Clinical features for diagnosis and management of patients with PRDM12 congenital insensitivity to pain.

This is the final version of the article. It first appeared from British Medical Journal Group via https://doi.org/10.1136/jmedgenet-2015-103646BACKGROUND: Congenital insensitivity to pain (CIP) is a rare extreme phenotype characterised by an inability to perceive pain present from birth due to lack of, or malfunction of, nociceptors. PRDM12 has recently been identified as a new gene that can cause CIP. The full phenotype and natural history have not yet been reported. METHODS: We have ascertained five adult patients and report their clinical features. RESULTS: Based on our findings, and those of previous patients, we describe the natural history of the PRDM12-CIP disorder, and derive diagnostic and management features to guide the clinical management of patients. CONCLUSIONS: PRDM12-CIP is a distinct and diagnosable disorder, and requires specific clinical management to minimise predictable complications.YCC was funded by the Cambridge NIHR BRC, and CGW by the UK MR

A miRNA Host Response Signature Accurately Discriminates Acute Respiratory Infection Etiologies

Background: Acute respiratory infections (ARIs) are the leading indication for antibacterial prescriptions despite a viral etiology in the majority of cases. The lack of available diagnostics to discriminate viral and bacterial etiologies contributes to this discordance. Recent efforts have focused on the host response as a source for novel diagnostic targets although none have explored the ability of host-derived microRNAs (miRNA) to discriminate between these etiologies.Methods: In this study, we compared host-derived miRNAs and mRNAs from human H3N2 influenza challenge subjects to those from patients with Streptococcus pneumoniae pneumonia. Sparse logistic regression models were used to generate miRNA signatures diagnostic of ARI etiologies. Generalized linear modeling of mRNAs to identify differentially expressed (DE) genes allowed analysis of potential miRNA:mRNA relationships. High likelihood miRNA:mRNA interactions were examined using binding target prediction and negative correlation to further explore potential changes in pathway regulation in response to infection.Results: The resultant miRNA signatures were highly accurate in discriminating ARI etiologies. Mean accuracy was 100% [88.8–100; 95% Confidence Interval (CI)] in discriminating the healthy state from S. pneumoniae pneumonia and 91.3% (72.0–98.9; 95% CI) in discriminating S. pneumoniae pneumonia from influenza infection. Subsequent differential mRNA gene expression analysis revealed alterations in regulatory networks consistent with known biology including immune cell activation and host response to viral infection. Negative correlation network analysis of miRNA:mRNA interactions revealed connections to pathways with known immunobiology such as interferon regulation and MAP kinase signaling.Conclusion: We have developed novel human host-response miRNA signatures for bacterial and viral ARI etiologies. miRNA host response signatures reveal accurate discrimination between S. pneumoniae pneumonia and influenza etiologies for ARI and integrated analyses of the host-pathogen interface are consistent with expected biology. These results highlight the differential miRNA host response to bacterial and viral etiologies of ARI, offering new opportunities to distinguish these entities

Human ASPM participates in spindle organisation, spindle orientation and cytokinesis

Background Mutations in the Abnormal Spindle Microcephaly related gene (ASPM) are the commonest cause of autosomal recessive primary microcephaly (MCPH) a disorder characterised by a small brain and associated mental retardation. ASPM encodes a mitotic spindle pole associated protein. It is suggested that the MCPH phenotype arises from proliferation defects in neural progenitor cells (NPC). Results We show that ASPM is a microtubule minus end-associated protein that is recruited in a microtubule-dependent manner to the pericentriolar matrix (PCM) at the spindle poles during mitosis. ASPM siRNA reduces ASPM protein at the spindle poles in cultured U2OS cells and severely perturbs a number of aspects of mitosis, including the orientation of the mitotic spindle, the main determinant of developmental asymmetrical cell division. The majority of ASPM depleted mitotic cells fail to complete cytokinesis. In MCPH patient fibroblasts we show that a pathogenic ASPM splice site mutation results in the expression of a novel variant protein lacking a tripeptide motif, a minimal alteration that correlates with a dramatic decrease in ASPM spindle pole localisation. Moreover, expression of dominant-negative ASPM C-terminal fragments cause severe spindle assembly defects and cytokinesis failure in cultured cells. Conclusions These observations indicate that ASPM participates in spindle organisation, spindle positioning and cytokinesis in all dividing cells and that the extreme C-terminus of the protein is required for ASPM localisation and function. Our data supports the hypothesis that the MCPH phenotype caused by ASPM mutation is a consequence of mitotic aberrations during neurogenesis. We propose the effects of ASPM mutation are tolerated in somatic cells but have profound consequences for the symmetrical division of NPCs, due to the unusual morphology of these cells. This antagonises the early expansion of the progenitor pool that underpins cortical neurogenesis, causing the MCPH phenotype

A Comprehensive Functional Analysis of NTRK1 Missense Mutations Causing Hereditary Sensory and Autonomic Neuropathy Type IV (HSAN IV).

