In an interview with Targeted Oncology, Katy Rezvani, MD, PhD, vice president and head, Institute for Cell Therapy Innovation and Discovery, Department of Institute for Cell Therapy Innovation and Discovery, Division of Cancer Medicine, The University of Texas MD Anderson Cancer Center, discusses the antitumor effectiveness of chimeric antigen receptor-natural killer (CAR-NK) cell therapy.
Watch the first part of Dr Rezvani’s interview.
Rezvani’s team employed a powerful and unbiased approach to identify the genetic levers that could make the NK cells resistant to the hostile conditions of the tumor microenvironment (TME).
To achieve this, they developed a strategy centered on a genome-wide CRISPR screen. This involved using a comprehensive CRISPR library that targets virtually every gene across the entire human genome. The core design principle was to ensure that only 1 gene was targeted and inactivated per individual NK cell. This created a vast, heterogeneous population of NK cells, each with a unique genetic modification.
They then subjected these genetically diverse NK cells to rigorous selective pressure, mimicking the challenges the cells would face in a solid tumor. The primary method involved a repeated challenge assay where the NK cells were continuously exposed to tumor cells. The fittest, most potent NK cells—those that successfully killed the tumor cells and continued to proliferate—would selectively grow out and dominate the culture.
By sequencing these surviving, dominant NK cells, Rezvani’s team could determine precisely which gene knockout conferred the survival and fitness advantage. This allowed them to identify specific genetic targets that, when inactivated, enhance NK cell killing and persistence in the presence of cancer.
In addition to the tumor challenge assay, Rezvani’s team exposed the NK cells to other key elements of the TME’s hostility. This was a critical step in isolating the regulators that govern TME resistance:
Hypoxia: Exposure to low oxygen levels.Acidity: Exposure to a low pH environment.Immunosuppressive Molecules: Direct exposure to potent inhibitory factors like TGF-β
The entire rationale behind this multi-pronged screening was the discovery of checkpoints or regulators that are critically involved in inducing suppression in NK cells under these diverse negative conditions. Isolating these key regulatory genes then provides a clear roadmap for the next-generation engineering of NK cell therapies.
The screen generated a wealth of data, confirming that a complex interplay of genetic factors governs NK cell resilience. Rezvani and her team found that some of the identified regulators had previously been implicated in CAR T cell function, such as MED12. However, they also identified other genes that appear to be more uniquely critical for NK cell biology, including ARIH2 and CCNC.
Furthermore, data revealed the potential for synergistic effects, showing that targeting a combination of genes—for instance, knocking out 2 specific genes simultaneously—was often more advantageous for NK cell fitness than targeting just 1.
Rezvani is highly optimistic that this comprehensive dataset will enable researchers to develop highly engineered NK cells that are significantly more effective at overcoming the immense hurdles posed by the TME. The ultimate goal is to translate these genetic insights into a robust cell therapy product that can effectively treat and ultimately cure solid tumors.
REFERENCE:Bierderstädt A, Basar R, Park JM, et al. Genome-wide CRISPR screens identify critical targets to enhance CAR-NK cell antitumor potency. Cancer Cell. 2025 Aug 18:S1535-6108(25)00327-7. doi: 10.1016/j.ccell.2025.07.021. Online ahead of print.