Kv1.3 blockade by ShK186 modulates CD4+ effector memory T-cell activity of patients with Granulomatosis with polyangiitis

OBJECTIVES: Granulomatosis with polyangiitis (GPA) is a chronic relapsing systemic autoimmune vasculitis. Current treatment of GPA is unsatisfactory as it relies on strong immunosuppressive regimens, with either cyclophosphamide or rituximab, that reduce the immunogenicity of several vaccines and are risk factors of severe form of COVID-19. This emphasizes the need to identify new drug target and to develop treatment strategies with less harmful side effects. Since CD4+ effector memory T cells (TEM) play a key role in the pathogenesis of GPA, we aimed in this study to modulate CD4+TEM cell activity via Kv1.3 blockade using the specific peptide inhibiter, ShK-186.METHODS: Peripheral blood of 27 GPA-patients in remission and 16 age- and sex-matched healthy controls (HCs) were pre-incubated in vitro in the presence or absence of ShK-186, followed by stimulation with PMA, calcium ionophore and brefeldin-A. The effect of ShK-186 on the cytokine production (IFNγ, TNFα, IL-4, IL-17, IL-21) within total and subsets of CD4+TH cells were assessed using flow cytometry.RESULTS: ShK-186 reduced the expression level of IFNγ, TNFα, IL-4, IL-17, and IL-21 in CD4+TH cells from GPA-patients in vitro. Further analysis performed on sorted CD4+T cell subsets, revealed that ShK-186 predominantly inhibited the cytokine production of CD4+TEM cells. ShK-186 treatment reduced the production of the pro-inflammatory cytokines to the level seen in CD4+ TH cells from HCs.CONCLUSIONS: Modulation of cellular effector function by ShK-186 may constitute a novel treatment strategy for GPA with high specificity and less harmful side effects.</p

Kv1.3 blockade by ShK186 modulates CD4+ effector memory T-cell activity of patients with Granulomatosis with polyangiitis

OBJECTIVES: Granulomatosis with polyangiitis (GPA) is a chronic relapsing systemic autoimmune vasculitis. Current treatment of GPA is unsatisfactory as it relies on strong immunosuppressive regimens, with either cyclophosphamide or rituximab, that reduce the immunogenicity of several vaccines and are risk factors of severe form of COVID-19. This emphasizes the need to identify new drug target and to develop treatment strategies with less harmful side effects. Since CD4+ effector memory T cells (TEM) play a key role in the pathogenesis of GPA, we aimed in this study to modulate CD4+TEM cell activity via Kv1.3 blockade using the specific peptide inhibiter, ShK-186.METHODS: Peripheral blood of 27 GPA-patients in remission and 16 age- and sex-matched healthy controls (HCs) were pre-incubated in vitro in the presence or absence of ShK-186, followed by stimulation with PMA, calcium ionophore and brefeldin-A. The effect of ShK-186 on the cytokine production (IFNγ, TNFα, IL-4, IL-17, IL-21) within total and subsets of CD4+TH cells were assessed using flow cytometry.RESULTS: ShK-186 reduced the expression level of IFNγ, TNFα, IL-4, IL-17, and IL-21 in CD4+TH cells from GPA-patients in vitro. Further analysis performed on sorted CD4+T cell subsets, revealed that ShK-186 predominantly inhibited the cytokine production of CD4+TEM cells. ShK-186 treatment reduced the production of the pro-inflammatory cytokines to the level seen in CD4+ TH cells from HCs.CONCLUSIONS: Modulation of cellular effector function by ShK-186 may constitute a novel treatment strategy for GPA with high specificity and less harmful side effects.</p

Kv1.3 blockade by ShK186 modulates CD4+ effector memory T-cell activity of patients with Granulomatosis with polyangiitis

OBJECTIVES: Granulomatosis with polyangiitis (GPA) is a chronic relapsing systemic autoimmune vasculitis. Current treatment of GPA is unsatisfactory as it relies on strong immunosuppressive regimens, with either cyclophosphamide or rituximab, that reduce the immunogenicity of several vaccines and are risk factors of severe form of COVID-19. This emphasizes the need to identify new drug target and to develop treatment strategies with less harmful side effects. Since CD4+ effector memory T cells (TEM) play a key role in the pathogenesis of GPA, we aimed in this study to modulate CD4+TEM cell activity via Kv1.3 blockade using the specific peptide inhibiter, ShK-186.METHODS: Peripheral blood of 27 GPA-patients in remission and 16 age- and sex-matched healthy controls (HCs) were pre-incubated in vitro in the presence or absence of ShK-186, followed by stimulation with PMA, calcium ionophore and brefeldin-A. The effect of ShK-186 on the cytokine production (IFNγ, TNFα, IL-4, IL-17, IL-21) within total and subsets of CD4+TH cells were assessed using flow cytometry.RESULTS: ShK-186 reduced the expression level of IFNγ, TNFα, IL-4, IL-17, and IL-21 in CD4+TH cells from GPA-patients in vitro. Further analysis performed on sorted CD4+T cell subsets, revealed that ShK-186 predominantly inhibited the cytokine production of CD4+TEM cells. ShK-186 treatment reduced the production of the pro-inflammatory cytokines to the level seen in CD4+ TH cells from HCs.CONCLUSIONS: Modulation of cellular effector function by ShK-186 may constitute a novel treatment strategy for GPA with high specificity and less harmful side effects.</p

