CXCL12 Chemokine Research Articles

The Absence of CXCL10 Activity Does Not Affect the Capability of CD8+ T Cells to Migrate and Eliminate the Tissue Cysts of Toxoplasma gondii from the Brains of Chronically Infected Mice

Toxoplasma gondii forms tissue cysts in neurons and astrocytes in the brain to establish chronic infection, and astrocytes express the CXCL10 chemokine in chronically infected mice. Since chemokines mediate the migration of T cells to attack their targets, and since CXCL10 plays key roles in T cell-mediated control of the proliferation of tachyzoites (the acute stage form) of T. gondii during the acute stage of infection, we examined whether CXCL10 is involved in recruiting anti-cyst CD8+ cytotoxic T cells to eliminate the cysts in their brains. We employed adoptive transfer of CD8+ immune T cells to infected, T cell-deficient SCID and RAG1−/− mice in combination with blocking CXCL10 activity by neutralizing antibody or a deletion of this chemokine gene. The treatment of chronically infected (infected and treated with sulfadiazine) SCID mice with the anti-CXCL10 antibody did not inhibit the recruitment of the transferred CD8+ T cells into their brains and the removal of cerebral T. gondii cysts by the T cells. In addition, the neutralization of CXCL10 did not reduce the cerebral expression of mRNA for the mediators (perforin and granzyme B [GzmB]) of the cytotoxic activity of CD8+ T cells in the SCID mice. Consistently, the adoptive transfer of CD8+ immune T cells to chronically infected RAG1−/−CXCL10−/− mice did not show any defects in recruiting the CD8+ T cells into their brains and eliminating the cysts when compared to infected RAG1−/− mice. The former rather displayed enhanced cyst removal with increased cerebral expression of GzmB mRNA. These results indicate that the absence of CXCL10 activity does not ablate the capability of CD8+ cytotoxic T cells to migrate into the brain and eliminate T. gondii cysts from the brains of chronically infected mice. These results also suggest that the immune system utilizes distinct chemokines to control T. gondii depending on the two different life cycle stages, tachyzoite and cyst, of this protozoan parasite.

Thymic epithelial cell responses to the friedelin triterpene in vitro

Thymic epithelial cells (TECs) dysfunction can lead to disorders in the adaptive immune response, resulting in immunodeficiency or autoimmune diseases. Therefore, the investigation of new drugs with immunomodulatory capacity can contribute to the development of strategies to improve thymic functions. In this context, this study aimed to investigate the in vitro effects of the pentacyclic triterpene friedelin (FD) on TEC biology. For this, murine 2BH4 cells were treated with 0.1 and 1 μM FD for 24 h. After treatment, fibronectin and laminin production was increased (16% and 37% respectively) by TECs, however it did not alter the expression of CXCL12 chemokine. The interaction between TEC and thymocytes was also evaluated, in which a greater adhesion (45%) and survival (228%) of thymocytes to treated-TECs was observed. MHC molecules were up-regulated by FD treatment plus thymocyte coculture. Based on these results it was possible to attest that FD has an important and promissory role in the physiology of murine TECs.

Increased expression of CXCL10 and CCL3 salivary gland chemokines in primary Sjögren's syndrome detected and systematically quantified using novel RNAscope® in situ hybridisation.

Primary Sjogren's syndrome is a chronic inflammatory disease characterised by the destruction of exocrine glands. We have previously shown significantly upregulated levels of CXCL10 and CCL3 chemokines in saliva from Sjogren's syndrome patients. In this study, we examined the expression pattern and localisation of these chemokines at the site of inflammation in patients' minor salivary glands using novel RNAscope® in situ hybridisation. Minor salivary glands from 33 primary Sjogren's syndrome patients and 22 non-Sjogren's syndrome sicca controls were included. The biopsies were formalin- fixed, paraffin-embedded and histopathologically evaluated. The CXCL10 and CCL3 mRNA expression in the glandular tissue was investigated using reverse transcription quantitative real-time polymerase chain reaction followed by RNAscope® in situ hybridisation. The mRNA expression of CXCL10 was higher than CCL3 in all patients. Significantly elevated expression of CXCL10 and CCL3 was detected in patients that also expressed autoantibody positivity and a positive biopsy for mononuclear cell infiltrates when compared to controls. CXCL10 was localised as clusters within focal infiltrates as well as adjacent to acinar and ductal epithelium, while CCL3 was expressed as scattered single mRNA molecules in focal infiltrates and in acinar cells. Our findings suggest CXCL10 as a possible disease biomarker in primary Sjogren's syndrome due to its upregulated expression in both saliva and minor salivary glands of patients and the localisation in the tissue. This should be re-assessed in a larger primary Sjogren's syndrome patient cohort, followed by additional functional studies to further validate its potential as a disease biomarker.

