Exome sequencing reveals variants in known and novel candidate genes for severe sperm motility disorders

Publisher Copyright: © The Author(s) 2021. Published by Oxford University Press on behalf of European Society of Human Reproduction and Embryology.STUDY QUESTION: What are the causative genetic variants in patients with male infertility due to severe sperm motility disorders? SUMMARY ANSWER: We identified high confidence disease-causing variants in multiple genes previously associated with severe sperm motility disorders in 10 out of 21 patients (48%) and variants in novel candidate genes in seven additional patients (33%). WHAT IS KNOWN ALREADY: Severe sperm motility disorders are a form of male infertility characterised by immotile sperm often in combination with a spectrum of structural abnormalities of the sperm flagellum that do not affect viability. Currently, depending on the clinical sub-categorisation, up to 50% of causality in patients with severe sperm motility disorders can be explained by pathogenic variants in at least 22 genes. STUDY DESIGN, SIZE, DURATION: We performed exome sequencing in 21 patients with severe sperm motility disorders from two different clinics. PARTICIPANTS/MATERIALS, SETTING, METHOD: Two groups of infertile men, one from Argentina (n = 9) and one from Australia (n = 12), with clinically defined severe sperm motility disorders (motility <5%) and normal morphology values of 0-4%, were included. All patients in the Argentine cohort were diagnosed with DFS-MMAF, based on light and transmission electron microscopy. Sperm ultrastructural information was not available for the Australian cohort. Exome sequencing was performed in all 21 patients and variants with an allele frequency of <1% in the gnomAD population were prioritised and interpreted. MAIN RESULTS AND ROLE OF CHANCE: In 10 of 21 patients (48%), we identified pathogenic variants in known sperm assembly genes: CFAP43 (3 patients); CFAP44 (2 patients), CFAP58 (1 patient), QRICH2 (2 patients), DNAH1 (1 patient) and DNAH6 (1 patient). The diagnostic rate did not differ markedly between the Argentinian and the Australian cohort (55% and 42%, respectively). Furthermore, we identified patients with variants in the novel human candidate sperm motility genes: DNAH12, DRC1, MDC1, PACRG, SSPL2C and TPTE2. One patient presented with variants in four candidate genes and it remains unclear which variants were responsible for the severe sperm motility defect in this patient.N/A. LIMITATIONS, REASONS FOR CAUTION: In this study, we described patients with either a homozygous or two heterozygous candidate pathogenic variants in genes linked to sperm motility disorders. Due to unavailability of parental DNA, we have not assessed the frequency of de novo or maternally inherited dominant variants and could not determine the parental origin of the mutations to establish in all cases that the mutations are present on both alleles. WIDER IMPLICATIONS OF THE FINDINGS: Our results confirm the likely causal role of variants in six known genes for sperm motility and we demonstrate that exome sequencing is an effective method to diagnose patients with severe sperm motility disorders (10/21 diagnosed; 48%). Furthermore, our analysis revealed six novel candidate genes for severe sperm motility disorders. Genome-wide sequencing of additional patient cohorts and re-analysis of exome data of currently unsolved cases may reveal additional variants in these novel candidate genes. STUDY FUNDING/COMPETING INTEREST(S): This project was supported in part by funding from the Australian National Health and Medical Research Council (APP1120356) to M.K.O.B., J.A.V. and R.I.M.L., The Netherlands Organisation for Scientific Research (918-15-667) to J.A.V., the Royal Society and Wolfson Foundation (WM160091) to J.A.V., as well as an Investigator Award in Science from the Wellcome Trust (209451) to J.A.V. and Grants from the National Research Council of Argentina (PIP 0900 and 4584) and ANPCyT (PICT 9591) to H.E.C. and a UUKi Rutherford Fund Fellowship awarded to B.J.H.publishersversionPeer reviewe

