Novel therapeutic peptides are increasingly making their way into clinical application. The cationic and amphipathic properties of certain peptides allow them to cross biological membranes in a non-disruptive way without apparent toxicity increasing drug bioavailability. By modifying the primary structure of the Limulus-derived LALF(32-51) peptide we designed a novel peptide, L-2, with antineoplastic effect and cell-penetrating capacity. Interestingly, L-2 induced cellular cytotoxicity in a variety of tumor cell lines and systemic injection into immunocompetent and nude mice bearing established solid tumor, resulted in substantial regression of the tumor mass and apoptosis. To isolate the gene transcripts specifically regulated by L-2 in tumor cells, we conducted suppressive subtractive hybridization (SSH) analysis and identified a set of genes involved in biological processes relevant to cancer biology. Our findings describe a novel peptide that modifies the gene expression of the tumor cells and exhibits antitumor effect in vivo, indicating that peptide L-2 is a potential candidate for anticancer therapy.
Objective. CIGB-228 is a novel therapeutic vaccine consisting of HLA-restricted HPV16 E7 epitope adjuvated with VSSP. This trial was designed to evaluate the toxicity, safety, immunogenicity, HPV clearance, and lesion regression. Methods. Seven women were entered. All were HLA-A2 positive, had biopsy-proven high-grade CIN, histologically positive for HPV16, and beared persistent postbiopsy lesions visible by digital colposcopy. HLA-A2 women with biopsy-proven high-grade CIN, HPV16-positive, and beared persistent postbiopsy lesions visible by digital colposcopy were vaccinated. One weekly injections of CIGB-228 vaccine was given for four weeks. Then, loop electrosurgical excision procedure (LEEP) of the transformation zone was performed. Study subjects were followed for 1 year after LEEP. Results. No toxicity beyond grade 1 was observed during and after the four vaccinations. Five of seven women had complete and partial regression. Cellular immune response was seen in all patients. HPV was cleared in three of the patients with complete response.
Conclusion. CIGB-228 vaccination was well tolerated and capable to induce IFNγ-associated T-cell response in women with high-grade CIN. In several patients, lesion regression and HPV clearance were observed.
SummaryHigh‐risk human papillomaviruses (HPVs) cause cervical cancer, and while there are good prophylactic vaccines on the market, these are ineffective against established infections, creating a clear need for therapeutic vaccines. The HPV E7 protein is one of the essential oncoproteins for the onset and maintenance of malignancy and is therefore an ideal therapeutic vaccine target. We fused the HPV‐16 E7 protein to the Limulus polyphemus antilipopolysaccharide factor (LALF
32‐51), a small hydrophobic peptide that can penetrate cell membranes and that has immunomodulatory properties. LALF
32‐51‐E7 was transiently expressed in Nicotiana benthamiana, and we previously determined that it accumulated better when targeted to chloroplasts compared to being localized in the cytoplasm. Subsequently, we aimed to prove whether LALF
32‐51‐E7 was indeed associated with the chloroplasts by determining its subcellular localization. The LALF
32‐51‐E7 gene was fused to one encoding enhanced GFP to generate a LG fusion protein, and localization was determined by confocal laser scanning microscopy and transmission electron microscopy (TEM). The fluorescence observed from chloroplast‐targeted LG was distinctively different from that of the cytoplasmic LG. Small spherical structures resembling protein bodies (PBs) were seen that clearly localized with the chloroplasts. Larger but less abundant PB‐like structures were also seen for the cytoplasmic LG. PB‐like structure formation was confirmed for both LG and LALF
32‐51‐E7 by TEM. LALF
32‐51‐E7 was indeed targeted to the chloroplasts by the chloroplast transit peptide used in this study, and it formed aggregated PB‐like structures. This study could open a new avenue for the use of LALF
32‐51 as a PB‐inducing peptide.
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