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In vitro and ex vivo evaluation of a multi-epitope heparinase vaccine for various malignancies.

Tang XD, Guo SL, Wang GZ, Li N, Wu YY, Fang DC, Fan YH, Yang SM - Cancer Sci. (2013)

Bottom Line: Previous studies have indicated that heparanase (Hpa) might represent a candidate universal tumor-associated antigen.The results showed that multi-epitope vaccines Hpa525 + 277 + 405 + 16 and Hpa8 + 310 + 315 + 363 induced higher Hpa-specific lysis of various cancer cells from different tissues in a HLA-A2-restricted and heparanase-specific manner compared with the single epitope vaccines Hpa525, Hpa277, Hpa405, Hpa16, Hpa8, Hpa310, Hpa315 and Hpa363, both in vitro and ex vivo.Therefore, the present study provides theoretical evidence for the use of heparanase multi-epitope vaccines for clinical application.

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology, Xinqiao Hospital, Third Military Medical University, Chongqing, China; Department of Respiratory and Critical Care Medicine, The First Affiliated Hospital, Chongqing Medical University, Chongqing, China.

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Heparanase specificity and HLA-A2 restriction of cytotoxic T lymphocytes (CTL) generated from different heparanase vaccines in vitro (A) and ex vivo (B). The CTL generated from the HLA-A2-restricted nonapeptide HIVpol(476-484)(ILLEPVHGV) derived from HIV served as the negative peptide control (NP). (A) The cytotoxic activity of CTL induced by heparanase multi-epitope vaccines and single epitope peptides against MCF-7 (A[a, b]), MCF-7/Hpa (A[c, d]), HepG2 (A[e, f]) and HepG2/HLA-A2 cells (A[g,h]) in vitro. (B) The cytotoxic activity of CTL induced by heparanase multi-epitope vaccines and single epitope peptides against MCF-7 (B[a, b]), MCF-7/Hpa (B[c, d]), HepG2 (B[e, f]), and HepG2/HLA-A2 cells (B[g, h]) ex vivo. E/T, effector/target.
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fig04: Heparanase specificity and HLA-A2 restriction of cytotoxic T lymphocytes (CTL) generated from different heparanase vaccines in vitro (A) and ex vivo (B). The CTL generated from the HLA-A2-restricted nonapeptide HIVpol(476-484)(ILLEPVHGV) derived from HIV served as the negative peptide control (NP). (A) The cytotoxic activity of CTL induced by heparanase multi-epitope vaccines and single epitope peptides against MCF-7 (A[a, b]), MCF-7/Hpa (A[c, d]), HepG2 (A[e, f]) and HepG2/HLA-A2 cells (A[g,h]) in vitro. (B) The cytotoxic activity of CTL induced by heparanase multi-epitope vaccines and single epitope peptides against MCF-7 (B[a, b]), MCF-7/Hpa (B[c, d]), HepG2 (B[e, f]), and HepG2/HLA-A2 cells (B[g, h]) ex vivo. E/T, effector/target.

Mentions: To further confirm the heparanase specificity of the CTL in vitro and ex vivo, we used the HLA-A2-positive, heparanase-negative breast cancer cell line MCF-7 (Fig.4). MCF-7 cells were transfected with recombinant, replication-defective adenovirus (Ad-Hpa) encoding a full-length cDNA of human heparanase at a multiplicity of infection of 200 and cultured for 2 days in fresh DMEM medium containing 10% FCS. In our previous study, western blot analysis showed that heparanase protein could be detected in MCF-7/Hpa cells, whereas in MCF-7 cells the expression of heparanase is very weak.19 Heparanase-specific CTL were generated using peptide-pulsed DC from HLA-A2-positive PBMC and DC from the bone marrow of C57BL/6-Tg transgenic mice. After being stimulated three times, the killing effect of CTL induced by Hpa525 + 277 + 405 + 16 and Hpa8 + 310 + 315 + 363 and their corresponding single peptides Hpa525, Hpa277, Hpa405, Hpa16, Hpa8, Hpa310, Hpa315 and Hpa363 were determined at various E/T ratios using a 4-h 51Cr release assay. The results showed that these heparanase-specific CTL could lyse MCF-7/Hpa cells, whereas no obvious lysis of MCF-7 cells was detected even at the highest E/T ratio. Furthermore, compared with the corresponding single heparanase peptides, heparanase multi-epitope vaccines elicited more robust specific lysis of MCF-7/Hpa cells (Fig.4). These results clearly indicate that the CTL specifically targeted heparanase peptides that were presented in the context of HLA-A2 and the multi-epitope vaccines of human heparanase could elicit much more potent killing effects compared with their corresponding single peptides.