Kv1.3 blockade by ShK186 modulates CD4+ effector memory T-cell activity of patients with Granulomatosis with polyangiitis

OBJECTIVES: Granulomatosis with polyangiitis (GPA) is a chronic relapsing systemic autoimmune vasculitis. Current treatment of GPA is unsatisfactory as it relies on strong immunosuppressive regimens, with either cyclophosphamide or rituximab, that reduce the immunogenicity of several vaccines and are risk factors of severe form of COVID-19. This emphasizes the need to identify new drug target and to develop treatment strategies with less harmful side effects. Since CD4+ effector memory T cells (TEM) play a key role in the pathogenesis of GPA, we aimed in this study to modulate CD4+TEM cell activity via Kv1.3 blockade using the specific peptide inhibiter, ShK-186.METHODS: Peripheral blood of 27 GPA-patients in remission and 16 age- and sex-matched healthy controls (HCs) were pre-incubated in vitro in the presence or absence of ShK-186, followed by stimulation with PMA, calcium ionophore and brefeldin-A. The effect of ShK-186 on the cytokine production (IFNγ, TNFα, IL-4, IL-17, IL-21) within total and subsets of CD4+TH cells were assessed using flow cytometry.RESULTS: ShK-186 reduced the expression level of IFNγ, TNFα, IL-4, IL-17, and IL-21 in CD4+TH cells from GPA-patients in vitro. Further analysis performed on sorted CD4+T cell subsets, revealed that ShK-186 predominantly inhibited the cytokine production of CD4+TEM cells. ShK-186 treatment reduced the production of the pro-inflammatory cytokines to the level seen in CD4+ TH cells from HCs.CONCLUSIONS: Modulation of cellular effector function by ShK-186 may constitute a novel treatment strategy for GPA with high specificity and less harmful side effects.</p

Kv1.3 blockade by ShK186 modulates CD4+ effector memory T-cell activity of patients with Granulomatosis with polyangiitis

OBJECTIVES: Granulomatosis with polyangiitis (GPA) is a chronic relapsing systemic autoimmune vasculitis. Current treatment of GPA is unsatisfactory as it relies on strong immunosuppressive regimens, with either cyclophosphamide or rituximab, that reduce the immunogenicity of several vaccines and are risk factors of severe form of COVID-19. This emphasizes the need to identify new drug target and to develop treatment strategies with less harmful side effects. Since CD4+ effector memory T cells (TEM) play a key role in the pathogenesis of GPA, we aimed in this study to modulate CD4+TEM cell activity via Kv1.3 blockade using the specific peptide inhibiter, ShK-186.METHODS: Peripheral blood of 27 GPA-patients in remission and 16 age- and sex-matched healthy controls (HCs) were pre-incubated in vitro in the presence or absence of ShK-186, followed by stimulation with PMA, calcium ionophore and brefeldin-A. The effect of ShK-186 on the cytokine production (IFNγ, TNFα, IL-4, IL-17, IL-21) within total and subsets of CD4+TH cells were assessed using flow cytometry.RESULTS: ShK-186 reduced the expression level of IFNγ, TNFα, IL-4, IL-17, and IL-21 in CD4+TH cells from GPA-patients in vitro. Further analysis performed on sorted CD4+T cell subsets, revealed that ShK-186 predominantly inhibited the cytokine production of CD4+TEM cells. ShK-186 treatment reduced the production of the pro-inflammatory cytokines to the level seen in CD4+ TH cells from HCs.CONCLUSIONS: Modulation of cellular effector function by ShK-186 may constitute a novel treatment strategy for GPA with high specificity and less harmful side effects.</p

High-density rhesus macaque oligonucleotide microarray design using early-stage rhesus genome sequence information and human genome annotations

