Bacillus Calmette-Guerin (BCG) induces superior anti-tumour responses by V delta 2(+) T cells compared with the aminobisphosphonate drug zoledronic acid

V delta 2(+) T cells can recognize malignantly transformed cells as well as those infected with mycobacteria. This cross-reactivity supports the idea of using mycobacteria to manipulate V delta 2(+) T cells in cancer immunotherapy. To date, therapeutic interventions using V delta 2(+) T cells in cancer have involved expanding these cells in or ex vivo using zoledronic acid (ZA). Here, we show that the mycobacterium Bacillus Calmette-Guerin (BCG) also causes V delta 2(+) T-cell expansion in vitro and that resulting V delta 2(+) cell populations are cytotoxic toward tumour cell lines. We show that both ZA and BCG-expanded V delta 2+ cells effectively killed both Daudi and THP-1 cells. THP-1 cell killing by both ZA and BCG-expanded V delta 2+ cells was enhanced by treatment of targets cells with ZA. Although no difference in cytotoxic activity between ZA- and BCG-expanded V delta 2+ cells was observed, BCG-expanded cells degranulated more and produced a more diverse range of cytokines upon tumour cell recognition compared to ZA-expanded cells. ZA-expanded V delta 2+ cells were shown to upregulate exhaustion marker CD57 to a greater extent than BCG-expanded V delta 2+ cells. Furthermore, ZA expansion was associated with upregulation of inhibitory markers PD-1 and TIM3 in a dose-dependent manner whereas PD-1 expression was not increased following expansion using BCG. Intradermal BCG vaccination of rhesus macaques caused in vivo expansion of V delta 2(+) cells. In combination with the aforementioned in vitro data, this finding suggests that BCG treatment could induce expansion of V delta 2(+) T cells with enhanced anti-tumour potential compared to ZA treatment and that either ZA or BCG could be used intratumourally as a means to potentiate stronger anti-tumour V delta 2(+) T-cell responses. Vd2+ gamma-delta T cells have been shown in vitroto elicit potent cytolytic responses toward tumour cells, suggesting the potential for these responses to be manipulated in cancer immunotherapy. We show that heat-killed Bacillus Calmette-Guerin (HKBCG) promotes the expansion of Vd2+ T cells both in vitroand in vivo. These HKBCG-expanded cells exhibited enhanced degranulation and cytokine production in tumour cells compared to cells expanded using bisphosphonates.