Induction of Translational Readthrough on Protein Tyrosine Phosphatases Targeted by Premature Termination Codon Mutations in Human Disease.

Nonsense mutations generating premature termination codons (PTCs) in various genes are frequently associated with somatic cancer and hereditary human diseases since PTCs commonly generate truncated proteins with defective or altered function. Induced translational readthrough during protein biosynthesis facilitates the incorporation of an amino acid at the position of a PTC, allowing the synthesis of a complete protein. This may evade the pathological effect of the PTC mutation and provide new therapeutic opportunities. Several protein tyrosine phosphatases (PTPs) genes are targeted by PTC in human disease, the tumor suppressor PTEN being the more prominent paradigm. Here, using PTEN and laforin as examples, two PTPs from the dual-specificity phosphatase subfamily, we describe methodologies to analyze in silico the distribution and frequency of pathogenic PTC in PTP genes. We also summarize laboratory protocols and technical notes to study the induced translational readthrough reconstitution of the synthesis of PTP targeted by PTC in association with disease in cellular models.

Estimation of carcass chemical composition in beef-on-dairy cattle using dual-energy X-ray absorptiometry (DXA) scans of cold half-carcass or 11th rib cut.

The aim of the present study was to estimate the chemical composition (water, lipid, protein, mineral, and energy contents) of carcasses measured postmortem using dual-energy X-ray absorptiometry (DXA) scans of cold half-carcass or 11th rib cut. One hundred and twenty beef-on-dairy (dam: Swiss Brown, sire: Angus, Limousin, or Simmental) bulls (n = 66), heifers (n = 42), and steers (n = 12) were included in the study. The reference carcass composition measured after grinding, homogenization, and chemical analyses was estimated from DXA variables using simple or multiple linear regressions with model training on 70% (n = 84) and validation on 30% (n = 36) of the observations. In the validation step, the estimates of water and protein masses from the half-carcass (R2 = 0.998 and 0.997; root mean square error of prediction [RMSEP], 1.0 and 0.5 kg, respectively) and 11th rib DXA scans (R2 = 0.997 and 0.996; RMSEP, 1.5 and 0.5 kg, respectively) were precise. Lipid mass was estimated precisely from the half-carcass DXA scan (R2 = 0.990; RMSEP = 1.0 kg) with a slightly lower precision from the 11th rib DXA scan (R2 = 0.968; RMSEP = 1.7 kg). Mineral mass was estimated from half-carcass (R²â€…= 0.975 and RMSEP = 0.3 kg) and 11th rib DXA scans (R2 = 0.947 and RMSEP = 0.4 kg). For the energy content, the R2 values ranged from 0.989 (11th rib DXA scan) to 0.996 (half-carcass DXA scan), and the RMSEP ranged from 36 (half-carcass) to 55 MJ (11th rib). The proportions of water, lipids, and energy in the carcasses were also precisely estimated (R2 ≥ 0.882) using either the half-carcass (RMSEP ≤ 1.0%) or 11th rib-cut DXA scans (RMSEP ≤ 1.3%). Precision was lower for the protein and mineral proportions (R2 ≤ 0.794, RMSEP ≤ 0.5%). The cattle category (sex and breed of sire) effect was observed only in some estimative models for proportions from the 11th rib cut. In conclusion, DXA imaging of either a cold half-carcass or 11th rib cut is a precise method for estimating the chemical composition of carcasses from beef-on-dairy cattle.

Assessment of the water, lipid, protein, mineral, and energy contents of beef carcass allows for an understanding of the bovine growth physiology and is key to determining the carcass's commercial value at the slaughterhouse. Direct measurement of the carcass chemical composition requires postmortem grinding and homogenization of a half-carcass to perform chemical analyses. This reference method is expensive, time-consuming, and destructive of edible meat. The aim of the present study was to develop an alternative and nondestructive method to determine carcass chemical composition based on image scans obtained using dual-energy X-ray absorptiometry (DXA). Equations were calibrated to estimate the carcass composition based on the DXA scans of a whole half-carcass or a single-rib cut in an accurate, precise, fast, and reproducible way. These were established for seven types of beef-on-dairy cattle of different sexes and breeds of sire, which are among the most commonly used in specialized beef-on-dairy fattening production systems worldwide.

Novel anti-PTEN C2 domain monoclonal antibodies to analyse the expression and function of PTEN isoform variants.

