Cold Plasma with Immunomodulatory Properties Has Significant Anti-Lymphoma Activities in Vitro and In Vivo

In recent years, cancer immunotherapy has revolutionized cancer care. Remarkable clinical efficacy and durable responses to checkpoint blockade antibodies or to genetically engineered T-cells (CAR T cell) have been observed in patients with multiple cancers. However, not all cancer patients benefit from these therapies and as such novel immunotherapeutic approaches are needed. The cold atmospheric plasma (CAP) is a form of near room temperature ionized gas, which has shown a promising application in cancer therapy given its antitumor effects in vitro as well as in vivo. For the first time, we have shown that upon CAP treatment, the viability of the immune cells remained unaffected. Strikingly, we observed a significantly stronger immune activation of those CAP treated cells when compare with helium gas control. First, we have demonstrated that even without LPS stimulation, in vitro exposure of peritoneal elicited macrophages (PEM) to CAP for 15 or 30 seconds was sufficient to trigger the production of the pro-inflammatory cytokines IL-12 and IL-6. In addition, decreased production of anti-inflammatory cytokine IL-10 and diminished the expression of PD-L1 were observed in CAP-treated PEM as well. It indicated that CAP treatment may potentially facilitate PEMs as better activator of T cells. Second, in lieu of the stimulatory effects of CAP upon PEM, we asked whether CAP could enhance T-cell activation. So we isolated T cells from spleens of C57BL/6 mice and exposed these T cells to CAP followed by anti-CD3/CD28 stimulation. Our study shown that the production of IL-2 and IFN-g were significantly increased by T-cells treated with CAP as compared with helium gas control. Furthermore, to gain insights into effects of CAP upon immune responses in vivo, we isolated lymph nodes from OTII mice and directly exposed these LNs with CAP, helium gas control or left untreated. Then CD4+ T cells were further isolated from the LNs and cultured with macrophages in the presence or absence of OVA peptide for 48 hours, respectively. Surprisingly, CD4+ T cells isolated from CAP-treated LNs displayed enhanced function indicated by its increased production of IL-2 and IFN-g. More importantly, strong in vivo anti-tumor effects were observed when adoptively transfer CAP exposed T cells to lymphoma bearing animals.

Taken together, we have shown for the first time an elevated immune-stimulatory effect of CAP upon both APCs and T cells in vitro and in vivo. Our finding may potentially shed light of a novel therapeutic approach for future cancer immunotherapy.