Clearance Of Apoptotic Bodies Research Articles

Galectin-3: action and clinical utility in chronic kidney disease.

Chronic kidney disease is a significant cause of morbidity and mortality worldwide. In recent years, Galectin-3 has been put forward as a potential biomarker of chronic kidney disease progression. This review aims to assess the clinical utility of Galectin-3 in various pathological processes leading up to chronic kidney disease such as diabetes and lupus nephritis. We conducted a systematic search on PubMed from inception to September 2023, using the search term ("Galectin-3" OR "gal-3") AND ("renal" OR "kidney"). Galectin-3 has been shown to be both pro-fibrotic and protective against renal fibrosis through various mechanisms such as apoptotic body clearance and modulation of the Wnt pathway. Studies have found associations between raised Galectin-3, incidence and progression of chronic kidney disease. In lupus nephritis, Galectin-3 may serve as a biomarker for lupus nephritis activity. Although Galectin-3 inhibits cystogenesis, there is no correlation between total kidney volume and Galectin-3 in polycystic kidney disease. The role of Galectin-3 in staging and prognostication of renal cell carcinoma is yet to be determined. Galectin-3 has potential in predicting chronic kidney disease progression, in combination with other biomarkers. However, more trials are required given that present studies demonstrate conflicting results on the relationship between Galectin-3 and clinical outcomes in chronic kidney disease patients of varying aetiologies.

Gas6/TAM system as potential biomarker for multiple sclerosis prognosis.

The protein growth arrest-specific 6 (Gas6) and its tyrosine kinase receptors Tyro-3, Axl, and Mer (TAM) are ubiquitous proteins involved in regulating inflammation and apoptotic body clearance. Multiple sclerosis (MS) is the most common inflammatory demyelinating disease of the central nervous system leading to progressive and irreversible disability if not diagnosed and treated promptly. Gas6 and TAM receptors have been associated with neuronal remyelination and stimulation of oligodendrocyte survival. However, few data are available regarding clinical correlation in MS patients. We aimed to evaluate soluble levels of these molecules in the cerebrospinal fluid (CSF) and serum at MS diagnosis and correlate them with short-term disease severity. In a prospective cohort study, we enrolled 64 patients with a diagnosis of clinical isolated syndrome (CIS), radiological isolated syndrome (RIS) and relapsing-remitting (RR) MS according to the McDonald 2017 Criteria. Before any treatment initiation, we sampled the serum and CSF, and collected clinical data: disease course, presence of gadolinium-enhancing lesions, and expanded disability status score (EDSS). At the last clinical follow-up, we assessed EDSS and calculated MS severity score (MSSS) and age-related MS severity (ARMSS). Gas6 and TAM receptors were determined using an ELISA kit (R&D Systems) and compared to neurofilament (NFLs) levels evaluated with SimplePlex™ fluorescence-based immunoassay. At diagnosis, serum sAxl was higher in patients receiving none or low-efficacy disease-modifying treatments (DMTs) versus patients with high-efficacy DMTs (p = 0.04). Higher CSF Gas6 and serum sAXL were associated with an EDSS<3 at diagnosis (p = 0.04; p = 0.037). Serum Gas6 correlates to a lower MSSS (r2 = -0.32, p = 0.01). Serum and CSF NFLs were confirmed as disability biomarkers in our cohort according to EDSS (p = 0.005; p = 0.002) and MSSS (r2= 0.27, p = 0.03; r2 = 0.39, p = 0.001). Results were corroborated using multivariate analysis. Our data suggest a protective role of Gas6 and its receptors in patients with MS and suitable severity disease biomarkers.

Serum GAS6, sAXL, IL-10, NO, and BCL-2 levels are decreased in patients with Behçet's disease.