Hereditary sensory and autonomic neuropathy type IV (HSAN IV) is an autosomal recessive disorder characterized by a complete lack of pain perception and anhidrosis. Here, we studied a cohort of seven patients with HSAN IV and describe a comprehensive functional analysis of seven novel NTRK1 missense mutations, c.1550G >A, c.1565G >A, c.1970T >C, c.2096T >C, c.2254T >A, c.2288G >C, and c.2311C >T, corresponding to p.G517E, p.G522E, p.L657P, p.I699T, p.C752S, p.C763S, and p.R771C, all of which were predicted pathogenic by in silico analysis. The results allowed us to assess the pathogenicity of each mutation and to gain novel insights into tropomyosin receptor kinase A (TRKA) downstream signaling. Each mutation was systematically analyzed for TRKA glycosylation states, intracellular and cell membrane expression patterns, nerve growth factor stimulated TRKA autophosphorylation, TRKA-Y496 phosphorylation, PLCγ activity, and neurite outgrowth. We showed a diverse range of functional effects: one mutation appeared fully functional, another had partial activity in all assays, one mutation affected only the PLCγ pathway and four mutations were proved null in all assays. Thus, we conclude that complete abolition of TRKA kinase activity is not the only pathogenic mechanism underlying HSAN IV. By corollary, the assessment of the clinical pathogenicity of HSAN IV mutations is more complex than initially predicted and requires a multifaceted approach.We acknowledge funding from the Medical Research Council (SSS and MSN) and Cambridge Biomedical Research Campus (Y-CC)

Potential Cost-effectiveness of Early Identification of Hospital-acquired Infection in Critically Ill Patients

Limitations in methods for the rapid diagnosis of hospital-acquired infections often delay initiation of effective antimicrobial therapy. New diagnostic approaches offer potential clinical and cost-related improvements in the management of these infections. We developed a decision modeling framework to assess the potential cost-effectiveness of a rapid biomarker assay to identify hospital-acquired infection in high-risk patients earlier than standard diagnostic testing. The framework includes parameters representing rates of infection, rates of delayed appropriate therapy, and impact of delayed therapy on mortality, along with assumptions about diagnostic test characteristics and their impact on delayed therapy and length of stay. Parameter estimates were based on contemporary, published studies and supplemented with data from a four-site, observational, clinical study. Extensive sensitivity analyses were performed. The base-case analysis assumed 17.6% of ventilated patients and 11.2% of nonventilated patients develop hospital-acquired infection and that 28.7% of patients with hospital-acquired infection experience delays in appropriate antibiotic therapy with standard care. We assumed this percentage decreased by 50% (to 14.4%) among patients with true-positive results and increased by 50% (to 43.1%) among patients with false-negative results using a hypothetical biomarker assay. Cost of testing was set at $110/d. In the base-case analysis, among ventilated patients, daily diagnostic testing starting on admission reduced inpatient mortality from 12.3 to 11.9$1,640 per patient, resulting in an incremental cost-effectiveness ratio of $21,389 per life-year saved. Among nonventilated patients, inpatient mortality decreased from 7.3 to 7.1$1,381 with diagnostic testing. The resulting incremental cost-effectiveness ratio was $42,325 per life-year saved. Threshold analyses revealed the probabilities of developing hospital-acquired infection in ventilated and nonventilated patients could be as low as 8.4 and 9.8$50,000 per life-year saved. Development and use of serial diagnostic testing that reduces the proportion of patients with delays in appropriate antibiotic therapy for hospital-acquired infections could reduce inpatient mortality. The model presented here offers a cost-effectiveness framework for future test development

Human Labor Pain Is Influenced by the Voltage-Gated Potassium Channel KV6.4 Subunit.

By studying healthy women who do not request analgesia during their first delivery, we investigate genetic effects on labor pain. Such women have normal sensory and psychometric test results, except for significantly higher cuff pressure pain. We find an excess of heterozygotes carrying the rare allele of SNP rs140124801 in KCNG4. The rare variant KV6.4-Met419 has a dominant-negative effect and cannot modulate the voltage dependence of KV2.1 inactivation because it fails to traffic to the plasma membrane. In vivo, Kcng4 (KV6.4) expression occurs in 40% of retrograde-labeled mouse uterine sensory neurons, all of which express KV2.1, and over 90% express the nociceptor genes Trpv1 and Scn10a. In neurons overexpressing KV6.4-Met419, the voltage dependence of inactivation for KV2.1 is more depolarized compared with neurons overexpressing KV6.4. Finally, KV6.4-Met419-overexpressing neurons have a higher action potential threshold. We conclude that KV6.4 can influence human labor pain by modulating the excitability of uterine nociceptors.MCL, DKM, DW, and CGW acknowledge funding from Addenbrooke’s Charitable Trust and the NIHR Cambridge Biomedical Research Centre. MN was funded by the Wellcome Trust (200183/Z/15/Z); JH and ESS by a Rosetrees Postdoctoral Grant (A1296) and the BBSRC (BB/R006210/1); GC and ESS by Versus Arthritis Grants (RG21973); VBL and FR by the Wellcome Trust (106262/Z/14/Z and 106263/Z/14/Z) and a joint MRC programme within the Metabolic Diseases Unit (MRC_MC_UU_12012/3). EF, GI and CB were funded by the Cambridge NIHR Biomedical Research Centre Integrative Genomics theme and LAP by a BBSRC-funded studentship (BB/M011194/1)

Evidence of a genetic background predisposing to complex regional pain syndrome type 1.

