Myeloid cells are recruited to damaged tissues where they can resolve infections and tumor growth or stimulate wound healing and tumor progression. Recruitment of these cells is regulated by ...integrins, a family of adhesion receptors that includes integrin CD11b. Here we report that, unexpectedly, integrin CD11b does not regulate myeloid cell recruitment to tumors but instead controls myeloid cell polarization and tumor growth. CD11b activation promotes pro-inflammatory macrophage polarization by stimulating expression of microRNA Let7a. In contrast, inhibition of CD11b prevents Let7a expression and induces cMyc expression, leading to immune suppressive macrophage polarization, vascular maturation, and accelerated tumor growth. Pharmacological activation of CD11b with a small molecule agonist, Leukadherin 1 (LA1), promotes pro-inflammatory macrophage polarization and suppresses tumor growth in animal models of murine and human cancer. These studies identify CD11b as negative regulator of immune suppression and a target for cancer immune therapy.
Podocyte injury is an early event in diabetic kidney disease and is a hallmark of glomerulopathy. MicroRNA-146a (miR-146a) is highly expressed in many cell types under homeostatic conditions, and ...plays an important anti-inflammatory role in myeloid cells. However, its role in podocytes is unclear. Here, we show that miR-146a expression levels decrease in the glomeruli of patients with type 2 diabetes (T2D), which correlates with increased albuminuria and glomerular damage. miR-146a levels are also significantly reduced in the glomeruli of albuminuric BTBR ob/ob mice, indicating its significant role in maintaining podocyte health. miR-146a-deficient mice (miR-146a−/−) showed accelerated development of glomerulopathy and albuminuria upon streptozotocin (STZ)-induced hyperglycemia. The miR-146a targets, Notch-1 and ErbB4, were also significantly up-regulated in the glomeruli of diabetic patients and mice, suggesting induction of the downstream TGFβ signaling. Treatment with a pan-ErbB kinase inhibitor erlotinib with nanomolar activity against ErbB4 significantly suppressed diabetic glomerular injury and albuminuria in both WT and miR-146a−/− animals. Treatment of podocytes in vitro with TGF-β1 resulted in increased expression of Notch-1, ErbB4, pErbB4, and pEGFR, the heterodimerization partner of ErbB4, suggesting increased ErbB4/EGFR signaling. TGF-β1 also increased levels of inflammatory cytokine monocyte chemoattractant protein-1 (MCP-1) and MCP-1 induced protein-1 (MCPIP1), a suppressor of miR-146a, suggesting an autocrine loop. Inhibition of ErbB4/EGFR with erlotinib co-treatment of podocytes suppressed this signaling. Our findings suggest a novel role for miR-146a in protecting against diabetic glomerulopathy and podocyte injury. They also point to ErbB4/EGFR as a novel, druggable target for therapeutic intervention, especially because several pan-ErbB inhibitors are clinically available.
Lung cancer is one of the leading causes of cancer-related deaths in the United States. A major hurdle for improved therapies is immune suppression mediated by the tumor and its microenvironment. The ...lung tumor microenvironment (TME) contains large numbers of tumor-associated macrophages (TAMs), which suppress the adaptive immune response, increase neo-vascularization of the tumor, and provide pro-tumor factors to promote tumor growth. CD11b is highly expressed on myeloid cells, including TAMs, where it forms a heterodimeric integrin receptor with CD18 (known as CD11b/CD18, Mac-1, CR3, and αMβ2), and plays an important role in recruitment and biological functions of these cells, and is a validated therapeutic target. Here, we describe our pre-clinical studies targeting CD11b in the context of lung cancer, using pharmacologic and genetic approaches that work via positive allosteric modulation of CD11b function. GB1275 is a novel small molecule modulator of CD11b that is currently in Phase 1/2 clinical development. We assess GB1275 treatment effects on tumor growth and immune infiltrates in the murine Lewis Lung Carcinoma (LLC) syngeneic tumor model. Additionally, as an orthogonal approach to determine mechanisms of action, we utilize our recently developed novel CD11b knock-in (KI) mouse that constitutively expresses CD11b containing an activating isoleucine to glycine substitution at residue 332 in the ligand binding CD11b A-domain (I332G) that acts as a positive allosteric modulator of CD11b activity. We report that pharmacologic modulation of CD11b with GB1275 significantly reduces LLC tumor growth. CD11b KI mice similarly show significant reduction in both the size and rate of LLC tumor growth, as compared to WT mice, mimicking our observed treatment effects with GB1275. Tumor profiling revealed a significant reduction in TAM infiltration in GB1275-treated and in CD11b KI mice, increase in the ratio of M1/M2-like TAMs, and concomitant increase in cytotoxic T cells. The profiling also showed a significant decrease in CCL2 levels and a concomitant reduction in Ly6C
monocytes in circulation in both groups. These findings suggest that positive allosteric modulation of CD11b reduces TAM density and reprograms them to enhance the adaptive immune response and is a novel therapeutic strategy against lung cancer.