Behçet's disease (BD) is an autoimmune chronic systemic inflammatory disease characterized by a versatile clinical spectrum. Growth arrest specific protein 6 (GAS6)/soluble AXL (sAXL) signaling pathway draws attention in the resolution of inflammation, and its deficiency is associated with chronic inflammatory, autoimmune diseases, as well as clearance of apoptotic cells by phagocytes - efferocytosis. In this study, it was aimed to investigate whether GAS6/sAXL, interleukin (IL)-10, nitric oxide (NO), and BCL-2 levels were associated with inflammation and efferocytosis contributes to the pathogenesis of BD. A total of 37 Behçet patients with ocular involvement and 30 healthy control subjects were included in this study. GAS6, sAXL, IL-10, NO, and BCL-2 levels were quantified using enzyme-linked immunosorbent assay (ELISA) method. Serum GAS6, sAXL, IL-10, NO, and BCL-2 levels were significantly lower in patients with BD compared to the controls (P < 0.005, P < 0.001, P < 0.001, P < 0.001, and P < 0.001, respectively). In correlation analysis, research parameters decreased in patients with BD was significantly correlated with each other: GAS6-IL-10 (r = 0.585, P < 0.001), GAS6-BCL-2 (r = 0.541, P < 0.001), sAXL-BCL-2 (r = 0.696, P < 0.001), IL-10-NO (r = 0.717, P < 0.001), IL-10-BCL-2 (r = 0.759, P < 0.001), and NO-BCL-2 (r = 0.541, P < 0.001). In conclusion, decreased serum BCL-2 level may be an indicator of increased apoptosis in these patients and decreased levels of GAS6/sAXL, IL-10, and NO may indicate insufficient clearance of apoptotic bodies released as a result of increased apoptosis in BD patients.

Lymphocyte apoptosis and its association with the inflammatory markers and disease severity in juvenile-onset systemic lupus erythematosus patients

BackgroundThe defective clearance of apoptotic bodies in juvenile-onset systemic lupus erythematosus (jSLE) potentially leads to the persistence of autoreactive lymphocytes and the perpetuation of the autoimmune response. These factors contribute to the disturbance in lymphocyte apoptosis and show potential as key determinants in the clinical course and severity of jSLE. This study evaluates the role of peripheral blood (PB) lymphocyte apoptosis in prognosis of jSLE and as a predictor for disease activity.MethodsThe study involved 100 jSLE patients and 50 healthy controls. Flow cytometry was used to analyze percentages of lymphocyte apoptosis in PB of all study participants. Plasma levels of pro-inflammatory cytokines were determined using ELISA.ResultsOur results showed that percentages of lymphocyte apoptosis in PB of jSLE patients are significantly higher than those of healthy controls. These percentages are significantly positively associated with disease activity of patients (SLEDAI-2 K). Furthermore, plasma cytokine levels (IL-17, IFN-γ and TNF-α) are significantly elevated in jSLE patients compared to their levels in healthy controls. Also, there are weak significant positive correlations between percentages of PB lymphocyte apoptosis and each of IL-17 and IFN-γ plasma levels in jSLE patients. Moreover, PB lymphocyte apoptosis percentages among jSLE patients are higher in the presence of some clinical and laboratory features than those in their absence.ConclusionPeripheral apoptotic lymphocytes could contribute to the prognosis of jSLE and could be used as a predictor for disease activity in jSLE patients.

Rare Autoimmune Diseases Role of Genetics - Example of Systemic Lupus Erythematosus

Systemic lupus erythematosus (SLE) presents a complex clinical landscape with diverse manifestations, suggesting a multifactorial etiology. However, the identification of rare monogenic forms of the disease has shed light on specific genetic defects underlying SLE pathogenesis, offering valuable insights into its underlying mechanisms and clinical heterogeneity. By categorizing these monogenic forms based on the implicated signaling pathways, such as apoptotic body clearance, type I interferon signaling, JAK-STAT pathway dysregulation, innate immune receptor dysfunction and lymphocytic abnormalities, a more nuanced understanding of SLE's molecular basis emerges. Particularly in pediatric populations, where monogenic forms are more prevalent, routine genetic testing becomes increasingly important, with a diagnostic yield of approximately 10% depending on the demographic and methodological factors involved. This approach not only enhances diagnostic accuracy but also informs personalized treatment strategies tailored to the specific molecular defects driving the disease phenotype.