PTEN is a major tumor suppressor gene frequently mutated in human tumors, and germline PTEN gene mutations are the molecular diagnostic of PTEN Hamartoma Tumor Syndrome (PHTS), a heterogeneous disorder that manifests with multiple hamartomas, cancer predisposition, and neurodevelopmental alterations. A diversity of translational and splicing PTEN isoforms exist, as well as PTEN C-terminal truncated variants generated by disease-associated nonsense mutations. However, most of the available anti-PTEN monoclonal antibodies (mAb) recognize epitopes at the PTEN C-terminal tail, which may introduce a bias in the analysis of the expression of PTEN isoforms and variants. We here describe the generation and precise characterization of anti-PTEN mAb recognizing the PTEN C2-domain, and their use to monitor the expression and function of PTEN isoforms and PTEN missense and nonsense mutations associated to disease. These anti-PTEN C2 domain mAb are suitable to study the pathogenicity of PTEN C-terminal truncations that retain stability and function but have lost the PTEN C-terminal epitopes. The use of well-defined anti-PTEN mAb recognizing distinct PTEN regions, as the ones here described, will help to understand the deleterious effects of specific PTEN mutations in human disease.

The expression pattern of pyruvate dehydrogenase kinases predicts prognosis and correlates with immune exhaustion in clear cell renal cell carcinoma.

Renal cancer cells constitute a paradigm of tumor cells with a glycolytic reprogramming which drives metabolic alterations favouring cell survival and transformation. We studied the expression and activity of pyruvate dehydrogenase kinases (PDK1-4), key enzymes of the energy metabolism, in renal cancer cells. We analysed the expression, subcellular distribution and clinicopathological correlations of PDK1-4 by immunohistochemistry of tumor tissue microarray samples from a cohort of 96 clear cell renal cell carcinoma (ccRCC) patients. Gene expression analysis was performed on whole tumor tissue sections of a subset of ccRCC samples. PDK2 and PDK3 protein expression in tumor cells correlated with lower patient overall survival, whereas PDK1 protein expression correlated with higher patient survival. Gene expression analysis revealed molecular association of PDK2 and PDK3 expression with PI3K signalling pathway, as well as with T cell infiltration and exhausted CD8 T cells. Inhibition of PDK by dichloroacetate in human renal cancer cell lines resulted in lower cell viability, which was accompanied by an increase in pAKT. Together, our findings suggest a differential role for PDK enzymes in ccRCC progression, and highlight PDK as actionable metabolic proteins in relation with PI3K signalling and exhausted CD8 T cells in ccRCC.

Functional analysis of PTEN variants of unknown significance from PHTS patients unveils complex patterns of PTEN biological activity in disease.

Heterozygous germline mutations in PTEN gene predispose to hamartomas and tumors in different tissues, as well as to neurodevelopmental disorders, and define at genetic level the PTEN Hamartoma Tumor Syndrome (PHTS). The major physiologic role of PTEN protein is the dephosphorylation of phosphatidylinositol (3,4,5)-trisphosphate (PIP3), counteracting the pro-oncogenic function of phosphatidylinositol 3-kinase (PI3K), and PTEN mutations in PHTS patients frequently abrogate PTEN PIP3 catalytic activity. PTEN also displays non-canonical PIP3-independent functions, but their involvement in PHTS pathogeny is less understood. We have previously identified and described, at clinical and genetic level, novel PTEN variants of unknown functional significance in PHTS patients. Here, we have performed an extensive functional characterization of these PTEN variants (c.77 C > T, p.(Thr26Ile), T26I; c.284 C > G, p.(Pro95Arg), P95R; c.529 T > A, p.(Tyr177Asn), Y177N; c.781 C > G, p.(Gln261Glu), Q261E; c.829 A > G, p.(Thr277Ala), T277A; and c.929 A > G, p.(Asp310Gly), D310G), including cell expression levels and protein stability, PIP3-phosphatase activity, and subcellular localization. In addition, caspase-3 cleavage analysis in cells has been assessed using a C2-domain caspase-3 cleavage-specific anti-PTEN antibody. We have found complex patterns of functional activity on PTEN variants, ranging from loss of PIP3-phosphatase activity, diminished protein expression and stability, and altered nuclear/cytoplasmic localization, to intact functional properties, when compared with PTEN wild type. Furthermore, we have found that PTEN cleavage at the C2-domain by the pro-apoptotic protease caspase-3 is diminished in specific PTEN PHTS variants. Our findings illustrate the multifaceted molecular features of pathogenic PTEN protein variants, which could account for the complexity in the genotype/phenotype manifestations of PHTS patients.

