Variable Alkaloid Defenses in the Dendrobatid Poison Frog Oophaga pumilio are Perceived as Differences in Palatability to Arthropods.

Conspicuously colored dendrobatid frogs sequester alkaloid defenses from dietary arthropods, resulting in considerable alkaloid variation among populations; however, little is known about how variation is perceived as a defense against predators. Previous studies have found variable alkaloids in the dendrobatid Oophaga pumilio to be associated with differences in toxicity to laboratory mice, suggesting variable defenses are important. Arthropods are natural predators that use chemoreception to detect prey, including frogs, and may therefore perceive variation in alkaloid profiles as differences in palatability. The goal of the present study is to determine how arthropods respond to variable alkaloid defenses in O. pumilio. Frog alkaloids were sampled from individual O. pumilio from ten geographic locations throughout the Bocas del Toro region of Panama and the Caribbean coast of Costa Rica. Alkaloid extracts were used in feeding bioassays with the vinegar fly Drosophila melanogaster and the ant Ectatomma ruidum. Both species of arthropods fed significantly less on frog alkaloid extracts when compared to controls, and differences in alkaloid palatability were observed among frog populations, as well as between sexes and life stages within a population. Differences in alkaloid quantity, richness, and type were the main predictors of arthropod palatability. Our findings also represent the first direct evidence of a palatability spectrum in a vertebrate that sequesters chemical defenses from dietary sources. Further, the presence of a palatability spectrum suggests that variable alkaloid defenses in O. pumilio are ecologically relevant and play an important role in natural predator-prey interactions, particularly with respect to arthropod predators.

Immune Therapy Targeting E6/E7 Oncogenes of Human Paillomavirus Type 6 (HPV-6) Reduces or Eliminates the Need for Surgical Intervention in the Treatment of HPV-6 Associated Recurrent Respiratory Papillomatosis.

: Background: Recurrent respiratory papillomatosis (RRP) is a rare disorder characterized by the generation of papillomas of the aerodigestive tract, usually associated with human papilloma virus (HPV) subtypes 6, 11. INO-3106 is a DNA plasmid-based immunotherapy targeting E6 and E7 proteins of HPV6, in order to create a robust immune T cell response. METHODS: Testing of INO-3016 in animal models confirmed immunogenicity of the DNA-based therapy. A single-site open-label Phase 1 study was initiated for patients with HPV6-positive RRP. Patients were dosed with INO-3106 with or without INO-9012, a DNA plasmid immunotherapy that encodes IL-12, delivered intramuscularly (IM) in combination with electroporation (EP) with the CELLECTRA® device. Patients received an escalating dose of INO-3106, 3 mg once and then 6 mg for three additional doses, each dose three weeks apart, with the third and fourth doses co-administered with INO-9012. The primary objective of the study was to evaluate the safety and tolerability of INO-3106 with and without INO-9012. The secondary objective was to determine cellular immune responses to INO-3106 with and without INO-9012. Exploratory objectives included preliminary clinical efficacy to the therapy. RESULTS: Three patients were enrolled in this study, of which two had RRP. Study therapy was well-tolerated, with no related serious adverse events and all related adverse events (AEs) were low-grade. Injection site pain was the most common related AE reported. Immunogenicity was evidenced by multiple immune assays showing engagement and expansion of an HPV6-specific cellular response, including cytotoxic T cells. Preliminary efficacy was demonstrated in patients with RRP in the form of reduction in need for surgical intervention for papilloma growth. Prior to intervention, both patients required surgical intervention approximately every 180 days. One patient demonstrated a greater than three-fold increase in surgery avoidance (584 days) and the other patient remains completely surgery-free as of the last contact at 915 days, a greater than 5-fold increase in surgery interval. CONCLUSION: INO-3106 with and without INO-9012 was well tolerated, immunogenic and demonstrated preliminary efficacy in patients with HPV6-associated RRP aerodigestive lesions. Further clinical study is indicated.

Immunotherapy Targeting HPV16/18 Generates Potent Immune Responses in HPV-Associated Head and Neck Cancer.

PURPOSE: Clinical responses with programmed death (PD-1) receptor-directed antibodies occur in about 20% of patients with advanced head and neck squamous cell cancer (HNSCCa). Viral neoantigens, such as the E6/E7 proteins of HPV16/18, are attractive targets for therapeutic immunization and offer an immune activation strategy that may be complementary to PD-1 inhibition. PATIENTS AND METHODS: We report phase Ib/II safety, tolerability, and immunogenicity results of immunotherapy with MEDI0457 (DNA immunotherapy targeting HPV16/18 E6/E7 with IL12 encoding plasmids) delivered by electroporation with CELLECTRA constant current device. Twenty-two patients with locally advanced, p16+ HNSCCa received MEDI0457. RESULTS: MEDI0457 was associated with mild injection site reactions, but no treatment-related grade 3-5 adverse events (AE) were noted. Eighteen of 21 evaluable patients showed elevated antigen-specific T-cell activity by IFNγ ELISpot, and persistent cellular responses surpassing 100 spot-forming units (SFUs)/106 peripheral blood mononuclear cells (PBMCs) were noted out to 1 year. Induction of HPV-specific CD8+ T cells was observed. MEDI0457 shifted the CD8+/FoxP3+ ratio in 4 of 5 post immunotherapy tumor samples and increased the number of perforin+ immune infiltrates in all 5 patients. One patient developed metastatic disease and was treated with anti-PD-1 therapy with a rapid and durable complete response. Flow-cytometric analyses revealed induction of HPV16-specific PD-1+ CD8+ T cells that were not found prior to MEDI0547 (0% vs. 1.8%). CONCLUSIONS: These data demonstrate that MEDI0457 can generate durable HPV16/18 antigen-specific peripheral and tumor immune responses. This approach may be used as a complementary strategy to PD-1/PD-L1 inhibition in HPV-associated HNSCCa to improve therapeutic outcomes.