Jigar V. Desai

Inhibition of B Cell Receptor Signaling by Ibrutinib in Primary CNS Lymphoma

Primary CNS lymphoma (PCNSL) harbors mutations that reinforce B cell receptor (BCR) signaling. Ibrutinib, a Brutons tyrosine kinase (BTK) inhibitor, targets BCR signaling and is particularly active in lymphomas with mutations altering the BCR subunit CD79B and MYD88. We performed a proof-of-concept phase Ib study of ibrutinib monotherapy followed by ibrutinib plus chemotherapy (DA-TEDDi-R). In 18 PCNSL patients, 94% showed tumor reductions with ibrutinib alone, including patients having PCNSL with CD79B and/or MYD88 mutations, and 86% of evaluable patients achieved complete remission with DA-TEDDi-R. Increased aspergillosis was observed with ibrutinib monotherapy and DA-TEDDi-R. Aspergillosis was linked to BTK-dependent fungal immunity in a murine model. PCNSL is highly dependent on BCR signaling, and ibrutinib appears to enhance the efficacy of chemotherapy.

An Ocular Commensal Protects against Corneal Infection by Driving an Interleukin-17 Response from Mucosal γδ T Cells

Summary Mucosal sites such as the intestine, oral cavity, nasopharynx, and vagina all have associated commensal flora. The surface of the eye is also a mucosal site, but proof of a living, resident ocular microbiome remains elusive. Here, we used a mouse model of ocular surface disease to reveal that commensals were present in the ocular mucosa and had functional immunological consequences. We isolated one such candidate commensal, Corynebacterium mastitidis, and showed that this organism elicited a commensal‐specific interleukin‐17 response from &ggr;&dgr; T cells in the ocular mucosa that was central to local immunity. The commensal‐specific response drove neutrophil recruitment and the release of antimicrobials into the tears and protected the eye from pathogenic Candida albicans or Pseudomonas aeruginosa infection. Our findings provide direct evidence that a resident commensal microbiome exists on the ocular surface and identify the cellular mechanisms underlying its effects on ocular immune homeostasis and host defense. Graphical Abstract Figure. No Caption available. HighlightsCorynebacterium mastitidis colonizes the mouse conjunctivaC. mastitidis induces interleukin‐17 production from mucosal &ggr;&dgr; T cellsIntroduction of the commensal to mice that lack it protects the eye from infectionTopical antibiotics cause the ocular surface to be more susceptible to infection &NA; Although the eye is a mucosal site, there has been a long‐standing controversy regarding whether a resident microbiome exists on the ocular surface. St. Leger et al. show that a microorganism that lives on the conjunctiva tunes local mucosal immunity and protects the eye from pathogenic infection.

VT-1161 protects mice against oropharyngeal candidiasis caused by fluconazole-susceptible and -resistant Candida albicans

Background Candida albicans, the most common human fungal pathogen, causes chronic mucosal infections in patients with inborn errors of IL-17 immunity that rely heavily on chronic, often lifelong, azole antifungal agents for treatment. However, a rise in azole resistance has predicated a need for developing new antifungal drugs. Objectives To test the in vitro and in vivo efficacy of VT-1161 and VT-1129 in the treatment of oropharyngeal candidiasis with azole-susceptible or -resistant C. albicans strains. Methods MICs of VT-1161, VT-1129 and nine licensed antifungal drugs were determined for 31 Candida clinical isolates. The drug concentrations in mouse serum and tongues were measured following oral administration. IL-17-signalling-deficient Act1-/- mice were infected with fluconazole-susceptible or fluconazole-resistant C. albicans strains, and the amount of mucosal fungal burden was determined after fluconazole or VT-1161 treatment. Results Fourteen isolates (45%) were not fluconazole susceptible (MIC ≥4 mg/L). VT-1161 and VT-1129 showed significant in vitro activity against the majority of the 31 mucosal clinical isolates (MIC50 0.03 and 0.06 mg/L, respectively), including Candida glabrata (MIC50, 0.125 and 0.25 mg/L, respectively). After oral doses, VT-1161 and VT-1129 concentrations in mouse serum and tongues were well above their MIC50 values. VT-1161 was highly effective as treatment of both fluconazole-susceptible and -resistant oropharyngeal candidiasis in Act1-/- mice. Conclusions VT-1129 and VT-1161 exhibit significant in vitro activity against Candida strains, including fluconazole-resistant C. albicans and C. glabrata. VT-1161 administration in mice results in significant mucosal drug accumulation and eradicates infection caused by fluconazole-susceptible and -resistant Candida strains.