Hopes on immunotherapy targeting B7-H3 in neuroblastoma.

Neuroblastoma is one of the most aggressive cancer forms in children, with highly heterogenous clinical manifestations ranging from spontaneous regression to high metastatic capacity. High-risk neuroblastoma has the highest mortality rates of all pediatric cancers, highlighting the urgent need for effective novel therapeutic interventions. B7-H3 immune checkpoint protein is highly expressed in neuroblastoma, and it is involved in oncogenic signaling, tumor cell plasticity, and drug resistance. Immunotherapies based on immune checkpoint inhibition have improved patient survival in several human cancers, and recent reports provide preclinical evidence on the benefits of targeting B7-H3 in neuroblastoma, with emphasis on novel CAR T/NK-cell approaches. Here, we summarize the current status of neuroblastoma targeted therapies, with a focus on B7-H3 as a promising novel immunoregulatory therapeutic target for high-risk neuroblastoma.

Correlation of expression of Major Vault Protein with androgen receptor and immune checkpoint protein B7-H3, and with poor prognosis in prostate cancer.

Prostate cancer diagnosis and early stratification is an important aspect to avoid undertreatment of high-risk prostate cancer patients. Major Vault Protein (MVP) has been proposed as a prognostic biomarker in prostate cancer. PTEN and the immune checkpoint protein B7-H3 interact with MVP and are important in prostate cancer progression and therapy response. We evaluated the expression of MVP by immunohistochemistry of tissue microarray samples from a retrospective cohort consisting of 119 prostate cancer patients. We correlated the protein expression of MVP with clinicopathological characteristics, and protein expression of androgen receptor (AR), PTEN, immune checkpoint proteins B7-H3 and PD-L1. We found MVP to be expressed in 53 % of prostate tumors, and correlated positively with biochemical recurrence (ρ = 0.211/p = 0.021). Furthermore, we found positive correlation of MVP expression with expression of AR (ρ = 0.244/p = 0.009) and the immune checkpoint protein B7-H3 (ρ = 0.200/p = 0.029), but not with PD-L1 (ρ = 0.152/p = 0.117) or PTEN expression (ρ = - 0.034/p = 0.721). Our findings support the notion that expression of MVP is associated with poor prognosis in prostate cancer. The correlation between MVP and immune checkpoint protein B7-H3 in prostate cancer suggests a role for MVP in immunoregulation and drug resistance.

Estimation of empty body and carcass chemical composition of lactating and growing cattle: comparison of imaging, adipose cellularity, and rib dissection methods.

The aim of present study was to compare in vivo and post mortem methods for estimating the empty body (EB) and carcass chemical compositions of Simmental lactating and growing cattle. Indirect methods were calibrated against the direct post mortem reference determination of chemical compositions of EB and carcass, determined after grinding and analyzing the water, lipid, protein, mineral masses, and energy content. The indirect methods applied to 12 lactating cows and 10 of their offspring were ultrasound (US), half-carcass and 11th rib dual-energy X-ray absorptiometry (DXA) scans, subcutaneous and perirenal adipose cell size (ACS), and dissection of the 11th rib. Additionally, three-dimensional (3D) images were captured for 8 cows. Multiple linear regressions with leave-one-out-cross-validations were tested between predictive variables derived from the methods tested, and the EB and carcass chemical compositions. Partial least square regressions were used to estimate body composition with morphological traits measured on 3D images. Body weight (BW) alone estimated the EB and carcass composition masses with a root mean squared error of prediction (RMSEP) for the EB from 1 kg for minerals to 12.4 kg for lipids, and for carcass from 0.9 kg for minerals to 7.8 kg for water. Subcutaneous adipose tissue thickness measured by US was the most accurate in vivo predictor when associated with BW to estimate chemical composition, with the EB lipid mass RMSEP = 11 kg and R 2 = 0.75; carcass water mass RMSEP = 6 kg and R 2 = 0.98; and carcass energy content RMSEP = 236 MJ and R 2 = 0.91. Post mortem, carcass lipid mass was best estimated by half-carcass DXA scan (RMSEP = 2 kg, R 2 = 0.98), 11th rib DXA scan (RMSEP = 3 kg, R 2 = 0.96), 11th rib dissection (RMSEP = 4 kg, R 2 = 0.92), and perirenal ACS (RMSEP = 6 kg, R 2 = 0.79) in this respective order. The results obtained by 11th rib DXA scan were accurate and close to the half-carcass DXA scan with a reduction in scan time. Morphological traits from 3D images delivered promising estimations of the cow EB and carcass chemical component masses with an error less than 13 kg for the EB lipid mass and than 740 MJ for the EB energy. Future research is required to test the 3D imaging method on a larger number of animals to confirm and quantify its interest in estimating body composition in living animals.

