bi-allelic loss-of-function germline variants in the base excision repair (BER) gene MBD4. We identified five individuals with bi-allelic MBD4 variants within four families and these individuals had a personal and/or family history of adenomatous colorectal polyposis, acute myeloid leukemia, and uveal melanoma. MBD4 encodes a glycosylase involved in repair of G:T mismatches resulting from deamination of 5'-methylcytosine. The colorectal adenomas from MBD4-deficient individuals showed a mutator phenotype attributable to mutational signature SBS1, consistent with the function of MBD4. MBD4-deficient polyps harbored somatic mutations in similar driver genes to sporadic colorectal tumors, although AMER1 mutations were more common and KRAS mutations less frequent. Our findings expand the role of BER deficiencies in tumor predisposition. Inclusion of MBD4 in genetic testing for polyposis and multi-tumor phenotypes is warranted to improve disease management.
Assuntos
Polipose Adenomatosa do Colo , Neoplasias Colorretais , Neoplasias Uveais , Polipose Adenomatosa do Colo/genética , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Endodesoxirribonucleases/genética , Predisposição Genética para Doença , Células Germinativas/patologia , Mutação em Linhagem Germinativa/genética , Humanos , Neoplasias Uveais/genéticaRESUMO
A large amount of circumstantial evidence has accumulated suggesting that Toll-like receptor (TLR) signals are involved in driving chronic lymphocytic leukemia (CLL) cell proliferation, but direct in vivo evidence for this is still lacking. We have now further addressed this possibility by pharmacologically inhibiting or genetically inactivating the TLR pathway in murine CLL and human Richter syndrome (RS) patient-derived xenograft (PDX) cells. Surprisingly, we show that pharmacologic inhibition of TLR signaling by treatment with an IRAK1/4 inhibitor delays the growth of the transplanted malignant cells in recipient mice, but genetic inactivation of the same pathway by CRISPR/Cas9-mediated disruption of IRAK4 or its proximal adaptor MyD88 has no effect. We further show that treatment with the IRAK1/4 inhibitor results in depletion of macrophages and demonstrate that these cells can support the survival and enhance the proliferation of both murine Eµ-TCL1 leukemia and human RS cells. We also show that genetic disruption of the B-cell receptor (BCR) by CRISPR/Cas9 editing of the immunoglobulin M constant region gene inhibits the growth of human RS-PDX cells in vivo, consistent with our previous finding with murine Eµ-TCL1 leukemia cells. Finally, we show that genetic disruption of IRAK4 does not result in negative selection of human CLL cell lines xenografted in immunodeficient mice. The obtained data suggest that TLR signals are unlikely to represent a major driver of CLL/RS cell proliferation and provide further evidence that signals from macrophages and the BCR promote the growth and survival of CLL and RS cells in vivo.
Assuntos
Leucemia Linfocítica Crônica de Células B , Linfoma Difuso de Grandes Células B , Humanos , Camundongos , Animais , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Quinases Associadas a Receptores de Interleucina-1/genética , Modelos Animais de Doenças , Receptores de Antígenos de Linfócitos B/metabolismo , Receptores Toll-Like , Macrófagos/metabolismoRESUMO
B-cell receptor (BCR) signals play a critical role in the pathogenesis of chronic lymphocytic leukemia (CLL), but their role in regulating CLL cell proliferation has still not been firmly established. Unlike normal B cells, CLL cells do not proliferate in vitro upon engagement of the BCR, suggesting that CLL cell proliferation is regulated by other signals from the microenvironment, such as those provided by Toll-like receptors or T cells. Here, we report that BCR engagement of human and murine CLL cells induces several positive regulators of the cell cycle, but simultaneously induces the negative regulators CDKN1A, CDKN2A, and CDKN2B, which block cell-cycle progression. We further show that introduction of genetic lesions that downregulate these cell-cycle inhibitors, such as inactivating lesions in CDKN2A, CDKN2B, and the CDKN1A regulator TP53, leads to more aggressive disease in a murine in vivo CLL model and spontaneous proliferation in vitro that is BCR dependent but independent of costimulatory signals. Importantly, inactivating lesions in CDKN2A, CDKN2B, and TP53 frequently co-occur in Richter syndrome (RS), and BCR stimulation of human RS cells with such lesions is sufficient to induce proliferation. We also show that tumor cells with combined TP53 and CDKN2A/2B abnormalities remain sensitive to BCR-inhibitor treatment and are synergistically sensitive to the combination of a BCR and cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitor both in vitro and in vivo. These data provide evidence that BCR signals are directly involved in driving CLL cell proliferation and reveal a novel mechanism of Richter transformation.