A de novo paradigm for male infertility

De novo mutations are known to play a prominent role in sporadic disorders with reduced fitness. We hypothesize that de novo mutations play an important role in severe male infertility and explain a portion of the genetic causes of this understudied disorder. To test this hypothesis, we utilize trio-based exome sequencing in a cohort of 185 infertile males and their unaffected parents. Following a systematic analysis, 29 of 145 rare (MAF < 0.1%) protein-altering de novo mutations are classified as possibly causative of the male infertility phenotype. We observed a significant enrichment of loss-of-function de novo mutations in loss-of-function-intolerant genes (p -value = 1.00 × 10 −5) in infertile men compared to controls. Additionally, we detected a significant increase in predicted pathogenic de novo missense mutations affecting missense-intolerant genes (p -value = 5.01 × 10 −4) in contrast to predicted benign de novo mutations. One gene we identify, RBM5, is an essential regulator of male germ cell pre-mRNA splicing and has been previously implicated in male infertility in mice. In a follow-up study, 6 rare pathogenic missense mutations affecting this gene are observed in a cohort of 2,506 infertile patients, whilst we find no such mutations in a cohort of 5,784 fertile men (p -value = 0.03). Our results provide evidence for the role of de novo mutations in severe male infertility and point to new candidate genes affecting fertility. Germline de novo mutations can impact individual fitness, but their role in human male infertility is understudied. Trio-based exome sequencing identifies many new candidate genes affecting male fertility, including an essential regulator of male germ cell pre-mRNA splicing

A de novo paradigm for male infertility

Funding Information: (DFG, CRU326) to C.F. and F.T. This project was also supported in part by funding from the Australian National Health and Medical Research Council (APP1120356) to M.K.O.B., by grants from the National Institutes of Health of the United States of America (R01HD078641 to D.F.C. and K.I.A., P50HD096723 to D.F.C.) and from the Biotechnology and Biological Sciences Research Council (BB/S008039/1) to D.J.E. Funding Information: We are grateful for the participation of all patients and their parents in this study. We thank Laurens van de Wiel (Radboudumc), Sebastian Judd-Mole (Monash University), Arron Scott and Bryan Hepworth (Newcastle University) for technical support, and Margot J Wyrwoll (University of Münster) for help with handling MERGE samples and data. This project was funded by The Netherlands Organization for Scientific Research (918-15-667) to J.A.V. as well as an Investigator Award in Science from the Wellcome Trust (209451) to J.A.V. a grant from the Catherine van Tussenbroek Foundation to M.S.O. a grant from MERCK to R.S. a UUKi Rutherford Fund Fellowship awarded to B.J.H. and the German Research Foundation Clinical Research Unit “Male Germ Cells” Publisher Copyright: © 2022, The Author(s).De novo mutations are known to play a prominent role in sporadic disorders with reduced fitness. We hypothesize that de novo mutations play an important role in severe male infertility and explain a portion of the genetic causes of this understudied disorder. To test this hypothesis, we utilize trio-based exome sequencing in a cohort of 185 infertile males and their unaffected parents. Following a systematic analysis, 29 of 145 rare (MAF < 0.1%) protein-altering de novo mutations are classified as possibly causative of the male infertility phenotype. We observed a significant enrichment of loss-of-function de novo mutations in loss-of-function-intolerant genes (p-value = 1.00 × 10−5) in infertile men compared to controls. Additionally, we detected a significant increase in predicted pathogenic de novo missense mutations affecting missense-intolerant genes (p-value = 5.01 × 10−4) in contrast to predicted benign de novo mutations. One gene we identify, RBM5, is an essential regulator of male germ cell pre-mRNA splicing and has been previously implicated in male infertility in mice. In a follow-up study, 6 rare pathogenic missense mutations affecting this gene are observed in a cohort of 2,506 infertile patients, whilst we find no such mutations in a cohort of 5,784 fertile men (p-value = 0.03). Our results provide evidence for the role of de novo mutations in severe male infertility and point to new candidate genes affecting fertility.publishersversionpublishe