Heterogeneous Expression and Subcellular Localization of Pyruvate Dehydrogenase Complex in Prostate Cancer.

Background: Pyruvate dehydrogenase (PDH) complex converts pyruvate into acetyl-CoA by pyruvate decarboxylation, which drives energy metabolism during cell growth, including prostate cancer (PCa) cell growth. The major catalytic subunit of PDH, PDHA1, is regulated by phosphorylation/dephosphorylation by pyruvate dehydrogenase kinases (PDKs) and pyruvate dehydrogenase phosphatases (PDPs). There are four kinases, PDK1, PDK2, PDK3 and PDK4, which can phosphorylate and inactivate PDH; and two phosphatases, PDP1 and PDP2, that dephosphorylate and activate PDH. Methods: We have analyzed by immunohistochemistry the expression and clinicopathological correlations of PDHA1, PDP1, PDP2, PDK1, PDK2, PDK3, and PDK4, as well as of androgen receptor (AR), in a retrospective PCa cohort of patients. A total of 120 PCa samples of representative tumor areas from all patients were included in tissue microarray (TMA) blocks for analysis. In addition, we studied the subcellular localization of PDK2 and PDK3, and the effects of the PDK inhibitor dichloroacetate (DCA) in the growth, proliferation, and mitochondrial respiration of PCa cells. Results: We found heterogeneous expression of the PDH complex components in PCa tumors. PDHA1, PDP1, PDK1, PDK2, and PDK4 expression correlated positively with AR expression. A significant correlation of PDK2 immunostaining with biochemical recurrence and disease-free survival was revealed. In PCa tissue specimens, PDK2 displayed cytoplasmic and nuclear immunostaining, whereas PDK1, PDK3 and PDK4 showed mostly cytoplasmic staining. In cells, ectopically expressed PDK2 and PDK3 were mainly localized in mitochondria compartments. An increase in maximal mitochondrial respiration was observed in PCa cells upon PDK inhibition by DCA, in parallel with less proliferative capacity. Conclusion: Our findings support the notion that expression of specific PDH complex components is related with AR signaling in PCa tumors. Furthermore, PDK2 expression associated with poor PCa prognosis. This highlights a potential for PDH complex components as targets for intervention in PCa.

B7-H4 Immune Checkpoint Protein Affects Viability and Targeted Therapy of Renal Cancer Cells.

Targeted therapy in combination with immune checkpoint inhibitors has been recently implemented in advanced or metastatic renal cancer treatment. However, many treated patients either do not respond or develop resistance to therapy, making alternative immune checkpoint-based immunotherapies of potential clinical benefit for specific groups of patients. In this study, we analyzed the global expression of B7 immune checkpoint family members (PD-L1, PD-L2, B7-H2, B7-H3, B7-H4, B7-H5, B7-H6, and B7-H7) in human renal cancer cells (Caki-1, A-498, and 786-O cell lines) upon treatment with clinically relevant targeted drugs, including tyrosine kinase inhibitors (Axitinib, Cabozantinib, and Lenvatinib) and mTOR inhibitors (Everolimus and Temsirolimus). Gene expression analysis by quantitative PCR revealed differential expression patterns of the B7 family members in renal cancer cell lines upon targeted drug treatments. B7-H4 gene expression was upregulated after treatment with various targeted drugs in Caki-1 and 786-O renal cancer cells. Knocking down the expression of B7-H4 by RNA interference (RNAi) using small interfering RNA (siRNA) decreased renal cancer cell viability and increased drug sensitivity. Our results suggest that B7-H4 expression is induced upon targeted therapy in renal cancer cells and highlight B7-H4 as an actionable immune checkpoint protein in combination with targeted therapy in advanced renal cancer cases resistant to current treatments.