Assuntos
Transformação Celular Neoplásica , Inibidor de Quinase Dependente de Ciclina p15 , Inibidor p16 de Quinase Dependente de Ciclina , Leucemia Linfocítica Crônica de Células B , Receptores de Antígenos de Linfócitos B , Transdução de Sinais , Proteína Supressora de Tumor p53 , Animais , Linhagem Celular Tumoral , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/imunologia , Inibidor de Quinase Dependente de Ciclina p15/genética , Inibidor de Quinase Dependente de Ciclina p15/imunologia , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/imunologia , Humanos , Leucemia Linfocítica Crônica de Células B/genética , Leucemia Linfocítica Crônica de Células B/imunologia , Camundongos , Receptores de Antígenos de Linfócitos B/genética , Receptores de Antígenos de Linfócitos B/imunologia , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/imunologiaRESUMO
Chronic lymphocytic leukemia (CLL) is characterized by the existence of subsets of patients with (quasi)identical, stereotyped B-cell receptor (BcR) immunoglobulins. Patients in certain major stereotyped subsets often display remarkably consistent clinicobiological profiles, suggesting that the study of BcR immunoglobulin stereotypy in CLL has important implications for understanding disease pathophysiology and refining clinical decision-making. Nevertheless, several issues remain open, especially pertaining to the actual frequency of BcR immunoglobulin stereotypy and major subsets, as well as the existence of higher-order connections between individual subsets. To address these issues, we investigated clonotypic IGHV-IGHD-IGHJ gene rearrangements in a series of 29 856 patients with CLL, by far the largest series worldwide. We report that the stereotyped fraction of CLL peaks at 41% of the entire cohort and that all 19 previously identified major subsets retained their relative size and ranking, while 10 new ones emerged; overall, major stereotyped subsets had a cumulative frequency of 13.5%. Higher-level relationships were evident between subsets, particularly for major stereotyped subsets with unmutated IGHV genes (U-CLL), for which close relations with other subsets, termed "satellites," were identified. Satellite subsets accounted for 3% of the entire cohort. These results confirm our previous notion that major subsets can be robustly identified and are consistent in relative size, hence representing distinct disease variants amenable to compartmentalized research with the potential of overcoming the pronounced heterogeneity of CLL. Furthermore, the existence of satellite subsets reveals a novel aspect of repertoire restriction with implications for refined molecular classification of CLL.
Assuntos
Cadeias Pesadas de Imunoglobulinas/genética , Região Variável de Imunoglobulina/genética , Leucemia Linfocítica Crônica de Células B/genética , Frequência do Gene , Rearranjo Gênico , Humanos , Hipermutação Somática de ImunoglobulinaRESUMO
The high recurrence rate and invasive diagnostic and monitoring methods in bladder cancer (BCa) clinical management require the development of new non-invasive molecular tools for early detection, particularly for low-grade and low-stage BCa as well as for risk stratification. By using an in-solution digestion method and label-free data-independent LC-MS/MS coupled with ion mobility, we profiled the BCa tissues from initiation to advanced stages and confidently identified and quantified 1619 proteins (≥2 peptides). A statistically significant difference in abundance (Anova ≤ 0.05) showed 494 proteins. Significant correlation with stage with steady up or down with BCa stages showed 15 proteins. Testing of NNMT, GALK1, and HTRA1 in urine samples showed excellent diagnostic potential for NNMT and GALK1 with AUC of 1.000 (95% CI: 1.000-1.000; p < 0.0001) and 0.801 (95% CI: 0.655-0.947; p < 0.0001), respectively. NNMT and GALK1 also showed very good potential in discriminating non-invasive low-grade from invasive high-grade BCa with AUC of 0.763 (95% CI: 0.606-0.921; p = 0.001) and 0.801 (95% CI: 0.653-0.950; p < 0.0001), respectively. The combination of NNMT and GALK1 increased prognostic accuracy (AUC = 0.813). Our results broaden the range of potential novel candidates for non-invasive BCa diagnosis and prognosis.
