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IsoPlexis Uses Single-Cell Platform to Assess T Cell Activity in CAR-T Cancer Therapy

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NEW YORK (GenomeWeb) – Single-cell omics firm IsoPlexis presented data at the American Association for Cancer Research annual meeting this week in Chicago demonstrating the ability of its platform to assess the activity of T cells used in adoptive cell transfer (ACT) cancer therapies.

The work was part of a larger study looking at the effectiveness of ACT in combination with the cytokine agonist NKTR-214. The study, led by Antoni Ribas, director of the tumor immunology program at the University of California Los Angeles' Jonsson Comprehensive Cancer Center, found that the ACT-NKTR-214 combination improved the induction of cytotoxic T cells and increased their polyfunctionality, indicating that addition of NKTR-214 could boost the performance of ACT.

NKTR-214 is produced by San Francisco-based biopharma company Nektar Therapeutics, which also participated in the study. In recently presented data from PhaseII trials, NKTR-214 enhanced the effect of Bristol-Myers Squibb's anti-PD-1 inhibitor Opdivo (nivolumab) in patients with melanoma, renal cell carcinoma, and non-small lung cancer.

Looking at mouse models of melanoma, the Ribas-led team compared the effects of NKTR-214 in boosting ACT to Interleukin-2 (IL-2), which is also known to activate cytotoxic T cells. Treatment with IL-2 comes with unwanted toxicity, however, and patients often relapse after an initial response.

Ribas and his colleagues used IsoPlexis's IsoCode system to assess the polyfunctionality of the induced T cells, the notion being that polyfunctional T cells — meaning those that secrete two or more cytokines per cell — are likely more effective at fighting cancer.

This idea stems from work in infectious disease, where, Ribas said, "there is a solid body of literature supporting [the idea] that polyfunctional T cells making multiple cytokines in response to their antigen are more effective in controlling viral infections."

"The data on antitumor responses is more indirect," he said, but, "I have no doubt that it is also the case."

"The finding in a preclinical model that the IsoPlexis assay can detect a remarkable increase in polyfunctionality in tumor-infiltrating lymphocytes recovered from tumors of mice treated with NKTR214 compared to IL-2 is the most conclusive data we have obtained in this model system," said Ribas, who is a member of IsoPlexis' scientific advisory board and has an equity stake in the company.

Launched in 2014 as a spinout from the lab of Rong Fan, associate professor of biomedical engineering at Yale University, IsoPlexis has made ACT its primary focus. The company offers a single-cell cytokine panel that researchers can use to identify expression patterns linked to patient response to such therapies.

The company's 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 far beyond the levels allowed by fluorescence readout alone.

In a 2014 study in Proceedings of the National Academy of Sciences, Fan and his co-authors showed the system could measure 42 immune effector proteins in single cells. Currently, the company sells a 32-plex single-cell cytokine panel, and in the work presented at AACR this week, Ribas and his colleagues used a 28-plex cytokine assay.

The study found that treatment with NKTR-214 significantly boosted the polyfunctionality of T cells compared to IL-2, leading to a tenfold increase in the percentage of polyfunctional cells.

Ribas said the IsoPlexis system offered advantages over approaches like flow cytometry in terms of assay simplicity and multiplexing capabilities. And, he added, while single-cell transcriptome assays from companies like NanoString and Fluidigm allow researchers to measure cytokine transcripts produced by individual T cells in a highly multiplexed manner, the production of these cytokine transcripts does not always mean these proteins will be produced and secreted.

Fluidigm's mass cytometry technology does allow multiplexing of dozens of protein measurements at the single-cell level and has been similarly employed to study the cytokine and chemokine production of T cells in ACT therapy. And, in fact, Ribas and his colleagues also used mass cytometry analysis in the AACR study, though they used it to look not at levels of cytokine expression but rather at the different T cell populations present in the mouse tumors and spleens after treatment with either ACT-NKTR-214 or ACT-IL-2.

According to Sean Mackay, Isoplexis's CEO and co-founder, the company's participation in the study stemmed from previous work the company did in collaboration with immunotherapy firm Kite Pharma (which is owned by Gilead Sciences), where Ribas is a member of the scientific advisory board.

IsoPlexis had previously shown the ability of its technology "to find highly multifunctional single cells in Kite Pharma's CAR-T trials," Mackay said. "So, the thought then was, let's apply our technology to try to detect if this adopted cell transfer [treatment] with this new [NKTR-214] therapy could create the same type of highly multifunctional single-cell subsets that we had previously associated with a good response to CAR-T."

Mackay noted that the IsoPlexis analysis found not only that addition of NKTR-214 increased the proportion of polyfunctional T cells, but the multiple cytokines produced by these polyfunctional cells were "the right type of cytokines."

They researchers "saw [expression] of cytokines that are extremely important for creating the kind of cytotoxic T cells they are looking for," he said, adding that this included cytokines like RANTES and granzyme B that have traditionally been challenging to measure at the single-cell level.

IsoPlexis is currently developing an automated version of the IsoCode platform that it hopes will help it further penetrate the ACT clinical trial market. The Branford, Connecticut-based company received a $1.8 million grant last summer from the National Cancer Institute's Small Business Innovation Research program to fund development of the system. It also closed a $13.5 million Series B round in September.

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