Effects of Chitosan and N-Succinyl Chitosan on Metabolic Disorders Caused by Oral Administration of Olanzapine in Mice.

The issue of human mental health is gaining more and more attention nowadays. However, most mental disorders are treated with antipsychotic drugs that cause weight gain and metabolic disorders, which include olanzapine (OLZ). The search for and development of natural compounds for the prevention of obesity when taking antipsychotic drugs is an urgent task. The biopolymer chitosan (Chi) and its derivatives have lipid-lowering and anti-diabetic properties, which makes them potential therapeutic substances for use in the treatment of metabolic disorders. The purpose of this work was to analyze the effect of the natural biopolymer Chi, its derivative N-succinyl chitosan (SuChi), and Orlistat (ORL) as a control on the effects caused by the intake of OLZ in a mouse model. Mice were fed with pearl barley porridge mixed with OLZ or combinations OLZ + Chi, OLZ + SuChi, or OLZ + ORL for 2 months. The weight, lipid profile, blood chemokines, expression of genes associated with appetite regulation, and behavior of the mice were analyzed in dynamics. For the first time, data were obtained on the effects of Chi and SuChi on metabolic changes during the co-administration of antipsychotics. Oral OLZ increased body weight, food and water intake, and glucose, triglyceride, and cholesterol levels in blood. ORL and SuChi better normalized lipid metabolism than Chi, decreasing triglyceride and cholesterol levels. OLZ decreased the production of all chemokines tested at the 4th week of treatment and increased CXCL1, CXCL13, and CCL22 chemokine levels at the 7th week. All of the supplements corrected the level of CXCL1, CXCL13, and CCL22 chemokines but did not recover suppressed chemokines. SuChi and ORL stimulated the expression of satiety associated proopiomelanocortin (POMC) and suppressed the appetite-stimulating Agouti-related protein (AgRP) genes. All supplements improved the locomotion of mice. Taken collectively, we found that SuChi more than Chi possessed an activity close to that of ORL, preventing metabolic disorders in mice fed with OLZ. As OLZ carries positive charge and SuChi is negatively charged, we hypothesized that SuChi's protective effect can be explained by electrostatic interaction between OLZ byproducts and SuChi in the gastrointestinal tract.

Structural insights into KSHV-GPCR constitutive activation and CXCL1 chemokine recognition

Kaposi's sarcoma-associated herpesvirus (KSHV) encodes a viral G protein-coupled receptor, KSHV-GPCR, that contributes to KSHV immune evasion and pathogenesis of Kaposi's sarcoma. KSHV-GPCR shares a high similarity with CXC chemokine receptors CXCR2 and can be activated by selected chemokine ligands. Like other herpesvirus-encoded GPCRs, KSHV-GPCR is characterized by its constitutive activity by coupling to various G proteins. We investigated the structural basis of ligand-dependent and constitutive activation of KSHV-GPCR, obtaining high-resolution cryo-EM structures of KSHV-GPCR-Gi complexes with and without the bound CXCL1 chemokine. Analysis of the apo-KSHV-GPCR-Gi structure (2.81 Å) unraveled the involvement of extracellular loop 2 in constitutive activation of the receptor. In comparison, the CXCL1-bound KSHV-GPCR-Gi structure (3.01 Å) showed a two-site binding mode and provided detailed information of CXCL1 binding to a chemokine receptor. The dual activation mechanism employed by KSHV-GPCR represents an evolutionary adaptation for immune evasion and contributes to the pathogenesis of Kaposi's sarcoma. Together with results from functional assays that confirmed the structural models, these findings may help to develop therapeutic strategies for KSHV infection.