Identification and Functional Analysis of a Novel CTNNB1 Mutation in Pediatric Medulloblastoma.

Medulloblastoma is the primary malignant tumor of the Central Nervous System (CNS) most common in pediatrics. We present here, the histological, molecular, and functional analysis of a cohort of 88 pediatric medulloblastoma tumor samples. The WNT-activated subgroup comprised 10% of our cohort, and all WNT-activated patients had exon 3 CTNNB1 mutations and were immunostained for nuclear ß-catenin. One novel heterozygous CTNNB1 mutation was found, which resulted in the deletion of ß-catenin Ser37 residue (ΔS37). The ΔS37 ß-catenin variant ectopically expressed in U2OS human osteosarcoma cells displayed higher protein expression levels than wild-type ß-catenin, and functional analysis disclosed gain-of-function properties in terms of elevated TCF/LEF transcriptional activity in cells. Our results suggest that the stabilization and nuclear accumulation of ΔS37 ß-catenin contributed to early medulloblastoma tumorigenesis.

Avaliação do consumo alimentar de pacientes em hemodiálise: comparação com recomendações do Kidney Disease Outcome Quality Initiative / Assessment of food consumption in hemodialysis patients: comparison with recommendations from the Kidney Disease Outcome Quality Initiative

Introdução: A análise da ingestão alimentar de pacientes com doença renal crônica (DRC) em tratamento dialítico é eficaz para fornecer informações a fim de auxiliar no diagnóstico nutricional e nortear as condutas dietéticas necessárias. Objetivo: Analisar a adequação do consumo alimentar de acordo com as recomendações publicadas pelo Kidney Disease Outcome Quality Initiative (KDOQI) para macro e micronutrientes, além de avaliar a capacidade antioxidante total da dieta (CATd), de portadores de DRC. Materiais e Métodos: Estudo transversal, com 60 voluntários portadores de DRC em tratamento hemodialítico de um Hospital Universitário de Juiz de Fora ­ MG. A caracterização da amostra foi feita no período de junho de 2019 a fevereiro de 2020, através da aplicação de um Questionário Quantitativo de Frequência Alimentar (QQFA) e de coleta de dados pessoais, clínicos e comorbidades. As análises estatísticas foram conduzidas utilizando-se o software SPSS, versão 20.0. As variáveis numéricas foram apresentadas na forma de média (± desvio-padrão), mediana, mínimo e máximo, enquanto as categóricas em frequência absoluta e relativa. Resultados: Observou-se que 43% e 55% dos pacientes apresentaram um consumo calórico e proteico acima do preconizado pela KDOQI, respectivamente. Para os micronutrientes, foi observado que 77% dos pacientes apresentaram consumo de cálcio abaixo do recomendado e de fósforo 65% maior do que a recomendação atual. Percebeu-se que 67% e 40% dos pacientes apresentaram adequado consumo de sódio e potássio. O perfil lipídico da dieta dos pacientes, demonstrou uma desproporção da razão ômega 6/ômega 3, além de um CATd de 4,05 mmol/dia. Conclusão: Através do presente estudo podemos concluir que o consumo alimentar dos pacientes em hemodiálise avaliados apresenta algumas inadequações em relação às recomendações propostas pela literatura.

Introduction: The analysis of food intake in patients with chronic kidney disease (CKD) undergoing dialysis is effective to provide information to assist in nutritional diagnosis and guide the necessary dietary behaviors. Objective: To analyze the adequacy of food consumption according to the recommendations published by the Kidney Disease Outcome Quality Initiative (KDOQI) for macro and micronutrients, in addition to evaluating the total antioxidant capacity of the diet (CATd) of patients with CKD. Materials and Methods: Cross-sectional study with 60 volunteers with CKD undergoing hemodialysis treatment at a University Hospital in Juiz de Fora ­ MG. The characterization of the sample was carried out from June 2019 to February 2020, through the application of a Quantitative Food Frequency Questionnaire (QQFA) and the collection of personal, clinical and comorbid data. Statistical analyzes were conducted using SPSS software, version 20.0. Numerical variables were presented as mean (± standard deviation), median, minimum and maximum, while categorical variables in absolute and relative frequency. Results: It was observed that 43% and 55% of the patients had a caloric and protein consumption above that recommended by the KDOQI, respectively. For micronutrients, it was observed that 77% of the patients had calcium intake below the recommended level and phosphorus consumption 65% higher than the current recommendation. It was noticed that 67% and 40% of the patients had adequate consumption of sodium and potassium. The lipid profile of the patients' diet showed a disproportion of the omega 6/omega 3 ratio, in addition to a CATd of 4.05 mmol/day. Conclusion: Through the present study we can conclude that the food consumption of the evaluated hemodialysis patients presents some inadequacies in relation to the recommendations proposed by the literature

