Follicular CD8+ T Cells Are Elevated in HIV Infection and Induce PD-L1 on B Cells.

Follicular CD8+CXCR5+ T cells are a specialized CD8+ T cell subset with unique follicular-homing capabilities that have been reported to display effector functions in viral immunity, tumor immunity, and autoimmunity. CD8+CXCR5+ T cells exhibit B cell helper functions and express CD40L, ICOS, programmed cell death protein 1 (PD-1), and BCL-6, the transcriptional regulator of CD4+CXCR5+ T follicular helper cells and of germinal center B cells. HIV is known to be sequestered in lymphoid follicles, and CD8+CXCR5+ T cell frequency is a marker for disease severity, given that HIV-infected patients with lower numbers of circulating CD8+CXCR5+ T cells display lower CD4+ T cell counts. Likewise, several groups have reported a direct correlation between the quantity of CD8+CXCR5+ T cells and suppression of HIV viral load. In this study, we observed elevated absolute numbers of CD8+CXCR5+ and CD8+CXCR5+BCL-6+PD-1+ T cells in the blood of HIV-infected participants of the Multicenter AIDS Cohort Study. We further demonstrated in vitro that activated human CD8+CXCR5+ T cells isolated from peripheral blood and tonsil from healthy donors show increased CD40L expression and induce the production of PD ligand 1 (PD-L1)+IgG+ B cells. Moreover, absolute numbers of CD8+CXCR5+ T cells significantly and positively correlated with numbers of PD-L1+ B cells found in blood of HIV-infected individuals. Altogether, these results show that activated CD8+CXCR5+ T cells have the ability to activate B cells and increase the percentage of PD-L1+ and PD-L1+IgG+ B cells, which provides insights into the early events of B cell activation and differentiation and may play a role in disease progression and lymphomagenesis in HIV-infected individuals.

Immunization of Mice with Virus-Like Vesicles of Kaposi Sarcoma-Associated Herpesvirus Reveals a Role for Antibodies Targeting ORF4 in Activating Complement-Mediated Neutralization.

Infection by Kaposi sarcoma-associated herpesvirus (KSHV) can cause severe consequences, such as cancers and lymphoproliferative diseases. Whole inactivated viruses (WIV) with chemically destroyed genetic materials have been used as antigens in several licensed vaccines. During KSHV productive replication, virus-like vesicles (VLVs) that lack capsids and viral genomes are generated along with virions. Here, we investigated the immunogenicity of KSHV VLVs produced from a viral mutant that was defective in capsid formation and DNA packaging. Mice immunized with adjuvanted VLVs generated KSHV-specific T cell and antibody responses. Neutralization of KSHV infection by the VLV immune serum was low but was markedly enhanced in the presence of the complement system. Complement-enhanced neutralization and complement deposition on KSHV-infected cells was dependent on antibodies targeting viral open reading frame 4 (ORF4). However, limited complement-mediated enhancement was detected in the sera of a small cohort of KSHV-infected humans which contained few neutralizing antibodies. Therefore, vaccination that induces antibody effector functions can potentially improve infection-induced humoral immunity. Overall, our study highlights a potential benefit of engaging complement-mediated antibody functions in future KSHV vaccine development. IMPORTANCE KSHV is a virus that can lead to cancer after infection. A vaccine that prevents KSHV infection or transmission would be helpful in preventing the development of these cancers. We investigated KSHV VLV as an immunogen for vaccination. We determined that antibodies targeting the viral protein ORF4 induced by VLV immunization could engage the complement system and neutralize viral infection. However, ORF4-specific antibodies were seldom detected in the sera of KSHV-infected humans. Moreover, these human sera did not potently trigger complement-mediated neutralization, indicating an improvement that immunization can confer. Our study suggests a new antibody-mediated mechanism to control KSHV infection and underscores the benefit of activating the complement system in a future KSHV vaccine.

Associations between per- and poly-fluoroalkyl substance (PFAS) exposure and immune responses among women in the California Teachers study: A cross-sectional evaluation.