In vitro and ex vivo evaluation of a multi-epitope heparinase vaccine for various malignancies.

Tang XD, Guo SL, Wang GZ, Li N, Wu YY, Fang DC, Fan YH, Yang SM - Cancer Sci. (2013)

Heparanase specificity and HLA-A2 restriction of cytotoxic T lymphocytes (CTL) generated from different heparanase vaccines in vitro (A) and ex vivo (B). The CTL generated from the HLA-A2-restricted nonapeptide HIVpol(476-484)(ILLEPVHGV) derived from HIV served as the negative peptide control (NP). (A) The cytotoxic activity of CTL induced by heparanase multi-epitope vaccines and single epitope peptides against MCF-7 (A[a, b]), MCF-7/Hpa (A[c, d]), HepG2 (A[e, f]) and HepG2/HLA-A2 cells (A[g,h]) in vitro. (B) The cytotoxic activity of CTL induced by heparanase multi-epitope vaccines and single epitope peptides against MCF-7 (B[a, b]), MCF-7/Hpa (B[c, d]), HepG2 (B[e, f]), and HepG2/HLA-A2 cells (B[g, h]) ex vivo. E/T, effector/target.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4317872&req=5

fig04: Heparanase specificity and HLA-A2 restriction of cytotoxic T lymphocytes (CTL) generated from different heparanase vaccines in vitro (A) and ex vivo (B). The CTL generated from the HLA-A2-restricted nonapeptide HIVpol(476-484)(ILLEPVHGV) derived from HIV served as the negative peptide control (NP). (A) The cytotoxic activity of CTL induced by heparanase multi-epitope vaccines and single epitope peptides against MCF-7 (A[a, b]), MCF-7/Hpa (A[c, d]), HepG2 (A[e, f]) and HepG2/HLA-A2 cells (A[g,h]) in vitro. (B) The cytotoxic activity of CTL induced by heparanase multi-epitope vaccines and single epitope peptides against MCF-7 (B[a, b]), MCF-7/Hpa (B[c, d]), HepG2 (B[e, f]), and HepG2/HLA-A2 cells (B[g, h]) ex vivo. E/T, effector/target.
Mentions: To further confirm the heparanase specificity of the CTL in vitro and ex vivo, we used the HLA-A2-positive, heparanase-negative breast cancer cell line MCF-7 (Fig.4). MCF-7 cells were transfected with recombinant, replication-defective adenovirus (Ad-Hpa) encoding a full-length cDNA of human heparanase at a multiplicity of infection of 200 and cultured for 2 days in fresh DMEM medium containing 10% FCS. In our previous study, western blot analysis showed that heparanase protein could be detected in MCF-7/Hpa cells, whereas in MCF-7 cells the expression of heparanase is very weak.19 Heparanase-specific CTL were generated using peptide-pulsed DC from HLA-A2-positive PBMC and DC from the bone marrow of C57BL/6-Tg transgenic mice. After being stimulated three times, the killing effect of CTL induced by Hpa525 + 277 + 405 + 16 and Hpa8 + 310 + 315 + 363 and their corresponding single peptides Hpa525, Hpa277, Hpa405, Hpa16, Hpa8, Hpa310, Hpa315 and Hpa363 were determined at various E/T ratios using a 4-h 51Cr release assay. The results showed that these heparanase-specific CTL could lyse MCF-7/Hpa cells, whereas no obvious lysis of MCF-7 cells was detected even at the highest E/T ratio. Furthermore, compared with the corresponding single heparanase peptides, heparanase multi-epitope vaccines elicited more robust specific lysis of MCF-7/Hpa cells (Fig.4). These results clearly indicate that the CTL specifically targeted heparanase peptides that were presented in the context of HLA-A2 and the multi-epitope vaccines of human heparanase could elicit much more potent killing effects compared with their corresponding single peptides.

Bottom Line: Previous studies have indicated that heparanase (Hpa) might represent a candidate universal tumor-associated antigen.The results showed that multi-epitope vaccines Hpa525 + 277 + 405 + 16 and Hpa8 + 310 + 315 + 363 induced higher Hpa-specific lysis of various cancer cells from different tissues in a HLA-A2-restricted and heparanase-specific manner compared with the single epitope vaccines Hpa525, Hpa277, Hpa405, Hpa16, Hpa8, Hpa310, Hpa315 and Hpa363, both in vitro and ex vivo.Therefore, the present study provides theoretical evidence for the use of heparanase multi-epitope vaccines for clinical application.

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology, Xinqiao Hospital, Third Military Medical University, Chongqing, China; Department of Respiratory and Critical Care Medicine, The First Affiliated Hospital, Chongqing Medical University, Chongqing, China.

Show MeSH
Related in: MedlinePlus