Effect of the Rubicon protein on LC3-associated phagocytosis by monocytes in the patients with severe atopic bronchial asthma

Atopic bronchial asthma is the most common and severe allergic disease among a wide range of similar diseases. The main pathogenesis of this disease is characterized by a disturbance of T lymphocyte homeostasis, which significantly worsens the general state of health. In atopic bronchial asthma, there is impaired process of T cell apoptosis. This entails dysregulation and maintenance of peripheral lymphocyte homeostasis. Normally, T cells must undergo apoptosis, and its products should be utilized by neighboring cells, or professional phagocytes: monocytes, macrophages, or dendritic cells. This process is altered in atopic bronchial asthma. The immune system disorders, such as autoimmunity, often result from dysregulation of lymphocyte apoptosis. This is especially true in cases of insufficient or missed clearance of apoptotic bodies. Recently, the research and medical communities pay much attention to efferocytosis, a form of phagocytosi which proceeds by removal of apoptotic cells by phagocytes by means of LC3-associated phagocytosis (LAP). This process initiates uptake of the particles due to interactions between the phagocyte plasma membrane receptors and apoptotic cell. Further on, a single-membrane phagosome is formed in the cell with the participation of certain autophagy proteins (Beclin-1, VPS34, UVRAG, ATG5, ATG12, ATG7, ATG4, ATG4, LC3). The phagosome is enriched with LC3 protein molecules and fused with lysosomes, in which the captured “cargo” is then lysed. As a part of our work, a detailed analysis of some key protein contents at the LAP pathway was carried out for peripheral blood monocytes of patients with severe bronchial asthma. It was found that the expression of Rubicon protein is increased, thus allowing to conclude that the LAP pathway is activated in monocytes of healthy donors, thus allowing phagocytosis of dying T cells. At the same time, the components characteristic of both autophagy and LC3-associated phagocytosis are activated in the monocytes of patients with severe atopic asthma. However, one should note that decreased expression of the Rubicon protein, a putative marker of LC3-associated phagocytosis, has been clearly confirmed.

Periodic bilayer organization in the complexes of Beta-2 Glycoprotein I with anionic lipid membranes

β2 glycoprotein I (β2GPI) is a soluble protein that participates in blood coagulation, clearance of apoptotic bodies and generation of antigens in antiphospholipid syndrome among many other functions. We studied the aggregates formed by β2GPI with the anionic phospholipids palmitoyloleoylphosphatidyl glycerol, dimyristoylphosphatidyl glycerol, dipalmitoylphosphatidyl glycerol and cardiolipin using small angle X-ray scattering. The complexes obtained in a medium containing 0.01 M NaCl showed Bragg peaks up to the sixth order in a well-defined integer sequence indicating the presence of a lamellar stacking with a periodicity of 17.8 nm and with largely reduced membrane fluctuations. Modeling the complex signal allowed us to conclude that the coherence length was only two bilayers and that about 15% of the total surface was actually stacked. The space between bilayers allows accommodating an extended β2GPI molecule making a bridge between the interacting bilayers. The interactions between membranes mediated by β2GPI was favored when the membranes were in the liquid crystalline state.

Targeting Tumor-Associated Macrophages by MMP2-Sensitive Apoptotic Body-Mimicking Nanoparticles.

Tumor-associated macrophages (TAMs), a major player in the tumor microenvironment, were recently recognized as a potential therapeutic target. To date, very few anticancer drugs or drug-delivery systems were designed to target the TAMs. Inspired by the "eat me" signal, phosphatidylserine (PS), mediated phagocytic clearance of apoptotic bodies, in this study, the matrix metalloproteinase 2 (MMP2)-sensitive PS-modified nanoparticles were developed. In the design, the PS is externalized to the nanoparticles' surface only when the nanoparticles reach the MMP2-overexpressing tumor site, allowing for the TAM-specific phagocytosis. The nanoparticles' excellent macrophage/TAM selectivity was observed in various biological models, including various cell lines, coculture cells, coculture cell spheroids, zebrafish, and tumor-bearing mice. The nanoparticles' TAM specificity remarkably enhanced the TAM depletion capability of the loaded model drug, dasatinib, resulting in the improved anticancer activity. The MMP2-sensitive apoptotic body-mimicking nanoparticles might be a promising delivery tool for TAM-centered cancer diagnoses and treatments.

TIM-1 As a Signal Receptor Triggers Dengue Virus-Induced Autophagy.

