FOXD3 Regulates VISTA Expression in Melanoma.

Immune checkpoint inhibitors have improved patient survival in melanoma, but the innate resistance of many patients necessitates the investigation of alternative immune targets. Many immune checkpoint proteins lack proper characterization, including V-domain Ig suppressor of T cell activation (VISTA). VISTA expression on immune cells can suppress T cell activity; however, few studies have investigated its expression and regulation in cancer cells. In this study, we observe that VISTA is expressed in melanoma patient samples and cell lines. Tumor cell-specific expression of VISTA promotes tumor onset in vivo, associated with increased intratumoral T regulatory cells, and enhanced PDL-1 expression on tumor-infiltrating macrophages. VISTA transcript levels are regulated by the stemness factor Forkhead box D3 (FOXD3). BRAF inhibition upregulates FOXD3 and reduces VISTA expression. Overall, this study demonstrates melanoma cell expression of VISTA and its regulation by FOXD3, contributing to the rationale for therapeutic strategies that combine targeted inhibitors with immune checkpoint blockade

Tumour micro-environment elicits innate resistance to RAF inhibitors through HGF secretion.

Drug resistance presents a challenge to the treatment of cancer patients. Many studies have focused on cell-autonomous mechanisms of drug resistance. By contrast, we proposed that the tumour micro-environment confers innate resistance to therapy. Here we developed a co-culture system to systematically assay the ability of 23 stromal cell types to influence the innate resistance of 45 cancer cell lines to 35 anticancer drugs. We found that stroma-mediated resistance is common, particularly to targeted agents. We characterized further the stroma-mediated resistance of BRAF-mutant melanoma to RAF inhibitors because most patients with this type of cancer show some degree of innate resistance. Proteomic analysis showed that stromal cell secretion of hepatocyte growth factor (HGF) resulted in activation of the HGF receptor MET, reactivation of the mitogen-activated protein kinase (MAPK) and phosphatidylinositol-3-OH kinase (PI(3)K)-AKT signalling pathways, and immediate resistance to RAF inhibition. Immunohistochemistry experiments confirmed stromal cell expression of HGF in patients with BRAF-mutant melanoma and showed a significant correlation between HGF expression by stromal cells and innate resistance to RAF inhibitor treatment. Dual inhibition of RAF and either HGF or MET resulted in reversal of drug resistance, suggesting RAF plus HGF or MET inhibitory combination therapy as a potential therapeutic strategy for BRAF-mutant melanoma. A similar resistance mechanism was uncovered in a subset of BRAF-mutant colorectal and glioblastoma cell lines. More generally, this study indicates that the systematic dissection of interactions between tumours and their micro-environment can uncover important mechanisms underlying drug resistance

Driving with retinitis pigmentosa

Background: To establish the proportion of patients with retinitis pigmentosa (RP) meeting the Australian fitness to drive (FTD) visual standards. Methodology: A prospective consecutive case series of patients with a clinical or genetic diagnosis of RP. Data on age at symptom onset, current driving status, inheritance pattern, better eye visual acuity (BEVA), binocular Esterman visual field (BEVF) parameters, genotype and ability to meet the driving standards based on BEVA and BEVF were collected. Outcome measures included the proportion of RP patients overall meeting the standards and clinical predictors for passing. A sub-analysis was performed on those RP patients who reported to drive. Change in BEVA and BEVF parameters across age in specific genotype groups was assessed. Results: Overall, 228 patients with RP had a BEVF assessment. Only 39% (89/228) met the driving standards. Younger age at the time of testing was the only significant predictor (p \u3c 0.01) for passing. Of the 55% of RP patients who reported to drive, 52% (65/125) met the standards, decreasing to 14% in the 56- to 65-year-old age group. RP patients harbouring mutations in HK1 or RHO genes may have slower rates of decline in their VF parameters. Conclusion: Nearly 40% of RP patients met the driving standards. However, almost 50% of RP drivers were unaware of their failure to meet the current standards. BEVF testing is essential in the assessment of RP patients who are still driving. Phenotype and genotype predictors for passing the standards warrant further investigation. Abbreviation: FTD, fitness to drive; IRD, inherited retinal disease; RP, retinitis pigmentosa; RHO, rhodopsin; HK1, hexokinase 1; PRPF31 pre-mRNA processing factor 31; RPGR, retinitis pigmentosa GTPase regulator; VF, visual field; BEVA, better eye visual acuity; BEVF, binocular Esterman visual field