Assuntos
Proteômica , Neoplasias da Bexiga Urinária , Humanos , Biomarcadores Tumorais/urina , Cromatografia Líquida , Detecção Precoce de Câncer , Nicotinamida N-Metiltransferase , Espectrometria de Massas em Tandem , Neoplasias da Bexiga Urinária/metabolismoRESUMO
BACKGROUND: There are little evidences about the therapeutic efficacy of different lipid-lowering agents in the reduction of elevated lipoprotein(a) [Lp(a)]. OBJECTIVE: testing the effect of different lipid-lowering agents on elevated Lp(a). METHODS: prospective interventional study performed in patients with CAD, or high CAD risk, with Lp(a), >50 mg/dL. Lp(a), total cholesterol (C), HDL-C, LDL-C, triglycerides (TGs), apolipoprotein (Apo) A1, Apo B, enzymes of myocyte and hepatic injury were comparatively analyzed between 4 lipid-lowering strategies: rosuvastatin (R group) 40 mg, atorvastatin (A group) 80 mg, atorvastatin 40 mg add-on micronized fenofibrate (A+F group), and atorvastatin 40 mg add-on 1 g extended-release niacin (A+ERN group). Comparison was made for their therapeutic efficacy on Lp(a), and safety. RESULTS: 87 patients with mean Lp(a) 94.6 ± 39.6 mg/dL were analyzed. Groups: 25 patients in the R, 22 in the A, 20 in the A+F and 20 in A+ERN group. Significant reduction in all lipid fractions in all treatment groups was reported after 6 months. The average reduction of Lp(a) was 15.9 ± 21.0 mg/dL, with: 18.2 ± 24.8 (P = 0.001) in the R group, 17.3 ± 10.4 (P = 0.001) in A+F, 19.5 ± 10.9 (P = 0.001) in A+ERN and the lowest in the A group (11.24 ± 22.91, P = 0.032). No adverse effects were observed in any of the treatment groups. CONCLUSIONS: When compared with atorvastatin, it seems that rosuvastatin can achieve more significant decrease of Lp(a).The efficacy of the second one can be increased by adding fibrate or ERN.
Assuntos
Atorvastatina/uso terapêutico , Doença da Artéria Coronariana/prevenção & controle , Hipolipemiantes/uso terapêutico , Lipoproteína(a)/sangue , Rosuvastatina Cálcica/uso terapêutico , Apolipoproteína A-I/sangue , Atorvastatina/administração & dosagem , Atorvastatina/efeitos adversos , Colesterol/sangue , Doença da Artéria Coronariana/sangue , Quimioterapia Combinada , Feminino , Fenofibrato/administração & dosagem , Fenofibrato/efeitos adversos , Fenofibrato/uso terapêutico , Humanos , Hipolipemiantes/administração & dosagem , Hipolipemiantes/efeitos adversos , Masculino , Pessoa de Meia-Idade , Niacina/administração & dosagem , Niacina/efeitos adversos , Niacina/uso terapêutico , Estudos Prospectivos , Rosuvastatina Cálcica/administração & dosagem , Rosuvastatina Cálcica/efeitos adversos , Resultado do Tratamento , Triglicerídeos/sangueRESUMO
BACKGROUND: It has been shown that single nucleotide polymorphisms (SNPs) near the interleukin 28B (IL28B) gene were associated with sustained virological response following standard antivirological treatment of chronic hepatitis C. OBJECTIVES: The aim of the study was to evaluate the association between SNPs near the IL28B gene and response to the treatment of chronic hepatitis C in hemodialysis patients. PATIENTS AND METHODS: The study group included 24 hemodialysis patients with chronic hepatitis C routinely treated with pegylated interferon α-2 a. HCV genotype 1 was the cause of chronic hepatitis C in all study participants. Sustained virological response was determined by an assay with a sensitivity of 20 IU/mL, 6 months after completion of the antivirological treatment. The genotyping of the three most widely studied IL28B gene polymorphisms (rs12979860, rs8099917, and rs12980275) was performed in all study participants. RESULTS: Sustained virological response was achieved in 50% of the treated patients. The treatment response was significantly associated with the CC genotype of rs12979860, TT genotype of rs8099917, and AA genotype of rs12980275 (p = 0.003, p = 0.009, and p = 0.012, respectively). CONCLUSIONS: The three most widely studied SNPs near the IL28B gene were associated with sustained virological response following antivirological treatment of chronic hepatitis C in hemodialysis patients.