Dengue virus (DENV) infection triggers the activation of autophagy to facilitate the viral replication cycle from various aspects. Although a number of stimulators are proposed to activate autophagy, none of them appears prior to the uncoating process. Given that T-cell immunoglobulin and mucin domain 1 (TIM-1) receptor is a putative DENV receptor and promotes apoptotic body clearance by autophagy induction, it raises the possibility that TIM-1 may participate in the activation of DENV-induced autophagy. In this study, confocal images first revealed the co-localization of TIM-1 with autophagosomes in DENV-induced autophagy rather than rapamycin-induced autophagy, suggesting the co-transportation of TIM-1 with DENV during infection. The treatment of siRNA to knockdown TIM-1 expression in DENV-infected GFP-microtubule-associated protein light chain 3 (LC3)-Huh7.5 cells revealed that TIM-1 is required not only for DENV cellular internalization but also for autophagy activation. Furthermore, knockdown p85, a subunit of phosphoinositide 3-kinases (PI3Ks), which is co-localized with TIM-1 at rab5-positive endosomes caused the reduction of autophagy, indicating that TIM-1-mediated DENV-induced autophagy requires p85. Taken together, the current study uncovered TIM-1 as a novel factor for triggering autophagy in DENV infection through TIM-1-p85 axis, in addition to serving as a DENV receptor.

Clearance of apoptotic bodies by neutrophils induce tissue regenerative responses

Clearance of apoptotic bodies by neutrophils induce tissue regenerative responses

Exploring the etiopathogenesis of systemic lupus erythematosus: a genetic perspective.

Systemic lupus erythematosus (SLE) is an autoimmune multi-organ disorder that presents itself in a thousand ways. Its clinical course is extremely unpredictable, which makes diagnosis and treatment a challenge for clinicians. It appears that the clinical course of SLE is determined by genetic material in combination with environmental factors. In this article, we review recent findings on the pathogenesis of SLE from the perspective of genetics, focusing on defects in the clearance of apoptotic bodies and immune complexes, on alterations in the innate immune system response, and on impaired pathways in the adaptive immune system. Furthermore, the major histocompatibility complex (MHC) and non-MHC genes discovered during genome-wide association studies (GWASs) in SLE patients are also evaluated. In addition, the effect of these polymorphisms on the function of their related transcripts and their association with the clinical manifestations of SLE and its pathophysiology are explained. Finally, the association of genetic polymorphisms with clinical responses to common medications used in the treatment of SLE is also discussed.

Structural Aspects of Carbohydrate Recognition Mechanisms of C-Type Lectins.

Carbohydrate recognition is an essential function occurring in all living organisms. Lectins are carbohydrate-binding proteins and are classified into several families. In mammals, Ca2+-dependent C-type lectins, such as β-galactoside-binding galectin and sialic acid-binding siglec, play crucial roles in the immune response and homeostasis. C-type lectins are abundant and diverse in animals. Their immunological activities include lymphocyte homing, pathogen recognition, and clearance of apoptotic bodies. C-type lectin domains are composed of 110-130 amino acid residues with highly conserved structural folds. Remarkably, individual lectins can accept a wide variety of sugar ligands and can distinguish subtle structural differences in closely related ligands. In addition, several C-type lectin-like proteins specifically bind to carbohydrate ligands in Ca2+-independent ways. The accumulated 3D structural evidence clarifies the unexpected structural versatility of C-type lectins underlying the variety of ligand binding modes. In this issue, we focus on the structural aspects of carbohydrate recognition mechanisms of C-type lectins and C-type lectin-like proteins.

Trichloroethene metabolite dichloroacetyl chloride induces apoptosis and compromises phagocytosis in Kupffer Cells: Activation of inflammasome and MAPKs.

Exposure to trichloroethene (TCE), an occupational and ubiquitous environmental contaminant, is associated with the development of several autoimmune diseases, including autoimmune hepatitis (AIH). However, mechanisms contributing to TCE-mediated AIH are not known. Earlier, we have shown that dichloroacetyl chloride (DCAC), one of the reactive metabolites of TCE with strong acylating capability, can elicit an autoimmune response at much lower dose than TCE in female MRL+/+ mice. Furthermore, Kupffer cells (KCs), the liver resident macrophages, are crucial for hepatic homeostasis, but can also participate in the immunopathogenesis of AIH. However, contribution of KCs in TCE-mediated AIH and the underlying mechanisms are not understood. We hypothesized that increased apoptosis and delayed clearance of apoptotic bodies, due to compromised KC function, will result in the breakdown of self-tolerance, autoimmunity, and ultimately AIH. Therefore, using an in vitro model of immortalized mouse KCs, we investigated the contribution of DCAC in TCE-mediated AIH. KCs were treated with different concentrations of DCAC and apoptosis was measured by Annexin V and PI staining. Also, the impact of DCAC on phagocytic potential of KCs was evaluated. Furthermore, markers of inflammasome (NLRP3 and caspase1) were analyzed by real-time PCR and Western blot analysis. DCAC treatment resulted in significantly increased early and late-stage apoptosis, accompanied with inflammasome activation (NLRP3 increases). DCAC treatment resulted in decreased phagocytic function of KCs in a dose-dependent manner, with reduced MFG-E8 levels (phagocytotic function). Furthermore, DCAC exposure led to induction of phos-ERK and phos-AKT signaling. These findings suggest that DCAC induces apoptosis and inflammasome activation, while compromising the phagocytic function of KCs. Our data support that increased apoptosis and impaired KC function by DCAC could be contributory to TCE-mediated AIH.

