Accumulating clinical evidences over recent years support the notion that the immune system can cure cancers. The potential of T cell therapy against cancer cells is documented by the persistent clinical responses observed after transfer of tumor specific T cells in some cancer patients. However, tumor-associated antigens are often self-antigens, and thus high-avidity tumor-specific lymphocytes are often deleted or anergized to prevent detrimental autoimmunity. This hurdle can be overcome by the transfer of high-avidity T cell receptor (TCR) genes isolated from rare tumor-specific lymphocytes into polyclonal T cells. Initial clinical studies have demonstrated the feasibility of TCR gene transfer, but the therapeutic results have been thus far suboptimal. This likely reflects limitations of current gene transfer approaches, which may fail to establish stable high-level TCR expression because the exogenous tumor-specific TCR α and β chains compete with endogenous TCR chains for surface expression. This problem is further aggravated by the potential for inappropriate pairing of exogenous and endogenous TCR chains, which leads to the assembly of novel TCRs with unpredictable, and possibly auto-reactive, specificities. Finally, the presence of endogenous TCR limits the use of TCR gene transfer in the allogeneic setting due to the risk of graft-versus-host disease (GvHD).
To overcome these limitations, in collaboration with the group of Chiara Bonini, San Raffaele Scientific Institute, we developed the first strategy based on engineered nucleases to edit T cell specificity at the DNA level. Our approach combines somatic knockout of the endogenous TCR genes by transient exposure to α and β chain specific Nucleases, with the introduction of the desired tumor-specific TCR by lentiviral vector gene transfer. The resulting ‘TCR-edited’ lymphocytes showed enhanced tumor killing activity with sharply reduced non-specific alloreactivity, as compared to matched cells undergoing conventional TCR gene transfer (Provasi*, Genovese* et al., Nature Medicine 2012). This work was the first proof that gene editing can be used to genetically re-write the endogenous antigen specificity of cytotoxic T cells and enable the feasibility of a safe allogeneic T cell transplantation, thus providing the basis for several other studies in the rapidly expanding cancer immunotherapy field, some of which already entered clinical testing.
To simplify the TCR gene editing procedure in view of clinical development, we developed the TCR single editing approach which enables a rapid generation of highly performing tumor specific T cells (Mastaglio, Genovese et al., Blood 2017). This innovative approach is now widely used in the immunotherapy field for generating allo-compatible T cells or to express CAR genes under the control of endogenous TCR promoter.
To simplify the TCR gene editing procedure in view of clinical development, we developed the TCR single editing approach which enables a rapid generation of highly performing tumor specific T cells (Mastaglio, Genovese et al., Blood 2017). This innovative approach is now widely used in the immunotherapy field for generating allo-compatible T cells or to express CAR genes under the control of endogenous TCR promoter.
Chimeric antigen receptor T cells (CAR-T), bispecific and toxin-conjugated antibodies represent emerging approaches that can overcome cancer immune-evasion and dampening of alloreactive immune responses. Ideally, the target antigen should be strictly tumor specific, to avoid toxicity to the healthy tissues, and essential for tumor survival, to reduce the occurrence of antigen-loss variants. Yet, despite great efforts in advancing immunophenotype profiling and high throughput genetic screening of cancer cells, the identification of targets remains a major hurdle to the full realization of the potential of these approaches. Moreover, although CAR T cell have demonstrated highly compelling efficacy in patients with CD19+ hematological malignancies, comparable therapeutic activity of CAR T cells has not yet been achieved in solid tumors, mainly because of difficulties of engineered T cells to circumvent the immunosuppressive tumor microenvironment. Thus innovative approaches and suitable models have to be exploited to broad the applicability of these revolutionary technologies for the treatment of highly aggressive malignancies.