Abstract LB-137: A new clinical-grade MEMS-based cell sorter for adoptive T-cell therapy
Authors: Yee Cassian , Farrar Erik , Grummitt Daryl , Harley John , Linton Jim , Foster John 
Published: August 2012

For advanced, refractory malignancies, adoptive T cell therapy represents an increasingly attractive modality with the potential for significant anti-tumor efficacy, minimal toxicity and longterm immunoprotection. This approach involves the ex vivo isolation and expansion of a patient's tumor-reactive T cells. Such T cells are rare, and difficult to isolate. Iterative cycles of in vitro re-stimulation, limiting dilution cloning and screening of hundreds to thousands of T cell colonies are required to select for a tumor-reactive population of T cells. Although remarkable responses have been achieved, the time and resource intensive nature of this process has limit the use of adoptive T cell therapy to a handful of centers with specialized facilities and personnel. A closed-system clinical-grade cell sorter that provides a high-speed, sterile and multi-parametric system for isolating low frequency antigen specific T-cells in a cost-effective, easy-to-use and high performance manner would be desirable. A personal-sized cell sorter was developed which uses classical Fluorescence Activated Cell Sorter (FACS) laser excitation and fluorescence emission for interrogation and identification of cells; however, the laser excitation and detection regions, sorting actuator, sample reservoirs, and all of the fluidic channels are contained in a single, closed, disposable, low cost cartridge. The heart of the sorting cartridge is the MEMS microchip which is the world's fastest valve which can open and close in about 15-20 usec allowing a sort rate of more than 100 million cells/hour Cell Sorting of Human Antigen-specific CD8 T cells We determined the lowest threshold limit for MEMS-based antigen-specific T cell single color sorting using fluorescent-conjugated peptide-MHC multimers using a panel of well-characterized tumor-associated antigen (TAA)-specific T cell clones established in our lab were titrated o autologous PBMCs. We demonstrate discrete recognition of the antigen-specific population and the ability to routinely achieve a sort purity of &gt; 85%, yield of &gt; 90% and viability of &gt; 95% starting from a T cell frequency of &lt; 0.5%, with retention of viability, cytolytic, cytokine-producing and proliferative functions. We present data demonstrating routine feasibiligy in achieving enrichment of rare antigen-specific human CD8 and CD4 T cell populations to purity of &gt; 85% with preserved functionality, enhanced viability over conventional stream in air sorters (where cells are subject to 10-50 fold greater shear forces), and expansion capacity after sort to &gt;2000 fold reducing time to treatment for patients from 84 to 24 days. The use of a completely self-contained, disposable cartridge system provides substantial cost savings and allows for multiple samples to be run by a single operator. Development of multicolor and tandem sorting to facilitate enrichment of large volume samples is being pursued.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-137. doi:1538-7445.AM2012-LB-137