Understanding the role of host immune responses in invasive candidiasis

Invasive candidiasis is the most common nosocomial bloodstream mycosis in the ICU affecting > 250,000 individuals annually worldwide [1]. It is most often caused by Candida albicans, a commensal yeast of mucosal surfaces, although infections by non-albicans species, including C. glabrata, C. tropicalis and C. parapsilosis are increasing [1]. The recent global emergence of C. auris raises public health concerns because of the species’ multidrug-resistance and resilience to antiseptics, its misidentification by routine microbiological techniques, and its persistence on human skin and hospital environment that causes hospital-associated horizontal transmission and outbreaks [2].

Microscopy of fungal biofilms

Fungal biofilms are heterogeneous, surface-associated colonies comprised of filamentous hyphae (chains of elongated cells), pseudohyphal cells, yeast-form cells, and various forms of extracellular matrix. When grown on a substratum under liquid culture medium, the microbial fungus Candida albicans forms dense biofilms that range in thickness from 100 to 600μm. Apical hyphae in the medium and invasive hyphae in the substratum may add greatly to the thickness and complexity of the biofilm. Because of the heterogeneity of the structure, and the large refractive index differences between cell walls, cytoplasm, and medium, fungal biofilms appear optically opaque. For fixed specimens that can be transferred out of an aqueous medium, refractive index matching methods provide a high degree of clarification. Confocal scanning, 2-photon scanning, or selective-plane illumination microscopy then can be used to obtain high-quality image data spanning the full thickness of the biofilm. Using refractive index matching and confocal microscopy, we have imaged many interesting features within wild-type, mutant, and engineered biofilms, including cellular phenotypes that vary with position, the effect of growth conditions, and gene expression through reporter constructs. This approach greatly expands the range of microscopical studies, allowing researchers to observe and quantify specific phenomena within medically or industrially relevant forms of microbial growth.

Aspergillosis, eosinophilic esophagitis, and allergic rhinitis in signal transducer and activator of transcription 3 haploinsufficiency

STAT3 haploinsufficiency caused by a novel STAT3 splice site mutation is associated with elevated IgE, allergic rhinitis, eosinophilic esophagitis, and invasive aspergillosis. This case expands our understanding of the spectrum of disease associated with STAT3 mutations.

VT-1598 inhibits the in vitro growth of mucosal Candida strains and protects against fluconazole-susceptible and -resistant oral candidiasis in IL-17 signalling-deficient mice

Background Chronic mucocutaneous candidiasis (CMC) treatment often induces drug resistance, posing long-term challenges. A novel broad-spectrum fungal CYP51 inhibitor, VT-1598, specifically targets fungal CYP51, but not human CYP enzymes. Objectives To determine the efficacy of VT-1598 in the treatment of oral Candida infection caused by fluconazole-susceptible and -resistant clinical isolates. Methods The MICs of VT-1598 and fluconazole for 28 Candida isolates recovered from patients with inherited CMC were determined using CLSI M27-A3 and M27-S4 guidelines. Plasma and tongue VT-1598 or fluconazole concentrations were measured in mice following oral administration to determine tissue distribution. Tongue fungal load was determined in IL-17 signalling-deficient Act1-/- mice following sublingual Candida albicans infection and oral treatment with fluconazole or VT-1598. Results Among the 28 Candida isolates, 10 (36%) had fluconazole MICs of ≥4 mg/L, whereas VT-1598 demonstrated potent in vitro activity against all isolates (MIC90, 0.125 mg/L). After oral administration, VT-1598 levels in mouse plasma and tongue were significantly greater than those of fluconazole. In vivo, VT-1598 exhibited significant efficacy against fluconazole-susceptible and -resistant C. albicans, even at low drug doses. Furthermore, after a 10 day washout period, tongue fungal burdens in fluconazole-treated mice returned to vehicle control levels, whereas, in contrast, they were undetectable in mice treated with VT-1598. Conclusions VT-1598 effectively controls in vitro growth of mucosally derived Candida clinical isolates, including fluconazole-resistant strains. In vivo, VT-1598 eliminates C. albicans, even after a long washout period or at low doses. Therefore, VT-1598 is a promising drug candidate that may significantly improve treatment options for CMC patients.