A de novo paradigm for male infertility

Genetics of Male Infertility Initiative (GEMINI) consortium: Donald F. Conrad, Liina Nagirnaja, Kenneth I. Aston, Douglas T. Carrell, James M. Hotaling, Timothy G. Jenkins, Rob McLachlan, Moira K. O’Bryan, Peter N. Schlegel, Michael L. Eisenberg, Jay I. Sandlow, Emily S. Jungheim, Kenan R. Omurtag, Alexandra M. Lopes, Susana Seixas, Filipa Carvalho, Susana Fernandes, Alberto Barros, João Gonçalves, Iris Caetano, Graça Pinto, Sónia Correia, Maris Laan, Margus Punab, Ewa Rajpert-De Meyts, Niels Jørgensen, Kristian Almstrup, Csilla G. Krausz & Keith A. Jarvi.De novo mutations are known to play a prominent role in sporadic disorders with reduced fitness. We hypothesize that de novo mutations play an important role in severe male infertility and explain a portion of the genetic causes of this understudied disorder. To test this hypothesis, we utilize trio-based exome sequencing in a cohort of 185 infertile males and their unaffected parents. Following a systematic analysis, 29 of 145 rare (MAF < 0.1%) protein-altering de novo mutations are classified as possibly causative of the male infertility phenotype. We observed a significant enrichment of loss-of-function de novo mutations in loss-of-function-intolerant genes (p-value = 1.00 × 10−5) in infertile men compared to controls. Additionally, we detected a significant increase in predicted pathogenic de novo missense mutations affecting missense-intolerant genes (p-value = 5.01 × 10−4) in contrast to predicted benign de novo mutations. One gene we identify, RBM5, is an essential regulator of male germ cell pre-mRNA splicing and has been previously implicated in male infertility in mice. In a follow-up study, 6 rare pathogenic missense mutations affecting this gene are observed in a cohort of 2,506 infertile patients, whilst we find no such mutations in a cohort of 5,784 fertile men (p-value = 0.03). Our results provide evidence for the role of de novo mutations in severe male infertility and point to new candidate genes affecting fertility.This project was funded by The Netherlands Organization for Scientific Research (918-15-667) to J.A.V. as well as an Investigator Award in Science from the Wellcome Trust (209451) to J.A.V. a grant from the Catherine van Tussenbroek Foundation to M.S.O. a grant from MERCK to R.S. a UUKi Rutherford Fund Fellowship awarded to B.J.H. and the German Research Foundation Clinical Research Unit “Male Germ Cells” (DFG, CRU326) to C.F. and F.T. This project was also supported in part by funding from the Australian National Health and Medical Research Council (APP1120356) to M.K.O.B., by grants from the National Institutes of Health of the United States of America (R01HD078641 to D.F.C. and K.I.A., P50HD096723 to D.F.C.) and from the Biotechnology and Biological Sciences Research Council (BB/S008039/1) to D.J.E.info:eu-repo/semantics/publishedVersio

Switching to dual/monotherapy determines an increase in CD8+ in HIV-infected individuals: An observational cohort study

Background: The CD4/CD8 ratio has been associated with the risk of AIDS and non-AIDS events. We describe trends in immunological parameters in people who underwent a switch to monotherapy or dual therapy, compared to a control group remaining on triple antiretroviral therapy (ART). Methods: We included patients in Icona who started a three-drug combination ART regimen from an ART-naïve status and achieved a viral load ≤ 50 copies/mL; they were subsequently switched to another triple or to a mono or double regimen. Standard linear regression at fixed points in time (12-24 months after the switch) and linear mixed model analysis with random intercepts and slopes were used to compare CD4 and CD8 counts and their ratio over time according to regimen types (triple vs. dual and vs. mono). Results: A total of 1241 patients were included; 1073 switched to triple regimens, 104 to dual (72 with 1 nucleoside reverse transcriptase inhibitor (NRTI), 32 NRTI-sparing), and 64 to monotherapy. At 12 months after the switch, for the multivariable linear regression the mean change in the log10 CD4/CD8 ratio for patients on dual therapy was -0.03 (95% confidence interval (CI) -0.05, -0.0002), and the mean change in CD8 count was +99 (95% CI +12.1, +186.3), taking those on triple therapy as reference. In contrast, there was no evidence for a difference in CD4 count change. When using all counts, there was evidence for a significant difference in the slope of the ratio and CD8 count between people who were switched to triple (points/year change ratio = +0.056, CD8 = -25.7) and those to dual regimen (ratio = -0.029, CD8 = +110.4). Conclusions: We found an increase in CD8 lymphocytes in people who were switched to dual regimens compared to those who were switched to triple. Patients on monotherapy did not show significant differences. The long-term implications of this difference should be ascertained

Gender differences in the use of cardiovascular interventions in HIV-positive persons; the D:A:D Study

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Effectiveness of dolutegravir-based regimens as either first-line or switch antiretroviral therapy: data from the Icona cohort

