Increased Soluble PD-1 Predicts Response to Nivolumab plus Ipilimumab in Melanoma

SIMPLE SUMMARY: Checkpoint inhibitors have emerged as an effective therapy for patients with metastatic melanoma significantly improving survival for these patients. Despite this, many patients do not respond to the therapy and no current biomarkers can identify responders from non-responders. Using machine learning, we analysed cytokine levels in serially collected liquid biopsy to identify cytokine changes associated with response to checkpoint inhibitors in advanced-stage melanoma patients. The results presented here highlight that serial measurements of cytokine levels are a strong predictor of treatment response. Particularly, we demonstrate that high increases of soluble PD-1 measured from baseline to on-treatment is significantly associated with superior PFS in patients treated with nivolumab plus ipililumab. These results suggest that monitoring cytokine levels using serial samples is informative of treatment response and can improve guidance of treatment modality and the outcome of cancer patients. ABSTRACT: Background: Checkpoint inhibitors have revolutionized the treatment of metastatic melanoma, yielding long-term survival in a considerable proportion of the patients. Yet, 40–60% of patients do not achieve a long-term benefit from such therapy, emphasizing the urgent need to identify biomarkers that can predict response to immunotherapy and guide patients for the best possible treatment. Here, we exploited an unsupervised machine learning approach to identify potential inflammatory cytokine signatures from liquid biopsies, which could predict response to immunotherapy in melanoma. Methods: We studied a cohort of 77 patients diagnosed with unresectable advanced-stage melanoma undergoing treatment with first-line nivolumab plus ipilimumab or pembrolizumab. Baseline and on-treatment plasma samples were tested for levels of PD-1, PD-L1, IFNγ, IFNβ, CCL20, CXCL5, CXCL10, IL6, IL8, IL10, MCP1, and TNFα and analyzed by Uniform Manifold Approximation and Projection (UMAP) dimension reduction method and k-means clustering analysis. Results: Interestingly, using UMAP analysis, we found that treatment-induced cytokine changes measured as a ratio between baseline and on-treatment samples correlated significantly to progression-free survival (PFS). For patients treated with nivolumab plus ipilimumab we identified a group of patients with superior PFS that were characterized by significantly higher baseline-to-on-treatment increments of PD-1, PD-L1, IFNγ, IL10, CXCL10, and TNFα compared to patients with worse PFS. Particularly, a high PD-1 increment was a strong individual predictor for superior PFS (HR = 0.13; 95% CI 0.034–0.49; p = 0.0026). In contrast, decreasing levels of IFNγ and IL6 and increasing levels of CXCL5 were associated with superior PFS in the pembrolizumab group, although none of the cytokines were individually predictors for PFS. Conclusions: In short, our study demonstrates that a high increment of PD-1 is associated with superior PFS in advanced-stage melanoma patients treated with nivolumab plus ipilimumab. In contrast, decreasing levels of IFNγ and IL6, and increasing levels of CXCL5 are associated with response to pembrolizumab. These results suggest that using serial samples to monitor changes in cytokine levels early during treatment is informative for treatment response

Frequency and Env determinants of HIV-1 subtype C strains from antiretroviral therapy-naive subjects that display incomplete inhibition by maraviroc

Background: Entry of human immunodeficiency virus type 1 (HIV-1) into cells involves the interaction of the viral gp120 envelope glycoproteins (Env) with cellular CD4 and a secondary coreceptor, which is typically one of the chemokine receptors CCR5 or CXCR4. CCR5-using (R5) HIV-1 strains that display reduced sensitivity to CCR5 antagonists can use antagonist-bound CCR5 for entry. In this study, we investigated whether naturally occurring gp120 alterations in HIV-1 subtype C (C-HIV) variants exist in antiretroviral therapy (ART)-naïve subjects that may influence their sensitivity to the CCR5 antagonist maraviroc (MVC). Results: Using a longitudinal panel of 244 R5 Envs cloned from 20 ART-naïve subjects with progressive C-HIV infection, we show that 40% of subjects (n = 8) harbored viruses that displayed incomplete inhibition by MVC, as shown by plateau's of reduced maximal percent inhibitions (MPIs). Specifically, when pseudotyped onto luciferase reporter viruses, 16 Envs exhibited MPIs below 98% in NP2-CCR5 cells (range 79.7-97.3%), which were lower still in 293-Affinofile cells that were engineered to express high levels of CCR5 (range 15.8-72.5%). We further show that Envs exhibiting reduced MPIs to MVC utilized MVC-bound CCR5 less efficiently than MVC-free CCR5, which is consistent with the mechanism of resistance to CCR5 antagonists that can occur in patients failing therapy. Mutagenesis studies identified strain-specific mutations in the gp120 V3 loop that contributed to reduced MPIs to MVC. Conclusions: The results of our study suggest that some ART-naïve subjects with C-HIV infection harbor HIV-1 with reduced MPIs to MVC, and demonstrate that the gp120 V3 loop region contributes to this phenotype

Human Cytomegalovirus Tegument Protein pp65 (pUL83) Dampens Type I Interferon Production by Inactivating the DNA Sensor cGAS without Affecting STING

The innate immune response plays a pivotal role during human cytomegalovirus (HCMV) primary infection. Indeed, HCMV infection of primary fibroblasts rapidly triggers strong induction of type I interferons (IFN-I) accompanied by proinflammatory cytokine release. Here, we show that primary human foreskin fibroblasts (HFFs) infected with a mutant HCMV TB40/E strain unable to express UL83-encoded pp65 (v65Stop) produce significantly higher IFN-\u3b2 levels than HFFs infected with the wild-type TB40/E strain or the pp65 revertant (v65Rev), suggesting that the tegument protein pp65 may dampen IFN-\u3b2 production. To clarify the mechanisms through which pp65 inhibits IFN-\u3b2 production, we analyzed the activation of the cGAS/STING/IRF3 axis in HFFs infected with either wild-type, v65Rev or the pp65-deficient mutant v65Stop. We found that pp65 selectively binds to cGAS and prevents its interaction with STING, thus inactivating the signaling pathway through the cGAS/STING/IRF3 axis. Consistently, addition of exogenous cGAMP to v65Rev infected cells triggered the production of IFN-\u3b2 levels similar to those observed with v65Stop infected cells confirming that pp65 inactivation of IFN-\u3b2 production occurs at the cGAS level. Notably, within the first 24 hours of HCMV infection, STING undergoes proteasome degradation independent of the presence or absence of pp65. Collectively, our data provide mechanistic insights into the interplay between HCMV pp65 and cGAS, leading to subsequent immune evasion by this prominent DNA virus.IMPORTANCE Primary human foreskin fibroblasts (HFFs) produce type I IFN (IFN-I) when infected with HCMV. However, we observed significantly higher IFN-\u3b2 levels when HFFs were infected with HCMV unable to express UL83-encoded pp65 (v65Stop), suggesting that pp65 (pUL83) may constitute a viral evasion factor. This study demonstrates that HCMV tegument protein pp65 inhibits IFN-\u3b2 production by binding and inactivating cGAS early during infection. In addition, this inhibitory activity specifically targets cGAS since it can be bypassed via the addition of exogenous cGAMP, even in presence of pp65. Notably, STING proteasome-mediated degradation was observed in both the presence and absence of pp65. Collectively, our data underscore the important role of tegument protein pp65 as a critical molecular hub in HCMV's evasion strategy to the innate immune response