Human defects in STAT3 promote oral mucosal fungal and bacterial dysbiosis

Studies in patients with genetic defects can provide unique insights regarding the role of specific genes and pathways in humans. Patients with defects in the Th17/IL-17 axis, such as patients harboring loss-of-function STAT3 mutations (autosomal-dominant hyper IgE syndrome; AD-HIES) present with recurrent oral fungal infections. Our studies aimed to comprehensively evaluate consequences of STAT3 deficiency on the oral commensal microbiome. We characterized fungal and bacterial communities in AD-HIES in the presence and absence of oral fungal infection compared with healthy volunteers. Analyses of oral mucosal fungal communities in AD-HIES revealed severe dysbiosis with dominance of Candida albicans (C. albicans) in actively infected patients and minimal representation of health-associated fungi and/or opportunists. Bacterial communities also displayed dysbiosis in AD-HIES, particularly in the setting of active Candida infection. Active candidiasis was associated with decreased microbial diversity and enrichment of the streptococci Streptococcus oralis (S. oralis) and S. mutans, suggesting an interkingdom interaction of C. albicans with oral streptococci. Increased abundance of S. mutans was consistent with susceptibility to dental caries in AD-HIES. Collectively, our findings illustrate a critical role for STAT3/Th17 in the containment of C. albicans as a commensal organism and an overall contribution in the establishment of fungal and bacterial oral commensal communities.

The Role of Neutrophils in Host Defense Against Invasive Fungal Infections

Purpose of ReviewInvasive fungal infections caused by the commensal yeast Candida and the ubiquitous, inhaled mold Aspergillus have emerged as major causes of morbidity and mortality in critically ill and immunosuppressed patient populations. Here, we review how neutrophils contribute to effective immunity against these infections.Recent FindingsStudies in mouse models of invasive candidiasis and aspergillosis and observations in hematological patients with chemotherapy-induced neutropenia and in patients with primary immunodeficiency disorders that manifest with these infections have highlighted the critical role of neutrophils and have identified key immune factors that promote neutrophil-mediated effective host defense against invasive fungal disease.SummaryNeutrophils are crucial in host protection against invasive candidiasis and aspergillosis. Recent advances in our understanding of the molecular cues that mediate protective neutrophil recruitment and effector function against these infections hold promise for developing immune-based strategies to improve the outcomes of affected patients.

VT-1598 Inhibits the in vitro Growth of Mucosal Candida Isolates and Protects Against Oropharyngeal Candidiasis in IL-17 Deficient Mice

Abstract Background Patients with chronic mucocutaneous candidiasis (CMC) often develop azole-resistant Candida infections, making treatment difficult due to lack of oral antifungal drug options. VT-1598 is a novel broad-spectrum fungal CYP51 inhibitor designed for exquisite selectivity for the fungal target versus human CYP enzymes to circumvent classic azole side effects like drug–drug interactions. We report the efficacy of VT-1598 in the treatment of oral Candida infection (including by azole-resistant strains). Methods The in vitro MIC values of 28 Candida species isolated from patients with CMC due to AIRE mutations were tested against VT-1598 and fluconazole (FLC), using CLSI broth microdilution M27-S4. Plasma VT-1598 levels were measured using LC–MS/MS with electrospray ionization. Tongue fungal load was determined in IL-17 deficient Act1- /- mice following sublingual C. albicans infection and once-daily oral treatment for 4 days with 25 mg/kg FLC or 3.2, 8, and 20 mg/kg VT-1598 starting 18 hours post-infection. Results Among 28 Candida isolates tested (22 C. albicans, three C. glabrata, and one each of C. utilis, C. dubiliensis, and C. krusei), 10 (36%) were not susceptible to FLC, based on CLSI breakpoints (>4 mg/ml). Remarkably, all 28 isolates were highly susceptible to VT-1598 (MIC50 and MIC90, 0.06 and 0.125 mg/ml, respectively). Oral administration of VT-1598 led to mean drug levels in mouse plasma (2.0, 3.0, and 11 mg/ml at the low, mid, and high doses, respectively) that were higher than the MIC values. In vivo, VT-1598 was significantly more effective, compared with FLC, against FLC-susceptible and -resistant C. albicans, and led to elimination of fungal growth even at the lowest tested dose (3.2 mg/kg). After a 10-day washout period from the last dose, mice treated with VT-1598 did not have mucosal fungal growth, while mice treated with FLC had tongue fungal loads similar to vehicle control. Conclusion VT-1598 shows in vitro activity against mucosally derived Candida, including FLC-resistant strains. In vivo, VT-1598 achieves high plasma concentrations and eliminates viable C. albicans, even at low doses and after an extended washout period. These data indicate that VT-1598 may be a significantly improved treatment option for patients with CMC. Disclosures C. M. Yates, Viamet Pharmaceuticals, Inc.: Employee, Salary. O. J. Cohen, Viamet Pharmaceuticals, Inc.: Employee, Salary. R. J. Schotzinger, Viamet Pharmaceuticals, Inc.: Employee, Salary. E. P. Garvey, Viamet Pharmaceuticals, Inc.: Employee, Salary. M. Lionakis, Viamet Pharmaceuticals: Research support (CRADA), Research support