Vγ9Vδ2 T cells - response to P. falciparum-derived phosphoantigens and potential for use in colon cancer immunotherapy 

Abstract: Vγ9Vδ2 T cell is the dominant circulating γδ T cell subset in humans, can expand massively upon malaria infection and are cytotoxic to cancer cells. Vγ9Vδ2 T cells are stimulated by phosphoantigens, primarily isoprenoid pyrophosphates like isopentenyl pyrophosphate and (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMBPP). Vγ9Vδ2 T cell from human blood were studied for proliferation, response to blood-stage malaria parasites and during colon cancer progression. Vγ9Vδ2 T cell proliferation was stimulated by media from P. falciparum-infected erythrocytes from all asexual blood stages - rings, trophozoites, schizonts and rupturing schizonts as well as sexual stage gametocytes assessed by the protocols we developed to obtain pure cultures of all stages. Further, we demonstrated that the molecules that stimulated the Vγ9Vδ2 T cell proliferation are phosphoantigens that are released from intact infected erythrocytes. This does not require schizont rupture.  Interestingly, the parasites consumed all the iron ion of hemoglobins during their development from the ring to the rupturing schizont stage. We found that an Anopheles gambiae immune cell line responds to HMBPP by activation of MAPK and PI3K signaling pathways. Moreover, transcription of dual oxidase and nitric oxide synthase was upregulated by addition of HMBPP in the midgut of Anopheles gambiae which increases cell tolerance to oxidative stress. A range of small isoprenoid pyrophosphates were found to stimulate proliferation of Vγ9Vδ2 T cells from PBMCs as was the isoprenoid monophosphate DMAP. However other isoprenoid monophosphates and alcohols did not. We found that cryopreserved unexpectedly increase the proliferation ability of HMBPP–stimulated PBMCs. To test the cytotoxicity of Vγ9Vδ2 T cells against adherent colon cancer cell lines, a flow cytometry-based assay was developed. Using the assay we found that proliferated Vγ9Vδ2 T cells are cytotoxcitic to various cancer cells and that HMBPP increases cytotoxicity towards adherent colon cancer cells. In a clinical study we found that Vγ9Vδ2 T cells could not always be proliferated from colon cancer patients and that the inflammatory homing receptor CXCR3 was expressed at higher levels in colon cancer patients than the control group. Moreover, at cancer stadium 4 a lower frequency of Vγ9Vδ2 T cells was more common than in the other groups.