👤 Jon R Skinner

Allergen-specific immunotherapy (AIT) is the only disease-modifying treatment in allergic airway diseases. Underlying immunological mechanisms and candidate biomarkers, which may be translated into predictive/surrogate measures of clinical efficacy, remain an active area of research. The aim of this study was to evaluate Pollinex Quattro (PQ) Grass AIT induced immunomodulatory mechanisms, based on transcriptome profiling of peripheral blood mononuclear cells. 119 subjects with grass pollen induced seasonal allergic rhinitis (SAR) were randomized in a 2:2:1:1 ratio to receive a cumulative dose of PQ Grass as a conventional or extended pre-seasonal regimen, placebo, or placebo with MicroCrystalline Tyrosine. Gene expression analysis was an exploratory endpoint evaluated in a subgroup of 30 subjects randomly selected from the four treatment arms. Samples were collected at three time points: screening (baseline), before the start of the grass pollen season and at the end of the season. This study was funded by the manufacturer of PQ. Transcriptome analysis demonstrated that the most significant changes in gene expression, for both treatment regimens, were at the end of the grass pollen season, with the main Th1 candidate molecules (IL-12A, IFNγ) upregulated and Th2 signature cytokines downregulated (IL-4, IL-13, IL-9) (p < .05). Canonical pathways analysis demonstrated Th1, Th2, Th17 and IL-17 as the most significantly enriched pathways based on absolute value of activation z-score (IzI score ≥ 2, p < .05). Upstream regulator analysis showed pronounced inhibition of pro-inflammatory allergic molecules IgE, IL-17A, IL-17F, IL-25 (IL-17E) (IzI score ≥ 2, FDR < 0.05) and activation of pro-tolerogenic molecules IL-12A, IL-27, IL-35 (EBI3) at the end of the grass pollen season. Peripheral blood mononuclear cells transcriptome profile showed an inhibition of Th2, Th17 pro-inflammatory allergic responses and immune deviation towards Th1 responses. PQ Grass extended regimen exhibited a superior mechanistic efficacy profile in comparison with PQ conventional regimen. Show less

Aotearoa/New Zealand has a multiethnic population. Patients with hypertrophic cardiomyopathy (HCM) are enrolled in the national Cardiac Inherited Diseases Registry New Zealand. Here, we report the characteristics of Cardiac Inherited Diseases Registry New Zealand HCM probands with and without pathogenic or likely pathogenic (P/LP) genetic variants for HCM, and assess genetic testing yield and variant spectrum by self-identified ethnicity. Probands with HCM and enrolled in Cardiac Inherited Diseases Registry New Zealand who have undergone clinical genetic testing over a 17-year period were included. Clinical data, family history, and genetic test results were analyzed. Of 336 probands, 121 (36%) were women, 220 (66%) were European ethnicity, 41 (12%) were Māori, 26 (8%) were Pacific people, and 49 (15%) were other ethnicities. Thirteen probands (4%) presented with sudden death and 19 (6%) with cardiac arrest. A total of 134 (40%) had a P/LP variant identified; most commonly in the Carriage of a P/LP variant in HCM probands is associated with presentation at younger age, and cardiac arrest or sudden death events. Māori or Pacific probands were less likely to have a P/LP variant identified than European or other ethnicity probands. Show less

Childhood cancer survivors (CCS) exhibit significantly increased chronic diseases and premature death. Abnormalities in DNA methylation are associated with development of chronic diseases and reduced life expectancy. We investigated the hypothesis that anti-cancer treatments are associated with long-term DNA methylation changes that could be key drivers of adverse late health effects. Genome-wide DNA methylation was assessed using MethylationEPIC arrays in paired samples (before/after therapy) from 32 childhood cancer patients. Separately, methylation was determined in 32 samples from different adult CCS (mean 22-years post-diagnosis) and compared with cancer-free controls (n = 284). Widespread DNA methylation changes were identified post-treatment in childhood cancer patients, including 146 differentially methylated regions (DMRs), which were consistently altered in the 32 post-treatment samples. Analysis of adult CCS identified matching methylation changes at 107/146 of the DMRs, suggesting potential long-term retention of post-therapy changes. Adult survivors also exhibited epigenetic age acceleration, independent of DMR methylation. Furthermore, altered methylation at the DUSP6 DMR was significantly associated with early mortality, suggesting altered methylation may be prognostic for some late adverse health effects in CCS. These novel methylation changes could serve as biomarkers for assessing normal cell toxicity in ongoing treatments and predicting long-term health outcomes in CCS. Show less

Therapeutics that inhibit IL-6 at different points in its signaling pathway are in clinical use, yet whether the immunological effects of these interventions differ based on their molecular target is unknown. We performed short-term interventions in individuals with type 1 diabetes using anti-IL-6 (siltuximab) or anti-IL-6 receptor (IL-6R; tocilizumab) therapies and investigated the impact of this in vivo blockade on T cell fate and function. Immune outcomes were influenced by the target of the therapeutic intervention (IL-6 versus IL-6R) and by peak drug concentration. Tocilizumab reduced ICOS expression on T follicular helper cell populations and T cell receptor-driven (TCR-driven) STAT3 phosphorylation. Siltuximab reversed resistance to Treg-mediated suppression and increased TCR-driven phosphorylated STAT3 and production of IL-10, IL-21, and IL-27 by T effectors. Together, these findings indicate that the context of IL-6 blockade in vivo drives distinct T cell-intrinsic changes that may influence therapeutic outcomes. Show less

