Abstract
Nitrogen-containing bisphosphonates (N-BP), including zoledronic acid (ZOL) and alendronate (ALD), have been proposed as sensitisers in γδ T cell immunotherapy in pre-clinical and clinical studies. Therapeutic efficacy of N-BPs is hampered by their rapid renal excretion and high affinity for bone. Liposomal formulations of N-BP have been proposed to improve accumulation in solid tumours. Liposomal ALD (L-ALD) has been suggested as a suitable alternative to liposomal ZOL (L-ZOL), due to unexpected mice death experienced in pre-clinical studies with the latter. Only one study so far has proven the therapeutic efficacy of L-ALD, in combination with γδ T cell immunotherapy, after intraperitoneal administration of γδ T cell resulting in delayed growth of ovarian cancer in mice. This study aims to assess the in vitro efficacy of L-ALD, in combination with γδ T cell immunotherapy, in a range of cancerous cell lines, using L-ZOL as a comparator. The therapeutic efficacy was tested in a pseudo-metastatic lung mouse model, following intravenous injection of γδ T cell, L-ALD or the combination. In vivo biocompatibility and organ biodistribution studies of L-N-BPs were undertaken simultaneously. Higher concentrations of L-ALD (40–60μM) than L-ZOL (3–10μM) were required to produce a comparative reduction in cell viability in vitro, when used in combination with γδ T cells. Significant inhibition of tumour growth was observed after treatment with both L-ALD and γδ T cells in pseudo-metastatic lung melanoma tumour-bearing mice after tail vein injection of both treatments, suggesting that therapeutically relevant concentrations of L-ALD and γδ T cell could be achieved in the tumour sites, resulting in significant delay in tumour growth.
Highlights
Circulating gamma delta T cells represent 1–10% of all peripheral blood T lymphocytes [1] and predominantly express the Vγ9Vδ2 T cell receptor (TCR) [2]
Methods were developed to quantify the amount of zoledronic acid (ZOL) and ALD encapsulated into liposomes, as described in the supplementary information, using UV–Vis or HPLC (Fig. S2), copper sulphate-based UV spectroscopy method or ophthalaldehyde (OPA)-based fluorescence methods (Fig. S3)
As we have shown in a pseudo-metastatic lung melanoma tumour model (Fig. 7), significant inhibition of tumour growth was observed when Liposomal ALD (L-ALD) was used in combination with γδ T cells
Summary
Circulating gamma delta (γδ) T cells represent 1–10% of all peripheral blood T lymphocytes [1] and predominantly express the Vγ9Vδ2 T cell receptor (TCR) [2]. They recognize non-peptide phosphoantigens (PAgs) such as isopentyl pyrophosphate (IPP) [3]. Nitrogen-containing bisphosphonates (N-BPs), such as zoledronic acid (ZOL) or alendronate (ALD), are known to inhibit farnesyl pyrophosphate (FPP) synthase, an enzyme in the mevalonate pathway, in cancer cells, causing intracellular accumulation of PAgs [12].
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