Transforming kidney transplant monitoring with urine CXCL9 and CXCL10: practical clinical implementation

In kidney transplant recipients, urine CXCL9 and CXCL10 (uCXCL9/10) chemokines have reached a sufficiently high level of evidence to be recommended by the European Society of Organ Transplantation for the monitoring of immune quiescence. To assess the risk of acute rejection (AR), the advantage of uCXCL9/10 is their cost-effectiveness and their high diagnostic performance. Here, we evaluated the feasibility of a next-generation immunoassay for quantifying uCXCL9/10 levels. It demonstrated high efficiency with minimal workflow and a 90-min time to result. Preanalytical studies indicated stability of uCXCL9/10 levels and analytical studies confirmed excellent linearity and precision. In a cohort of 1048 samples collected at biopsy, the results correlated significantly with ELISA quantification and were integrated into a previously validated 8-parameter urine chemokine model. The next generation immunoassay achieved an accuracy of 0.84 for AR diagnosis. This study validates this technology as a robust, locally available and unexpensive platform and marks a significant step towards the widespread implementation of uCXCL9/10, for immune quiescence monitoring. Therefore, we developed an open-access web application using uCXCL9/10 to calculate AR risk and improve clinical decision-making to perform biopsy, ushering in a new era in kidney transplantation, where personalized, data-driven care becomes the norm.

CXCL12 chemokine dimer signaling modulates acute myelogenous leukemia cell migration through altered receptor internalization.

Acute myeloid leukemia (AML) is a malignancy of immature myeloid blast cells with stem-like and chemoresistant cells being retained in the bone marrow through CXCL12-CXCR4 signaling. Current CXCR4 inhibitors mobilize AML cells into the bloodstream where they become more chemosensitive have failed to improve patient survival, likely reflecting persistent receptor localization on target cells. Here we characterize the signaling properties of CXCL12-locked dimer (CXCL12-LD), a bioengineered variant of the dimeric CXCL12 structure. CXCL12-LD binding resulted in lower levels of G protein, β-arrestin, and intracellular calcium mobilization, consistent with the locked dimer being a partial agonist of CXCR4. Further, CXCL12-LD failed to induce chemotaxis in AML cells. Despite these partial agonist properties, CXCL12-LD increased CXCR4 internalization compared to wildtype and locked-monomer forms of CXCL12. Analysis of a previously published AML transcriptomic data showed CXCR4 positive AML cells co-express genes involved in chemoresistance and maintenance of a blast-like state. The CXCL12-LD partial agonist effectively mobilized stem cells into the bloodstream in mice suggesting a potential role for their use in targeting CXCR4. Together, our results suggest that enhanced internalization by CXCL12-LD partial agonist signaling can avoid pharmacodynamic tolerance and may identify new avenues to better target GPCRs.

A subpopulation of human bone marrow erythroid cells displays a myeloid gene expression signature similar to that of classic monocytes.

Erythroid cells, serving as progenitors and precursors to erythrocytes responsible for oxygen transport, were shown to exhibit an immunosuppressive and immunoregulatory phenotype. Previous investigations from our research group have revealed an antimicrobial gene expression profile within murine bone marrow erythroid cells which suggested a role for erythroid cells in innate immunity. In the present study, we focused on elucidating the characteristics of human bone marrow erythroid cells through comprehensive analyses, including NanoString gene expression profiling utilizing the Immune Response V2 panel, a BioPlex examination of chemokine and TGF-beta family proteins secretion, and analysis of publicly available single-cell RNA-seq data. Our findings demonstrate that an erythroid cell subpopulation manifests a myeloid-like gene expression signature comprised of antibacterial immunity and neutrophil chemotaxis genes which suggests an involvement of human erythroid cells in the innate immunity. Furthermore, we found that human erythroid cells secreted CCL22, CCL24, CXCL5, CXCL8, and MIF chemokines. The ability of human erythroid cells to express these chemokines might facilitate the restriction of immune cells in the bone marrow under normal conditions or contribute to the ability of erythroid cells to induce local immunosuppression by recruiting immune cells in their immediate vicinity in case of extramedullary hematopoiesis.

