In a new approach to CAR T cell engineering, Smith et al. skip the petri dish and head straight to the mouse. Using a stable, easy-to-manufacture nanoparticle gene delivery system, circulating T cells can be quickly programmed in vivo with CD19-targeted CARs.
The nanoparticles were designed to contain two co-encapsulated DNA plasmids, one encoding a 194-1BBz CAR for leukaemia treatment, and the second a hyperactive iPB7 transposase to achieve integration and persistent CAR expression in actively dividing T cells. The plasmids were packaged with a PBAE polymer functionalized with peptides containing microtubule-associated sequences and nuclear localization signals to facilitate nuclear import of their genetic cargo. The nanoparticles were specifically targeted to T cells by encapsulating them with conjugates of polyglutamic acid and anti-CD3e antibody fragments.
When administered in vitro or intravenously into immunocompetent B-ALL bearing mice, nanoparticles were selectively taken up by T cells (particularly naive T cells, the predominant T cell phenotype in peripheral blood) within two hours, and CARs were expressed on the treated cells’ surfaces in as little as 30 hours. Following stable somatic integration of the CAR gene and a period of rapid proliferation, which was dependent on their interaction with tumor antigens, the functionality and antitumor efficacy of the nanoparticle-transduced CAR T cells were comparable to that of conventional CAR T cells transduced with a lentiviral vector. Notably, many nanoparticle-programmed effector T cells eventually acquired a central memory phenotype.
Some off-target nanoparticle uptake by phagocytes was observed, but the transgenes were not strongly expressed nor did the phagocytes rapidly proliferate. No significant toxicities were observed, indicating that this method of systemic CAR gene delivery is likely to be safe.
If this method proves safe and effective in clinical trials, it would bypass the need for patient lymphodepletion, as well as the ex-vivo processes of traditional CAR T cell production, which could make CAR T cell treatment easier, cheaper, and more broadly applicable. Additionally, nanoparticles can be stored in lyophilized form, which would allow them to be manufactured off site, stored, and shipped worldwide. One caveat to note is that nanocarrier-transduced CAR T cells will likely face the same hurdles as traditional CAR T cells when it comes to tackling the challenge of accessing and eliminating solid tumors.
by Lauren Hitchings