Bidirectional interactions between the immune system and the nervous system are increasingly appreciated as playing a pathogenic role in chronic pain. Unraveling the mechanisms by which inflammatory ...pain is mediated through communication between nerves and immune cells may lead to exciting new strategies for therapeutic intervention. In this narrative review, we focus on the role of macrophages in the pathogenesis of osteoarthritis (OA) pain. From regulating homeostasis to conducting phagocytosis, and from inducing inflammation to resolving it, macrophages are plastic cells that are highly adaptable to their environment. They rely on communicating with the environment through cytokines, growth factors, neuropeptides, and other signals to respond to inflammation or injury. The contribution of macrophages to OA joint damage has garnered much attention in recent years. Here, we discuss how macrophages may participate in the initiation and maintenance of pain in OA. We aim to summarize what is currently known about macrophages in OA pain and identify important gaps in the field to fuel future investigations.
Introduction: Diabetic foot ulcer (DFU) prevalence is as high as 25% and 40-80% of DFUs become infected (DFI). About 20% of infected ulcers will spread to bone causing diabetic foot osteomyelitis ...(DFO). DFU costs Medicare $9-13 billion/year. The most expensive costs associated with DFU are inpatient costs and hospital admissions. DFO costs are driven mostly by surgical procedures. DFU patients have a 3-year cumulative mortality rate of 28% and rates approaching 50% in amputated patients.
Areas covered: This review will summarize the current health and economic burden of DFO covering management, epidemiology, and copious costs associated with DFO. The review began by searching PubMed and Cochrane databases for various terms including, 'diabetic osteomyelitis costs,' 'diabetic foot infection,' and 'diabetes and antibiotics.' Additionally, references from retrieved publications were reviewed. The global burden of DFU calls for investigating new therapeutic options.
Expert opinion: For DFI, anti-biofilm agents have had success because they directly deliver antimicrobials to the infection site. For DFO, intraosseous (I/O) antibiotic therapy similarly bypasses the issue of vascular disease, will likely have improved therapeutic efficacy, and reduced costs for DFO patients. I/O antibiotic therapy has had clinical success in one case report already, and may significantly improve the lives of those afflicted with DFO.
Diabetic glomerular injury is a major complication of diabetes mellitus and is the leading cause of end stage renal disease (ESRD). Healthy podocytes are essential for glomerular function and health. ...Injury or loss of these cells results in increased proteinuria and kidney dysfunction and is a common finding in various glomerulopathies. Thus, mechanistic understanding of pathways that protect podocytes from damage are essential for development of future therapeutics. MicroRNA-146a (miR-146a) is a negative regulator of inflammation and is highly expressed in myeloid cells and podocytes. We previously reported that miR-146a levels are significantly reduced in the glomeruli of patients with diabetic nephropathy (DN). Here we report generation of mice with selective deletion of miR-146a in podocytes and use of these mice in models of glomerular injury. Induction of glomerular injury in C57BL/6 wildtype mice (WT) and podocyte-specific miR-146a knockout (Pod-miR146a
) animals
administration of low-dose lipopolysaccharide (LPS) or nephrotoxic serum (NTS) resulted in increased proteinuria in the knockout mice, suggesting that podocyte-expressed miR-146a protects these cells, and thus glomeruli, from damage. Furthermore, induction of hyperglycemia using streptozotocin (STZ) also resulted in an accelerated development of glomerulopathy and a rapid increase in proteinuria in the knockout animals, as compared to the WT animals, further confirming the protective role of podocyte-expressed miR-146a. We also confirmed that the direct miR-146a target, ErbB4, was significantly upregulated in the diseased glomeruli and erlotinib, an ErbB4 and EGFR inhibitor, reducedits upregulation and the proteinuria in treated animals. Primary miR146
podocytes from these animals also showed a basally upregulated TGFβ-Smad3 signaling
. Taken together, this study shows that podocyte-specific miR-146a is imperative for protecting podocytes from glomerular damage,
modulation of ErbB4/EGFR, TGFβ, and linked downstream signaling.
