Heat shock protein derived from a non-autologous tumour can be used as an anti-tumour vaccine

Abstract

Antigenic cross-reactivity between certain tumours has allowed the development of more widely applicable, major histocompatibility complex-disparate (allogeneic) whole-cell vaccines. This principle should also allow heat shock proteins (hsp) derived from certain tumours (and carrying cross-reactive antigens) to be used as vaccines to generate anti-tumour immunity in a range of cancer patients. Here, hsp70 derived from gp70-antigen+ B16 melanoma generated cytotoxic-T-lymphocyte-mediated immune protection in BALB/c mice against challenge with gp70-antigen+ CT26 colorectal tumour cells. Using ovalbumin as a model tumour antigen, it is shown that hsp70 enhances peptide re-presentation by dendritic cells via class I over equimolar whole ovalbumin antigen. However, while transfection of tumour cells with inducible hsp70 increases hsp yield from tumours, it does not enhance antigen recognition via purified hsp70 nor via whole cells or their lysate.

Figure 1

Expression by tumours of gp70. Tumour lines were examined by RT-PCR for expression of gp70, with GAPDH used as a control. 0 = no DNA added.

Figure 2

Purification of hsp70 from tumours. The hsp70 was extracted from tumour cells and examined by SDS–PAGE and Western blotting. Lane 1 shows a typical SDS–PAGE of hsp70 extracted from B16. Lanes 2 and 3 show Western blots of hsp70 extracted from B16-OVA and B16-OVA-hsp70, respectively.

Figure 3

Immunity following vaccination. BALB/c mice (n = 4 per group) were vaccinated or were mock-vaccinated (PBS), and were subsequently challenged with a tumorigenic dose (2 × 10⁵) of live CT26 cells (except d); % tumour-free survival is shown. Significance of protection: with (a) CT26 cells P = 0·064, (b) B16 cells P = 0·029, (c) hsp70 derived from B16 P = 0·031, (d) hsp70 derived from B16 and challenged with RENCA P = 1. (e) CT26 tumour volume on day 17 post-challenge, following vaccination with hsp70 from B16 compared to hsp70 from K1735 P = 0·02*. (f) Vaccination with hsp70 elicited similar CTL activity (% specific lysis at 50 : 1 effector to CT26 target ratio) to vaccination with CT26 cells.

Figure 4

Antigen presentation to T cells by DC can be mediated via purified hsp70, but not direct presentation by tumour cells. (a) The sensitivity of the detection of OVA peptide SIINFEKL by B3Z T-cell hybridoma on DC2.4 was determined, and compared to presentation by processing whole OVA protein. A405 gives the β-galactosidase activity in T cells which is linked to their interleukin-2 expression/activation. (b) The hsp70 derived from B16-OVA or B16-OVA transfected with inducible hsp70 (B16-OVA-hsp) elicited recognition by B3Z. Likewise (c) B16-OVA and B16-OVA-hsp whole cells were recognized equally by B3Z T-cell hybridoma.