INTRODUCTION: Per- and polyfluoroalkyl substances (PFAS) are persistent environmental contaminants that have been linked to a number of health outcomes, including those related to immune dysfunction. However, there are limited numbers of epidemiological-based studies that directly examine the association between PFAS exposure and immune responses. METHODS: In this cross-sectional study nested in the California Teachers Study cohort, we measured nine PFAS analytes in serum. Of the 9 analytes, we further evaluated four (PFHxS [perfluorohexane sulfonate], PFNA [perfluorononanoic acid], PFOA [perfluorooctanoic acid], PFOS [perfluorooctanesulfonic acid]) that had detection levels of > 80 %, in relation to 16 systemic inflammatory/immune markers and corresponding immune pathways (Th1 [pro-inflammatory/macrophage activation], B-cell activation, and T-cell activation). Study participants (n = 722) were female, completed a questionnaire regarding various health measures and behaviors, and donated a blood sample between 2013-2016. The association between PFAS analytes and individual immune markers and pathways were evaluated by calculating odds ratios (OR) and 95 % confidence intervals (CI) in a logistic regression model. PFAS analytes were evaluated both as a dichotomous exposure (above or below the respective median) and as a continuous variable (per 1 unit increase [ng/mL]). RESULTS: The prevalence of detecting any PFAS analyte rose with increasing age, with the highest PFAS prevalence observed among those aged 75 + years and the lowest PFAS prevalence observed among those aged 40-49 years (study participant age range: 40-95 years). Significant associations with BAFF (B-cell activating factor) levels above the median were observed among participants with elevated (defined as above the median) levels of PFHxS (OR=1.53), PFOA (OR=1.43), and PFOS (OR=1.40). Similarly, there were statistically significant associations between elevated levels of PFHxS and TNFRII (tumor necrosis factor receptor 2) levels (OR=1.78) and IL2Rα (interleukin 2 receptor subunit alpha) levels (OR=1.48). We also observed significant inverse associations between elevated PFNA and sCD14 (soluble cluster of differentiation 14) (OR=0.73). No significant associations were observed between elevated PFNA and any immune marker. Evaluation of PFAS exposures as continuous exposures in association with dichotomized cytokines were generally consistent with the dichotomized associations. CONCLUSIONS: PFAS exposure was associated with altered levels of circulating inflammatory/immune markers; the associations were specific to PFAS analyte and immune marker. If validated, our results may suggest potential immune mechanisms underlying associations between the different PFAS analytes and adverse health outcomes.

Host characteristics associated with serologic inflammatory biomarkers in women.

BACKGROUND: There is growing evidence that exposure to low-grade inflammation may be associated with adverse health outcomes. METHODS: We conducted a cross-sectional study within the California Teachers Study prospective cohort, among female participants who had completed a questionnaire that asked about their health behaviors (e.g., diabetes, physical activity, body mass index, medication use) and who had donated blood within a year of their questionnaire. 822 women with stored serum were evaluated for 16 immune biomarkers. In addition, four immune pathways were constructed: Th1, pro-inflammatory/macrophage activation, B-cell activation, and T-cell activation. Odds ratios (ORs) and 95% confidence intervals (CI) for the association between host characteristics and immune biomarkers were assessed using logistic regression models. RESULT: Compared to women of a normal BMI, obese women (>30 kg/m2) were positively associated with sTNFR2, CD27, IL6, CXCL13, sIL-2Rα, and IL6Ra levels above the median, with odds ratios ranging from 1.5 to 6.0. The pro-inflammatory/macrophage activation pathway was positively associated with diabetes (OR = 2.12, 95% CI = 1.14-3.95), fueled by individual associations between diabetes and sTNF-R2, TNFα and sCD27. Physical activity was inversely associated with sTNF-R2, TNFα, CXCL13, IL6, IL10, and IFN-γ levels, particularly for the highest category of activity (5.88+ hours/week) (ORs = 0.32-0.69). In pathway-based analyses, the Th1 pathway which includes decreased levels of IL4 and IL10 was positively associated with elevated physical activity (OR = 1.5). In contrast, the pro-inflammatory, B- and T-cell activation pathways were positively associated with higher BMI (OR ranging from 1.6 to 3) and inversely associated with increasing levels of physical activity. CONCLUSIONS: Several host characteristics were associated with circulating levels of immune biomarkers, including markers of inflammation. Further understanding of associations between immune marker profiles with human disease are warranted.

HIV-infection and cocaine use regulate semen extracellular vesicles proteome and miRNAome in a manner that mediates strategic monocyte haptotaxis governed by miR-128 network.

