Induction of anti-EGFR immune response with mimotopes identified from a phage display peptide library by panitumumab

Abstract

The epidermal growth factor receptor (EGFR) is overexpressed in several epithelial tumors. Anti-EGFR humanized monoclonal antibodies, cetuximab and panitumumab, in combination with chemotherapy have improved the prognosis for patients with wild-type RAS tumors. To identify mimotopes of EGFR and develop mimotope-based EGFR vaccines, we screened a phage display peptide library with panitumumab. Two EGFR mimotopes P19 and P26, which could be recognized by panitumumab specifically, were isolated. To enhance the immune responses, we generated recombinant proteins of P19 or P26 fused to a heat-shock cognate protein 70 (Hsc70), and evaluated the efficacy of Hsc70-P19 and Hsc70-P26 as vaccines in vivo. Immunization with Hsc70-P19 or Hsc70-P26 fusion protein stimulated the immune system to produce specific antibodies against peptides as well as EGFR. Moreover, antibodies elicited against mimotopes could induce antibody-dependent cellular cytotoxicity (ADCC), complement-dependent cytotoxicity (CDC), and inhibit the proliferation of EGFR-overexpressing A431 cells. Treatment with Hsc70-P19 and Hsc70-P26 significantly reduced tumor growth in BALB/c transplantable lung cancer models. Although there was no sequence homology between the phage-derived peptides and EGFR by alignments, both peptides mimic the conformational structure of EGFR binding to panitumumab. In conclusion, the mimotopes we identified from phage display peptide library could be promising candidate vaccines for active anti-EGFR immunotherapy against cancers.

CENPH Inhibits Rapamycin Sensitivity by Regulating GOLPH3-dependent mTOR Signaling Pathway in Colorectal Cancer

“…The MTT assay was performed to determine the cell proliferation and cell sensitivity to rapamycin as previously described 26 , 27 . Briefly, cells (0.5×10 4 cells/well) transfected with plasmids or siRNAs were seeded in 96-well plates and incubated overnight.…”

“…The recombinant His-CENPH and GST-GOLPH3 protein were expressed and purified according to manufacturer's instructions (Amersham). GST pull-down and His-tag pull-down assays were performed as described previously 27 , 28 . Briefly, E. coli cell lysates containing GST and His-tag fusion protein were prepared in lysis buffer (1% Triton X-100, 1 mmol/L PMSF, and 1 mg/mL lysozyme in PBS) followed by sonication.…”

Phage-Displayed Peptides for Targeting Tyrosine Kinase Membrane Receptors in Cancer Therapy

“…The recombinant fusion protein of P19/P26/heat shock-related protein (HSP) 70, used for immunization, induced the production of specific antibodies against the peptides and EGFR. This allowed consideration of these peptides as mimotopes suitable for vaccines for anti-EGFR immunotherapy in tumors [ 113 ]. HER2 is a 185-kDa transmembrane glycoprotein with an extracellular domain of 105 kDa (p105HER2 ECD).…”

Ligand-Targeted Delivery of Photosensitizers for Cancer Treatment

“…In situ phage display studies include what was reported by Li et al over a decade ago and allowed the discovery of the GE11 peptide (YHWYGYTPQNVI) [ 39 ], still largely used to target EGFR [ 118 , 119 , 120 ]. The EGFR mimotopes, P26 (VPGWSQAFMALA) and P19 (DTDWVRMRDSAR), were also recently identified [ 121 ]. Other examples include the LS-7 (LQNAPRS) peptide which targets the CSC-associated marker CD133 [ 122 ] and the new nanobody VUN100 for targeting the G-protein coupled receptor homolog US28 that is found to be overexpressed in glioblastoma [ 123 ].…”