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IsoPlexis Single-Cell Platform Links T Cell Functionality to Patient Response to CAR T Therapy


NEW YORK (GenomeWeb) – A team led by researchers from Kite Pharma, Isoplexis, and the National Institutes of Health has found that the polyfunctionality of T cells introduced to patients undergoing chimeric antigen receptor (CAR) T cell therapy for non-Hodgkin lymphoma correlates with treatment response.

Detailed in a study published this week in Blood, the findings indicate the importance of polyfunctional T cells in determining the efficacy of CAR T therapy and the usefulness of single-cell cytokine analysis for assessing this polyfunctionality, said Sean Mackay, the CEO and co-founder of IsoPlexis, whose IsoCode single-cell analysis platform was used in the research.

Developed in the lab of Rong Fan, associate professor of biomedical engineering at Yale University and an author on the Blood paper, the IsoCode platform uses microchips featuring arrays of thousands of microchambers that isolate individual cells from samples of interest. These chambers are then sealed with a slide patterned with groups of antibodies in a number of different spatially isolated lines. This allows the researchers to identify proteins based on the color of fluorescence produced upon binding and the location on the slide where the binding event occurs. In this way, they can multiplex well beyond the levels allowed by fluorescence readout alone.

The technology underpinning the system was originally developed in the lab of California Institute of Technology researcher James Heath, where Fan was a postdoc. Fan, Heath, and their collaborators have demonstrated the platform's ability to measure a variety of proteins at the single-cell level, including phosphoproteins and various immune factors. IsoPlexis's lead product is a 32-cytokine panel, which it used in the Blood study and which it is promoting as a tool for helping to optimize adoptive cell transfer (ACT) cancer therapies.

One of the main aspects of such therapies the company and its collaborators are exploring is the role of multifunctional T cells — defined as T cells expressing multiple cytokines and chemokines — in ACT treatments. Past studies in infectious disease have found that multifunctional T cells are more effective at controlling viral infections, which suggests that a similar dynamic could be at play in cancer immunotherapies.

The results of the Blood study would seem to bear this out, as the researchers found that T cell functionality was linked to response of lymphoma patients to CAR T therapy, with the median polyfunctionality strength index (percentage of polyfunctional cells, multiplied by the mean fluorescence intensity (MFI) of the proteins secreted by those cells) scoring twice as high in responders versus non-responders.

IsoPlexis and its collaborators had previously demonstrated the IsoCode platform's ability to assess levels of T cell polyfunctionality, but McKay said that this was the first time they had tied these measurements to actual patient clinical response to ACT.

Also of note, he said, was the fact that other measurements of the infused T cells, including measurement of phenotypic cell surface markers by flow cytometry and bulk measurements of cytokine and chemokine secretions were not associated with clinical outcomes, suggesting, he said, the importance of making these measurements at the single-cell level.

"I think that supports that original thesis that underlies our company and that is being explored alternatively in single-cell genomics, that retaining a large degree of information while also being able to profile individual cells and capture their heterogeneity is going to be very meaningful for clinical work down the road," McKay said.

In the study, the researchers looked at a cohort of 22 lymphoma patients, evaluating CAR T products from 20 of them. Using the IsoCode system, they measured levels of 32 cytokines at the single-cell level in samples of roughly 2,000 cells per patient.

Mackay said the company has found that sampling around 2,000 cells provides a reproducible measurement "of the full spectrum of heterogeneous functional cell responses that we would expect to see across all the CAR T cells infused in each patient."

That data is based on validation studies the company has run that found "that we observe the same or similar cytokine-secreting single-cell subsets from a patient across multiple replicates so long as we gather more than 1,000 cells per patient," he said. "We have run replicate CAR T assays from the same donor in the past and achieved low variability across the replicates, [providing] further validation that we are capturing the full profile of these cells."

The researchers found 20 to 25 percent of all CAR T product cells were polyfunctional and noted that the proteins secreted by these polyfunctional cells spanned "effector molecules (granzyme B), stimulatory/immune modulating cytokines (IFN-γ, IL-5), and chemokines (IL-8, MIP-1α)."

They also observed that polyfunctional T cells expressing the cytokine IL-17A were linked to increased levels of neurotoxicity.

Mackay said polyfunctionality data like that generated in the Blood study could potentially be used by drugmakers to inform their manufacturing processes to generate the most effective CAR T products.

"A biopharma might optimize the manufacturing components in their process to achieve products with higher potential for efficacy, for example, with high PSI and specific cytokine signatures at the single-cell level, as well as lower toxicity based on existing polyfunctional data," he said.

Mackay suggested that as the ACT field evolves, such measurements could be used in the clinic to help predict patient response or side effects. Single-cell data "may provide actionable information to help clinicians to monitor and proactively address the highest risk patients in either category as they first receive the therapy," he said.

IsoPlexis is currently developing an automated version of the IsoCode platform that it hopes will help it further penetrate the ACT clinical trial market.