BACKGROUND: Extracellular vesicles (EVs) are regulators of cell-cell interactions and mediators of horizontal transfer of bioactive molecules between cells. EV-mediated cell-cell interactions play roles in physiological and pathophysiological processes, which maybe modulated by exposure to pathogens and cocaine use. However, the effect of pathogens and cocaine use on EV composition and function are not fully understood. RESULTS: Here, we used systems biology and multi-omics analysis to show that HIV infection (HIV +) and cocaine (COC) use (COC +) promote the release of semen-derived EVs (SEV) with dysregulated extracellular proteome (exProtein), miRNAome (exmiR), and exmiR networks. Integrating SEV proteome and miRNAome revealed a significant decrease in the enrichment of disease-associated, brain-enriched, and HIV-associated miR-128-3p (miR-128) in HIV + COC + SEV with a concomitant increase in miR-128 targets-PEAK1 and RND3/RhoE. Using two-dimensional-substrate single cell haptotaxis, we observed that in the presence of HIV + COC + SEV, contact guidance provided by the extracellular matrix (ECM, collagen type 1) network facilitated far-ranging haptotactic cues that guided monocytes over longer distances. Functionalizing SEV with a miR-128 mimic revealed that the strategic changes in monocyte haptotaxis are in large part the result of SEV-associated miR-128. CONCLUSIONS: We propose that compositionally and functionally distinct HIV + COC + and HIV-COC- SEVs and their exmiR networks may provide cells relevant but divergent haptotactic guidance in the absence of chemotactic cues, under both physiological and pathophysiological conditions.

CloneRetriever: An Automated Algorithm to Identify Clonal B and T Cell Gene Rearrangements by Next-Generation Sequencing for the Diagnosis of Lymphoid Malignancies.

BACKGROUND: Clonal immunoglobulin and T-cell receptor rearrangements serve as tumor-specific markers that have become mainstays of the diagnosis and monitoring of lymphoid malignancy. Next-generation sequencing (NGS) techniques targeting these loci have been successfully applied to lymphoblastic leukemia and multiple myeloma for minimal residual disease detection. However, adoption of NGS for primary diagnosis remains limited. METHODS: We addressed the bioinformatics challenges associated with immune cell sequencing and clone detection by designing a novel web tool, CloneRetriever (CR), which uses machine-learning principles to generate clone classification schemes that are customizable, and can be applied to large datasets. CR has 2 applications-a "validation" mode to derive a clonality classifier, and a "live" mode to screen for clones by applying a validated and/or customized classifier. In this study, CR-generated multiple classifiers using 2 datasets comprising 106 annotated patient samples. A custom classifier was then applied to 36 unannotated samples. RESULTS: The optimal classifier for clonality required clonal dominance ≥4.5× above background, read representation ≥8% of all reads, and technical replicate agreement. Depending on the dataset and analysis step, the optimal algorithm yielded sensitivities of 81%-90%, specificities of 97%-100%, areas under the curve of 91%-94%, positive predictive values of 92-100%, and negative predictive values of 88%-98%. Customization of the algorithms yielded 95%-100% concordance with gold-standard clonality determination, including rescue of indeterminate samples. Application to a set of unknowns showed concordance rates of 83%-96%. CONCLUSIONS: CR is an out-of-the-box ready and user-friendly software designed to identify clonal rearrangements in large NGS datasets for the diagnosis of lymphoid malignancies.

The Role of Microbial Translocation and Immune Activation in AIDS-Associated Non-Hodgkin Lymphoma Pathogenesis: What Have We Learned?

Human immunodeficiency virus (HIV) infection is associated with greatly increased risk for development of non-Hodgkin lymphoma (NHL). Nearly all acquired immunodeficiency syndrome (AIDS)-associated NHL (AIDS-NHL) is of B-cell origin. Two major mechanisms are believed to contribute to the genesis of AIDS-NHL: (1) loss of immunoregulation of Epstein-Barr virus (EBV)+ B cells, resulting from impaired T-cell function late in the course of HIV disease and (2) chronic B-cell activation, leading to DNA-modifying events that contribute to oncogene mutations/ translocations. HIV infection has long been known to be associated with chronic inflammation and polyclonal B-cell activation, and more recently, microbial translocation. Microbial translocation is bacterial product leakage from gut lumen into the peripheral circulation, resulting in high levels of lipopolysaccharide (LPS) in the peripheral circulation, leading to chronic immune activation and inflammation. We review recent literature linking microbial translocation to lymphom-agenesis. This includes epidemiological studies of biomarkers of microbial translocation with risk of AIDS-NHL and emerging data on the mechanisms by which microbial translocation may lead to AIDS-NHL development.

Urbanization and anticoagulant poisons promote immune dysfunction in bobcats.

Understanding how human activities influence immune response to environmental stressors can support biodiversity conservation across increasingly urbanizing landscapes. We studied a bobcat (Lynx rufus) population in urban southern California that experienced a rapid population decline from 2002-2005 due to notoedric mange. Because anticoagulant rodenticide (AR) exposure was an underlying complication in mange deaths, we aimed to understand sublethal contributions of urbanization and ARs on 65 biochemical markers of immune and organ function. Variance in immunological variables was primarily associated with AR exposure and secondarily with urbanization. Use of urban habitat and AR exposure has pervasive, complex and predictable effects on biochemical markers of immune and organ function in free-ranging bobcats that include impacts on neutrophil, lymphocyte and cytokine populations, total bilirubin and phosphorus. We find evidence of both inflammatory response and immune suppression associated with urban land use and rat poison exposure that could influence susceptibility to opportunistic infections. Consequently, AR exposure may influence mortality and has population-level effects, as previous work in the focal population has revealed substantial mortality caused by mange infection. The secondary effects of anticoagulant exposure may be a worldwide, largely unrecognized problem affecting a variety of vertebrate species in human-dominated environments.