156INTRODUCTION: Concerns about dolutegravir (DTG) tolerability in the real-life setting have recently arisen. We aimed to estimate the risk of treatment discontinuation and virological failure of DTG-based regimens from a large cohort of HIV-infected individuals. METHODS: We performed a multicentre, observational study including all antiretroviral therapy (ART)-naïve and virologically suppressed treatment-experienced (TE) patients from the Icona (Italian Cohort Naïve Antiretrovirals) cohort who started, for the first time, a DTG-based regimen from January 2015 to December 2017. We estimated the cumulative risk of DTG discontinuation regardless of the reason and for toxicity, and of virological failure using Kaplan-Meier curves. We used Cox regression model to investigate predictors of DTG discontinuation. RESULTS: About 1679 individuals (932 ART-naïve, 747 TE) were included. The one- and two-year probabilities (95% CI) of DTG discontinuation were 6.7% (4.9 to 8.4) and 11.5% (8.7 to 14.3) for ART-naïve and 6.6% (4.6 to 8.6) and 7.6% (5.4 to 9.8) for TE subjects. In both ART-naïve and TE patients, discontinuations of DTG were mainly driven by toxicity with an estimated risk (95% CI) of 4.0% (2.6 to 5.4) and 2.5% (1.3 to 3.6) by one year and 5.6% (3.8 to 7.5) and 4.0% (2.4 to 5.6) by two years respectively. Neuropsychiatric events were the main reason for stopping DTG in both ART-naïve (2.1%) and TE (1.7%) patients. In ART-naïve, a concomitant AIDS diagnosis predicted the risk of discontinuing DTG for any reason (adjusted relative hazard (aRH) = 3.38, p = 0.001), whereas starting DTG in combination with abacavir (ABC) was associated with a higher risk of discontinuing because of toxicity (aRH = 3.30, p = 0.009). TE patients starting a DTG-based dual therapy compared to a triple therapy had a lower risk of discontinuation for any reason (adjusted hazard ratio (aHR) = 2.50, p = 0.037 for ABC-based triple-therapies, aHR = 3.56, p = 0.012 for tenofovir-based) and for toxicity (aHR = 5.26, p = 0.030 for ABC-based, aHR = 6.60, p = 0.024 for tenofovir-based). The one- and two-year probabilities (95% CI) of virological failure were 1.2% (0.3 to 2.0) and 4.6% (2.7 to 6.5) in the ART naïve group and 2.2% (1.0 to 3.3) and 2.9% (1.5 to 4.3) in the TE group. CONCLUSIONS: In this large cohort, DTG showed excellent efficacy and optimal tolerability both as first-line and switching ART. The low risk of treatment-limiting toxicities in ART-naïve as well as in treated individuals reassures on the use of DTG in everyday clinical practicenonenoneMondi A.; Cozzi-Lepri A.; Tavelli A.; Rusconi S.; Vichi F.; Ceccherini-Silberstein F.; Calcagno A.; De Luca A.; Maggiolo F.; Marchetti G.; Antinori A.; d'Arminio Monforte A.; Andreoni M.; Castagna A.; Castelli F.; Cauda R.; Di Perri G.; Galli M.; Iardino R.; Ippolito G.; azzarin A.; Rezza G.; von Schloesser F.; Viale P.; Ceccherini-Silberstein F.; Cozzi-Lepri A.; Girardi E.; Lo Caputo S.; Mussini C.; Puoti M.; Perno C.F.; Balotta C.; Bandera A.; Bonora S.; Borderi M.; Capetti A.; Capobianchi M.R.; Cicalini S.; Cingolani A.; Cinque P.; Di Biagio A.; Gianotti N.; Gori A.; Guaraldi G.; Lapadula G.; Lichtner M.; Madeddu G.; Maggiolo F.; Monno L.; Nozza S.; Quiros Roldan E.; Rossotti R.; Rusconi S.; Santoro M.M.; aracino A.; Sarmati L.; Fanti I.; Galli L.; Lorenzini P.; Rodano' A.; Macchia M.; Tavelli A.; Carletti