BACKGROUND: Until recently, few genomic reagents specific for non-human primate research have been available. To address this need, we have constructed a macaque-specific high-density oligonucleotide microarray by using highly fragmented low-pass sequence contigs from the rhesus genome project together with the detailed sequence and exon structure of the human genome. Using this method, we designed oligonucleotide probes to over 17,000 distinct rhesus/human gene orthologs and increased by four-fold the number of available genes relative to our first-generation expressed sequence tag (EST)-derived array. RESULTS: We constructed a database containing 248,000 exon sequences from 23,000 human RefSeq genes and compared each human exon with its best matching sequence in the January 2005 version of the rhesus genome project list of 486,000 DNA contigs. Best matching rhesus exon sequences for each of the 23,000 human genes were then concatenated in the proper order and orientation to produce a rhesus "virtual transcriptome." Microarray probes were designed, one per gene, to the region closest to the 3' untranslated region (UTR) of each rhesus virtual transcript. Each probe was compared to a composite rhesus/human transcript database to test for cross-hybridization potential yielding a final probe set representing 18,296 rhesus/human gene orthologs, including transcript variants, and over 17,000 distinct genes. We hybridized mRNA from rhesus brain and spleen to both the EST- and genome-derived microarrays. Besides four-fold greater gene coverage, the genome-derived array also showed greater mean signal intensities for genes present on both arrays. Genome-derived probes showed 99.4% identity when compared to 4,767 rhesus GenBank sequence tag site (STS) sequences indicating that early stage low-pass versions of complex genomes are of sufficient quality to yield valuable functional genomic information when combined with finished genome information from a closely related species. CONCLUSION: The number of different genes represented on microarrays for unfinished genomes can be greatly increased by matching known gene transcript annotations from a closely related species with sequence data from the unfinished genome. Signal intensity on both EST- and genome-derived arrays was highly correlated with probe distance from the 3' UTR, information often missing from ESTs yet present in early-stage genome projects

Analysis of the Macaca mulatta transcriptome and the sequence divergence between Macaca and human

We report the initial sequencing and comparative analysis of the Macaca mulatta transcriptome. Cloned sequences from 11 tissues, nine animals, and three species (M. mulatta, M. fascicularis, and M. nemestrina) were sampled, resulting in the generation of 48,642 sequence reads. These data represent an initial sampling of the putative rhesus orthologs for 6,216 human genes. Mean nucleotide diversity within M. mulatta and sequence divergence among M. fascicularis, M. nemestrina, and M. mulatta are also reported

Safety and pharmacodynamics of dalazatide, a Kv1.3 channel inhibitor, in the treatment of plaque psoriasis: A randomized phase 1b trial

<div><p>Background</p><p>Dalazatide is a specific inhibitor of the Kv1.3 potassium channel. The expression and function of Kv1.3 channels are required for the function of chronically activated memory T cells, which have been shown to be key mediators of autoimmune diseases, including psoriasis.</p><p>Objective</p><p>The primary objective was to evaluate the safety of repeat doses of dalazatide in adult patients with mild-to-moderate plaque psoriasis. Secondary objectives were to evaluate clinical proof of concept and the effects of dalazatide on mediators of inflammation in the blood and on chronically activated memory T cell populations.</p><p>Methods</p><p>Patients (<i>n</i> = 24) were randomized 5:5:2 to receive dalazatide at 30 mcg/dose, 60 mcg/dose, or placebo twice weekly by subcutaneous injection (9 doses total). Safety was assessed on the basis of physical and neurological examination and laboratory testing. Clinical assessments included body-surface area affected, Psoriasis Area and Severity Index (PASI), and investigator and patient questionnaires.</p><p>Results</p><p>The most common adverse events were temporary mild (Grade 1) hypoesthesia (<i>n</i> = 20; 75% placebo, 85% dalazatide) and paresthesia (<i>n</i> = 15; 25% placebo, 70% dalazatide) involving the hands, feet, or perioral area. Nine of 10 patients in the 60 mcg/dose group had a reduction in their PASI score between baseline and Day 32, and the mean reduction in PASI score was significant in this group (<i>P</i> < 0.01). Dalazatide treatment reduced the plasma levels of multiple inflammation markers and reduced the expression of T cell activation markers on peripheral blood memory T cells.</p><p>Limitations</p><p>The study was small and drug treatment was for a short duration (4 weeks).</p><p>Conclusion</p><p>This study indicates that dalazatide is generally well tolerated and can improve psoriatic skin lesions by modulating T cell surface and activation marker expression and inhibiting mediators of inflammation in the blood. Larger studies of longer duration are warranted.</p></div