BST2/Tetherin is constitutively expressed on human thymocytes with the phenotype and function of Treg cells.

In contrast to peripheral plasmacytoid DCs (pDCs), thymic pDCs constitutively express low levels of IFN-α. This leads to induction of interferon secondary genes (ISGs) in medullary thymocytes, raising the question whether IFN-α may play a role in T-cell development. When characterizing further differences between peripheral and thymic pDCs, we found that thymic pDCs have a phenotype consistent with an "activated signature" including expression of TNF-α and bone marrow stromal cell antigen 2 (BST2), but no expression of ILT7. Given that BST2 is induced by IFN-α, and IFN-α secretion is controlled by interaction between ILT7 and BST2, this regulatory pathway is apparently lost in thymic pDCs. Further, we also show that BST2 is constitutively expressed on a subset of medullary thymocytes at the mRNA and protein level reflecting a history of IFN-α transduced signals. The majority of BST2(+) thymocytes express CCR5 rendering them prevalent targets for R5-tropic HIV infection. Moreover, BST2(+) thymocytes express Foxp3 and CD25, consistent with the phenotype of natural Treg cells, and exert suppressive activity as they impair the proliferation of autologous CD3(+) thymocytes. Collectively, our results suggest that low levels of IFN-α secreted by thymic pDCs play an important role in the development of natural Treg cells.

Characterization of unique B-cell populations in the circulation of people living with HIV prior to non-Hodgkin lymphoma diagnosis.

People living with HIV (PLWH) are at higher risk of developing lymphoma. In this study, we performed cytometry by time-of-flight (CyTOF) on peripheral blood mononuclear cells of cART-naïve HIV+ individuals and cART-naïve HIV+ individuals prior to AIDS-associated non-Hodgkin lymphoma (pre-NHL) diagnosis. Participants were enrolled in the Los Angeles site of the MACS/WIHS Combined Cohort Study (MWCCS). Uniform Manifold Approximation and Projection (UMAP) and unsupervised clustering analysis were performed to identify differences in the expression of B-cell activation markers and/or oncogenic markers associated with lymphomagenesis. CD10+CD27- B cells, CD20+CD27- B cells, and B-cell populations with aberrant features (CD20+CD27+CXCR4+CD71+ B cells and CD20+CXCR4+cMYC+ B cells) were significantly elevated in HIV+ cART-naïve compared to HIV-negative samples. CD20+CD27+CD24+CXCR4+CXCR5+ B cells, CD20+CD27+CD10+CD24+CXCR4+cMYC+ B cells, and a cluster of CD20+CXCR4hiCD27-CD24+CXCR5+CD40+CD4+AICDA+ B cells were significantly elevated in HIV+ pre-NHL (cART-naïve) compared to HIV+ cART-naïve samples. A potentially clonal cluster of CD20+CXCR4+CXCR5+cMYC+AICDA+ B cells and a cluster of germinal center B-cell-like cells (CD19-CD20+CXCR4+Bcl-6+PD-L1+cMYC+) were also found in the circulation of HIV+ pre-NHL (cART-naïve) samples. Moreover, significantly elevated clusters of CD19+CD24hiCD38hi cMYC+ AICDA+ B regulatory cells were identified in HIV+ pre-NHL (cART-naïve) compared to HIV+ cART-naïve samples. The present study identifies unique B-cell subsets in PLWH with potential pre-malignant features that may contribute to the development of pre-tumor B cells in PLWH and that may play a role in lymphomagenesis.

Extracellular vesicles as biomarkers for AIDS-associated non-Hodgkin lymphoma risk.