F.; Carrara S.; Di Caro A.; Graziano S.; Petrone F.; Prota G.; Quartu S.; Truffa S.; Giacometti I.A.; Costantini A.; Barocci V.; Angarano G.; Fabrizio C.; Suardi C.; i V.; Verucchi G.; Castelnuovo F.; Minardi C.; Menzaghi B.; Abeli C.; Cacopardo B.; Celesia B.; Vecchiet J.; Falasca K.; Pan A.; Lorenzotti S.; Sighinolfi L.; Segala D.; Blanc P.; Cassola G.; Viscoli C.; lessandrini A.; Bobbio N.; Mazzarello G.; Pozzetto I.; Bonfanti P.; Molteni C.; Chiodera A.; Milini P.; Nunnari G.; Pellicano G.; Rizzardini G.; Bai F.; Moioli M.C.; Piolini R.; Ridolfo A.L.; Salpietro S.; Tincati C.; Puzzolante C.; Migliorino C.; Sangiovanni V.; Borgia G.; Esposito V.; Di Martino F.; Gentile I.; Maddaloni L.; Cattelan A.M.; Marinello S.; Cascio A.; Colomba C.; Baldelli F.; Schiaroli E.; Parruti G.; Sozio F.; Magnani G.; Ursitti M.A.; Cristaudo A.; Vullo V.; Acinapura R.; Baldin G.; Capozzi M.; Rivano Capparucia M.; Iaiani G.; atini A.; Mastrorosa I.; Plazzi M.M.; Savinelli S.; Vergori A.; Cecchetto M.; Viviani F.; Bagella P.; Rossetti B.; Fontana Del Vecchio R.; Francisci D.; Di Giuli C.; Caramello P.; Orofino G.C.; Sciandra M.; Bassetti M.; ondero A.; Pellizzer G.; Manfrin V.; Starnini G.; alungo A.Mondi, A.; Cozzi-Lepri, A.; Tavelli, A.; Rusconi, S.; Vichi, F.; Ceccherini-Silberstein, F.; Calcagno, A.; De Luca, A.; Maggiolo, F.; Marchetti, G.; Antinori, A.; d'Arminio Monforte, A.; Andreoni, M.; Castagna, A.; Castelli, F.; Cauda, R.; Di Perri, G.; Galli, M.; Iardino, R.; Ippolito, G.; Azzarin, A.; Rezza, G.; von Schloesser, F.; Viale, P.; Ceccherini-Silberstein, F.; Cozzi-Lepri, A.; Girardi, E.; Lo Caputo, S.; Mussini, C.; Puoti, M.; Perno, C. F.; Balotta, C.; Bandera, A.; Bonora, S.; Borderi, M.; Capetti, A.; Capobianchi, M. R.; Cicalini, S.; Cingolani, A.; Cinque, P.; Di Biagio, A.; Gianotti, N.; Gori, A.; Guaraldi, G.; Lapadula, G.; Lichtner, M.; Madeddu, G.; Maggiolo, F.; Monno, L.; Nozza, S.; Quiros Roldan, E.; Rossotti, R.; Rusconi, S.; Santoro, M. M.; Aracino, A.; Sarmati, L.; Fanti, I.; Galli, L.; Lorenzini, P.; Rodano', A.; Macchia, M.; Tavelli, A.; Carletti, F.; Carrara, S.; Di Caro, A.; Graziano, S.; Petrone, F.; Prota, G.; Quartu, S.; Truffa, S.; Giacometti, I. A.; Costantini, A.; Barocci, V.; Angarano, G.; Fabrizio, C.; Suardi, C.; I, V.; Verucchi, G.; Castelnuovo, F.; Minardi, C.; Menzaghi, B.; Abeli, C.; Cacopardo, B.; Celesia, B.; Vecchiet, J.; Falasca, K.; Pan, A.; Lorenzotti, S.; Sighinolfi, L.; Segala, D.; Blanc, P.; Cassola, G.; Viscoli, C.; Lessandrini, A.; Bobbio, N.; Mazzarello, G.; Pozzetto, I.; Bonfanti, P.; Molteni, C.; Chiodera, A.; Milini, P.; Nunnari, G.; Pellicano, G.; Rizzardini, G.; Bai, F.; Moioli, M. C.; Piolini, R.; Ridolfo, A. L.; Salpietro, S.; Tincati, C.; Puzzolante, C.; Migliorino, C.; Sangiovanni, V.; Borgia, G.; Esposito, V.; Di Martino, F.; Gentile, I.; Maddaloni, L.; Cattelan, A. M.; Marinello, S.; Cascio, A.; Colomba, C.; Baldelli, F.; Schiaroli, E.; Parruti, G.; Sozio, F.; Magnani, G.; Ursitti, M. A.; Cristaudo, A.; Vullo, V.; Acinapura, R.; Baldin, G.; Capozzi, M.; Rivano Capparucia, M.; Iaiani, G.; Atini, A.; Mastrorosa, I.; Plazzi, M. M.; Savinelli, S.; Vergori, A.; Cecchetto, M.; Viviani, F.; Bagella, P.; Rossetti, B.; Fontana Del Vecchio, R.; Francisci, D.; Di Giuli, C.; Caramello, P.; Orofino, G. C.; Sciandra, M.; Bassetti, M.; Ondero, A.; Pellizzer, G.; Manfrin, V.; Starnini, G.; Alungo, A

