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SARS-CoV-2 has continued spreading around the world in recent years since the initial outbreak in 2019, frequently developing into new variants with greater human infectious capacity. SARS-CoV-2 and its mutants use the angiotensin-converting enzyme 2 (ACE2) as a cellular entry receptor, which has triggered several therapeutic strategies against COVID-19 relying on the use of ACE2 recombinant proteins as decoy receptors. In this work, we propose an ACE2 silent Fc fusion protein (ACE2-hFcLALA) as a candidate therapy against COVID-19. This fusion protein was able to block the binding of SARS-CoV-2 RBD to ACE2 receptor as measured by ELISA and flow cytometry inhibition assays. Moreover, we used classical neutralization assays and a progeny neutralization assay to show that the ACE2-hFcLALA fusion protein is capable of neutralizing the authentic virus. Additionally, we found that this fusion protein was more effective in preventing in vitro infection with different variants of interest (alpha, beta, delta, and omicron) compared to the D614G strain. Our results suggest the potential of this molecule to be used in both therapeutic and preventive settings against current and emerging mutants that use ACE2 as a gateway to human cells.

Despite the cumulative evidence supporting HER3 as a target for antitumor therapies, no agents targeting HER3 have been approved for cancer treatment. Most of the agents evaluated in preclinical and clinical trials have been specific monoclonal antibodies (MAbs), with few examples of active immunotherapy directed against this receptor. However, some cancer vaccine formats may generate polyclonal antibodies (PAbs) that replicate the diverse effector mechanisms of MAbs, including ligand neutralization and receptor degradation. In this study, we developed a protein subunit-based monovalent vaccine candidate targeting the extracellular domain (ECD) of HER3. Immunization of mice with a formulation targeting murine ErbB3-ECD successfully overcome tolerance to this self-antigen, inducing high titers of ErbB3-specific PAbs. The antitumor potential of this formulation and the induced PAbs was demonstrated in vivo and in vitro in an ErbB3-overexpressing 3LL-D122-derived tumor model. The immunogenicity of the HER3-ECD-based vaccine candidate was confirmed by the induction of high titers of HER3-specific PAbs. Consistent with the initial results, HER3-ECD-targeting PAbs were cytotoxic in several human epithelial tumor cell lines and exerted antitumor effects in vivo. These results support the value of HER3 as a tumor antigen and the effector mechanisms of HER3-specific therapeutic MAbs, while suggesting the potential of the proposed vaccine candidate for the treatment of HER3-expressing carcinomas.

BackgroundAntitumor therapies targeting HER1/EGFR and HER2, such as monoclonal antibodies (MAbs) and tyrosine-kinase inhibitors (TKIs), have demonstrated a significant clinical benefit, but the emergence of resistance limits long-term efficacy. While secondary HER1 mutations confer tolerance to TKI, compensatory upregulation of HER2 drives resistance to anti-HER1 MAbs, which identifies MAb combinations targeting both receptors as an attractive therapeutic strategy. Nevertheless, toxicity hampers the clinical validation of this approach. Alternatively, cancer vaccines may induce antibodies directed against several antigens with less concern about induced toxicity.MethodsPolyclonal antibodies (PAbs) targeting HER1 and HER2 were induced in mice or rabbits through immunization. Recognition of different epitopes on targets by PAbs was validated by phage-display technology. Receptor downregulation was evaluated by flow cytometry, immunofluorescence, and Western blot. MTT assays assessed cytotoxicity, while the antitumor effect of PAbs was assayed in nude mice.ResultsPAbs promoted degradation of HER1 and HER2 regarding clinical MAbs or their combinations. As a result, inhibition of cytotoxicity on tumor cell lines was improved, even in the presence of oncogenic mutations in HER1, as well as in cetuximab-insensitive cells. Accordingly, the antitumor effect of vaccination-induced PAbs was observed in lung tumor lines representative of sensitivity or resistance to HER1 targeting therapies.ConclusionsImmunization against HER1 and HER2 receptors offers an alternative to passive administration of combinations of MAbs, since vaccination-induced PAbs promote the downregulation of both receptors and they have a higher impact on the survival of tumor cells.