Introduction: Extracellular vesicles are membrane-bound structures secreted into the extracellular milieu by cells and can carry bioactive molecules. There is emerging evidence suggesting that EVs play a role in the diagnosis, treatment, and prognosis of certain cancers. In this study, we investigate the association of EVs bearing PD-L1 and molecules important in B-cell activation and differentiation with AIDS-NHL risk. Methods: EVs were isolated from archived serum collected prior to the diagnosis of AIDS-NHL in cases (N = 51) and matched HIV+ controls (N = 52) who were men enrolled in the Los Angeles site of the MACS/WIHS Combined Cohort Study (MWCCS). Serum specimens of AIDS-NHL cases were collected at a mean time of 1.25 years (range of 2 to 36 months) prior to an AIDS-NHL diagnosis. The expression of PD-L1 and other molecules on EVs (CD40, CD40L, TNF-RII, IL-6Rα, B7-H3, ICAM-1, and FasL) were quantified by Luminex multiplex assay. Results and discussion: We observed significantly higher levels of EVs bearing PD-L1, CD40, TNF-RII and/or IL-6Rα in AIDS-NHL cases compared with controls. Using multivariate conditional logistic regression models adjusted for age and CD4+ T-cell count, we found that EVs bearing PD-L1 (OR = 1.93; 95% CI: 1.10 - 3.38), CD40 (OR = 1.97, 95% CI: 1.09 - 3.58), TNF-RII (OR = 5.06; 95% CI: 1.99 - 12.85) and/or IL-6Rα (OR = 4.67; 95% CI: 1.40 - 15.53) were significantly and positively associated with AIDS-NHL risk. In addition, EVs bearing these molecules were significantly and positively associated with non-CNS lymphoma: PD-L1 (OR = 1.94; 95% CI: 1.01 - 3.72); CD40 (OR = 2.66; 95% CI: 1.12 - 6.35); TNF-RII (OR = 9.64; 95% CI: 2.52 - 36.86); IL-6Rα (OR = 8.34; 95% CI: 1.73 - 40.15). These findings suggest that EVs bearing PD-L1, CD40, TNF-RII and/or IL-6Rα could serve as biomarkers for the early detection of NHL in PLWH.

Characterization of T Follicular Helper Cells and T Follicular Regulatory Cells in HIV-Infected and Sero-Negative Individuals.

Humoral immune response is important in fighting pathogens by the production of specific antibodies by B cells. In germinal centers, T follicular helper (TFH) cells provide important help to B-cell antibody production but also contribute to HIV persistence. T follicular regulatory (TFR) cells, which inhibit the function of TFH cells, express similar surface markers. Since FOXP3 is the only marker that distinguishes TFR from TFH cells it is unknown whether the increase in TFH cells observed in HIV infection and HIV persistence may be partly due to an increase in TFR cells. Using multicolor flow cytometry to detect TFH and TFR cells in cryopreserved peripheral blood mononuclear cells from HIV-infected and non-infected participants in the UCLA Multicenter AIDS Cohort Study (MACS), we identified CD3+CXCR5+CD4+CD8-BCL6+ peripheral blood TFH (pTFH) cells and CD3+CXCR5+CD4+CD8-FOXP3+ peripheral blood TFR (pTFR) cells. Unlike TFR cells in germinal centers, pTFR cells do not express B cell lymphoma 6 (BCL6), a TFH cell master transcriptional regulator. Our major findings are that the frequency of pTFH cells, but not pTFR cells was higher in HIV-infected participants of the MACS and that pTFH cells expressed less CCR5 in HIV-infected MACS participants. Constitutive expression of CCR5 in TFR cells supports their potential to contribute to HIV persistence.

Circulating markers of microbial translocation and host response to bacteria with risk of colorectal cancer: a prospective, nested case-control study in men.

BACKGROUND: Gut microbial dysbiosis contributes to colorectal cancer (CRC) pathogenesis, possibly mediated in part by increased intestinal permeability to endotoxin lipopolysaccharide (LPS), microbial translocation, and subsequent endotoxemia and inflammation. However, epidemiologic evidence linking circulating markers of microbial translocation with CRC risk is limited. METHODS: We conducted a prospective, nested case-control study of 261 incident CRC cases and 261 controls (matched on age and time of blood draw) among 18,159 men with pre-diagnostic blood specimens in the Health Professionals Follow-Up Study (1993-2009). We examined three complementary markers of microbial translocation and host response to bacteria, including LPS-binding protein (LBP), soluble CD14 (sCD14), and endotoxincore antibody (EndoCAb) immunoglobulin M (IgM), with subsequent risk of CRC. Unconditional logistic regressions were used to estimate odds ratios (ORs) and 95% confidence intervals (CIs). FINDINGS: Pre-diagnostic circulating levels of sCD14 were associated with a higher risk of incident CRC. Compared to men in the lowest quartile, the multivariable OR was 1.90 (95% CI, 1.13-3.22) for men in the highest quartile (OR per standard deviation [SD] increase, 1.28; 95%CI 1.06-1.53; Ptrend = 0.01). This positive association remained similar after adjusting for C-reactive protein, interleukin-6, and soluble tumor necrosis factor receptor-2, and within strata of putative CRC risk factors. We also observed a suggestive inverse association between EndoCAb IgM and risk of CRC (OR per SD increase, 0.84; 95%CI 0.69-1.02; Ptrend = 0.09). INTERPRETATION: Microbial translocation and host response to bacteria, as reflected by sCD14, is associated with risk of incident CRC in men. FUNDING: US National Institutes of Health.

