Dual Targeting of Prostate Cancer with an off-the-shelf CAR NK-92 cells

Prostate cancer is the second most common malignancy in men globally and the fourth most common cancer worldwide, according to 2022 data. This disease is highly heterogeneous and can be influenced by a combination of genetic and environmental risk factors. Prostate cancer treatment poses a major clinical challenge due to its high heterogeneity, frequent recurrence and development of treatment resistance. Immunotherapy has recently emerged as a promising therapeutic strategy in the oncology field. Unlike traditional treatments, which often lack specificity and are often toxic, immunotherapy seek to potentiate the natural antitumor activity of the immune system, thereby offering greater specificity and reduced off-target effects. Among immunotherapeutic approaches, adoptive cell therapy (ACT) has shown remarkable potential for a range of malignancies. ACT is based on the infusion of immune cells (usually derived from the patient) that are engineered to recognise and destroy tumour cells. The most common ACT used in clinical practice is CAR-T cell therapy, particularly successful for haematological malignancies. Nevertheless, this therapy is often limited by challenges such as graft-versus-host-disease, laborious and time-consuming manufacturing, and high production costs. To overcome these limitations, CAR engineered natural killer (NK) cells have emerged as an attractive alternative to CAR-T cell therapy. Particularly, the NK92 cell line offers the unique possibility to generate an “off-the-shelf” therapeutic product. This immortalised cell line, firstly isolated from a patient with non-Hodgkin lymphoma, allows reproducible and repeated clinical applications without the need for cost-intensive cell purification methods. The goal of this project is to engineer NK92 cells with two distinct chimeric antigen receptors (CARs) targeting two antigens that are consistently overexpressed in prostate cancer: hPSMA and hPSCA. The rationale behind this dual-targeting strategy is to reduce the risk of tumour escape, a mechanism by which malignant cells downregulate a specific antigen to evade immune surveillance. By simultaneously addressing multiple antigens, the risk of tumour adaptation and resistance may be reduced, thereby improving the efficacy of immunotherapeutic interventions. In addition, this strategy allows targeting of two clinically relevant antigens, ensuring applicability to tumors expressing either one or both of them. In this work, we demonstrated that Dual CAR-engineered NK92 cells display high and specific recognition not only of prostate cancer cell lines expressing a single target antigen (either hPSMA or hPSCA), but also of those co-expressing both antigens, thereby confirming their versatility potential to overcome tumour heterogeneity. In conclusion, the preliminary data obtained with this dual-targeting approach suggest a clear advantage over NK92 cells engineered with a single CAR, which are limited to recognizing only one antigen and are therefore more susceptible to tumour escape mechanisms. Furthermore, by simultaneously targeting two clinically relevant antigens, this strategy enables a broader and potentially more effective antitumor response, even when the dominant antigen expression varies among patients. While these findings are promising, additional studies are needed to corroboratethem both in vitro and in vivo.