Immunotherapy with dendritic cells in combination with cytotoxic chemotherapy may eliminate minimal disease burden by generating cytotoxic T lymphocytes (CTLs). Improving the cytosolic bioavailability of tumor-specific antigens to improve access to HLA class I molecules would result in better generation of cytotoxic T lymphocytes. Numerous cell-penetrating domains (CPDs) are known to carry linked heterologous antigens through the plasma membrane into the intracellular compartment.
In a study reported in JAMA Surgery, Batchu and colleagues assessed whether fusing melanoma antigen family A,3 (MAGE-A3), a tumor-specific cancer-testis antigen, with a cell-penetrating domain would increase the cytosolic bioavailability of MAGE-A3. MAGE-3 was amplified by polymerase chain reaction and cloned in frame with a cell-penetrating domain (YARKARRQARR) at the amino-terminal end and hexahistidine at the carboxy-terminal end to generate CPD–MAGE-A3.
Western blot analysis with MAGE-A3 antibodies recognized both MAGE-A3 and CPD–MAGE-A3 proteins, whereas analysis with cell-penetrating domain antibodies recognized only CPD–MAGE-A3. Purified CPD–MAGE-A3 exhibited improved dendritic cell membrane penetration vs MAGE-A3 alone. The finding on flow cytometry of high expression of unique dendritic cell markers (CD80, CD83, CD86, and HLA-DR) on dendritic cells was consistent with a mature dendritic cell phenotype, indicating that pulsing with CPD–MAGE-A3 did not alter the repertoire of cell-surface antigens required for T-cell activation.
The investigators concluded, “We have demonstrated for the first time, to our knowledge, that cloning and purification of MAGE-A3 with CPD enhances its cytosolic bioavailability in [dendritic cells] without altering cell-surface antigens, potentially making it a more potent therapeutic cancer vaccine compared with existing MAGE-A3 protein and peptide vaccines.” ■
Batchu RB, et al: JAMA Surg 149:451-457, 2014.