BackgroundComplex regional pain syndrome type 1 (CRPS-1) is a rare, disabling and sometimes chronic disorder usually arising after a trauma. This exploratory study examined whether patients with chronic CRPS-1 have a different genetic profile compared with those who do not have the condition.MethodsExome sequencing was performed to seek altered non-synonymous SNP allele frequencies in a discovery cohort of well-characterised patients with chronic CRPS-1 (n=34) compared with population databases. Identified SNP alleles were confirmed by Sanger sequencing and sought in a replication cohort (n=50). Gene expression of peripheral blood macrophages was assessed.ResultsIn the discovery cohort, the rare allele frequencies of four non-synonymous SNPs were statistically increased. The replication cohort confirmed this finding. In a chronic pain cohort, these alleles were not overexpressed. In total, 25 out of 84 (29.8%) patients with CRPS-1 expressed a rare allele. The SNPs were rs41289586 in ANO10, rs28360457 in P2RX7, rs1126930 in PRKAG1 and rs80308281 in SLC12A9. Males were more likely than females to have a rare SNP allele, 8 out of 14 (57.1%) vs 17 out of 70 (24.3%) (Fisher's p=0.023). ANO10, P2RX7, PRKAG1 and SLC12A9 were all expressed in macrophages from healthy human controls.ConclusionA single SNP in each of the genes ANO10, P2RX7, PRKAG1 and SLC12A9 was associated with developing chronic CRPS-1, with more males than females expressing these rare alleles. Our work suggests the possibility that a permissive genetic background is an important factor in the development of CRPS-1

Human genetic and metabolite variation reveals that methylthioadenosine is a prognostic biomarker and an inflammatory regulator in sepsis.

Sepsis is a deleterious inflammatory response to infection with high mortality. Reliable sepsis biomarkers could improve diagnosis, prognosis, and treatment. Integration of human genetics, patient metabolite and cytokine measurements, and testing in a mouse model demonstrate that the methionine salvage pathway is a regulator of sepsis that can accurately predict prognosis in patients. Pathway-based genome-wide association analysis of nontyphoidal Salmonella bacteremia showed a strong enrichment for single-nucleotide polymorphisms near the components of the methionine salvage pathway. Measurement of the pathway's substrate, methylthioadenosine (MTA), in two cohorts of sepsis patients demonstrated increased plasma MTA in nonsurvivors. Plasma MTA was correlated with levels of inflammatory cytokines, indicating that elevated MTA marks a subset of patients with excessive inflammation. A machine-learning model combining MTA and other variables yielded approximately 80% accuracy (area under the curve) in predicting death. Furthermore, mice infected with Salmonella had prolonged survival when MTA was administered before infection, suggesting that manipulating MTA levels could regulate the severity of the inflammatory response. Our results demonstrate how combining genetic data, biomolecule measurements, and animal models can shape our understanding of disease and lead to new biomarkers for patient stratification and potential therapeutic targeting

Gene Expression-Based Classifiers Identify Staphylococcus aureus Infection in Mice and Humans

Staphylococcus aureus causes a spectrum of human infection. Diagnostic delays and uncertainty lead to treatment delays and inappropriate antibiotic use. A growing literature suggests the host’s inflammatory response to the pathogen represents a potential tool to improve upon current diagnostics. The hypothesis of this study is that the host responds differently to S. aureus than to E. coli infection in a quantifiable way, providing a new diagnostic avenue. This study uses Bayesian sparse factor modeling and penalized binary regression to define peripheral blood gene-expression classifiers of murine and human S. aureus infection. The murine-derived classifier distinguished S. aureus infection from healthy controls and Escherichia coli-infected mice across a range of conditions (mouse and bacterial strain, time post infection) and was validated in outbred mice (AUC>0.97). A S. aureus classifier derived from a cohort of 94 human subjects distinguished S. aureus blood stream infection (BSI) from healthy subjects (AUC 0.99) and E. coli BSI (AUC 0.84). Murine and human responses to S. aureus infection share common biological pathways, allowing the murine model to classify S. aureus BSI in humans (AUC 0.84). Both murine and human S. aureus classifiers were validated in an independent human cohort (AUC 0.95 and 0.92, respectively). The approach described here lends insight into the conserved and disparate pathways utilized by mice and humans in response to these infections. Furthermore, this study advances our understanding of S. aureus infection; the host response to it; and identifies new diagnostic and therapeutic avenues