Predictive Value of Serum Biomarkers for Response of Limited-Stage AIDS-Associated Kaposi Sarcoma to Antiretroviral Therapy With or Without Concomitant Chemotherapy in Resource-Limited Settings.

BACKGROUND: Guidelines for limited-stage human immunodeficiency virus-associated Kaposi sarcoma (AIDS/KS) recommend antiretroviral therapy (ART) as initial treatment. However, many such individuals show worsening KS and require additional chemotherapy. Methods to identify such patients are lacking. SETTING: We studied whether serum levels of biomarkers associated with angiogenesis, systemic inflammation, and immune activation, which are elevated in HIV-infected individuals and implicated in the development of KS, could prospectively identify individuals with limited-stage AIDS-KS who would benefit from chemotherapy administered with ART. METHODS: Serum specimens were obtained from participants in a randomized trial evaluating the value of adding oral etoposide chemotherapy to ART in treatment-naïve people with limited-stage AIDS-KS in resource-limited settings. Serum biomarkers of inflammation (C-reactive protein [CRP], interleukin [IL]-6, IL-8, IL-10, granulocyte colony stimulating factor, soluble tumor necrosis factor receptor-2), immune system activation (soluble IL-2 receptor alfa, C-X-C motif chemokine ligand 10/interferon gamma-induced protein 10, C-C motif ligand 2/monocyte chemoattractant protein 1), and angiogenesis (vascular endothelial growth factor, matrix metalloproteinase-2, -9, endoglin, hepatocyte growth factor) were measured at entry to determine whether baseline levels are associated with KS response. On-treatment changes in biomarker levels were determined to assess how etoposide modifies the effects of ART. RESULTS: Pretreatment CRP and IL-10 were higher in those whose KS progressed, and lowest in those who had good clinical responses. Pretreatment CRP, IL-6, and soluble tumor necrosis factor receptor-2 showed significant associations with KS progression at the week-48 primary endpoint. Immediate etoposide led to lower inflammation biomarker levels compared with ART alone. Early KS progression was associated with elevated pretreatment levels of inflammation-associated biomarkers and increasing levels post-treatment. CONCLUSIONS: Quantifying serum biomarkers, especially CRP, may help identify persons with AIDS-KS who would benefit from early introduction of chemotherapy in addition to ART.

Plasma extracellular vesicles bearing PD-L1, CD40, CD40L or TNF-RII are significantly reduced after treatment of AIDS-NHL.

Emerging evidence shows that tumor cells secrete extracellular vesicles (EVs) that carry bioactive cell surface markers, such as programmed death-ligand 1 (PD-L1), which can modulate immune responses and inhibit anti-tumor responses, potentially playing a role in lymphomagenesis and in promoting the growth of these cancers. In this study, we investigated the role of EVs expressing cell surface molecules associated with B cell activation and immune regulation. We measured levels of EVs derived from plasma from 57 subjects with AIDS-related non-Hodgkin lymphoma (AIDS-NHL) enrolled in the AIDS Malignancies Consortium (AMC) 034 clinical trial at baseline and post-treatment with rituximab plus concurrent infusional EPOCH chemotherapy. We found that plasma levels of EVs expressing PD-L1, CD40, CD40L or TNF-RII were significantly reduced after cancer treatment. AIDS-NHL patients with the diffuse large B cell lymphoma (DLBCL) tumor subtype had decreased plasma levels of EVs bearing PD-L1, compared to those with Burkitt's lymphoma. CD40, CD40L and TNF-RII-expressing EVs showed a significant positive correlation with plasma levels of IL-10, CXCL13, sCD25, sTNF-RII and IL-18. Our results suggest that patients with AIDS-NHL have higher levels of EVs expressing PD-L1, CD40, CD40L or TNF-RII in circulation before cancer treatment and that levels of these EVs are associated with levels of biomarkers of microbial translocation and inflammation.

Comparative outcomes for mature T-cell and NK/T-cell lymphomas in people with and without HIV and to AIDS-defining lymphomas.