Expression of lymphotoxin-αβ on antigen-specific T cells is required for DC function

During an immune response, activated antigen (Ag)-specific T cells condition dendritic cells (DCs) to enhance DC function and survival within the inflamed draining lymph node (LN). It has been difficult to ascertain the role of the tumor necrosis factor (TNF) superfamily member lymphotoxin-αβ (LTαβ) in this process because signaling through the LTβ-receptor (LTβR) controls multiple aspects of lymphoid tissue organization. To resolve this, we have used an in vivo system where the expression of TNF family ligands is manipulated only on the Ag-specific T cells that interact with and condition Ag-bearing DCs. We report that LTαβ is a critical participant required for optimal DC function, independent of its described role in maintaining lymphoid tissue organization. In the absence of LTαβ or CD40L on Ag-specific T cells, DC dysfunction could be rescued in vivo via CD40 or LTβR stimulation, respectively, suggesting that these two pathways cooperate for optimal DC conditioning

Global distribution of invasive serotype 35D streptococcus pneumoniae isolates following introduction of 13-valent pneumococcal conjugate vaccine

A newly recognized pneumococcal serotype 35D, which differs from the 35B polysaccharide in structure and serology by not binding to factor serum 35a, was recently reported. The genetic basis for this distinctive serology is due to the presence of an inactivating mutation in wciG, which encodes an O-acetyltransferase responsible for O-acetylation of a galactofuranose. Here, we assessed the genomic data of a worldwide pneumococcal collection to identify serotype 35D isolates and understand their geographical distribution, genetic background and invasiveness potential. Of 21,980 pneumococcal isolates, 444 were originally typed as serotype 35B by PneumoCaT. Analysis of wciGrevealed 23 isolates from carriage (n=4) and disease (n=19) with partial or complete loss-of-funtion mutations, including mutations resulting in pre-mature stop codons (n=22) and an in-frame mutation (n=1). These were selected for further analysis. The putative 35D isolates were geographically widespread and 65.2% (15/23) of them was recovered after PCV13 introduction. Compared with serotype 35B, putative serotype 35D isolates have higher invasive disease potentials based on odds ratio (OR) (11.58; 95% CI, 1.42-94.19 vs 0.61; 95% CI, 0.40-0.92) and a higher prevalence of macrolide resistance mediated by mefA (26.1% vs 7.6%, p=0.009). Using Quellung, 50% (10/20) of viable isolates were serotype 35D, 25% (5/20) serotype 35B, and 25% (5/20) a mixture of 35B/35D. The discrepancy between phenotype and genotype requires further investigation. These findings illustrated a global distribution of an invasive serotype 35D among young children post-PCV13 introduction and underlined the invasive potential conferred by the loss of O-acetylation in the pneumococcal capsule

Genotype-specific lesion growth rates in stargardt disease

Reported growth rates (GR) of atrophic lesions in Stargardt disease (STGD1) vary widely. In the present study, we report the longitudinal natural history of patients with confirmed bial-lelic ABCA4 mutations from five genotype groups: c.6079C \u3e T, c.[2588G \u3e C;5603A \u3e T], c.3113C \u3e T, c.5882G \u3e A and c.5603A \u3e T. Fundus autofluorescence (AF) 30◦ × 30◦ images were manually seg-mented for boundaries of definitely decreased autofluorescence (DDAF). The primary outcome was the effective radius GR across five genotype groups. The age of DDAF formation in each eye was calculated using the x-intercept of the DDAF effective radius against age. Discordance between age at DDAF formation and symptom onset was compared. A total of 75 eyes from 39 STGD1 patients (17 male [44%]; mean ± SD age 45 ± 19 years; range 21–86) were recruited. Patients with c.3113C \u3e T or c.6079C \u3e T had a significantly faster effective radius GR at 0.17 mm/year (95% CI 0.12 to 0.22; p \u3c 0.001 and 0.14 to 0.21; p \u3c 0.001) respectively, as compared to those patients harbouring c.5882G \u3e A at 0.06 mm/year (95% CI 0.03–0.09), respectively. Future clinical trial design should consider the effect of genotype on the effective radius GR and the timing of DDAF formation relative to symptom onset

Lymphocytes from rheumatoid arthritis patients have elevated levels of intracellular peroxiredoxin 2, and a greater frequency of cells with exofacial peroxiredoxin 2, compared with healthy human lymphocytes

