Serological cloning of cancer/testis antigens expressed in prostate cancer using cDNA phage surface display.
Serological cloning of tumor-associated antigens (TAAs) using patient autoantibodies and tumor cDNA expression libraries (SEREX) has identified a wide array of tumor proteins eliciting B-cell responses in patients. However, alternative cloning strategies with the possibility of high throughput analysis of patient sera and tumor libraries may be of interest. We explored the pJuFo phage surface display system, allowing display of recombinant tumor proteins on the surface of M13 filamentous phage, for cloning of TAAs in prostate cancer (PC). Control experiments established that after a few rounds of selection on immobilized specific IgG, a high degree of enrichment of seroreactive clones was achieved. With an increasing number of selection rounds, a higher yield of positive clones was offset by an apparent loss of diversity in the repertoire of selected clones. Using autologous patient serum IgG in a combined biopanning and immunoscreening approach, we identified 13 different TAAs. Three of these (NY-ESO-1, Lage-1, and Xage-1) were known members of the cancer/testis family of TAAs, and one other protein had previously been isolated by SEREX in cancer types other than PC. Specific IgG responses against NY-ESO-1 were found in sera from 4/20 patients with hormone refractory PC, against Lage-1 in 3/20, and Xage-1 in 1/20. No reactivity against the remaining proteins was detected in other PC patients, and none of the TAAs reacted with serum from healthy subjects. The results demonstrate that phage surface display combined with postselection immunoscreening is suitable for cloning a diverse repertoire of TAAs from tumor tissue cDNA libraries. Furthermore, candidate TAAs for vaccine development of PC were identified.
Fosså
Alexander
A
Department of Immunology, Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway. alexanfo@labmed.uio.no
Alsøe
Lene
L
Crameri
Reto
R
Funderud
Steinar
S
Gaudernack
Gustav
G
Smeland
Erlend B
EB
D000951:Antigens, Neoplasm / Q000235:genetics* / Q000276:immunology; D000954:Antigens, Surface; D017104:Bacteriophage M13; D015153:Blotting, Western; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics*; D004797:Enzyme-Linked Immunosorbent Assay; D015723:Gene Library; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology; D008207:Lymphatic Metastasis; D008297:Male; D008565:Membrane Proteins / Q000276:immunology; D019151:Peptide Library; D011471:Prostatic Neoplasms / Q000235:genetics*; D011994:Recombinant Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism; D013737:Testis / Q000276:immunology*
D000951:Antigens, Neoplasm; D000954:Antigens, Surface; C104948:CTAG1B protein, human; C510162:CTAG2 protein, human; D018076:DNA, Complementary; D007074:Immunoglobulin G; D008565:Membrane Proteins; D019151:Peptide Library; D011994:Recombinant Proteins; C413656:XAGE1A protein, human