Transplantation of alveolar macrophages improves the efficacy of endothelial progenitor cell therapy in mouse model of bronchopulmonary dysplasia.

Bronchopulmonary dysplasia (BPD) is a severe complication of preterm births, which develops due to exposure to supplemental oxygen and mechanical ventilation. Published studies demonstrated that the number of endothelial progenitor cells (EPC) is decreased in mouse and human BPD lungs and that adoptive transfer of EPC is an effective approach in reversing the hyperoxia-induced lung damage in mouse model of BPD. Recent advancements in macrophage biology identified the specific subtypes of circulating and resident macrophages mediating the developmental and regenerative functions in the lungs. Several studies reported the successful application of macrophage therapy in accelerating the regenerative capacity of damaged tissues and enhancing the therapeutic efficacy of other transplantable progenitor cells. In the present study, we explored the efficacy of combined cell therapy with EPC and resident alveolar macrophages (rAM) in hyperoxia-induced BPD mouse model. rAM and EPC were purified from neonatal mouse lungs and were used for adoptive transfer to the recipient neonatal mice exposed to hyperoxia. Adoptive transfer of rAM alone did not result in engraftment of donor rAM into the lung tissue but increased the mRNA level and protein concentration of proangiogenic CXCL12 chemokine in recipient mouse lungs. Depletion of rAM by chlodronate-liposomes decreased the retention of donor EPC after their transplantation into hyperoxia-injured lungs. Adoptive transfer of rAM in combination with EPC enhanced the therapeutic efficacy of EPC as evidenced by increased retention of EPC, increased capillary density, improved arterial oxygenation, and alveolarization in hyperoxia-injured lungs. Dual therapy with EPC and rAM has promise in human BPD.NEW & NOTEWORTHY Recent studies demonstrated that transplantation of lung-resident endothelial progenitor cells (EPC) is an effective therapy in mouse model of bronchopulmonary dysplasia (BPD). However, key factors regulating the efficacy of EPC are unknown. Herein, we demonstrate that transplantation of tissue-resident alveolar macrophages (rAM) increases CXCL12 expression in neonatal mouse lungs. rAM are required for retention of donor EPC in hyperoxia-injured lungs. Co-transplantation of rAM and EPC improves the efficacy of EPC therapy in mouse BPD model.

Immune mechanisms and predictive biomarkers related to neoadjuvant immunotherapy response in stage III melanoma

The treatment for stage III melanoma has advanced significantly, nevertheless, a substantial proportion of patients experience relapse. Neoadjuvant immune checkpoint blockade has emerged as a promising approach, allowing early micrometastatic disease treatment, reduction of tumor burden before surgery, and enhanced tumor-specific T-cell responses. However, not all patients respond to treatment, highlighting the need for understanding immune mechanisms behind failure and identification of predictive markers. Here we performed a robust evaluation of systemic and tumoral immune profiles in a well-defined cohort of advanced melanoma patients treated with immune checkpoint inhibitors. Elevated CTACK and CXCL9 chemokines pre-treatment suggested their potential as predictive tools for treatment response. Furthermore, CD95 expression in CD8+ T lymphocytes surfaced as a favorable prognostic indicator, while PD-1, CD161, and PD-L2 exhibited correlations with worst outcomes. These findings shed light on the intricate interplay between immune markers and melanoma response to neoadjuvant immune checkpoint therapy, offering insights into personalized treatment strategies.

Impact of Cerebrospinal Fluid Leukocyte Infiltration and Neuroimmmune Mediators on Survival with HIV-Associated Cryptococcal Meningitis.

