PMID- 24671426
OWN - NLM
STAT- MEDLINE
DCOM- 20140915
LR  - 20140522
IS  - 2168-6262 (Electronic)
IS  - 2168-6254 (Linking)
VI  - 149
IP  - 5
DP  - 2014 May
TI  - MAGE-A3 with cell-penetrating domain as an efficient therapeutic cancer vaccine.
PG  - 451-7
LID - 10.1001/jamasurg.2013.4113 [doi]
AB  - IMPORTANCE: In conjunction with chemotherapy, immunotherapy with dendritic cells 
      (DCs) may eliminate minimal disease burden by generating cytotoxic T lymphocytes. 
      Enhanced cytosolic bioavailability of tumor-specific antigens improves access to 
      human leukocyte antigen (HLA) class I molecules for more efficient cytotoxic T 
      lymphocyte generation. Various cell-penetrating domains (CPDs) are known to ferry 
      covalently linked heterologous antigens to the intracellular compartment by 
      traversing the plasma membrane. OBJECTIVE: To determine whether generating 
      melanoma antigen family A, 3 (MAGE-A3), a tumor-specific cancer-testis antigen, 
      as a fusion protein with CPD will enhance the cytosolic bioavailability of 
      MAGE-A3. DESIGN: MAGE-A3 was amplified by polymerase chain reaction using 
      complementary DNA from renal tissue and cloned in frame with a CPD (YARKARRQARR) 
      at the amino-terminal end and hexahistidine at the carboxy-terminal end to 
      generate CPD-MAGE-A3 in a pQE-70 expression vector. Cultures were grown in 
      Escherichia coli BL21 Star (DE3-pLysS) cells followed by nickel-nitrilotriacetic 
      acid affinity purification of recombinant proteins. MAIN OUTCOMES AND MEASURES: 
      Measurement of DC membrane penetration of CPD-MAGE-A3 vs MAGE-A3 and 
      determination of the effect of CPD-MAGE-A3 pulsing on DC phenotypic expression of 
      cell-surface antigens. RESULTS: Media composition and 
      isopropyl-d-thiogalactosidase induction were optimized to achieve high levels of 
      protein expression followed by purification. Western blot analysis with MAGE-A3 
      antibodies recognized both MAGE-A3 and CPD-MAGE-A3 proteins, while CPD antibodies 
      recognized only CPD-MAGE-A3. Purified CPD-MAGE-A3 exhibited more efficient DC 
      membrane penetration than did MAGE-A3 alone as confirmed by immunofluorescence 
      analysis. High-level expression of several unique DC markers (CD80, CD83, CD86, 
      and HLA-DR) by flow cytometry was consistent with a mature DC phenotype, 
      indicating that pulsing with CPD-MAGE-A3 did not alter specific cell-surface 
      antigens required for T-cell activation. CONCLUSIONS AND RELEVANCE: We have 
      demonstrated for the first time, to our knowledge, that cloning and purification 
      of MAGE-A3 with CPD enhances its cytosolic bioavailability in DCs without 
      altering cell-surface antigens, potentially making it a more potent therapeutic 
      cancer vaccine compared with existing MAGE-A3 protein and peptide vaccines.
FAU - Batchu, Ramesh B
AU  - Batchu RB
AD  - Laboratory of Surgical Oncology and Developmental Therapeutics, Department of 
      Surgery, Wayne State University, Detroit, Michigan2John D. Dingell VA Medical 
      Center, Detroit, Michigan.
FAU - Gruzdyn, Oksana
AU  - Gruzdyn O
AD  - Laboratory of Surgical Oncology and Developmental Therapeutics, Department of 
      Surgery, Wayne State University, Detroit, Michigan2John D. Dingell VA Medical 
      Center, Detroit, Michigan.
FAU - Potti, Ravindra B
AU  - Potti RB
AD  - Department of Biotechnology, Srinidhi Institute of Science and Technology, 
      Hyderabad, India4Virocan Therapeutics Pvt Ltd, Guntur, India.
FAU - Weaver, Donald W
AU  - Weaver DW
AD  - Laboratory of Surgical Oncology and Developmental Therapeutics, Department of 
      Surgery, Wayne State University, Detroit, Michigan.
FAU - Gruber, Scott A
AU  - Gruber SA
AD  - Laboratory of Surgical Oncology and Developmental Therapeutics, Department of 
      Surgery, Wayne State University, Detroit, Michigan2John D. Dingell VA Medical 
      Center, Detroit, Michigan.
LA  - eng
PT  - Journal Article
PL  - United States
TA  - JAMA Surg
JT  - JAMA surgery
JID - 101589553
RN  - 0 (Antigens, Neoplasm)
RN  - 0 (Cancer Vaccines)
RN  - 0 (Cell-Penetrating Peptides)
RN  - 0 (MAGEA3 protein, human)
RN  - 0 (Neoplasm Proteins)
SB  - IM
CIN - JAMA Surg. 2014 May;149(5):457-8. PMID: 24671363
MH  - Antigens, Neoplasm/immunology/*therapeutic use
MH  - Biological Availability
MH  - Cancer Vaccines/immunology/*pharmacokinetics/*therapeutic use
MH  - Cell Membrane Permeability/*drug effects/immunology
MH  - Cell-Penetrating Peptides
MH  - Cloning, Molecular
MH  - Cytosol/*drug effects/*immunology
MH  - Dendritic Cells/*drug effects/*immunology
MH  - Drug Screening Assays, Antitumor
MH  - Humans
MH  - Neoplasm Proteins/immunology/*pharmacokinetics/*therapeutic use
MH  - T-Lymphocytes, Cytotoxic/drug effects/immunology
EDAT- 2014/03/29 06:00
MHDA- 2014/09/16 06:00
CRDT- 2014/03/28 06:00
PHST- 2014/03/28 06:00 [entrez]
PHST- 2014/03/29 06:00 [pubmed]
PHST- 2014/09/16 06:00 [medline]
AID - 1850879 [pii]
AID - 10.1001/jamasurg.2013.4113 [doi]
PST - ppublish
SO  - JAMA Surg. 2014 May;149(5):451-7. doi: 10.1001/jamasurg.2013.4113.