Effectiveness of dolutegravir-based regimens as either first-line or switch antiretroviral therapy: data from the Icona cohort

Introduction: Concerns about dolutegravir (DTG) tolerability in the real-life setting have recently arisen. We aimed to estimate the risk of treatment discontinuation and virological failure of DTG-based regimens from a large cohort of HIV-infected individuals. Methods: We performed a multicentre, observational study including all antiretroviral therapy (ART)-na\uefve and virologically suppressed treatment-experienced (TE) patients from the Icona (Italian Cohort Na\uefve Antiretrovirals) cohort who started, for the first time, a DTG-based regimen from January 2015 to December 2017. We estimated the cumulative risk of DTG discontinuation regardless of the reason and for toxicity, and of virological failure using Kaplan\u2013Meier curves. We used Cox regression model to investigate predictors of DTG discontinuation. Results: About 1679 individuals (932 ART-na\uefve, 747 TE) were included. The one- and two-year probabilities (95% CI) of DTG discontinuation were 6.7% (4.9 to 8.4) and 11.5% (8.7 to 14.3) for ART-na\uefve and 6.6% (4.6 to 8.6) and 7.6% (5.4 to 9.8) for TE subjects. In both ART-na\uefve and TE patients, discontinuations of DTG were mainly driven by toxicity with an estimated risk (95% CI) of 4.0% (2.6 to 5.4) and 2.5% (1.3 to 3.6) by one year and 5.6% (3.8 to 7.5) and 4.0% (2.4 to 5.6) by two years respectively. Neuropsychiatric events were the main reason for stopping DTG in both ART-na\uefve (2.1%) and TE (1.7%) patients. In ART-na\uefve, a concomitant AIDS diagnosis predicted the risk of discontinuing DTG for any reason (adjusted relative hazard (aRH)&nbsp;=&nbsp;3.38, p&nbsp;=&nbsp;0.001), whereas starting DTG in combination with abacavir (ABC) was associated with a higher risk of discontinuing because of toxicity (aRH&nbsp;=&nbsp;3.30, p&nbsp;=&nbsp;0.009). TE patients starting a DTG-based dual therapy compared to a triple therapy had a lower risk of discontinuation for any reason (adjusted hazard ratio (aHR)&nbsp;=&nbsp;2.50, p&nbsp;=&nbsp;0.037 for ABC-based triple-therapies, aHR&nbsp;=&nbsp;3.56, p&nbsp;=&nbsp;0.012 for tenofovir-based) and for toxicity (aHR&nbsp;=&nbsp;5.26, p&nbsp;=&nbsp;0.030 for ABC-based, aHR&nbsp;=&nbsp;6.60, p&nbsp;=&nbsp;0.024 for tenofovir-based). The one- and two-year probabilities (95% CI) of virological failure were 1.2% (0.3 to 2.0) and 4.6% (2.7 to 6.5) in the ART na\uefve group and 2.2% (1.0 to 3.3) and 2.9% (1.5 to 4.3) in the TE group. Conclusions: In this large cohort, DTG showed excellent efficacy and optimal tolerability both as first-line and switching ART. The low risk of treatment-limiting toxicities in ART-na\uefve as well as in treated individuals reassures on the use of DTG in everyday clinical practice