Vaccination strategy to target lysyl oxidase-like 4 in dendritic cell based immunotherapy for head and neck cancer

Abstract

Abstract. Overexpression of lysyl oxidase (LOX) is associated with the invasive potential of metastatic breast and head and neck cancer (HNC) cells and reduced metastasis-free and overall survival. Recently, we have demonstrated up-regulation of a new member of the LOX family, lysyl oxidase-like 4 (LOXL4), in invasive HNC revealed a significant correlation between LOXL4 expression and local lymph node metastases and higher tumour stages. The objective of this study was to examine whether cellular LOXL4 may provide an effective target for cell-meditated immunotherapy in invasive tumours associated with LOXL4 overexpression. As a feasibility study we expressed LOXL4 mRNA in immature dendritic cells derived from human peripheral blood mononuclear cells (PBMC). LOXL4 protein expression was ascertained using Western blotting and immunocytochemistry with polyclonal rabbit anti-LOXL4 antibody. The successfully transfected immature dendritic cells (DCs) were induced to mature with GM-CSF, IL-4, IL-1ß, TNF-α, IL-6, and PGE2, and then used to stimulate T cell enriched non-adherent fraction of PBMC. LOXL4 specific T cell stimulation induced cytotoxic T lymphocyte (CTL) response was monitored using IFN-γ secretion from the non-adherent PBMC fraction exposed to mature, LOXL4 transfected DCs acting as the antigen presenting target cells. LOXL4-DC stimulated T cells produced higher IFN-γ secretion compared to unstimulated T cells and T cells stimulated with untransfected DCs, in the presence of the pan-DR-epitope (PADRE). These initial results demonstrated the potential for LOXL4-transfected DCs to serve as efficient tumour vaccine and support their suitability as a vaccination strategy applicable to cancer patients with tumour specific up-regulation of LOXL4.

Lysyl oxidase like‐4 monoclonal antibody demonstrates therapeutic effect against head and neck squamous cell carcinoma cells and xenografts

“…We have found selective upregulation and amplification of the LOXL4 gene, overexpression of LOXL4 protein in primary and metastatic HNSCC cell lines and in patients with invasive HNSCC which correlated with local lymph node metastases versus primary tumor types and higher tumor stages . We also developed an affinity purified polyclonal antibody raised against the 85 kD LOXL4 antigen to demonstrate its expression in HNSCC tissues of different grading and staging, and studied its involvement in the structural organisation of squamous epithelia . So far, most of the described antibodies against HNSCC showed considerable cross reactivity with normal tissue or showed only reactivity with SCC of distinct sites of origin .…”

“…16,17 We also developed an affinity purified polyclonal antibody raised against the 85 kD LOXL4 antigen to demonstrate its expression in HNSCC tissues of different grading and staging, and studied its involvement in the structural organisation of squamous epithelia. 18 So far, most of the described antibodies against HNSCC [19][20][21][22][23][24][25] showed considerable cross reactivity with normal tissue or showed only reactivity with SCC of distinct sites of origin. 26 The first successful radioimmunotherapy results for HNSCC were shown with the E48 mAb in the early 1990 sec.…”

Artificial human antigen‐presenting cells are superior to dendritic cells at inducing cytotoxic T‐cell responses

“…Peptides and PLGA polymer HLA-A*0201 restricted peptides of Mart-1 [27][28][29][30][31][32][33][34][35] (Mart-1, AAGIGILTV), 16 Survivin 95-104 (SV95, ELTLGEFLKL) 17 were synthesized by GenScript Corporation (Piscataway, NJ). The purity and identity of the peptides were determined using HPLC analysis and mass spectrometry.…”

“…The in vitro controlled release of peptide was completed as we previously described with minor modification. 18 Mart-1 [27][28][29][30][31][32][33][34][35] or Survivin 95-104 peptide-loaded PLGA-NPs aliquots were suspended in PBS (pH 7Á4) containing 0Á01% sodium azide, and the suspension was incubated at 37°in a shaking incubator (60 r.p.m.). Triplicates were set up for each time-point.…”

Gene alterations in head and neck carcinomas and their role in promoting malignant behavior (Review)

“…However, the growth of existing metastases was not affected (67). In a feasibility study, we transfected cloned LOXL4-mRNA into immature dendritic cells and showed that cellular expression of LOXL4 could provide an effective target for cell mediated immunotherapy (68). Another important aspect of metastasis formation is the phenomenon of directional movement, which is well characterized for many cells, particularly leukocytes.…”