Background The use of checkpoint inhibitors has revolutionized cancer therapy. Unfortunately, these therapies often cause immune-related adverse effects, largely due to a lack of tumor specificity. Methods We stained human natural killer cells using fusion proteins composed of the extracellular portion of various tumor markers fused to the Fc portion of human IgG1, and identified Nectin4 as a novel TIGIT ligand. Next, we generated a novel Nectin4 blocking antibody and demonstrated its efficacy as a checkpoint inhibitor in killing assays and in vivo. Results We identify Nectin4 to be a novel ligand of TIGIT. We showed that, as opposed to all other known TIGIT ligands, which bind also additional receptors, Nectin4 interacts only with TIGIT. We show that the TIGIT-Nectin4 interaction inhibits natural killer cell activity, a critical part of the innate immune response. Finally, we developed blocking Nectin4 antibodies and demonstrated that they enhance tumor killing in vitro and in vivo. Conclusion We discovered that Nectin4 is a novel ligand for TIGIT and demonstrated that specific antibodies against it enhance tumor cell killing in vitro and in vivo. Since Nectin4 is expressed almost exclusively on tumor cells, our Nectin4-blocking antibodies represent a combination of cancer specificity and immune checkpoint activity, which may prove more effective and safe for cancer immunotherapy.

The poliovirus receptor (PVR) is a ubiquitously expressed glycoprotein involved in cellular adhesion and immune response. It engages the activating receptor DNAX accessory molecule (DNAM)-1, the inhibitory receptor TIGIT, and the CD96 receptor with both activating and inhibitory functions. Human cytomegalovirus (HCMV) down-regulates PVR expression, but the significance of this viral function in vivo remains unknown. Here, we demonstrate that mouse CMV (MCMV) also down-regulates the surface PVR. The m20.1 protein of MCMV retains PVR in the endoplasmic reticulum and promotes its degradation. A MCMV mutant lacking the PVR inhibitor was attenuated in normal mice but not in mice lacking DNAM-1. This attenuation was partially reversed by NK cell depletion, whereas the simultaneous depletion of mononuclear phagocytes abolished the virus control. This effect was associated with the increased expression of DNAM-1, whereas TIGIT and CD96 were absent on these cells. An increased level of proinflammatory cytokines in sera of mice infected with the virus lacking the m20.1 and an increased production of iNOS by inflammatory monocytes was observed. Blocking of CCL2 or the inhibition of iNOS significantly increased titer of the virus lacking m20.1. In this study, we have demonstrated that inflammatory monocytes, together with NK cells, are essential in the early control of CMV through the DNAM-1–PVR pathway.

Numerous viruses hijack cellular protein trafficking pathways to mediate cell entry or to rearrange membrane structures thereby promoting viral replication and antagonizing the immune response. Adaptor protein complexes (AP), which mediate protein sorting in endocytic and secretory transport pathways, are one of the conserved viral targets with many viruses possessing AP-interacting motifs. We present here different mechanisms of viral interference with AP complexes and the functional consequences that allow for efficient viral propagation and evasion of host immune defense. The ubiquity of this phenomenon is evidenced by the fact that there are representatives for AP interference in all major viral families, covered in this review. The best described examples are interactions of human immunodeficiency virus and human herpesviruses with AP complexes. Several other viruses, like Ebola, Nipah, and SARS-CoV-2, are pointed out as high priority disease-causative agents supporting the need for deeper understanding of virus-AP interplay which can be exploited in the design of novel antiviral therapies.

Studies assessing the dynamics and duration of antibody responses following SARS-CoV-2 infection or vaccination are an invaluable tool for vaccination schedule planning, assessment of risk groups and management of pandemics. In this study, we developed and employed ELISA assays to analyze the humoral responses to Nucleocapsid and Spike proteins in vaccinated health-care workers (HCW) and critically ill COVID-19 patients. Sera of more than 1000 HCWs and critically ill patients from the Clinical Hospital Center Rijeka were tested across a one-year period, encompassing the spread of major SARS-CoV-2 variants of concern (VOCs). We observed 97% of seroconversion in HCW cohort as well as sustained anti-Spike antibody response in vaccinees for more than 6 months. In contrast, the infection-induced anti-Nucleocapsid response was waning significantly in a six-month period. Furthermore, a substantial decrease in vaccinees’ anti-Spike antibodies binding to Spike protein of Omicron VOC was also observed. Critically ill COVID-19 patients had higher levels of anti-Spike and anti-Nucleocapsid antibodies compared to HCWs. No significant differences in anti-Spike and anti-Nucleocapsid antibody levels between the critically ill COVID-19 patients that were on non-invasive oxygen supplementation and those on invasive ventilation support were observed. However, stronger anti-Spike, but not anti-Nucleocapsid, antibody response correlated with a better disease outcome in the cohort of patients on invasive ventilation support. Altogether, our results contribute to the growing pool of data on humoral responses to SARS-CoV-2 infection and vaccination.

The global pandemic caused by SARS-CoV-2 is a major public health problem. Virus entry occurs via binding to ACE2. Five SARS-CoV-2 variants of concern (VOCs) were reported so far, all having immune escape characteristics. Infection with the current VOC Omicron was noticed in immunized and recovered individuals; therefore, the development of new treatments against VOC infections is urgently needed. Most approved mAbs treatments against SARS-CoV-2 are directed against the spike protein of the original virus and are therefore inefficient against Omicron. Here, we report on the generation of hACE2.16, an anti-ACE2 antibody that recognizes and blocks ACE2-RBD binding without affecting ACE2 enzymatic activity. We demonstrate that hACE2.16 binding to ACE2 does not affect its surface expression and that hACE2.16 blocks infection and virus production of various VOCs including Omicron BA.1 and BA.2. hACE2.16 might, therefore, be an efficient treatment against all VOCs, the current and probably also future ones.