Assuntos
Antivirais/uso terapêutico , Hepacivirus/genética , Hepatite C Crônica/tratamento farmacológico , Interferon-alfa/uso terapêutico , Interleucinas/genética , Polietilenoglicóis/uso terapêutico , Ribavirina/uso terapêutico , Adulto , Quimioterapia Combinada , Feminino , Frequência do Gene , Genótipo , Humanos , Interferons , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Proteínas Recombinantes/uso terapêutico , Diálise Renal/métodos , Carga ViralRESUMO
Occult hepatitis C virus (HCV) infection is characterized by the presence of HCV RNA in the liver cells or peripheral blood mononuclear cells of the patients whose serum samples test negative for HCV RNA, with or without presence of HCV antibodies. The present study reviews the existing literature on the persistence of occult hepatitis C virus infection, with description of the clinical characteristics and methods for identification of occult hepatitis C. Occult hepatitis C virus infection was detected in patients with abnormal results of liver function tests of unknown origin, with HCV antibodies and HCV RNA negativity in serum, and also in patients with spontaneous or treatment-induced recovery from hepatitis C. The viral replication in the liver cells and/or peripheral blood mononuclear cells was present in all clinical presentations of occult hepatitis C. The peripheral blood mononuclear cells represent an extra-hepatic site of HCV replication. The reason why HCV RNA was not detectable in the serum of patients with occult hepatitis C, could be the low number of circulating viral particles not detectable by the diagnostic tests with low sensitivity. It is uncertain whether occult hepatitis C is a different clinical entity or just a form of chronic hepatitis C virus infection. Data accumulated over the last decade demonstrated that an effective approach to the diagnosis of HCV infection would be the implementation of more sensitive HCV RNA diagnostic assays, and also, examination of the presence of viral particles in the cells of the immune system.
Assuntos
Hepacivirus/imunologia , Hepatite C/imunologia , Hepacivirus/genética , Hepatite C/sangue , Hepatite C/diagnóstico , Anticorpos Anti-Hepatite C/sangue , Humanos , RNA Viral/sangue , Diálise RenalRESUMO
Loss-of-function mutations in NFKBIE, which encodes for the NF-κB inhibitor IκBε, are frequent in chronic lymphocytic leukemia (CLL) and certain other B-cell malignancies and have been associated with accelerated disease progression and inferior responses to chemotherapy. Using in vitro and in vivo murine models and primary patient samples, we now show that NFKBIE-mutated CLL cells are selected by microenvironmental signals that activate the NF-κB pathway and induce alterations within the tumor microenvironment that can allow for immune escape, including expansion of CD8+ T-cells with an exhausted phenotype and increased PD-L1 expression on the malignant B-cells. Consistent with the latter observations, we find increased expression of exhaustion markers on T-cells from patients with NFKBIE-mutated CLL. In addition, we show that NFKBIE-mutated murine CLL cells display selective resistance to ibrutinib and report inferior outcomes to ibrutinib treatment in NFKBIE-mutated CLL patients. These findings suggest that NFKBIE mutations can contribute to CLL progression through multiple mechanisms, including a bidirectional crosstalk with the microenvironment and reduced sensitivity to BTK inhibitor treatment.
Assuntos
Adenina , Leucemia Linfocítica Crônica de Células B , Mutação , Piperidinas , Evasão Tumoral , Microambiente Tumoral , Animais , Humanos , Camundongos , Adenina/análogos & derivados , Adenina/farmacologia , Linfócitos T CD8-Positivos/imunologia , Leucemia Linfocítica Crônica de Células B/genética , Leucemia Linfocítica Crônica de Células B/patologia , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Leucemia Linfocítica Crônica de Células B/imunologia , NF-kappa B/metabolismo , Piperidinas/farmacologia , Piperidinas/uso terapêutico , Pirazóis/farmacologia , Pirazóis/uso terapêutico , Pirimidinas/farmacologia , Pirimidinas/uso terapêutico , Evasão Tumoral/genética , Microambiente Tumoral/imunologiaRESUMO
Manganese superoxide dismutase (MnSOD) is the most effective antioxidant enzyme in mitochondria and protects cells from reactive oxygen species-induced oxidative damage. The aim of this study was to investigate the association between MnSOD Ala-9 Val gene polymorphism and prostate cancer (PCa) risk in Turkish men with prostate cancer. 33 patients with PCa and 81 control individuals were included in the study. We observed an association between MnSOD Ala/Ala frequency and a higher PCa risk. In addition, we found that the increased risk of early-onset PCa (under age of 65) in the men homozygous for Ala allele was higher than the men homozygous for Val allele. However, we determined that MnSOD Ala-9 Val genotype was not associated with the aggressiveness of the disease. The results of our study suggest that MnSOD Ala/Ala genotype may influence on early-onset of PCa patients, but no effect on subsequent development of the disease in Turkish men. However, our study has a limitation that is small numbers of individuals for cases and controls. Therefore, the presented study limited our statistical power to fully investigate the gene polymorphism on cancer risk.