There are no studies comparing the prognosis for mature T-cell lymphoma (TCL) in people with HIV (PWH) to people without HIV (PWoH) and to AIDS-defining B-cell lymphomas (A-BCLs) in the modern antiretroviral therapy era. North American AIDS Cohort Collaboration on Research and Design and Comprehensive Oncology Measures for Peripheral T-cell Lymphoma Treatment are cohorts that enroll patients diagnosed with HIV and TCL, respectively. In our study, 52, 64, 101, 500, and 246 PWH with histologic confirmation of TCL, primary central nervous system lymphoma, Burkitt's lymphoma, diffuse large B-cell lymphoma (DLBCL), and Hodgkin's lymphoma (HL), respectively, and 450 TCLs without HIV were eligible for analysis. At the time of TCL diagnosis, anaplastic large-cell lymphoma (ALCL) was the most common TCL subtype within PWH. Although PWH with TCL diagnosed between 1996 and 2009 experienced a low 5-year survival probability at 0.23 (95% confidence interval [CI]: 0.13, 0.41), we observed a marked improvement in their survival when diagnosed between 2010 and 2016 (0.69; 95% CI: 0.48, 1; P = .04) in contrast to TCLs among PWoH (0.45; 95% CI: 0.41, 0.51; P = .53). Similarly, PWH with ALCLs diagnosed between 1996 and 2009 were associated with a conspicuously inferior 5-year survival probability (0.17; 95% CI: 0.07, 0.42) and consistently lagged behind A-BCL subtypes such as Burkitt's (0.43; 95% CI:0.33, 0.57; P = .09) and DLBCL (0.17; 95% CI: 0.06, 0.46; P = .11) and behind HL (0.57; 95% CI: 0.50, 0.65; P < .0001). Despite a small number, those diagnosed between 2010 and 2016 experienced a remarkable improvement in survival (0.67; 95% CI: 0.3, 1) in comparison with PWoH (0.76; 95% CI: 0.66, 0.87; P = .58). Thus, our analysis confirms improved overall survival for aggressive B- and T-cell malignancies among PWH in the last decade.

Immune Activation and Microbial Translocation as Prognostic Biomarkers for AIDS-Related Non-Hodgkin Lymphoma in the AMC-034 Study.

PURPOSE: AIDS-related non-Hodgkin lymphoma (ARL) is the most common cancer in HIV-infected individuals in the United States and other countries in which HIV-positive persons have access to effective combination antiretroviral therapy (cART). Our prior work showed that pretreatment/postdiagnosis plasma levels of some cytokines, such as IL6, IL10, and CXCL13, have the potential to serve as indicators of clinical response to treatment and survival in ARL. The aims of this study were to identify novel prognostic biomarkers for response to treatment and/or survival in persons with ARL, including biomarkers of microbial translocation and inflammation. EXPERIMENTAL DESIGN: We quantified plasma levels of several biomarkers (sCD14, LBP, FABP2, EndoCab IgM, IL18, CCL2/MCP-1, sCD163, IP-10/CXCL10, TARC/CCL17, TNFα, BAFF/BLyS, sTNFRII, sCD44, and sIL2Rα/sCD25) by multiplexed immunometric assays (Luminex) or ELISA in plasma specimens obtained from ARL patients enrolled in the AMC-034 trial, which compared infusional combination chemotherapy (EPOCH: etoposide, vincristine, doxorubicin, cyclophosphamide, and prednisone) with concurrent or sequential rituximab. Plasma was collected prior to the initiation of therapy (n = 57) and after treatment initiation (n = 55). RESULTS: We found that several biomarkers decreased significantly after treatment, including TNFα, sCD25, LBP, and TARC (CCL17). Moreover, pretreatment plasma levels of BAFF, sCD14, sTNFRII, and CCL2/MCP-1 were univariately associated with overall survival, and pretreatment levels of BAFF, sTNFRII, and CCL2/MCP-1 were also associated with progression-free survival. CONCLUSIONS: Our results suggest that patients with ARL who responded to therapy had lower pretreatment levels of inflammation and microbial translocation as compared with those who did not respond optimally.

Self-Reported Cannabis Use and Markers of Inflammation in Men Who Have Sex With Men With and Without HIV.