Copy-number variant (CNV) analysis is increasingly performed in genetic diagnostics. We leveraged recent gene curation efforts and technical standards for interpretation and reporting of CNVs to characterize clinically relevant CNVs in patients with inherited heart disease and sudden cardiac death. Exome sequencing data were analyzed for CNVs using eXome-Hidden Markov Model tool in 48 established disease genes. CNV breakpoint junctions were characterized. CNVs were classified using the American College of Medical Genetics and Genomics technical standards. We identified eight CNVs in 690 unrelated probands (1.2%). Characterization of breakpoint junctions revealed nonhomologous end joining was responsible for four deletions, whereas one duplication was caused by nonallelic homologous recombination between duplicated sequences in MYH6 and MYH7. Identifying the precise breakpoint junctions determined the genomic involvement and proved useful for interpreting the clinical relevance of CNVs. Three large deletions involving TTN, MYBPC3, and KCNH2 were classified as pathogenic in three patients. Haplotype analysis of a deletion in ACTN2, found in two families, suggests the deletion was caused by an ancestral event. CNVs infrequently cause inherited heart diseases and should be investigated when standard genetic testing does not reveal a genetic diagnosis. Show less

Bisphosphonate drugs such as zoledronic acid (ZOL), used for the treatment of common bone disorders, target the skeleton and inhibit bone resorption by preventing the prenylation of small GTPases in bone-destroying osteoclasts. Increasing evidence indicates that bisphosphonates also have pleiotropic effects outside the skeleton, most likely via cells of the monocyte/macrophage lineage exposed to nanomolar circulating drug concentrations. However, no effects of such low concentrations of ZOL have been reported using existing approaches. We have optimized a highly sensitive in vitro prenylation assay utilizing recombinant geranylgeranyltransferases to enable the detection of subtle effects of ZOL on the prenylation of Rab- and Rho-family GTPases. Using this assay, we found for the first time that concentrations of ZOL as low as 10nM caused inhibition of Rab prenylation in J774 macrophages following prolonged cell culture. By combining the assay with quantitative mass spectrometry we identified an accumulation of 18 different unprenylated Rab proteins in J774 cells after nanomolar ZOL treatment, with a >7-fold increase in the unprenylated form of Rab proteins associated with the endophagosome pathway (Rab1, Rab5, Rab6, Rab7, Rab11, Rab14 and Rab21). Finally, we also detected a clear effect of subcutaneous ZOL administration in vivo on the prenylation of Rab1A, Rab5B, Rab7A and Rab14 in mouse peritoneal macrophages, confirming that systemic treatment with bisphosphonate drug can inhibit prenylation in myeloid cells in vivo outside the skeleton. These observations begin a new era in defining the precise pharmacological actions of bisphosphonate drugs on the prenylation of small GTPases in vivo. Show less

The authors have previously shown that the clinically relevant concentrations of inhalational anesthetics dose-dependently inhibit the postsynaptic density protein (PSD)-95, Dlg, and ZO-1 domain-mediated protein interactions between N-methyl-d-aspartate receptors and PSD-95/synaptic-associated protein (SAP) 90 or PSD-93/Chapsyn-110 and that the knockdown of spinal PSD-95/SAP90 significantly reduces the minimum alveolar concentration (MAC) for isoflurane in rats. The authors designed antisense oligodeoxynucleotides based on the mouse PSD-95/SAP90 and PSD-93/Chapsyn-110 messenger RNAs that correspond to their PSD-95, Dlg, and ZO-1 domain nucleotides and can specifically knock down the respective proteins. The authors intrathecally injected antisense oligodeoxynucleotides into wild-type and PSD-93/Chapsyn-110 knockout mice to investigate the effect of PSD-95/SAP90 and/or PSD-93/Chapsyn-110 deficiency on halothane anesthesia. Both PSD-95/SAP90 and PSD-93/Chapsyn-110 antisense oligodeoxynucleotides caused a dose-dependent and significant reduction in the MAC of halothane in wild-type mice. The intrathecal injection of PSD-95/SAP90 antisense oligodeoxynucleotide at different doses (25 and 50 microg) reduced halothane MAC by 40 and 55%, respectively, and intrathecal injection of PSD-93/Chapsyn-110 antisense oligodeoxynucleotide at different doses (12 and 24 microg) reduced halothane MAC by 25 and 53%, respectively. The PSD-95/SAP90 antisense oligodeoxynucleotide showed similar effect on halothane MAC in wild-type and PSD-93/Chapsyn-110 knockout mice, suggesting that the combination of PSD-95/SAP90 knockdown with PSD-93/Chapsyn-110 deletion did not have an additive effect on halothane anesthesia. The current results indicate that PSD-95/SAP90 and PSD-93/Chapsyn-110 are involved in the molecular mechanisms of halothane anesthesia and that the functional role of PSD-95/SAP90 in halothane anesthesia is not enhanced after PSD-93/Chapsyn-110 deletion. Show less