AbstractPeroxiredoxin 2 has immune regulatory functions, but its expression in human peripheral blood lymphocytes and levels in extracellular fluid in healthy subjects and rheumatoid arthritis patients are poorly described. In the present study, the median intracellular peroxiredoxin 2 protein content of lymphocytes from rheumatoid arthritis patients was more than two-fold higher compared with healthy subjects’ lymphocytes. Intracellular peroxiredoxin 3 levels were similar in healthy and rheumatoid arthritis lymphocytes. Flow cytometry detected peroxiredoxin 2 on the surface of ca. 8% of T cells and ca. 56% of B cells (median % values) of all subjects analyzed. Exofacial thioredoxin-1 was also observed. In the total lymphocyte population from rheumatoid arthritis patients, few cells (median, 6%) displayed surface peroxiredoxin 2. In contrast, a significantly increased proportion of interleukin-17+ve lymphocytes were exofacially peroxiredoxin 2+ve (median, 39%). Prdx2 was also detected in human extracellular fluids. We suggest that crucial inflammatory cell subsets, i.e. interleukin-17+ve T cells, exhibit increased exofacial redox-regulating enzymes and that peroxiredoxin 2 may be involved in the persistence of pro-inflammatory cells in chronic inflammation

Interpreting ambiguous ‘trace’ results in Schistosoma mansoni CCA Tests: Estimating sensitivity and specificity of ambiguous results with no gold standard

Background The development of new diagnostics is an important tool in the fight against disease. Latent Class Analysis (LCA) is used to estimate the sensitivity and specificity of tests in the absence of a gold standard. The main field diagnostic for Schistosoma mansoni infection, Kato-Katz (KK), is not very sensitive at low infection intensities. A point-of-care circulating cathodic antigen (CCA) test has been shown to be more sensitive than KK. However, CCA can return an ambiguous ‘trace’ result between ‘positive’ and ‘negative’, and much debate has focused on interpretation of traces results. Methodology/Principle findings We show how LCA can be extended to include ambiguous trace results and analyse S. mansoni studies from both Côte d’Ivoire (CdI) and Uganda. We compare the diagnostic performance of KK and CCA and the observed results by each test to the estimated infection prevalence in the population. Prevalence by KK was higher in CdI (13.4%) than in Uganda (6.1%), but prevalence by CCA was similar between countries, both when trace was assumed to be negative (CCAtn: 11.7% in CdI and 9.7% in Uganda) and positive (CCAtp: 20.1% in CdI and 22.5% in Uganda). The estimated sensitivity of CCA was more consistent between countries than the estimated sensitivity of KK, and estimated infection prevalence did not significantly differ between CdI (20.5%) and Uganda (19.1%). The prevalence by CCA with trace as positive did not differ significantly from estimates of infection prevalence in either country, whereas both KK and CCA with trace as negative significantly underestimated infection prevalence in both countries. Conclusions Incorporation of ambiguous results into an LCA enables the effect of different treatment thresholds to be directly assessed and is applicable in many fields. Our results showed that CCA with trace as positive most accurately estimated infection prevalence

Sibling concordance in symptom onset and atrophy growth rates in Stargardt disease using ultra-widefield fundus autofluorescence

Purpose: To investigate concordance in symptom onset, area of dark autofluorescence (DAF), and growth rate (GR) between Stargardt disease siblings at an age-matched time point. Methods: In this retrospective longitudinal study of sibling pairs with identical biallelic ABCA4 variants, age at symptom onset, best-corrected visual acuity, atrophy area, and effective radius of DAF on ultra-widefield fundus autofluorescence were recorded. Absolute intersibling differences for both eyes were compared with absolute interocular differences using the Mann-Whitney test. Results: Overall 39 patients from 19 families were recruited. In 16 families, age-matched best-corrected visual acuity and DAF were compared between siblings. In 8 families, DAF GR was compared. The median (range) absolute difference in age at symptom onset between siblings was 3 (0-35) years. Absolute intersibling differences in age-matched best-corrected visual acuity were greater than interocular differences (P = 0.01). Similarly, absolute intersibling differences in DAF area and radius were greater than interocular differences (P = 0.04 for area and P = 0.001 for radius). Differences between absolute interocular and intersibling GR were not statistically significant (P = 0.44 for area GR and P = 0.61 for radius GR). Conclusion: There was significant discordance in age-matched best-corrected visual acuity and DAF beyond the expected limits of interocular asymmetry. Lack of significant intersibling differences in GR warrants further investigation