Assuntos
Estudos de Associação Genética , Neoplasias da Próstata/genética , Superóxido Dismutase/genética , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Neoplasias da Próstata/patologia , Fatores de Risco , TurquiaRESUMO
The advancements in proteomics have provided a better understanding of the functionality of apolipoproteins and lipoprotein-associated proteins, with the HDL lipoprotein fraction being the most studied. The focus of this study was to evaluate the HDL proteome in dyslipidemic subjects without an established cardiovascular disease, as well as to test whether rosuvastatin treatment alters the HDL proteome. Patients with primary hypercholesterolemia or mixed dyslipidemia were assigned to 20 mg/day rosuvastatin and blood samples were drawn at study entry and after 12 weeks of treatment. A label-free LC-MS/MS protein profiling was conducted, coupled with bioinformatics analysis. Sixty-nine HDL proteins were identified, belonging to four main biological function clusters: lipid transport and metabolism; platelet activation, degranulation, and aggregation, wound response and wound healing; immune response; inflammatory and acute phase response. Five HDL proteins showed statistically significant differences in the abundance (Anova ≤ 0.05), before and after rosuvastatin treatment. Platelet factor 4 variant (PF4V1), Pregnancy-specific beta-1-glycoprotein 2 (PSG2), Profilin-1 (PFN1) and Keratin type II cytoskeletal 2 epidermal (KRT2) showed decreased expressions, while Integrin alpha-IIb (ITGA2B) showed an increased expression after treatment with rosuvastatin. The ELISA validation of PFN1 segregated the subjects into responders and non-responders, as PFN1 levels after rosuvastatin were shown to mostly depend on the subjects' inflammatory phenotype. Findings from this study introduce novel insights into the HDL proteome and statin pleiotropism.
Assuntos
Inibidores de Hidroximetilglutaril-CoA Redutases , Proteoma , Feminino , Gravidez , Humanos , Rosuvastatina Cálcica/uso terapêutico , Cromatografia Líquida , Espectrometria de Massas em Tandem , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , ProfilinasRESUMO
Genomic epidemiology has proven to be a useful tool for investigating pandemic outbreaks and tracking pathogen spread and evolution. This study describes the circulation of SARS-CoV-2 strains in N. Macedonia during a period of one year, encompassing three waves of the COVID-19 pandemic. A certain percentage (2-3%) of positive cases were continuously selected and analyzed by whole genome sequencing (WGS) technology. Using this approach, a total of 337 SARS-CoV-2 genomes were sequenced and 26 different lineages belonging to 7 clades were detected. During the first wave of the pandemic, the most dominant lineage was B.1.1, followed by B.1.1.70, which became the most dominant in the second wave. The B.1.1.7 lineage completely overpassed all other variants in the third wave. Our study strengthens the notion that the progression of COVID-19 pandemic is associated with emergence of new SARS-CoV-2 variants with increased virulence. The measure of the impact of this viral dynamic on the spread of the pandemic should be evaluated in association with other factors that might influence the transmission.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , COVID-19/epidemiologia , COVID-19/virologia , Pandemias , SARS-CoV-2/genéticaRESUMO
Reviewing the past and the present status of personalized medicine, the hope and promise from several years ago was critically compared to what is really achieved to tailor the drug treatment according to the patient's individuality. The basis for consideration is what we know about the variant of the disease the patient is suffering from, and about the mechanisms influencing the plasma concentration-time profile, such as activity of metabolizing enzymes and transporters. In cancer treatment, drugs are currently selected regarding molecular properties of the cancer tissue, eg, expressing receptors such as HER2 receptor. Currently diagnostic tests are available allowing to detect somatic cell mutations that can be used to guide drug selection. Unfortunately, tumor heterogeneity and developing resistance by further mutations may limit the success of the therapy determined by molecular diagnostics. The present status can be described that in drug kinetics we know the influencing factors and we understand the mechanisms. However, only in a few cases the genetic background is the main determinant of kinetic variability, and environmental and other factors have an additional important role. Therefore, much more has to be done before we can translate the accumulating knowledge into a benefit for the patient. Only then, we can speak about personalized medicine.
