## 4 Preclinical and Clinical Experience: The Lesson Learned  
\(\gamma\delta\) T cells have been expanded ex vivo and studied in pre-clinical cancer models by numerous groups and their cytotoxicity was demonstrated against a variety of cancer cells derived from breast tumor, lung carcinoma, and liver cancer [140]. A study by [PERSON] et al. showed that \(\gamma\delta\) T cells are capable of recognizing and lysing prostate cancer cells via innate mechanisms independent of MHC [141]. Due to their high frequency in peripheral blood and easy expansion, V\(\gamma\)9V82 subset of \(\gamma\delta\) T cells have been predominantly studied in vitro and in xenograft mouse models of a range of tumors. In general, the best treatment outcomes were achieved when V\(\gamma\)9V82 cells were expanded ex vivo prior to infusion, when \(\gamma\delta\) T cell infusion was performed at the same time as tumor cell implantation, or at early timepoints such as once tumors were first palpable (\(<\)100 mm\({}^{3}\) volume), and when the repeated administration of pAg or N-BP drugs plus cytokine (typically IL-2) was implemented. A handful of clinical trials using \(\gamma\delta\) T cells for cancer treatment have been performed and several are in progress. Immunotherapies based on V\(\gamma\)9V82 T cells have been examined for hematological malignancies (NCT02656147), head and neck cancer [142], hepatocellular carcinoma (NCT00562666), renal carcinoma [70; 77], mammary carcinoma [143], prostate cancer [66], neuroblastoma [144], and lung cancer [145; 146] among others. These clinical studies confirmed the clinical benefit of approaches aiming at activating the anti-tumor cytotoxicity of V\(\gamma\)9V82 T cells. Notably, they revealed that adoptive transfer of \(\gamma\delta\) T cells is safe and feasible [147]. In most \(\gamma\delta\) T cell's clinical trials, objective responses were observed but the rates of complete remissions were low and long-term disease-free survival data were minimal. The details of different approaches used in \(\gamma\delta\) T cells clinical trials are beyond the scope of this review, but the lessons we learned from them are briefly described. The overall analysis of the clinical studies shows the safety profile of \(\gamma\delta\) T cell therapies. However, the efficacy was highly variable. Additionally, it is difficult to compare the efficacy since there is immense variation in the utilized strategies including the different protocols used to expand \(\gamma\delta\) T cells (ex vivo or in vivo) or the delivery method (e.g., ZOL with or without IL-2 or other cytokines, \(\gamma\delta\) T cells alone, or in combination with stimulating drugs). The immunotherapies using pAgs or N-BPs and IL-2 have been successful in increasing the number of circulating V\(\gamma\)9V\(\delta\)2 T cells; nevertheless, there is limited evidence of \(\gamma\delta\) T cells infiltration into the target tissue. A reduction in the response of circulating V\(\gamma\)9V\(\delta\)2 T cells to the pAgs injection have been observed which may be due to activation-induced anergy [148]. This anergy may have occurred after \(\gamma\delta\) T cell exposure to the inadequate activatory signals and suboptimal pAgs concentration; although, repeated stimulation of V\(\gamma\)9V\(\delta\)2 T cells by pAgs may also cause terminal differentiation and exhaustion which impedes \(\gamma\delta\) T cell function [149]. The low potency of pAg-based clinical trials may also be attributed to the poor systemic availability of pAg due to their rapid clearance from plasma. Researchers have attempted to devise new delivering strategies to increase the treatment potency. Local administration of pAgs for eliciting a potent tumor immunity via intratumoral, peritumoral, and intranodal injections have already been tested for various types of cancers. Intranodal injection of pAgs near a tumor mass which allows for activation of \(\gamma\delta\) T cells outside of the immunosuppressive tumor microenvironment, may improve the treatment efficacy [18].