Clinical significance of neutrophils functional activity in patients&#x0D; with systemic lupus erythematosus.

There is a revival of interest to the role of innate immunity in SLE and neutrophils as its main cellular components. Lupus neutrophils are characterized by impaired phagocytic capabilities, enhanced apoptosis and NETosis, an alternative mechanism ofneutrophil cell death, abnormal clearance ofapoptotic bodies and NETosis products. Therefore, investigation ofpathogenetic processes involving neutrophils is of immense importance, it may help in searching for potential therapeutic targets, understanding the mechanisms of drugs action already used in clinical practice and optimizing the therapeutic regimens by developing predictive factors. Free radical production is one of the crucial neutrophil functions. Thus, the study of neutrophil radical-producing function as a component of their functional status seems expedient. To attain this objective, a new method of luminol-enhanced chemiluminescence with a preliminary two-step stimulation by phorbol-12-myristate-13 acetate and formyl-methionyl-leucyl-phenylalanine was developed. Neutrophil radical-producing activity was evaluated in 46 SLE patients compared to 87 healthy controls. Increased spontaneous production of reactive oxygen species and more pronounced response to the PMA and fMLP effects were found. It was shown by higher specific peak and specific integral neutrophil activity, comparing to the control group. Interleukin-lfi for SLE patients’ neutrophils appeared to be both a pronounced stimulus and a priming agent, it was only a weak priming agent in healthy controls. These results are consistent with research data showing increased functional activity of neutrophils in SLE associated with increased production of pro-inflammatory cytokines, as well as increased production of neutrophil extracellular traps in the absence of additional stimulation, which may be a consequence of cytokine-primed condition.

Whole Exome Sequencing in Early-onset Systemic Lupus Erythematosus.

Systemic lupus erythematosus (SLE) is a multisystem autoimmune disorder. Early-onset, familial, and/or syndromic SLE may reveal monogenic pathologies. The aim of this study was to examine genetic associations in patients with early-onset or familial SLE. We enrolled 7 SLE cases (from different families) with disease onset ≤ 5 years of age and family history consistent with an autosomal recessive inheritance. Whole exome sequencing (WES) was performed in 6 index cases. Suspected variants were confirmed by Sanger sequencing. We did not perform WES in 1 patient who had features similar to the first 3 cases; only the exons of C1QA, C1QB, and C1QC were screened with Sanger sequencing. We demonstrated 2 novel and 3 previously reported variants in genes associated with SLE: a homozygous non-sense alteration (c.622C>T/p.Gln208Ter) in C1QA in 2 patients; homozygous non-sense alteration (c.79C>T/p.Gln27Ter) in C1QC in 1 (novel variant); homozygous missense alteration (c.100G>A/p.Gly34Arg) in C1QC in 1; homozygous missense alteration (c.1945G>C/p.Ala649Pro) in C1S in 1 (novel variant); and homozygous frameshift alteration (c.289_290delAC/p.Thr97Ilefs*2) in DNASE1L3 in 1 patient. Further, in 1 patient, we determined a strong candidate variant in HDAC7 (histone decetylase 7). Five patients had homozygous alterations in genes coding early complement proteins. This may lead to decreased clearance of apoptotic bodies. One patient had DNASE1L3 variant, which functions in the clearance of self-antigens. In 1 patient, we determined a novel gene that may be important in SLE pathogenesis. We suggest that monogenic causes/associations should be sought in early-onset and/or familial SLE.