Assuntos
Farmacogenética , Farmacocinética , Medicina de Precisão , Biomarcadores , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Doenças Cardiovasculares/tratamento farmacológico , Doenças Cardiovasculares/genética , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Aprovação de Drogas , Testes Genéticos , Genômica , Humanos , Terapia de Alvo Molecular , Medicina de Precisão/economiaRESUMO
AIM: To implement molecular analysis in the clinical diagnosis and management of Lynch syndrome (LS). METHODS: We analyzed the mutations in MLH1 and MSH2 in the selected LS families from the Republic of Macedonia. RESULTS: We performed the very first genetic identification of LS families and characterized a novel mutation. The novel nonsense germline point mutation c.392C>G in the codon 131 of MLH1(S131X) was identified as the underlying genetic cause of LS in three families. The haplotype analysis suggested a founder effect of this mutation in our population. CONCLUSION: We expect to detect the mutation in other LS patients from the region, and recommend cost-effective screening for this mutation by restriction fragment length polymorphism-polymerase chain reaction or DNA sequencing of MLH1 Exon5 prior to full genetic testing in all LS suspects of Macedonian ancestry.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Neoplasias Colorretais Hereditárias sem Polipose/genética , Mutação em Linhagem Germinativa , Mutação de Sentido Incorreto , Proteínas Nucleares/genética , Mutação Puntual , Sequência de Bases , Análise Mutacional de DNA , Feminino , Efeito Fundador , Haplótipos , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase Multiplex , Proteína 1 Homóloga a MutL , Proteína 2 Homóloga a MutS/genética , Linhagem , Estudos Prospectivos , República da Macedônia do Norte , Análise de Sequência de DNARESUMO
As the currently available tests for the clinical management of prostate cancer (PCa) are still far from providing precise diagnosis and risk stratification, the identification of new molecular marker(s) remains a pertinent clinical need. Candidate PCa biomarkers from the published proteomic comparative studies of prostate tissue (2002-2020) were collected and systematically evaluated. AZGP1, MDH2, FABP5, ENO1, GSTP1, GSTM2, and EZR were chosen for further evaluation in the urine of 85 PCa patients and controls using ELISA. Statistically significant differences in protein levels between PCa and BPH showed FABP5 (p = 0.019) and ENO1 (p = 0.015). A biomarker panel based on the combination of FABP5, ENO1, and PSA provided the highest accuracy (AUC = 0.795) for PCa detection. The combination of FABP5, EZR, AZGP1, and MDH2 showed AUC = 0.889 in PCa prognosis, with 85.29% of the samples correctly classified into low and high Gleason score (GS) groups. The addition of PSA to the panel slightly increased the AUC to 0.914. AZGP1, FABP5, and EZR showed significant correlation with GS, stage, and percentage of positive biopsy cores. Although validation using larger patient cohorts will be necessary to establish the credibility of the proposed biomarker panels in a clinical context, this study opens a way for the further testing of more high-quality proteomics biomarkers, which could ultimately add value to the clinical management of PCa.
RESUMO
Statins have shown anti-inflammatory pleiotropic effects in subjects with/at risk of cardiovascular disease. The aim of this study was to evaluate the inflammatory/immunomodulatory properties of rosuvastatin in subjects at low-to-moderate cardiovascular risk. Data was collected from patients' records, physical examination and blood sampling. Subjects were assigned to rosuvastatin 20 mg per day. Rosuvastatin significantly decreased C-reactive protein (p = 0.045), and increased vascular endothelial growth factor (p = 0.004) and epidermal growth factor (p = 0.009). A multivariate analysis identified total cholesterol (p = 0.027) and vascular endothelial growth factor (p = 0.011) to be independently associated with rosuvastatin treatment. Given beneficial/harmful role of growth factors, vascular endothelial growth factor (VEGF) and epidermal growth factor (EGF), in cardiovascular disease, one would suggest the need for routine monitoring of growth factor levels, especially in patients on long-term statin therapy.
