Interleukin-12 inhibits liver-specific drug-inducible systems in vivo

Abstract

Drug-inducible systems allow modulation of the duration and intensity of cytokine expression in liver immuno-based gene therapy protocols. However, the biological activity of the transgene may influence their function. We have analyzed the kinetics of interleukin-12 (IL-12) expression controlled by the doxycycline (Dox)-and the mifepristone (Mif)-dependent systems using two long-term expressing vectors directed to liver: a plasmid administered by hydrodynamic injection and a high-capacity adenoviral vector. Daily administration of Dox or Mif was associated with a progressive loss of inducibility and a decrease of murine IL-12 production. This inhibition occurred at the transcriptional level and was probably caused by an interferon (IFN)-g-mediated downmodulation of liverspecific promoters that control the expression of transactivators in these systems. Genome-wide expression microarrays studies revealed a parallel downregulation of liverspecific genes in mice overexpressing murine IL-12. However, a promoter naturally induced by IL-12 was also inhibited by this cytokine when placed in a plasmid vector. Interestingly, treatment with sodium butyrate, a class I/II histone deacetylase inhibitor, was able to rescue liverspecific promoter activity solely in the vector. We conclude that biologically active IL-12 can transiently inhibit the function of drug-inducible systems in non-integrative DNA vectors by reducing promoter activity, probably through IFN-g and protein deacetylation-dependent mechanisms.

Oxaliplatin in combination with liver-specific expression of interleukin 12 reduces the immunosuppressive microenvironment of tumours and eradicates metastatic colorectal cancer in mice

“…We had previously determined in our MC38 syngeneic tumour model that seric IL-12 <20 ng/ml has no antitumour effect, and levels >700 ng/ml are associated with toxicity 27. The two main obstacles to keeping to this therapeutic range are the variability in transduction among individuals and the progressive silencing of the drug-inducible system mediated by the immune system 28. We have focused on the favourable dose–response effect of the Mif-inducible system (figure 1A) to design a new induction protocol.…”

“…In contrast with a fixed amount of inducer (constant protocol), doubling the dose of Mif every 3 days contributes to the stabilisation of IL-12 levels for at least 10 days. Since downregulation of the system is due to a reversible inhibition of the transthyretin promoter,28 the induction protocols can be repeated after a resting period of at least 2 weeks in order to obtain several cycles of IL-12 expression (figure 2B). We have verified that the system is functional for more than 5 months, with a slow decrease in the intensity of expression in each cycle owing to the non-integrative nature of adenoviral vectors.…”

“…The possibility of controlling the intensity of IL-12 expression by adjusting the dose of inducer makes this approach safer and more clinically applicable. However, the function of drug-inducible expression systems is influenced by the immunostimulatory properties of IL-12 acting at the transcriptional level 28. This causes a progressive but reversible inhibition of transgene expression that might limit the therapeutic effect of this approach, especially in the case of pre-established liver metastases 29 30.…”

Inhibition of nuclear delivery of plasmid DNA and transcription by interferon γ: hurdles to be overcome for sustained gene therapy

“…[16][17][18][19][20][21][22][23] Although the mechanisms underlying these suppressive events have not fully been elucidated and would depend on the type of vectors and/or promoters, it is suggested that cytokines, including tumor necrosis factor (TNF)-a and IFN-g, are involved in the suppression. 20,24,25 Such cytokines are induced as a biological response to viral and non-viral vectors, including plasmid DNA/cationic liposome complexes or lipoplex.…”

Immunochemotherapy against colon cancer by gene transfer of interleukin-12 in combination with oxaliplatin

“…In addition, a step-wise increase of inducer is needed to cope with the transient silencing of liver-specific promoters caused by gamma-interferon. 8 We found that this individualized protocol allowed several cycles of IL-12 expression in the therapeutic range. 4,7 Using a syngeneic model of liver metastases in mice, we observed that a single cycle consisting on 10 daily inductions significantly extended the survival of animals and achieved eradication of hepatic tumors in 50% of them (Fig.…”