Cryptococcal meningitis remains a prominent cause of death in persons with advanced HIV disease. CSF leukocyte infiltration predicts survival at 18 weeks; however, how CSF immune response relates to CSF leukocyte infiltration is unknown. We enrolled 401 adults with HIV-associated cryptococcal meningitis in Uganda who received amphotericin and fluconazole induction therapy. We assessed the association of CSF leukocytes, chemokine, and cytokine responses with 18-week survival. Participants with CSF leukocytes ≥50/μL, had higher probability 68% (52/77) of 18-week survival compared with 52% (151/292) 18-week survival in those with ≤50 cells/μL (Hazard Ratio=1.63, 95% confidence intervals 1.14-2.23; p=0.008). Survival was also associated with higher expression of T helper (Th)-1, Th17 cytokines, and immune regulatory elements. CSF levels of Programmed Death-1 Ligand, CXCL10, and Interleukin (IL)-2 independently predicted survival. In multivariate analysis, CSF leukocytes were inversely associated with CSF fungal burden and positively associated with CSF protein, interferon-gamma (IFN-γ), IL-17A, tumor necrosis factor (TNF)-α, and peripheral blood CD4+ and CD8+ T cells expression. 18-week survival after diagnosis of cryptococcal meningitis was associated with higher CSF leukocytes at baseline with greater T helper 1 (IFN-γ, IL-2 and TNF-α cytokines), T helper 17 (IL-17A cytokine) and CXCR3+ T cell (CXCL10 chemokine) responses. These results highlight the interdependent contribution of soluble and cellular immune responses in predicting survival with HIV-associated cryptococcal meningitis.

Identification of CXCL9 chemokine as a potential biomarker for assessing clinical severity in COVID-19 patients.

The severity and clinical outcome of COVID-19 depend on virus-specific factors and the host's inflammatory response. Identifying biomarkers of severe COVID-19 is a crucial condition and predicts disease severity. This study enrolled a total of 167 patients with COVID-19. These patients were categorized into three groups based on the severity of the disease: moderate course - 78 individuals, severe course - 52 individuals, and extremely severe course - 37 individuals. We analyzed chemokines (IP-10, CXCL9, CCL17) and cytokine IL28B levels using the enzyme immunoassay (EIA) method. CXCL9 levels were increased in severe and extremely severe cases compared to moderate ones. The CCL17 chemokine demonstrated significant elevation in severe cases. However, there was no significant difference in the level of IP-10, and IL28B in the compared groups. Our findings suggest that CXCL9 and CCL17 chemokines could be used as biomarkers to assess the clinical status of patients with COVID-19 and can relate to disease severity. These biomarkers could aid in identifying patients at high risk for severe disease and help guide clinical decision-making for the effective management of COVID-19.

Early cognitive dysfunction development immunological predictors in the ischemic stroke acute period

The purpose of the study is activity identification CXC family cytokines in patients with cognitive impairments in the acute period of ischemic stroke.Material and methods. 78 patients with diagnosis “Ischemic stroke” were examined. Depending on cognitive impairment (Montreal Cognitive Assessment (MoCA)) patients were divided into two groups: the 1st group — 58 patients with cognitive decline (MoCA ≤ 25 points); the 2nd group — 20 patients without cognitive decline. Neuropsychologic testing was performed on the second day of hospitalisation and included episodic memory, executive function, speech, gnosis, praxis and IQCODE parameters examination. Laboratory diagnosis consisted of level assessing of CXC family chemokines (CXCL10, CXCL11, CXCL9, CXCL1, CXCL8) and TNF-α cytokine in patients’ plasma on the second day of hospitalisation. Statistical analysis was employed using the Python programing language and its libraries Pandas and SciPy.Results. Statistical analysis revealed the highest level of IP-10/CXCL10 chemokines (p = 0.002) and Gro-a/CXCL1 (p = 0.044) in patients of the 1st group, statistically significant correlations of MoCA and IQCODE with IP-10/CXCL10 and Gro-a/CXCL1 concentrations, correlations of IP-10/CXCL10 concentrations with semantic information processing functions (r = –0.512), subject gnosis (r = –0.211), memory (r = 0.275), speech (r = –0.400), and Gro-a/ CXCL1 level with semantic information processing (r = –0.418).Conclusion. The study of chemokines of the CXC cluster represents a relevant and promising direction in the diagnosis and assessment of progression of early post-stroke cognitive impairment of mixed genesis due to minimal invasiveness and high specificity. Further studies are needed to verify CXCL chemokines, particularly IP-10/CXCL10 and Gro-a/CXCL1 as potential molecular markers of neurological damage in neurodegenerative and inflammatory diseases of the central nervous system.