Assuntos
Doenças Cardiovasculares , Inibidores de Hidroximetilglutaril-CoA Redutases , Rosuvastatina Cálcica , Humanos , Anti-Inflamatórios/uso terapêutico , Doenças Cardiovasculares/prevenção & controle , Família de Proteínas EGF , Fatores de Risco de Doenças Cardíacas , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Fatores de Risco , Rosuvastatina Cálcica/uso terapêutico , Fator A de Crescimento do Endotélio VascularRESUMO
Understanding molecular mechanisms that underpin azoospermia and discovery of biomarkers that could enable reliable, non-invasive diagnosis are highly needed. Using label-free data-independent LC-MS/MS acquisition coupled with ion mobility, we compared the FFPE testicular proteome of patients with obstructive (OA) and non-obstructive azoospermia (NOA) subtypes hypospermatogenesis (Hyp) and Sertoli cell-only syndrome (SCO). Out of 2044 proteins identified based on ≥2 peptides, 61 proteins had the power to quantitatively discriminate OA from NOA and 30 to quantitatively discriminate SCO from Hyp and OA. Among these, H1-6, RANBP1 and TKTL2 showed superior potential for quantitative discrimination among OA, Hyp and SCO. Integrin signaling pathway, adherens junction, planar cell polarity/convergent extension pathway and Dectin-1 mediated noncanonical NF-kB signaling were significantly associated with the proteins that could discriminate OA from NOA. Comparison with 2 transcriptome datasets revealed 278 and 55 co-differentially expressed proteins/genes with statistically significant positive correlation. Gene expression analysis by qPCR of 6 genes (H1-6, RANBP1, TKTL2, TKTL1, H2BC1, and ACTL7B) with the highest discriminatory power on protein level and the same regulation trend with transcriptomic datasets, confirmed the proteomics results. In summary, our results suggest some underlying pathways in azoospermia and broaden the range of potential novel candidates for diagnosis. SIGNIFICANCE: Using a comparative proteomics approach on testicular tissue we have identified several pathways associated with azoospermia and a number of testis-specific and germ cell-specific proteins that have the potential to pinpoint the type of spermatogenesis failure. Furthermore, comparison with transcriptomics datasets based on genome-wide gene expression analyses of human testis specimens from azoospermia patients identified proteins that could discriminate between obstructive and non-obstructive azoospermia subtypes on both protein and mRNA levels. Up to our knowledge, this is the first integrated comparative analysis of proteomics and transcriptomics data from testicular tissues. We believe that the data from our study contributes significantly to increase the knowledge of molecular mechanisms of azoospermia and pave the way for new investigations in regards to non-invasive diagnosis.
Assuntos
Azoospermia , Oligospermia , Azoospermia/diagnóstico , Biomarcadores/metabolismo , Cromatografia Líquida , Humanos , Masculino , Oligospermia/genética , Oligospermia/metabolismo , Proteômica , Espectrometria de Massas em Tandem , Testículo/metabolismo , Transcetolase/metabolismoRESUMO
Background: Although vast clinical evidence supports the oxidative CVD hypothesis, little is known on the effects of statins on LDL/HDL oxidative functionality. Therefore, the aim of this study was to evaluate the antioxidative effects of rosuvastatin by monitoring the susceptibility of LDL to oxidation and the antioxidative HDL potential in low-to-moderate CV risk subjects. Methods: 40 adult ambulatory patients (aged 53.8±10.9 years, 27 women and 13 men) were included in the study. Data was collected from patients' records, physical examination, and blood sampling. Subjects were prescribed rosuvastatin at 20mg/day. Traditional risk-factors/indicators, lipid parameters, inflammatory/immune markers, LDL susceptibility to oxidation and HDL antioxidative potential were monitored and statistically analyzed with t-test, Chi-square test, one-way ANOVA, Mann-Whitney, and Kruskal-Wallis tests. Multivariate logistic regression analyses were made. Results were considered significant when p≤0.05. Results: 67% of the patients showed lower susceptibility of LDL to oxidation after rosuvastatin treatment (p=0.03), with no significant effect on baseline LDL oxidation and lag time. All three LDL oxidative indices were seen to be dependent on the subjects' lipid profile, hemoglobin levels and the IL-1α and IL-8 pro-inflammatory marker levels. 53% of the patients showed higher HDL antioxidative capacity after treatment, but without statistical significance (p=0.07). Increased antioxidative potential of HDL with rosuvastatin treatment was more likely in males (OR=9.350; p=0.010), and subjects achieving lower post-treatment CV relative risk levels (higher CV risk reduction) (OR=0.338; p=0.027). Conclusions: This study suggests the need of a comprehensive approach when investigating oxidative stress and LDL/HDL functions, especially in low-to-moderate CVD risk subjects.