Human IgM monoclonal antibodies block HIV-transmission to immune cells in cervico-vaginal tissues and across polarized epithelial cells in vitro

The importance of natural IgM antibodies in protection against infections is still emerging and these antibodies have a potential role in the maintenance of homeostasis through clearance of apoptotic bodies, complement-dependent mechanisms, inflammation and exclusion of misfolded proteins. Natural IgM act as a first line of defence against unknown hazardous factors and are present in most vertebrates. We investigated the functional capacity of anti-HIV-1 IgM monoclonal antibodies, from a combinatorial Fab library derived from healthy individuals, and evaluated their protective role in inhibiting HIV-1 in vitro when passing across the human mucosal epithelial barrier. Primary HIV-1 isolates were efficiently transmitted over the tight polarized epithelial cells when added to their apical surface. Efficient inhibition of HIV-1 transmission was achieved when anti-HIV-1 IgM monoclonal antibodies were added to the basolateral side of the cells. Two of these human IgM MoAbs had the ability to neutralize HIV and reduced infection of dendritic cells in primary cervico-vaginal tissue biopsies in vitro. This indicates a potential role of natural IgM antibodies in the reduction of HIV-1 transmission in mucosal tissues and improve our understanding of how natural IgM antibodies against a neutralizing epitope could interfere with viral transmission.

Abstract 428: An Unexpected Role for ACC in Lipid Droplet Formation in Macrophages in Response to Acellular Adipocyte Fat

In adipose tissues of obese mice, macrophages accumulate in crown-like structures (CLS) formed around dying adipocytes. The CLS macrophages are thought to clear the dead adipocytes by exophagy, which involves exocytosis of lysosomes and digestion of apoptotic adipocytes. After digesting the plasma membrane and other cytosolic contents of the adipocyte, CLS macrophages come in contact with the large lipid droplet, however, it is unknown how macrophages response to such acellular adipocytes. We hypothesized that the acelular adipocytes in CLSs promote inflammation and metabolically activate the macrophages to promote clearance of the lipid. To mimic the in vivo scenario, we exposed cultured murine bone marrow-derived macrophages to lipid droplets isolated from the adipocytes of high-fat diet-induced obese C57BL/6 mice. In response to adipocyte lipid, macrophages accumulated lipid droplets, which were similar in size and number to lipid droplets of CLS macrophages in vivo . Acellular lipid exposure significantly increased TNF-α gene expression, which was suppressed by the CD36 inhibitor sulfo-N-succinimidyl oleate, supporting CD36-mediated inflammatory response. Moreover, lipid exposure significantly lowered metabolic activity of macrophages and impaired clearance of apoptotic bodies from 3T3-L1 adipocytes. Using specific inhibitors, unexpectedly we found that lipid droplet accumulation in macrophages was independent of CD36 activity or processes involved in exophagy such as exocytosis of lysosomes, extracellular lipase activity, lipolysis and phagocytosis. Interestingly, lipid droplet accumulation was dependent on acetyl-CoA carboxylase (ACC) as determined by use of ACC inhibitors soraphen A and TOFA, and siRNA knock-down of ACC in macrophages. Furthermore, confocal microscopy of whole mount adipose tissue revealed expression of ACC in CLS macrophages. Altogether, using a novel in vitro model we demonstrate that acelular adipocytes suppress metabolic activity, but induce inflammation and de novo lipogenesis-mediated lipid droplet formation in macrophages.

Gas6 /TAM system: physiological insights and diseases

Growth arrest specific 6 (Gas6) is a multimodular circulating protein, the actions are mediated by the interaction with three transmembrane tyrosine kinase receptors: Tyro3, Axl, and MerTK, named TAM. Their regulatory roles are prominent in the mature immune, reproductive, hematopoietic, vascular, and nervous systems. Gas6 /TAM system plays a role in the regulation of inflammation, coagulation, cell growth, and clearance of apoptotic bodies. Deficiencies in TAM signaling are thought to contribute to chronic inflammatory and autoimmune disease in humans, and aberrantly elevated TAM signaling is strongly associated with cancer progression, and metastasis. We review the function of the Gas6/TAM system and the current evidence supporting its potential role in the pathogenesis of different diseases.

