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BACKGROUND: The CD47-signal regulatory protein alpha (SIRPα) 'don't eat me' signalling axis is perhaps the most prominent innate immune checkpoint to date. However, from initial clinical trials, it is evident that monotherapy with CD47-SIRPα blocking has a limited therapeutic effect at the maximum tolerated dose. Furthermore, treatment is associated with severe side effects, most notably anaemia, that are attributable to the ubiquitous expression of CD47. Nevertheless, promising clinical responses have been reported upon combination with the tumour-targeting antibody rituximab or azacytidine, although toxicity issues still hamper clinical application.MAIN BODY: Here, we discuss the current state of CD47-SIRPα blocking therapy with a focus on limitations of current strategies, such as depletion of red blood cells. Subsequently, we focus on innovations designed to overcome these limitations. These include novel antibody formats designed to selectively target CD47 on tumour cells as well as tumour-targeted bispecific antibodies with improved selectivity. In addition, the rationale and outcome of combinatorial approaches to improve the therapeutic effect of CD47 blockade are discussed. Such combinations include those with tumour-targeted opsonizing antibodies, systemic therapy, epigenetic drugs, other immunomodulatory T-cell-targeted therapeutics or dual immunomodulatory CD47 bispecific antibodies.CONCLUSION: With these advances in the design of CD47-SIRPα-targeting therapeutic strategies and increasing insight into the mechanism of action of this innate checkpoint, including the role of adaptive immunity, further advances in the clinical application of this checkpoint can be anticipated.

BACKGROUND: The CD47-signal regulatory protein alpha (SIRPα) 'don't eat me' signalling axis is perhaps the most prominent innate immune checkpoint to date. However, from initial clinical trials, it is evident that monotherapy with CD47-SIRPα blocking has a limited therapeutic effect at the maximum tolerated dose. Furthermore, treatment is associated with severe side effects, most notably anaemia, that are attributable to the ubiquitous expression of CD47. Nevertheless, promising clinical responses have been reported upon combination with the tumour-targeting antibody rituximab or azacytidine, although toxicity issues still hamper clinical application. MAIN BODY: Here, we discuss the current state of CD47-SIRPα blocking therapy with a focus on limitations of current strategies, such as depletion of red blood cells. Subsequently, we focus on innovations designed to overcome these limitations. These include novel antibody formats designed to selectively target CD47 on tumour cells as well as tumour-targeted bispecific antibodies with improved selectivity. In addition, the rationale and outcome of combinatorial approaches to improve the therapeutic effect of CD47 blockade are discussed. Such combinations include those with tumour-targeted opsonizing antibodies, systemic therapy, epigenetic drugs, other immunomodulatory T-cell-targeted therapeutics or dual immunomodulatory CD47 bispecific antibodies. CONCLUSION: With these advances in the design of CD47-SIRPα-targeting therapeutic strategies and increasing insight into the mechanism of action of this innate checkpoint, including the role of adaptive immunity, further advances in the clinical application of this checkpoint can be anticipated.

Background and Aims: The macrophage “don’t eat me” pathway CD47/SIRPα is a target for promising new immunotherapy. We hypothesized that a soluble variant of SIRPα is present in the blood and may function as a biomarker. Methods: Monocyte derived macrophages (MDMs) from human buffy-coats were stimulated into macrophage subtypes by LPS and IFN-γ (M1), IL-4 and IL-13 (M2a), IL-10 (M2c) and investigated using flow cytometry. Soluble SIRPα (sSIRPα) was measured in cell cultures and serum by Western blotting and an optimized ELISA. Serum samples were obtained from 120 healthy individuals and from 8 individuals challenged by an LPS injection. Results: All macrophage phenotypes expressed SIRPα by flowcytometry, and sSIRPα was present in all culture supernatants including unstimulated cells. M1 macrophages expressed the lowest level of SIRPαand released the highest level of sSIRPα (p < 0.05). In vivo, the serum level of sSIRPα increased significantly (p < 0.0001) after an LPS challenge in humans. The median concentration in healthy individuals was 28.7 µg/L (19.8–41.1, 95% reference interval), and 20.5 µg/L in an IFCC certified serum reference material. The protein was stable in serum for prolonged storage and repeated freeze/thawing. Conclusions: We demonstrate that sSIRPα is produced constitutively and the concentration increases upon macrophage activation both in vitro and in vivo. It is present in human serum where it may function as a biomarker for the activity of tumor-associated macrophages (TAMs), and for monitoring the effect of immunotherapy.