Advancing chemokine research: the molecular function of CXCL8.

CXCL8 and other chemokines have been implicated in tissue inflammation and are attractive candidates for therapeutic targeting to treat human disease.

Abstract PS03-04: Tumor immune microenvironment modulates resistance to estrogen suppression in ER+ breast cancer

Abstract Despite major advances in the treatment of estrogen receptor positive (ER+) breast cancer (BC), advanced disease continues to be the main cause of death from this disease. The role of the tumor immune microenvironment (TIME) in the progression of ER+ BC and its response to treatment is not completely understood. The aim of this study was to elucidate the role the TIME in the response of ER+ tumors to estrogen deprivation (ED). We collected samples from 215 postmenopausal patients with stage I-III ER+ BC, treated with letrozole for 2-4 weeks, to induce ED. We used AQUA on pre-treatment biopsies and on-treatment surgical biopsies to assess ER, PR, HER2, and Ki67. Then, we categorized the patients’ response to ED based on the Ki67 score in on-treatment samples: ED-sensitive (ED-S) if natural log (ln) of the Ki67 score ≤1.0 or ≤2.7% Ki67+ cells vs. ED-resistant (ED-R) if ln ≥2.0 or ≥7.4% Ki67+ cells. Firstly, we assessed TIME composition by investigating stromal tumor-infiltrating lymphocytes (sTILs) in H&E-stained FFPE of on-treatment tumor sections. ED-R tumors exhibited a significantly higher stromal TILs score (p=0.0001) relative to ED-S tumors. We next prepared tissue microarrays from 227 (on-treatment) surgical sections and subjected them to cyclic immunofluorescence (CycIF) with 38 antibodies to examine their intra-tumoral immune cell infiltration. From each tumor core, we segmented single cells and labeled them as immune, cancer, or stromal cells based on expression of a set of markers. Briefly, cells that stained strongly for CD45 (leukocyte marker), and/or CD4 (T lymphocyte marker), or CD68 (macrophage marker) were labeled immune cells. CD45/CD4/CD68-negative cells were categorized as tumor if E-cadherin and/or cytokeratin-positive, or stromal cells if -negative. Next, we assessed cell specific spatial enrichment by quantifying the expression of immune markers in the area immediately adjacent to each tumor cell. Immune-suppressive T-reg (FOXP3+) cells were enriched in the ED-S tumors (p=0.0004), as well as PD1+ (exhausted) T cells (p=0.0004), and CD68+ cells (p < 0.0001) compared to ED-R tumors. ED-R tumors exhibited higher CD20+ B cells (p < 0.0001), higher CD8+ T cells (p=0.0329), in addition to higher CD45+ cells (p < 0.0001), compared to ED-S tumors. RNA-sequencing of the same surgical samples showed a higher T cells cytolytic score in ED-R relative to ED-S (p=0.0058), suggesting enhanced CD8+ T cells activity, in addition to their higher infiltration. We are currently analyzing letrozole-induced changes in TIME composition using Geomix digital spatial profiler in paired pre- and on-treatment biopsies from ED-R and ED-S tumors and will be presented at the meeting. Consistent with the CycIF findings, GSEA of hallmark gene signatures from bulk RNA-sequencing of treated tumors revealed that immune-related gene sets, such as “IFN α response”, “IFNɣ response”, and “allograft rejection” were upregulated in ED-R vs. ED-S cancers. ED-R tumors showed enrichment of CXCL9, CXCL10, and CXCL11 chemokines and their receptor, CXCR3. Publicly available datasets of patients with ER+ breast cancers showed that higher expression CXCL9 (HR 1.36; p=0.016), CXCL10 (HR 1.71; p< 0.0001), and CXCL11 (HR 1.5; p=0.0016) are predictive of shorter relapse-free survival on antiestrogen therapy. We are currently investigating whether these chemokines play a causal role in resistance to ED, and if this is phenocopied by co-cultures of ER+ BC cells and CD8+ T-cells. Conclusions ED-resistant tumors are enriched with stromal TILs and exhibit higher immune cell intra-tumoral infiltration and CD8+T cells cytolytic activity compared to ER+ tumors sensitive to estrogen suppression. In contrast, ED-S tumors showed a more immunosuppressed milieu. The role of CXCL9, CXCL10 and CXCL11 in inducing resistance to ED warrants further investigation. Citation Format: Fabiana Napolitano, Yunguan Wang, Dhivya Sudhan, Paula Gonzalez-Ericsson, Luigi Formisano, Lei Guo, M Rosario Chica-Parrado, Chang-Ching Lin, Kyung-Min Lee, Hongli Ma, Nathaniel Evans, Alberto Servetto, Saurabh Mendiratta, Spencer Barnes, Yisheng Fang, Lin Xu, Justin Balko, Gordon Mills, Marilyne Labrie, Ariella Hanker, Carlos Arteaga. Tumor immune microenvironment modulates resistance to estrogen suppression in ER+ breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PS03-04.

