By making NK cells insensitive to tumor-secreted TGF-β, scientists have improved their efficacy against this deadly hepatocellular carcinoma (HCC) [1].
Liver cancer and TGF-β
Cancer cells, which are supposed to be vulnerable to the immune system, develop various defensive mechanisms to avoid detection and decrease immune cells’ fitness and viability. In solid tumors, the dense tumor microenvironment (TME) has a particularly strong dampening effect on immune activity.
HCC is the most widespread form of liver cancer. It is also very deadly, with a 5-year survival rate of around 20%. Many therapies have been tried and failed against HCC.
HCC is also one of the cancers that is characterized by overexpression of transforming growth factor beta (TGF-β). As its name hints at, TGF-β drives tumor growth, but it also interferes with the activity of immune cells [2], such as natural killer (NK) cells, which are part of the innate immune system.
Natural born killers, improved
In this new study, scientists from the University of California – San Diego combined several cutting-edge techniques to try and overcome this problem. To begin with, they used cellular reprogramming to manufacture brand-new NK cells. This involves producing induced pluripotent stem cells (iPSCs) from differentiated cells and their further re-differentiating them into NK cells.
However, there was a twist: in those iPSCs, genetic changes were introduced to make them resistant to TGF-β, either by knocking out TGF-β receptor 2 (TGFBR2) or by expressing its double-negative version, which also blocks TGF-β signaling. The researchers assumed that such cells would be less susceptible to the inhibitory effect of TGF-β secreted by HCC cells.
Their gamble seemed to pay off: the resulting NK cells showed superior cytotoxicity in an in vitro model of HCC, compared to wild type NK cells. They also retained functional activity when pre-treated with TGF-β, unlike wild type NKs. The only problem was that iPSCs lacking functional TGF-β receptors reproduced slower (since TGF-β boosts growth).
The researchers tried to further improve the efficacy of their modified NK cells by arming them with chimeric antigen receptors (CARs) that are routinely used against HCC. CARs are usually associated with T cells, but they can be introduced to other types of immune cells to direct them towards antigen-expressing cancer cells.
NK cells with TGFB2R knocked out and without CAR expression killed HCC cells better than CAR-expressing wild type NK cells. There was some synergy between CARs and TGFB2R knockout, albeit not a strong one. Importantly, when stressed with TGF-β, only TGFB2R-knockout cells, with or without CARs, retained their cytotoxicity, while CAR-only cells experienced quick loss of function.
Longer survival in vivo
The researchers observed similar results in a mouse model of HCC: TGFB2R knockout seemed to contribute the most toward NK’s anti-cancer activity, with or without CARs, markedly improving survival. NKs that only carried CARs were much less effective.
“These studies demonstrate that it is crucial to block transforming growth factor beta — at least for NK cells, but I also think it’s true for CAR T cells,” said Dan Kaufman, the lead author of this study. “If you unleash NK cells by blocking this inhibitory pathway, they should kill cancer quite nicely. Anyone developing such therapies for solid tumors should be working to inhibit transforming growth factor beta activity to improve cancer-killing and attain effective anti-tumor activity.”
NK cells are generally easier to mass-produce from iPSCs than T cells because the latter require more intricate differentiation protocols and thymus-like conditions to mature. Genetic modifications are also easier to perform in NK cells. NK cells require less personalization and can be used in donor-derived (allogeneic) settings more readily than T cells. As part of the innate immune system, NK cells have a broad range of activity against tumor cells without requiring prior sensitization to specific antigens. This all adds to NK cells’ promise in future anti-cancer therapies [3].
These studies demonstrate that even with NK cells engineered to express tumor-specific CARs, the immunosuppressive activity of TGF-β prevented effective anti-tumor activity if TGF-β activity is not inhibited. To disrupt TGF-β signaling, we utilized 2 different strategies via either deletion of TGFBR2 or over expression of the DN form of TGFBR2 in iPSC-derived NK cells. Both TGFBR2-KO and TGFBR2-DN NK cells demonstrated effective anti-HCC activity even without CAR expression, while CAR-NK cells without inhibition of TGF-β activity had little anti-HCC activity.
Literature
[1] Thangaraj, J. L., Coffey, M., Lopez, E., & Kaufman, D. S. (2024). Disruption of TGF-β signaling pathway is required to mediate effective killing of hepatocellular carcinoma by human iPSC-derived NK cells. Cell Stem Cell.
[2] Baba, A. B., Rah, B., Bhat, G. R., Mushtaq, I., Parveen, S., Hassan, R., … & Afroze, D. (2022). Transforming growth factor-beta (TGF-β) signaling in cancer-A betrayal within. Frontiers in pharmacology, 13, 791272.
[3] Goldenson, B. H., Hor, P., & Kaufman, D. S. (2022). iPSC-derived natural killer cell therapies-expansion and targeting. Frontiers in immunology, 13, 841107.