Abstract
Lung cancer is the leading cause of cancer related deaths in the United States. With a 5-year survival rate of 18%, lung cancer patients are in immediate need of new therapeutic options. A ...major problem facing immunotherapy for cancer is the active immune suppression by the tumor. Tumors show presence of large numbers of tumor associated macrophages (TAMs), which suppress the adaptive immune response, increase neo-vascularization to the tumor, and promote tumor survival. Integrin CD11b is highly expressed on TAMs and is essential for their recruitment and biological functions. Reasoning that CD11b activity is important for controlling tumor growth, we developed a novel small molecule agonist called leuhadherin-1 (LA1), which activates CD11b. We found that CD11b activation by LA1 significantly reduced tumor growth in wild type mice bearing Lewis Lung Carcinoma (LLC) tumors as early as one week post treatment. Conversely, LLC tumors grew at a faster rate in CD11b-/- mice, compared to wild type mice, showing that CD11b is important for controlling tumor growth. Importantly, LA1 treatment in tumor bearing CD11b-/- mice did not show any efficacy, demonstrating the specificity of LA1 to CD11b. These data suggest that CD11b activation via LA1 modulates TAMs in tumors and is a novel therapeutic strategy against cancer.
Citation Format: Terese Geraghty, Anugraha Rajagopalan, Samia Khan, Judith Varner, Vineet Gupta. Pharmacologic activation of integrin CD11b as a novel therapeutic strategy against lung cancer abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 137.
Osteoarthritis (OA) is a leading cause of chronic pain, yet OA pain management remains poor. Age is the strongest predictor of OA development, and mechanisms driving OA pain are unclear. We undertook ...this study to characterize age-associated changes in knee OA, pain-related behaviors, and dorsal root ganglion (DRG) molecular phenotypes in mice of both sexes.
Male or female C57BL/6 mice 6 or 20 months of age were evaluated for histopathologic knee OA, pain-related behaviors, and L3-L5 DRG immune characterization via flow cytometry. DRG gene expression in older mice and humans was also examined.
Male mice at 20 months of age had worse cartilage degeneration than 6-month-old mice. Older female mouse knees showed increased cartilage degeneration but to a lesser degree than those of male mice. Older mice of both sexes had worse mechanical allodynia, knee hyperalgesia, and grip strength compared to younger mice. For both sexes, DRGs from older mice showed decreased CD45+ cells and a significant increase in F4/80+ macrophages and CD11c+ dendritic cells. Older male mouse DRGs showed increased expression of Ccl2 and Ccl5, and older female mouse DRGs showed increased Cxcr4 and Ccl3 expression compared to 6-month-old mouse DRGs, among other differentially expressed genes. Human DRG analysis from 6 individuals >80 years of age revealed elevated CCL2 in men compared to women, whereas CCL3 was higher in DRGs from women.
We found that aging in male and female mice is accompanied by mild knee OA, mechanical sensitization, and changes to immune cell populations in the DRG, suggesting novel avenues for development of OA therapies.
Abstract
Lung cancer is the leading cause of cancer-related deaths in the United States. With a 5-year survival rate at 19%, lung cancer patients are urgently in need of new therapeutic options. A ...major problem facing cancer therapy is the immune suppression by the tumor and its subsequent escape from the immune response, which is mediated by the tumor microenvironment. Tumors have large numbers of myeloid-derived suppressor cells (MDSCs), which suppress the adaptive immune response, increase neovascularization of the tumor, and promote tumor survival. Integrin heterodimer CD11b/CD18 (aka Mac-1 or αMβ2) is highly expressed on MDSCs and plays an important role in innate cell recruitment and other biologic functions. GB1275 (previously known as ADH503) is a novel small-molecule CD11b allosteric modulator. In experimental models of pancreatic cancer, GB1275 reduces MDSC infiltration and M2 macrophage polarization leading to tumor growth inhibition. GB1275 is currently in phase 1/2 clinical development in patients with pancreatic, esophageal, gastric/GEJ, triple-negative breast, castration-resistant prostate, or microsatellite stable colorectal cancer. To investigate the impact of CD11b modulation in the context of lung cancer, GB1275 treatment effects on tumor growth and immune infiltrates were assessed in the Lewis Lung Carcinoma (LLC) syngeneic tumor model, propagated in C57BL/6 wild-type (WT) mice. Additionally, we developed a transgenic CD11b knockin (KI) mouse that renders CD11b in a partially active state. CD11b KI mice were engineered to express partially active CD11b by introducing a point mutation at position 332 causing an isoleucine to glycine switch (I332G). We studied LLC tumor growth in CD11b KI mice as an orthogonal approach to test the mechanism of action. We will present data showing that pharmacologic modulation of CD11b with GB1275 significantly reduces LLC tumor growth. Concomitantly, we will present our studies with the CD11b KI mice that show a significant reduction in both the size and the rate of LLC tumor growth, as compared to the WT mice, mimicking treatment effects with GB1275. Our comprehensive pharmacologic and genetic studies show that CD11b activation reprograms MDSCs to enhance the adaptive immune response and is a novel strategy against lung cancer.