Neuroinflammation generated by HIV-infected microglia promotes dysfunction and death of neurons in human brain organoids.

Despite the success of combination antiretroviral therapy (ART) for individuals living with HIV, mild forms of HIV-associated neurocognitive disorder (HAND) continue to occur. Brain microglia form the principal target for HIV infection in the brain. It remains unknown how infection of these cells leads to neuroinflammation, neuronal dysfunction, and/or death observed in HAND. Utilizing two different inducible pluripotent stem cell-derived brain organoid models (cerebral and choroid plexus [ChP] organoids) containing microglia, we investigated the pathogenic changes associated with HIV infection. Infection of microglia was associated with a sharp increase in CCL2 and CXCL10 chemokine gene expression and the activation of many type I interferon stimulated genes (MX1, ISG15, ISG20, IFI27, IFITM3 and others). Production of the proinflammatory chemokines persisted at low levels after treatment of the cell cultures with ART, consistent with the persistence of mild HAND following clinical introduction of ART. Expression of multiple members of the S100 family of inflammatory genes sharply increased following HIV infection of microglia measured by single-cell RNA-seq. However, S100 gene expression was not limited to microglia but was also detected more broadly in uninfected stromal cells, mature and immature ChP cells, neural progenitor cells and importantly in bystander neurons suggesting propagation of the inflammatory response to bystander cells. Neurotransmitter transporter expression declined in uninfected neurons, accompanied by increased expression of genes promoting cellular senescence and cell death. Together, these studies underscore how an inflammatory response generated in HIV-infected microglia is propagated to multiple uninfected bystander cells ultimately resulting in the dysfunction and death of bystander neurons.

Therapeutic prime/pull vaccination of HSV-2-infected guinea pigs with the ribonucleotide reductase 2 (RR2) protein and CXCL11 chemokine boosts antiviral local tissue-resident and effector memory CD4+ and CD8+ T cells and protects against recurrent genital herpes.

