Dovepress newly diagnosed cases.67,68 In tumors that overexpress EGFR, up to 75% of cases have rearrangements of the EGFR gene that lead to the expression of mutant forms.15,69–72 The most common EGFR mutation is EGFRvIII which results from an in-frame deletion of 267 amino acids in the extracellular domain (Figure 1).73 This receptor has constitutive tyrosine kinase activity and has important pro-oncogenic effects including enhancing proliferation, radio- and chemotherapeutic resistance, and migration, while inhibiting apoptosis.22–27,74,75 While 37%–86% of cells within EGFRvIII-expressing tumors express this receptor, EGFRvIII positive cells are able to secrete membrane-derived microvesicles with EGFRvIII which then merge with the plasma membranes of negative cells, conferring the same oncogenic advantages.76,77 As EGFRvIII contains an antigenic junction with a novel glycine residue and is not expressed on normal tissues, it is an effective target for immunotherapy.12 A variety of immunotherapies targeting EGFRvIII are currently under investigation including monoclonal antibodies, dendritic cell vaccination therapy, genetically modified T cells, and peptide vaccines. Various naked monoclonal antibodies have been shown to be specific for EGFRvIII and are able to elicit antitumor activity via Fc- and Fab-mediated activity.78–82 Monoclonal antibodies conjugated to toxins have also demonstrated signif icant cytotoxic activity against EGFRvIII-expressing tumors.83,84 Dendritic cell vaccination utilizes the antigen-presenting properties of dendritic cells to initiate antitumor responses. In vivo and human studies have demonstrated peptide-pulsed dendritic cells to induce EGFRvIII-specific cell-mediated immunity.85 More recently, genetically engineered T cells which express chimeric immune receptors have been shown to specifically lyse EGFRvIII-expressing gliomas cells in in vitro and in vivo studies.86 While these therapies are all attractive therapeutic modalities, peptide vaccines are one of the most studied and well understood immunotherapies. The most promising peptide vaccinate utilizes a peptide derived from the novel fusion junction amino acid sequence of EGFRvIII. This vaccine consists of PEPvII (H-Leu-Glu-Glu-Lys-Lys- Gln-Asn-Tyr-Val-Val-Thr-Asp-His-Cys-OH), an EGFRvIII- specific 14-mer peptide, and KLH.28 As it is able to activate humoral and cellular immunoreactivity, and has been shown to induce EGFRvIII-specific immune responses in preclinical and clinical studies.29,87–89 Detection of EGFRvIII mutations Due to the potential prognostic and therapeutic importance of EGFRvIII, its efficient detection is necessary for both Core Evidence 2012:7 Rindopepimut for the treatment of glioblastoma laboratory and clinical evaluation. As one of the most common methods of tissue preservation is formalin fixation plus paraffin embedding (FFPE), immunohistochemistry (IHC) is widely used as an accurate and reliable method for detecting EGFRvIII expression in stored samples.8,10,19,90,91 A variety of monoclonal and polyclonal antibodies have been developed which specifically recognize EGFRvIII and are commonly used for evaluating its expression in clinical studies. EGFRvIII can also be detected in fresh frozen and FFPE tissue using real-time reverse transcription-polymerase chain reaction (RT-PCR) and Southern blot assays.90,91 Preclinical studies Monoclonal antibodies targeted to EGFRvIII have shown to exhibit effective antitumor activity in in vitro and in vivo models. Treatment with unarmed murine IgG2a (Y10) and IgG1 (L8A4) monoclonal antibodies targeting EGFRvIII sig- nificantly inhibited tumor growth, though only treatment with IgG2a resulted in tumor-free survival after treatment was dis- continued.82 Though intraperitoneal therapy did not increase the median survival of mice with intracranial EGFRvIII B16 mela- nomas, single intratumoral injections of Y10 increased survival by 286%, with 26% of mice becoming long-term survivors (P , 0.001). The in vivo mechanism of action ofY10 was seen to be Fc receptor-dependent while being independent of T cells, NK cells, granulocytes, and complement. Immunotherapy using PEPvIII-KLH has also been effective against intracranial EGFRvIII-expressing tumors. Immunization of mice with DCs mixed with PEPvIII-KLH resulted in an approximately 600% increase in median survival, with 63% becoming long-term survivors.92 Also treatment with this mixture resulted in long-lasting protection as all rechallenged mice survived. Immunization with DCs mixed with PEPvIII-KLH produced EGFRvIII-specific IgG2a antibodies, correlating with antitumor activity. Another study demonstrated that a single dose of PEPvIII-KLH in Freund’s complete adjuvant increased median survival by 26% in mice with EGFRvIII-expressing intracerebral tumors (P = 0.0076).89 It was also seen that EGFRvIII expression was lost in 80% of relapsing tumors after vaccination, suggesting that EGFRvIII-negative escape is a potential mechanism of treatment failure (wild-type EGFR status was unknown in this study). Vaccination with PEPvIII-KLH resulted in humoral responses that could mediate antibody-dependent cellular cytotoxicity (ADCC), with the transfer of serum from vaccinated mice inducing antitumoral activity. The efficacy of PEPvIII-KLH was also seen to be dependent on CD8+ T and NK cells. submit your manuscript | www.dovepress.com Dovepress 97