This abstract is also being presented as Poster B71.
Citation Format: Terese D. Geraghty, Anugraha Rajagopalan, Rabail Aslam, Vineet Gupta. Modulation of integrin CD11b as a novel therapeutic strategy against lung cancer abstract. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr PR4.
Globally, cancer is among the leading causes of death with 1 in 6 deaths due to cancer. Lung cancer is leading cause of cancer-related death accounting for more deaths than breast, prostate, and ...colon cancer combined. Current treatment options for lung cancer patients vary by stage and type of disease, but include surgical resection whenever possible, chemotherapy and radiotherapy when applicable. These treatments help control disease progression, but lack specificity and have unwanted side effects. Retraining the immune system to respond to cancer is an attractive alternative to chemo- or radio- therapies because it has the ability to circumvent some of these severe side effects. For example, immune checkpoint inhibitors (e.g., anti-PD1, anti-CTLA-4, etc.), work by augmenting the anti-tumor T cell response, and have had a significant impact on improving the survival of lung cancer patients. Although, these new therapies have done well to improve lung cancer patient survival, there is still more room for development of targeted therapeutics that improve patient outcomes.Beyond T cell targeted immunotherapy, there are increasing efforts to find other immune cells as targets for new immunotherapies. Lung tumors have large numbers of tumor associated macrophages (TAMs) which suppress the adaptive immune response, increase neo-vascularization of the tumor, and secrete pro-tumor factors to promote tumor growth. Integrin CD11b/CD18 (also known as Mac-1, CR3 and αMβ2) is highly expressed on myeloid cells, including TAMs, and plays an important role in innate cell recruitment and other biological functions. GB1275 (previously known as ADH-503) is a novel small molecule CD11b allosteric modulator. GB1275 is currently in Phase 1/2 clinical development for various cancer types. Here, we describe our pre-clinical studies to investigate the impact of CD11b modulation in the context of lung cancer.To assess the role of CD11b in regulating lung tumor growth, we studied lung tumor growth in the absence of CD11b using CD11b knockout (KO) mice, by pharmacologically modulating CD11b in WT mice by treatment with GB1275, or in the context of genetically modulated CD11b using CD11b knock-in (KI) mice. All tumors were propagated in the Lewis lung carcinoma (LLC) syngeneic tumor model. LLC tumors grew faster and had heavier tumor burden in CD11b KO mice as compared to WT controls. In contrast to CD11b KO mice, pharmacologic modulation of CD11b by GB1275 treatment significantly reduced tumor growth. We developed a transgenic CD11b knock-in (KI) mouse that renders CD11b in a partially active state by introducing a point mutation at residue 332 of the ligand binding CD11b A/I-domain, changing isoleucine to glycine (I332G). CD11b KI mice showed a significant reduction in both the size and the rate of LLC tumor growth, as compared to the WT mice, mimicking treatment effects with GB1275. Tumor immune profiling showed a significant reduction in monocyte, granulocyte, and mature TAM infiltration in GB1275-treated and CD11b KI tumors. Dendritic cell populations changed to enhance antigen cross presentation as a result of CD11b modulation. There was an increase in the frequency of CD8+ cytotoxic T lymphocytes and decrease of FoxP3+ regulatory T cells in GB1275-treated and CD11b KI tumors. CD11b modulation affected TAM polarization manifested by increased frequencies of M1-like F4/80+MHCII+ TAMs and decreased M2-like F4/80+CD206+ TAMs and from gene expression profiling of CD11b-modulated macrophages. CD11b-modulated macrophages showed significant downregulation of Ccl2 mRNA levels, which resulted in less CCL2 protein secreted into circulation. Taken together, these results indicate that CD11b modulation not only reprograms TAMs to enhance the T cell response, but also works by reducing trafficking of new TAMs in the lung tumor microenvironment to reduce the overall inflammatory burden.Overall, CD11b modulation affects both monocyte/macrophage recruitment to lung tumors and reprograms TAMs to enhance the anti-tumor immune response. There are many antagonism approaches to inhibit pro-tumoral TAMs, but here we describe an agonistic approach via CD11b modulation. As CD11b+ cells are highly abundant in human lung tumors, targeting CD11b via allosteric modulation is a novel therapeutic strategy against lung cancer.
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