Expert Case Study: Probing Patient Responses to Cell Therapy for Safer, More Effective Treatments

In association with Dr. Theodore S. Nowicki, an assistant professor in the departments of Pediatric Hematology Oncology and Microbiology, Immunology and Molecular Genetics at University of California Los Angeles’ David Geffen School of Medicine, with a focus on T cell-based cancer immunotherapy.

All statements made by Dr. Nowicki are his own as an individual. The University of California Los Angeles does not endorse any products and was not involved in this particular program.

Making Cell Therapies Work Better for More People

Cell therapies are continuing to grow in importance, particularly for patients with intractable cancers who have failed all other treatments. Chimeric antigen receptor (CAR)-T cell therapies have been successful – even curative – for many of these patients, but researchers are seeking to broaden the impact for more patients, and to reduce the incidence of common adverse events.

“In the broadest sense, my lab’s focus is to understand what distinguishes responders from non-responders, and patients who do not experience toxicity from those who do,” said Dr. Theodore Nowicki, MD, PhD, a pediatric hematologist/oncologist and immunologist. He combines these disciplines in his research into CAR-T and T-cell receptor (TCR)-T cell therapies at University of California Los Angeles’ David Geffen School of Medicine. “We look at clinical samples, we look at clinical cohorts, but we also look at in vitro and animal models to try to understand the differences between groups – and then, of course, make more non-responders into responders, and reduce toxicity.”

Teasing Out the Causes of Neurotoxicity in CAR-T Patients

Dr. Nowicki’s group recently focused on immune effector cell-associated neurotoxicity syndrome (ICANS), a common and sometimes-fatal CAR-T complication. Clinical symptoms of ICANS are very similar to hypophosphatemia, caused when sudden spikes in metabolic demand for phosphorus deplete levels in the blood – a state seen frequently in patients with sepsis or refeeding syndrome.

To explore the connection, his team began a series of in vitro experiments demonstrating that CAR-T cell engagement with their target antigens led to extracellular phosphorus consumption. The researchers ran single-cell cytokine secretion assays to correlate the increase with the cells actually doing effector metabolism, using his lab’s IsoSpark. “It allowed us to quickly and robustly classify what was going on in the antigen-dependent effector metabolism of the CAR-T cells,” Nowicki said.

With this mechanistic evidence in hand, the group performed a retrospective analysis of patients treated with CAR-T cell therapy, and correlated drops in serum phosphorus levels with onset and severity of ICANS. The results were published in October in Cancer Immunology Research.

“The next step is actually identifying a causal relationship, which would be a profound story to tell,” said Nowicki. CAR-T cell adverse events like cytokine release syndrome can be treated with biologics, but ICANS has been a challenge. “If we do prove that we can actually mitigate or even prevent neurologic toxicity just by phosphorus supplementation, that is such an easy fix.”

IsoPlexis’ Single-Cell Cytokine Secretion Assays

“We were able to see cytokines that are created by CAR-T cells on a single-cell level, and also classify such a diverse array. That really allows us to get a much more holistic approach to classifying all of the cytokinetic polyfunctionality that these CAR-T cells are undergoing,” Nowicki added. “We could have done the experiments with single-cytokine assays, but obviously you might miss a lot.”

Nowicki was familiar with the IsoSpark and acquired one as he built his lab. “We wanted to be a really T cell biology-focused lab, and a cell therapy-focused lab. This platform made a lot of sense to feature heavily in a lot of the projects,” he said. “It allows us to assay a large number of analytes very quickly. But also, doing them in a single-cell level is vitally important, because with other assays, you can miss a lot if it’s just cytokines hanging out in the extracellular culture media.”

Getting the Most out of Clinical Samples

Single-cell proteomics is increasingly important for his lab’s work, including an ongoing trial with TCR-T cells, and comparing clinical samples of responders and non-responders. “Clinical samples are very finite, very precious, and once they’re gone, they’re gone,” said Nowicki. “Being able to get as much data as possible out of a single sample is a really important thing to plan towards. There are things that you really can only catch when you’re looking at a single-cell resolution of the data, and things that you would otherwise miss if you were looking at bulk data.”

Next Steps

For future experiments, Nowicki is considering additional tools that would expand his lab’s ability to look at signaling networks at the protein level with transcriptomics and functional proteomics, including Duomic, a solution in-development at IsoPlexis. “It’s exciting to see where the platform is going to go in the future as well.”

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