Protein Tyrosine Phosphatases in Neuroblastoma: Emerging Roles as Biomarkers and Therapeutic Targets.

Neuroblastoma is a type of cancer intimately related with early development and differentiation of neuroendocrine cells, and constitutes one of the pediatric cancers with higher incidence and mortality. Protein tyrosine phosphatases (PTPs) are key regulators of cell growth and differentiation by their direct effect on tyrosine dephosphorylation of specific protein substrates, exerting major functions in the modulation of intracellular signaling during neuron development in response to external cues driving cell proliferation, survival, and differentiation. We review here the current knowledge on the role of PTPs in neuroblastoma cell growth, survival, and differentiation. The potential of PTPs as biomarkers and molecular targets for inhibition in neuroblastoma therapies is discussed.

(Pro)renin Receptor Is a Novel Independent Prognostic Marker in Invasive Urothelial Carcinoma of the Bladder.

(Pro)renin receptor (PRR) is being investigated in several malignancies as it activates pathogenic pathways that contribute to cell proliferation, immunosuppressive microenvironments, and acquisition of aggressive neoplastic phenotypes. Its implication in urothelial cancer (UC) has not been evaluated so far. We retrospectively evaluate the prognostic role of PRR expression in a series of patients with invasive UC treated with radical cystectomy and other clinical and histopathological parameters including p53, markers of immune-checkpoint inhibition, and basal and luminal phenotypes evaluated by tissue microarray. Cox regression analyses using stepwise selection evaluated candidate prognostic factors and disease-specific survival. PRR was expressed in 77.3% of the primary tumors and in 70% of positive lymph nodes. PRR expression correlated with age (p = 0.006) and was associated with lower preoperatively hemoglobin levels. No other statistical association was evidenced with clinical and pathological variables (gender, ASA score, Charlson comorbidity index, grade, pT, pN) or immunohistochemical expressions evaluated (CK20, GA-TA3, CK5/6, CD44, PD-L1, PD-1, B7-H3, VISTA, and p53). PRR expression in primary tumors was associated with worse survival (log-rank, p = 0.008). Cox regression revealed that PRR expression (HR 1.85, 95% CI 1.22-2.8), pT (HR 7.02, 95% CI 2.68-18.39), pN (HR 2.3, 95% CI 1.27-4.19), and p53 expression (HR 1.95, 95% CI 1.1-3.45) were independent prognostic factors in this series. In conclusion, we describe PRR protein and its prognostic role in invasive UC for the first time. Likely mechanisms involved are MAPK/ERK activation, Wnt/ß-catenin signaling, and v-ATPAse function.

Immune checkpoint B7-H3 protein expression is associated with poor outcome and androgen receptor status in prostate cancer.

BACKGROUND: Novel immune checkpoint-based immunotherapies may benefit specific groups of prostate cancer patients who are resistant to other treatments. METHODS: We analyzed by immunohistochemistry the expression of B7-H3, PD-L1/B7-H1, and androgen receptor (AR) in tissue samples from 120 prostate adenocarcinoma patients treated with radical prostatectomy in Spain, and from 206 prostate adenocarcinoma patients treated with radical prostatectomy in Norway. RESULTS: B7-H3 expression correlated positively with AR expression and was associated with biochemical recurrence in the Spanish cohort, but PD-L1 expression correlated with neither of them. Findings for B7-H3 were validated in the Norwegian cohort, where B7-H3 expression correlated positively with Gleason grade, surgical margins, seminal vesicle invasion, and CAPRA-S risk group, and was associated with clinical recurrence. High B7-H3 expression in the Norwegian cohort was also consistent with positive AR expression. CONCLUSION: These results suggest distinct clinical relevance of the two immune checkpoint proteins PD-L1 and B7-H3 in prostate cancer. Our findings highlight B7-H3 as an actionable novel immune checkpoint protein in prostate cancer.