Classic drug development strategies have failed to meet the urgent clinical needs in treating infections with Gram-negative bacteria. Repurposing drugs can lead to timely availability of new antibiotics, accelerated by existing safety profiles. Glatiramer acetate (GA) is a widely used and safe formulation for treatment of multiple sclerosis. It contains a large diversity of essentially isomeric polypeptides with the cationic and amphiphilic character of many antimicrobial peptides (AMP). Here, we report that GA is antibacterial, targeting Gram-negative organisms with higher activity towards Pseudomonas aeruginosa than the naturally-occurring AMP LL-37 in human plasma. As judged from flow cytometric assays, bacterial killing by GA occurred within minutes. Laboratory strains of Escherichia coli and P. aeruginosa were killed by a process of condensing intracellular contents. Efficient killing by GA was also demonstrated in Acinetobacter baumannii clinical isolates and approximately 50% of clinical isolates of P. aeruginosa from chronic airway infection in CF patients. By contrast, the Gram-positive Staphylococcus aureus cells appeared to be protected from GA by an increased formation of nm-scale particulates. Our data identify GA as an attractive drug repurposing candidate to treat infections with Gram-negative bacteria.

Background Fascioliasis is an infectious disease caused by parasites Fasciola hepatica and F. gigantica. Humans are infected by the consumption of vegetables and water contaminated with the infective form of the parasite. Materials and methods In this study, an IgM-ELISA with the cysteine proteinase Fas2 antigen was evaluated with sera from 76 patients infected with F. hepatica, 24 patients with other parasite infections and 84 healthy volunteers. Results IgM-ELISA resulted in 43% positives in F. hepatica patients with positive serology to Fas2-ELISA, but no positives resulted from testing healthy volunteers and individuals infected with other parasites. The IgM-ELISA diagnostic parameters showed a sensitivity of 43.4% (95% CI 0.321-0.553), specificity of 100% (95% CI 0.957-1), and no cross-reactivity with other parasitic infection. Interference by rheumatoid factor in the IgM immunoassay was controlled by treating sera with rheumatoid factor absorbent before testing. Conclusions Fas2 antigen is detected by circulating IgM in patients infected with F. hepatica and IgM-ELISA using Fas2 appears as a specific immunoassay to detect the acute phase of the acute phase of F. hepatica infection in humans.

Tumor necrosis factor (TNF) has been implicated in inflammation-associated tumor progression. Although multiple reports identified a role for TNF signaling in established cancers, few studies have assessed the impact of TNF blockade on early tumor formation promotion. We aimed at exploring the effects of TNF neutralization in a preclinical mouse model of lung carcinogenesis. For this, Balb/c mice (n = 42) received four weekly intraperitoneal urethane injections (1 g/kg) and twice-weekly intraperitoneal soluble TNF receptor (etanercept; 10 mg/kg) administered during tumor initiation/promotion, tumor progression, or continuously (months 1, 6, and 1-8 after urethane start, respectively). Lung oncogenesis was assessed after 8 months. In separate short-term studies, Balb/c mice (n = 21) received a single control or urethane injection followed by twice-weekly intraperitoneal control or sTNFR:Fc injections. Lung inflammation was assessed after 1 week. We found that sTNFR:Fc treatment during tumor initiation/promotion resulted in a significant reduction of tumor number but not dimensions. However, sTNFR:Fc administered during tumor progression did not impact tumor multiplicity but significantly decreased tumor diameter. Continued sTNFR:Fc administration was effective in halting both respiratory tumor formation and progression in response to urethane. This favorable impact was associated with impaired cellular proliferation and new vessel formation in lung tumors. In addition, TNF neutralization altered the lung inflammatory response to urethane, evidenced by reductions in TNF and macrophage and increases in interferon γ and interleukin 10 content of the air spaces. sTNFR:Fc treatment of RAW264.7 macrophages downregulated TNF and enhanced interferon γ and interleukin 10 expression. In conclusion, TNF neutralization is effective against urethane-induced lung oncogenesis in mice and could present a lung chemoprevention strategy worth testing clinically.

Immunomodulatory properties of 10 dairy propionibacteria, analyzed on human peripheral blood mononuclear cells (PBMCs), revealed a highly strain-dependent induction of anti-inflammatory cytokine interleukin 10 (IL-10). Two selected strains of Propionibacterium freudenreichii showed a protective effect against two models of colitis in mice, suggesting a probiotic potential predicted by immune-based selection criteria for these cheese starter bacteria.

Interferon-beta (IFN-beta) achieves its beneficial effect on multiple sclerosis (MS) via anti-inflammatory properties. In this study, we assessed the expression of the brain-derived neurotrophic factor (BDNF) in peripheral blood mononuclear cells (PBMC) from relapsing-remitting multiple sclerosis (RRMS) patients treated or not with IFN-beta. Intracellular BDNF was measured by Western blot and ELISA and compared with serum BDNF. We found higher levels of BDNF in PBMC of IFN-beta-treated versus non-treated patients, whereas serum levels of BDNF were similar. We hypothesize that the increased intracellular BDNF secondary to IFN-beta is not released in the periphery. This release is probably not tissue specific but in MS patients, BDNF could be specifically delivered by PBMC at the site of re-activation, i.e. within the central nervous system.