Abstract B006: Functional antagonism of VSIG8-mediated immune suppression by oral VISTA agents

Abstract With the remarkable success of antibodies focusing on PD-1/PD-L1, the immune checkpoint blockade approach has established itself as a cornerstone to cancer therapy. While PD-1/PD-L1 antibodies primarily focus on T cells to achieve antitumor efficacy, other cells in the tumor microenvironment such as myeloid cells, including MDSCs, also play a role in immune evasion, thus contributing to the lack of response in 70-80% of patients. To overcome the immune resistance induced by MDSCs, V-domain Ig suppressor of T-cell activation (VISTA) expressed predominantly on myeloid cells and tumor-infiltrating lymphocytes is considered as an ideal target. Recent findings also support the role of VISTA pathway in clearance of apoptotic bodies and prevention of autoimmunity. VISTA is reported to mediate immune suppression through homophillic interaction as well as interaction with V-Set and immunoglobulin domain containing 8 (VSIG8). We sought to discover and develop an orally available, small-molecule VISTA antagonist targeting both VISTA and VSIG8 pathways. Unlike antibodies targeting VISTA in early clinical trials, an oral agent potentially offers the convenience and flexibility to adjust the dose and schedule to address any emergent adverse events and ease of combination therapy. Since VISTA belongs to B7 family and the extracellular Ig domain of VISTA shares significant sequence homology with the B7 family ligands PD-L1 and PD-L2, a focused library of compounds mimicking the interaction of checkpoint proteins of B7 family was designed and synthesized. Screening and analysis of the resulting library led to the identification of hits capable of functional disruption of the checkpoint protein(s) signaling, depending upon the pockets of sequence similarity of interacting proteins. Further optimization resulted in lead compounds targeting both VISTA and VSIG8 signaling pathways with desirable drug-like properties including good oral bioavailability. Potent functional activity comparable to that obtained with an anti-VISTA or anti-VSIG8 antibody in rescuing effector functions was observed with the lead compound along with selectivity against other immune checkpoint proteins. An advanced lead compound exhibited sustained immune PD in tumor-bearing animals including desirable impact on myeloid and T cells in both circulation and tumor. The advanced lead compound also exhibited significant efficacy in syngeneic preclinical tumor models of melanoma and colon cancers upon once-a-day oral dosing with excellent tolerability. Further development of the oral VISTA antagonist is under way towards advancing it to the clinic. Citation Format: Pottayil G. Sasikumar, Sudarshan S. Naremaddepalli, Raghuveer K. Ramachandra, Nagesh Gowda, Manikyala Rao Yerramsetti, Srinivasa Rao Bandireddy, Sreenivas Adurthi, Jiju Mani, Rashmi Nair, Amit A. Dhudashia, Samiulla S. Dodheri, Nagaraj M. Gowda, Murali Ramachandra. Functional antagonism of VSIG8-mediated immune suppression by oral VISTA agents [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr B006.

Cell Death in the Vessel Wall: The Good, the Bad, the Ugly.

Atherosclerotic vascular disease results from an imbalance of inflammatory and vascular cell accumulation and removal in the neointimal space. When pathways that promote cell recruitment, survival, and proliferation are favored over to those that activate cell death, egress, and clearance, the plaque expands. In contrast, programmed cell death and the efficient clearance of apoptotic bodies by efferocytosis reduce lesion cellularity and promote a reparative environment and lesion stability. However, should these carefully balanced pathways become disturbed, lesions can accumulate cell debris, damaged associated molecular patterns, and arrested macrophages, all contributing to the proinflammatory environment and lesion instability. Here, we review the latest understanding of how cell death in the vessel wall directly coordinates the development of atherosclerosis, and what molecular signals are orchestrating these pathways. We discuss the necessity of cell death and the ways in which the execution of different forms of cell death can direct different outcomes in the plaque, and how promoting the effective clearance of dead cells from the lesion is looking like a promising therapeutic path forward. In the setting of excess cholesterol and fat, inflammatory pathways of the innate immune system are aberrantly activated. It is widely believed that oxidative modifications of the phospholipid and protein moieties of low- and very–low-density lipoproteins trigger conserved nonspecific scavenging pathways in macrophages and dendritic cells. This initiates what is at first considered a protective pathway to remove unwanted and potentially cytotoxic remnants and debris. However, during the decades of life wherein circulating low- and very–low-density lipoprotein cholesterol are in excess, this protective response turns into a chronic state of inflammation that recruits additional inflammatory cells and activates vascular cells to promote lesion expansion. During the progression of atherosclerosis, like in many other diseases, there is a constant turnover of cells within and surrounding the plaque. The identification …