MMADHC premature termination codons in the pathogenesis of cobalamin D disorder: Potential of translational readthrough reconstitution.

Mutations in the MMADHC gene cause cobalamin D disorder (cblD), an autosomal recessive inborn disease with defects in intracellular cobalamin (cbl, vitamin B12) metabolism. CblD patients present methylmalonic aciduria (MMA), homocystinuria (HC), or combined MMA/HC, and usually suffer developmental delay and cognitive deficits. The most frequent MMADHC genetic alterations associated with disease generate MMADHC truncated proteins, in many cases due to mutations that create premature termination codons (PTC). In this study, we have performed a comprehensive and global characterization of MMADHC protein variants generated by all annotated MMADHC PTC mutations in cblD patients, and analyzed the potential of inducible translational PTC readthrough to reconstitute MMADHC biosynthesis. MMADHC protein truncation caused by disease-associated PTC differentially affected the alternative usage of translation initiation sites, protein abundance, and subcellular localization of MMADHC. Aminoglycoside compounds induced translational PTC readthrough of MMADHC truncated variants, allowing the biosynthesis of full-length MMADHC in a PTC-specific manner. Our results suggest that translational PTC readthrough-based interventions could complement current therapies for cblD patients carrying specific MMADHC PTC mutations.

A global analysis of the reconstitution of PTEN function by translational readthrough of PTEN pathogenic premature termination codons.

The PTEN tumor suppressor gene is mutated with high incidence in tumors and in the germline of patients with cancer predisposition or with macrocephaly associated with autism. PTEN nonsense mutations generating premature termination codons (PTC) and producing nonfunctional truncated PTEN proteins are frequent in association with human disease. However, there are no studies addressing the restoration of full-length PTEN proteins from the PTC-mutated PTEN gene by translational readthrough. Here, we have performed a global translational and functional readthrough analysis of the complete collection of PTEN PTC somatic or hereditary mutations found in tumors or in the germline of patients (disease-associated PTEN PTCome), and we set standards for the analysis of the potential of readthrough functional reconstitution in disease-relevant genes. Our analysis indicates that prevalent pathogenic PTEN PTC mutations are susceptible to PTEN functional restoration in response to readthrough-inducing compounds. Comprehensive readthrough analyses of disease-associated PTComes will be valuable tools for the implementation of readthrough-based precision interventions in specific groups of patients.

Estimation of dairy goat body composition: A direct calibration and comparison of eight methods.

The objective was to compare eight methods for estimation of dairy goat body composition, by calibrating against chemical composition (water, lipid, protein, mineral and energy) measured post-mortem. The methods tested on 20 Alpine goats were body condition score (BCS), 3-dimension imaging (3D) automatic assessment of BCS or whole body scan, ultrasound, computer tomography (CT), adipose cell diameter, deuterium oxide dilution space (D2OS) and bioelectrical impedance spectroscopy (BIS). Regressions were tested between predictive variates derived from the methods and empty body (EB) composition. The best equations for estimation of EB lipid mass included BW combined with i) perirenal adipose tissue mass and cell diameter (R2 = 0.95, residual standard deviation, rSD = 0.57 kg), ii) volume of fatty tissues measured by CT (R2 = 0.92, rSD = 0.76 kg), iii) D2OS (R2 = 0.91, rSD = 0.85 kg), and iv) resistance at infinite frequency from BIS (R2 = 0.87, rSD = 1.09 kg). The D2OS combined with BW provided the best equation for EB protein mass (R2 = 0.97, rSD = 0.17 kg), whereas BW alone provided a fair estimate (R2 = 0.92, rSD = 0.25 kg). Sternal BCS combined with BW provided good estimation of EB lipid and protein mass (R2 = 0.80 and 0.95, rSD = 1.27 and 0.22 kg, respectively). Compared to manual BCS, BCS by 3D slightly decreased the precision of the predictive equation for EB lipid (R2 = 0.74, rSD = 1.46 kg), and did not improve the estimation of EB protein compared with BW alone. Ultrasound measurements and whole body 3D imaging methods were not satisfactory estimators of body composition (R2 ≤ 0.40). Further developments in body composition techniques may contribute for high-throughput phenotyping of robustness.