Accumulation of the COMMD1 protein as a druggable pharmacology event to target cancer cells has not been evaluated so far in cancer animal models. We have previously demonstrated that a second-generation peptide, with cell-penetrating capacity, termed CIGB-552, was able to induce apoptosis mediated by stabilization of COMMD1. Here, we explore the antitumor effect by subcutaneous administration of CIGB-552 in a therapeutic schedule. Outstandingly, a significant delay of tumor growth was observed at 0.2 and 0.7 mg/kg (p < 0.01) or 1.4 mg/kg (p < 0.001) after CIGB-552 administration in both syngeneic murine tumors and patient-derived xenograft models. Furthermore, we evidenced that (131)I-CIGB-552 peptide was actually accumulated in the tumors after administration by subcutaneous route. A typical serine-proteases degradation pattern for CIGB-552 in BALB/c mice serum was identified. Further, biological characterization of the main metabolites of the peptide CIGB-552 suggests that the cell-penetrating capacity plays an important role in the cytotoxic activity. This report is the first in describing the antitumor effect induced by systemic administration of a peptide that targets COMMD1 for stabilization. Moreover, our data reinforce the perspectives of CIGB-552 for cancer targeted therapy.
The effects of lysine N(epsilon)-trimethylation at selected positions of the antimicrobial cecropin A-melittin hybrid peptide KWKLFKKIGAVLKVL-amide have been studied. All five monotrimethylated, four bis-trimethylated plus the per-trimethylated analogues have been synthesized and tested for antimicrobial activity on Leishmania parasites and on Gram-positive and -negative bacteria, as well as for hemolysis of sheep erythrocytes as a measure of cytotoxicity. The impact of trimethylation on the solution conformation of selected analogues has been evaluated by NMR, which indicates a slight decrease in the alpha-helical content of the modified peptides, particularly in the N-terminal region. Trimethylation also enhances the proteolytic stability of mono- and bis-trimethylated analogues by 2-3-fold. Although it tends to lower antimicrobial activity in absolute terms, trimethylation causes an even higher decrease in hemolytic activity and therefore results in improved selectivity for several analogues. The monotrimethylated analogue at position 6 shows the overall best selectivity against both the Leishmania donovani protozoan and Acinetobacter baumannii, a Gram-negative bacterium of increasing clinical concern.
The effort to develop vaccines based on economically accessible technological platforms available by developing countries vaccine manufacturers is essential to extend the immunization to the whole world population and to achieve the desired herd immunity, necessary to end the COVID–19 pandemic. Here we report on the development of a SARS–CoV–2 receptor–binding domain (RBD) protein, expressed in yeast Pichia pastoris. The RBD was modified with the addition of flexible N– and C–terminal amino acid extensions aimed to modulate the protein/protein interactions and facilitate protein purification. Fermentation with yeast extract culture medium yielded 30–40 mg/L. After purification by immobilized metal ion affinity chromatography and hydrophobic interaction chromatography, the RBD protein was characterized by mass–spectrometry, circular dichroism, and binding affinity to angiotensin–converting enzyme 2 (ACE2) receptor. The recombinant protein shows high antigenicity with convalescent human sera and also with sera from individuals vaccinated with the Pfizer–BioNTech mRNA or Sputnik V adenoviral–based vaccines. The RBD protein stimulates IFNγ, IL–2, IL–6, IL–4, and TNFα in mice secreting splenocytes from PBMC and lung, CD3+ enriched cells. Immunogenicity studies with 50 μg of the recombinant RBD formulated with alum, induce high levels of binding antibodies in mice and non–human primates, assessed by ELISA plates covered with RBD protein expressed in HEK293T cells. The mouse sera inhibited the RBD binding to ACE2 receptor in an in–vitro test and show neutralization of SARS–CoV–2 infection of Vero E6 cells. These data suggest that the RBD recombinant protein expressed in yeast P. pastoris is suitable as a vaccine candidate against COVID–19.
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