Assuntos
Doenças Cardiovasculares , Fluorbenzenos , Adulto , Antioxidantes/uso terapêutico , Biomarcadores , Doenças Cardiovasculares/prevenção & controle , HDL-Colesterol , Feminino , Fluorbenzenos/efeitos adversos , Fatores de Risco de Doenças Cardíacas , Humanos , Masculino , Pirimidinas/efeitos adversos , Fatores de Risco , Rosuvastatina Cálcica/efeitos adversos , Sulfonamidas/uso terapêuticoRESUMO
Carnitine palmitoyltransferase II deficiency (CPT II) is an autosomal recessive inherited disorder of long-chain fatty acid oxidation in the mitochondrial matrix, resulting in an inability to utilize fat for energy in cells. The most frequent myopathic form occurs in young adults and is associated with recurrent episodes of exercise-induced rhabdomyolysis. The myopathic form is caused by the Ser113Leu mutation of the CPT II gene. Rarely, massive rhabdomyolysis could be complicated by acute kidney injury (AKI), cardiomyopathy, and respiratory insufficiency. We present a case of an 18-year old male with myalgia, muscular weakness, and dark-colored urine after prolonged exercise and a recent mildSARS-CoV-2infection. Massive rhabdomyolysis was diagnosed with markedly increased serum concentrations of myoglobin and creatine kinase, with normal kidney function. The patient experienced two similar episodes in the years 2017 and 2018, with rhabdomyolysis and AKI treated with hemodialysis. After excluding autoimmune and infectious diseases as causes of recurrent rhabdomyolysis, the patient was genetically tested and Ser113Leu mutation of the CPT II gene was confirmed. When a patient presents with myalgia and dark-colored urine triggered by minor physical activities, genetic testing for possible CPT II deficiency should be initiated. TheSARS-CoV-2infection could be a factor that triggers the occurrence of rhabdomyolysis and aggravates the severity of the attack in patients with CPT II deficiency.
Assuntos
COVID-19 , Carnitina O-Palmitoiltransferase , Humanos , Adolescente , Carnitina O-Palmitoiltransferase/genética , COVID-19/complicações , SARS-CoV-2 , Exercício FísicoRESUMO
The proteomics of formalin-fixed, paraffin-embedded (FFPE) samples has advanced significantly during the last two decades, but there are many protocols and few studies comparing them directly. There is no consensus on the most effective protocol for shotgun proteomic analysis. We compared the in-solution digestion with RapiGest and Filter Aided Sample Preparation (FASP) of FFPE prostate tissues stored 7 years and mirroring fresh frozen samples, using two label-free data-independent LC-MS/MS acquisitions. RapiGest identified more proteins than FASP, with almost identical numbers of proteins from fresh and FFPE tissues and 69% overlap, good preservation of high-MW proteins, no bias regarding isoelectric point, and greater technical reproducibility. On the other hand, FASP yielded 20% fewer protein identifications in FFPE than in fresh tissue, with 64-69% overlap, depletion of proteins >70 kDa, lower efficiency in acidic and neutral range, and lower technical reproducibility. Both protocols showed highly similar subcellular compartments distribution, highly similar percentages of extracted unique peptides from FFPE and fresh tissues and high positive correlation between the absolute quantitation values of fresh and FFPE proteins. In conclusion, RapiGest extraction of FFPE tissues delivers a proteome that closely resembles the fresh frozen proteome and should be preferred over FASP in biomarker and quantification studies. SIGNIFICANCE: Here we analyzed the performance of two sample preparation methods for shotgun proteomic analysis of FFPE tissues to give a comprehensive overview of the obtained proteomes and the resemblance to its matching fresh frozen counterparts. These findings give us better understanding towards competent proteomics analysis of FFPE tissues. It is hoped that it will encourage further assessments of available protocols before establishing the most effective protocol for shotgun proteomic FFPE tissue analysis.