Background: Chronic inflammation contributes to aging and organ dysfunction in the general population, and is a particularly important determinant of morbidity and mortality among people with HIV (PWH). The effect of cannabis use on chronic inflammation is not well understood among PWH, who use cannabis more frequently than the general population. Materials and Methods: We evaluated participants in the Multicenter AIDS Cohort Study (MACS) beginning in 2004 with available data on cannabis use and inflammatory biomarkers. Associations of current cannabis use with plasma concentrations of inflammatory markers were adjusted for hepatitis C, tobacco smoking, and comorbidities. Markers were analyzed individually and in exploratory factor analysis (EFA). Results: We included 1352 men within the MACS. Twenty-seven percent of HIV-negative men, 41% of HIV viremic men, and 35% of virologically suppressed men reported cannabis use at baseline. Among cannabis users, 20-25% in all groups defined by HIV serostatus were daily users, and the same proportion reported weekly use. The remaining ∼50% of users in all groups reported monthly or less frequent use. Four biomarker groupings were identified by EFA: Factor 1: immune activation markers; Factor 2: proinflammatory cytokines; Factor 3: Th1- and Th2-promoting cytokines; and Factor 4: inflammatory chemokines. In EFA, daily users had 30% higher levels of Factor 2 biomarkers than nonusers (p=0.03); this was the only statistically significant difference by cannabis use status. Among individual markers, concentrations of IL-1ß, IL-2, IL-6, and IL-8 (Factor 2); IL-10 (Factor 3); and BAFF (Factor 1) were higher (p<0.05) among daily cannabis users than among nonusers, after adjusting for HIV serostatus and other covariates. Discussion: Associations between daily cannabis use and proinflammatory biomarker levels did not differ by HIV serostatus. Further prospective studies with measured cannabis components are needed to clarify the impact of these compounds on inflammation. Our findings can facilitate for hypothesis generation and selection of biomarkers to include in such studies.

Targeting TfR1 with the ch128.1/IgG1 Antibody Inhibits EBV-driven Lymphomagenesis in Immunosuppressed Mice Bearing EBV+ Human Primary B-cells.

Epstein-Barr virus (EBV) is a human gammaherpesvirus associated with the development of hematopoietic cancers of B-lymphocyte origin, including AIDS-related non-Hodgkin lymphoma (AIDS-NHL). Primary infection of B-cells with EBV results in their polyclonal activation and immortalization. The transferrin receptor 1 (TfR1), also known as CD71, is important for iron uptake and regulation of cellular proliferation. TfR1 is highly expressed in proliferating cells, including activated lymphocytes and malignant cells. We developed a mouse/human chimeric antibody targeting TfR1 (ch128.1/IgG1) that has previously shown significant antitumor activity in immunosuppressed mouse models bearing human malignant B-cells, including multiple myeloma and AIDS-NHL cells. In this article, we examined the effect of targeting TfR1 to inhibit EBV-driven activation and growth of human B-cells in vivo using an immunodeficient NOD.Cg-Prkdcscid Il2rgtm1Wjl /SzJ [NOD/SCID gamma (NSG)] mouse model. Mice were implanted with T-cell-depleted, human peripheral blood mononuclear cells (PBMCs), either without EBV (EBV-), or exposed to EBV in vitro (EBV+), intravenously via the tail vein. Mice implanted with EBV+ cells and treated with an IgG1 control antibody (400 µg/mouse) developed lymphoma-like growths of human B-cell origin that were EBV+, whereas mice implanted with EBV+ cells and treated with ch128.1/IgG1 (400 µg/mouse) showed increased survival and significantly reduced inflammation and B-cell activation. These results indicate that ch128.1/IgG1 is effective at preventing the growth of EBV+ human B-cell tumors in vivo, thus, indicating that there is significant potential for agents targeting TfR1 as therapeutic strategies to prevent the development of EBV-associated B-cell malignancies. SIGNIFICANCE: An anti-TfR1 antibody, ch128.1/IgG1, effectively inhibits the activation, growth, and immortalization of EBV+ human B-cells in vivo, as well as the development of these cells into lymphoma-like tumors in immunodeficient mice.

Lymphoma-Associated Biomarkers Are Increased in Current Smokers in Twin Pairs Discordant for Smoking.

Smoking is associated with a moderate increased risk of Hodgkin and follicular lymphoma. To understand why, we examined lymphoma-related biomarker levels among 134 smoking and non-smoking twins (67 pairs) ascertained from the Finnish Twin Cohort. Previously collected frozen serum samples were tested for cotinine to validate self-reported smoking history. In total, 27 immune biomarkers were assayed using the Luminex Multiplex platform (R & D Systems). Current and non-current smokers were defined by a serum cotinine concentration of >3.08 ng/mL and ≤3.08 ng/mL, respectively. Associations between biomarkers and smoking were assessed using linear mixed models to estimate beta coefficients and standard errors, adjusting for age, sex and twin pair as a random effect. There were 55 never smokers, 43 current smokers and 36 former smokers. CCL17/TARC, sgp130, haptoglobin, B-cell activating factor (BAFF) and monocyte chemoattractant protein-1 (MCP1) were significantly (p < 0.05) associated with current smoking and correlated with increasing cotinine concentrations (Ptrend < 0.05). The strongest association was observed for CCL17/TARC (Ptrend = 0.0001). Immune biomarker levels were similar in former and never smokers. Current smoking is associated with increased levels of lymphoma-associated biomarkers, suggesting a possible mechanism for the link between smoking and risk of these two B-cell lymphomas.