Cancer vaccine strategies and studies of human thioredoxin reductase splice variants

Abstract

Cancer involves abnormal, uncontrolled proliferation of cells, and evasion from the immune system. Immune evasion can be caused by defects in one or more of the components of the major histocompatibility complex class I antigen machinery, e.g. the beta2-microglobulin (beta2m) molecule. Here, it was demonstrated that vaccination of mice with beta2m-deficient dendritic cells, prior to a challenge with a tumorigenic dose of beta2mfree tumor cells of syngeneic origin, protected the vaccinated mice from tumor development. Antitumor immune reactions depend upon the cytokine composition in the tumor microenvironment. An efficient antitumor immune response correlates with local high-level expression of the granulocyte-macrophage colony-stimulating factor (GM-CSF) cytokine. A heat-inducible DNA vector for amplified GM-CSF expression was herein constructed, which showed almost three-fold greater expression levels upon induction by hyperthermia than a prototypic CMV promoter construct, while both constructs had similar basal levels.

Thioredoxin reductase (TrxR) is an essential redox-active selenoenzyme important for human health and disease. For example, TrxR is involved in many steps of tumorigenesis, with TrxR expression often upregulated in tumors. Moreover, TrxR is known to be a target of platinum-based drugs, which are used in anticancer therapy. Data presented here show that in organotypic cochlear cultures, direct exposure of equimolar concentrations of the frequently ototoxic platinum-based drug cisplatin and the rarely ototoxic oxaliplatin both targeted cochlear TrxR and caused hair loss to similar degrees. These findings emphasize the importance of understanding pharmacodynamics (oxaliplatin does not reach the inner ear in vivo) and may add to improve therapeutic strategies in order to lower patient toxicity caused by platinumbased drugs.

TrxR1 is the most abundant TrxR isoenzyme, which is predominantly found in the cytosol. The gene encoding TrxR1 TXNRD1 harbors a complex genomic structure, leading to numerous splice variants. Peculiarly, the v3 splice variant encompasses a unique glutaredoxin domain, transcription of which is guided by an alternative promoter, which is located upstream of the TrxR1 core promoter. Expression of v3 was herein found in the developing human cochlea and the Leydig cells of the testis. Transcripts encoding v3 were detected in human heart, liver, spleen, ovary, kidney and pancreas, as well as several cancer cell lines. Several cell stressors (including starvation, hypoxia, etoposide, rapamycin, nocodazole) increased v3 promoter activity and v3 expression, while simultaneously repressing the TrxR1 core promoter activity and expression of the classical form of TrxR1. Translation of v3 was found to be IRESdependent. In particular, prolonged starvation induced expression of v3, dynamic formation of membrane protrusions to which endogenous v3 was localized and an increase in cell motility, all of which correlated in time. Recombinant overexpression of v3 in transfected cell lines induced a similar phenotype, with the dynamic formation of membrane protrusions. The data presented herein indicate that the formation of membrane protrusions and increased cell motility are linked with each other and with the induction of v3 expression.

In conclusion, cell-based and DNA vector-based cancer vaccine strategies were studied with the potential to be used in therapeutic cancer vaccine approaches. The splice variant v3 may possibly be targeted in anticancer therapy to interfere with cancer cell motility and ultimately thus, the formation of metastases.