With more than 1000 cell and gene therapies currently in development, the number of trials is quickly increasing. CAR-T therapies are changing the landscape of precision medicine in oncology, but production comes with its own set of challenges. Because CAR-T therapies are highly personalized and use a patient’s own T cells, one of the most significant challenges in its development is characterizing the quality and potency of the products after production. To address these challenges, the CliniMACS Prodigy® and IsoCode Single-Cell Chip are enabling the automation of this process and empowering researchers to achieve critical correlates to in vivo data.
IsoPlexis Recognizes the Complexity of Producing Each Cell and Defines the Functional Quality of the Cellular Components in Terms of Its Function & Fitness
The IsoPlexis system detects the specific cytokines secreted from each single cell and identifies polyfunctional cells and cell subsets (those that secrete two or more cytokines), which have correlated to response in multiple studies. The IsoPlexis platform can also detect, within a phenotypically identical sample, functionally heterogeneous and potent cells, which are missed by Flow and ELISA (Figure 1). This significant finding demonstrates that not all T cells within a single sample function in the same way. The IsoLight system’s deep functional profiling, along with the ability to correlate its metrics to actual patient response, enables cell therapy manufacturers to ensure that a cell product is potent enough during development and enables optimization of critical cell therapy product characteristics.
Figure 1. Phenotypically identical cells, though functionally heterogeneous.
IsoPlexis’ Commitment to Automating the Precision CAR-T Bioprocessing Workflow & Defining T Cell Quality/Fitness with Miltenyi Biotec
Here, we discuss a case study where researchers used the next generation automation system from Miltenyi Biotec upstream and next generation automation from IsoPlexis downstream to identify the quality and fitness of the cell therapy product. The functional quality of CAR-T products is crucial for this type of cell-based immunotherapy, and when researchers can quantify the cytokine output from each cell, it enables optimization of the manufacturing processes to significantly improve product quality.
The primary challenge today is how to provide an objective functional readout to differentiate between bioprocessing methods and justify whether newly developed methods are functionally superior to alternative methods. In this study, The CliniMACS Prodigy® was used to automate the workflow for producing CAR-T products (Figure 2). Two production methods were used. The first method (M1) concluded with a wash step at day 5, while the second method (M2) concluded at day 3. This study identified which method produced higher functional quality cell therapy product in terms of fitness and potency.
Figure 2. Automated end-to-end CAR-T cell production and single-cell analysis with Miltenyi Biotec CliniMACS Prodigy® and IsoPlexis’ IsoLight system.
Why Precision CAR-T Functional Quality Analysis Matters in Bioprocessing
IsoPlexis’ Polyfunctional Strength Index (PSI) was used to assess the two methods. PSI simultaneously encompasses the breadth of cytokine secretions (polyfunctionality, or single CAR-T cells that secrete two or more cytokines) and the intensity of cytokine secretions (secretion amounts), which are both found at a single-cell level across 30+ cytokines. This unique metric enables evaluation of the potency of a cell product prior to application as a therapy. A recent commentary on CAR-T strength and polyfunctionality in Blood stated, “they have showed that by using single-cell analyses, heterogeneity may actually improve clinical outcomes.”
Increased cell quantities and viabilities do not always correlate to enhanced cellular function. By using the IsoLight platform to identify polyfunctional subsets within the products generated by the two bioprocessing methods, a higher degree of immune fitness was identified in the CAR-T products produced with M2, identifiable via elevated effector and stimulatory multiplexed cytokine signatures. This revealed that the M2 resulted in improved immune fitness of CAR-T products in both potency and diversity.
Why Automation Matters in the End-to-End Workflow
The need for automation is urgent in the cell therapy industry, given the need to streamline both product generation and analyses. In order to simplify the cell processing workflow, Miltenyi Biotec’s CliniMACS Prodigy® automates everything from cell separation and cell culture to formulation of the final cell product. When used with IsoPlexis’ IsoCode single-cell cytokine detection system, human involvement can be minimized throughout several different steps given that the IsoLight automates all fluidics & ELISA reagents for each cell, thereby increasing the efficiency of the bioprocessing workflow and accelerating the development of engineered cell products.
For more details on this end-to-end process, download our Tech Note, “Getting the most out of your CAR-T bioprocessing with the CliniMACS Prodigy® & IsoCode Single-Cell Chip.”