Following acute herpes simplex virus type 2 (HSV-2) infection, the virus undergoes an asymptomatic latent infection of sensory neurons of dorsal root ganglia (DRG). Chemical and physical stress cause intermittent virus reactivation from latently infected DRG and recurrent virus shedding in the genital mucosal epithelium causing genital herpes in symptomatic patients. While T cells appear to play a role in controlling virus reactivation from DRG and reducing the severity of recurrent genital herpes, the mechanisms for recruiting these T cells into DRG and the vaginal mucosa (VM) remain to be fully elucidated. The present study investigates the effect of CXCL9, CXCL10, and CXCL11 T-cell-attracting chemokines on the frequency and function of DRG- and VM-resident CD4+ and CD8+ T cells and its effect on the frequency and severity of recurrent genital herpes in the recurrent herpes guinea pig model. HSV-2 latent-infected guinea pigs were immunized intramuscularly with the HSV-2 ribonucleotide reductase 2 (RR2) protein (Prime) and subsequently treated intravaginally with the neurotropic adeno-associated virus type 8 expressing CXCL9, CXCL10, or CXCL11 chemokines to recruit CD4+ and CD8+ T cells into the infected DRG and VM (Pull). Compared to the RR2 therapeutic vaccine alone, the RR2/CXCL11 prime/pull therapeutic vaccine significantly increased the frequencies of functional tissue-resident and effector memory CD4+ and CD8+ T cells in both DRG and VM tissues. This was associated with less virus in the healed genital mucosal epithelium and reduced frequency and severity of recurrent genital herpes. These findings confirm the role of local DRG- and VM-resident CD4+ and CD8+ T cells in reducing virus shedding at the vaginal site of infection and the severity of recurrent genital herpes and propose the novel prime-pull vaccine strategy to protect against recurrent genital herpes.IMPORTANCEThe present study investigates the novel prime/pull therapeutic vaccine strategy to protect against recurrent genital herpes using the latently infected guinea pig model. In this study, we used the strategy that involves immunization of herpes simplex virus type 2-infected guinea pigs using a recombinantly expressed herpes tegument protein-ribonucleotide reductase 2 (RR2; prime), followed by intravaginal treatment with the neurotropic adeno-associated virus type 8 expressing CXCL9, CXCL10, or CXCL11 T-cell-attracting chemokines to recruit T cells into the infected dorsal root ganglia (DRG) and vaginal mucosa (VM) (pull). We show that the RR2/CXCL11 prime-pull therapeutic vaccine strategy elicited a significant reduction in virus shedding in the vaginal mucosa and decreased the severity and frequency of recurrent genital herpes. This protection was associated with increased frequencies of functional tissue-resident (TRM cells) and effector (TEM cells) memory CD4+ and CD8+ T cells infiltrating latently infected DRG tissues and the healed regions of the vaginal mucosa. These findings shed light on the role of tissue-resident and effector memory CD4+ and CD8+ T cells in DRG tissues and the VM in protection against recurrent genital herpes and propose the prime-pull therapeutic vaccine strategy in combating genital herpes.

Chemokine-derived oncolytic peptide induces immunogenic cancer cell death and significantly suppresses tumor growth

Chemokinostatin-1 (CKS1) is a 24-mer peptide originally discovered as an anti-angiogenic peptide derived from the CXCL1 chemokine. Here, we demonstrate that CKS1 acts not only as an anti-angiogenic peptide but also as an oncolytic peptide due to its structural and physical properties. CKS1 induced both necrotic and apoptotic cell death specifically in cancer cells while showing minimal toxicity in non-cancerous cells. Mechanistically, CKS1 disrupted the cell membrane of cancer cells quickly after treatment and activated the apoptotic pathway at later time points. Furthermore, immunogenic molecules were released from CKS1-treated cells, indicating that CKS1 induces immunogenic cell death. CKS1 effectively suppressed tumor growth in vivo. Collectively, these data demonstrate that CKS1 functions as an oncolytic peptide and has a therapeutic potential to treat cancer.

Neutrophil Membrane-Camouflaged Polyprodrug Nanomedicine for Inflammation Suppression in Ischemic Stroke Therapy.

Neuroinflammation has emerged as a major concern in ischemic stroke therapy because it exacebates neurological dysfunction and suppresses neurological recovery after ischemia/reperfusion. Fingolimod hydrochloride (FTY720) is an FDA-approved anti-inflammatory drug which exhibits potential neuroprotective effects in ischemic brain parenchyma. However, delivering a sufficient amount of FTY720 through the blood-brain barrier into brain lesions without inducing severe cardiovascular side effects remains challenging. Here, a neutrophil membrane-camouflaged polyprodrug nanomedicine that can migrate into ischemic brain tissues and in situ release FTY720 in response to elevated levels of reactive oxygen species. This nanomedicine delivers 15.2-fold more FTY720 into the ischemic brain and significantly reduces the risk of cardiotoxicity and infection compared with intravenously administered free drug. In addition, single-cell RNA-sequencing analysis identifies that the nanomedicine attenuates poststroke inflammation by reprogramming microglia toward anti-inflammatory phenotypes, which is realized via modulating Cebpb-regulated activation of NLRP3 inflammasomes and secretion of CXCL2 chemokine. This study offers new insights into the design and fabrication of polyprodrug nanomedicines for effective suppression of inflammation in ischemic stroke therapy.

Detection of CXCL10 chemokine level in serum of patients with warts

Detection of CXCL10 chemokine level in serum of patients with warts