NZ28-induced inhibition of HSF1, SP1 and NF-κB triggers the loss of the natural killer cell-activating ligands MICA/B on human tumor cells.

Daniela Schilling,Sarah Konrad,Annett Kühnel,Fabian Tetzlaff,Gabriele Multhoff

doi:10.1007/s00262-015-1665-9

Abstract

The activity of natural killer (NK) cells is regulated by activating and inhibiting receptors, whereby the C-type lectin natural killer group 2D (NKG2D) receptor serves as the major activating receptor on NK cells which recognizes major histocompatibility class I chain-related proteins A and B (MICA/B). The MICA/B expression has been described to be regulated by the transcription factor heat shock factor 1 (HSF1). Inhibition of heat shock protein 90 (Hsp90) is known to induce the heat shock response via activation of HSF1 which is associated with tumor development, metastasis and therapy resistance and also with an increased susceptibility to NK cell-mediated lysis. Therefore, we compared the effects of Hsp90 inhibitor NVP-AUY922, HSF1 inhibitor NZ28 and HSF1 knockdown on the sensitivity of lung (H1339) and breast (MDA-MB-231, T47D) cancer cells to NK cell-mediated cytotoxicity and the expression of the NKG2D ligands MICA/B. Although NVP-AUY922 activates HSF1, neither the MICA/B surface density on tumor cells nor their susceptibility to NK cell-mediated lysis was affected. A single knockdown of HSF1 by shRNA decreased the surface expression of MICB but not that of MICA, and thereby, the NK cell-mediated lysis was only partially blocked. In contrast, NZ28 completely blocked the MICA/B membrane expression on tumor cells and thereby strongly inhibited the NK cell-mediated cytotoxicity. This effect might be explained by a simultaneous inhibition of the transcription factors HSF1, Sp1 and NF-κB by NZ28. These findings suggest that new anticancer therapeutics should be investigated with respect to their effects on the innate immune system.Electronic supplementary materialThe online version of this article (doi:10.1007/s00262-015-1665-9) contains supplementary material, which is available to authorized users.

Highlights

Inhibition of heat shock protein 90 (Hsp90) which promotes tumor cell survival and proliferation by stabilizing multiple oncogenic client proteins is a promising concept to overcome resistance of tumor cells to anticancer therapies
To investigate whether inhibitors of the stress response can modulate the sensitivity of human tumor cells to natural killer (NK) cellmediated cytotoxicity, heat shock factor 1 (HSF1) inhibitor NZ28 and/or Hsp90 inhibitor NVP-AUY922-treated tumor cells were used as target cells for IL-2 (100 IU/ml)-activated NK cells
NK cells were stimulated for 4 days with 100 U/ml IL-2. a The expression of CD94, CD56 and natural killer group 2D receptor (NKG2D) on NK cells was determined by FACS analysis

Summary

Introduction

Inhibition of heat shock protein 90 (Hsp90) which promotes tumor cell survival and proliferation by stabilizing multiple oncogenic client proteins is a promising concept to overcome resistance of tumor cells to anticancer therapies. An enhanced sensitivity of tumor cells to NK cell-mediated lysis has been attributed to an increased membrane expression density of the NKG2D ligands MICA and MICB [6,7,8,9]. A treatment with TNF-α induces binding of the transcription factor NF-κB to the MICA promoter and thereby causes an up-regulated expression of MICA [15]. Our data demonstrate that Hsp inhibition alters neither the MICA/B surface density nor the sensitivity of the tumor cells to NK cell-mediated lysis. A knockdown of HSF1 decreases the membrane expression of MICB but not that of MICA, whereas a treatment with NZ28 inhibits the expression of both, MICA and MICB on the surface of the investigated tumor cells. We could show that NZ28 inhibits HSF1 and other transcription factors such as NF-κB and Sp1 which are responsible for the expression of MICA/B

Materials and methods

Results

Discussion