Abstract B31: Detection of viable tumor cells from cryopreserved buffy coat using the VTX-1 Liquid Biopsy Platform

Abstract

Abstract Introduction: Several circulating tumor cell (CTC) technologies have been developed amidst growing interests in the “liquid biopsy” field. The fully automated VTX-1 platform is developed to gently isolate intact CTCs from whole blood with high recovery and viability, compatible with many applications (1-3). However, many CTC technologies require processing of fresh patient samples at off-site facilities. Here we present encouraging preliminary results showing viable cell isolation by VTX-1 from cryopreserved buffy coat, using a telomerase-selective replicating adenovirus (TelomeScan). This widens potential applications of viable CTC detection for retrospective clinical studies and drug development. Methods: Buffy coat preparation: Buffy coat was prepared by centrifugation of healthy donor blood collected in EDTA, heparin, or heparin CPT vacutainers. Cell spiking and processing: 1) Cancer cells were spiked into freshly prepared buffy coat and processed with the Vortex chip at different dilutions. 2) Cancer cells were labeled and spiked into healthy donor blood, and buffy coat was prepared and processed by VTX-1 either immediately or following cryopreservation up to 1 week. Imaging was evaluated for cell recovery. Viable, telomerase-activated tumor cell detection: VTX-1 enriched cells were directly collected in chamber slides and infected with the TelomeScan adenovirus (OBP-401) for 24 h (4). The percentage of GFP (+), viable, telomerase-expressing cells was evaluated alongside cell recovery. Results: MCF7 cells were spiked into freshly prepared buffy coat, resuspended in 8mL, and processed at 5X, 10X, and 20X dilutions. The best compromise between recovery, WBC count, and processing time was obtained for the 10X dilution with 46% recovery. The majority of spiked cells (MCF7 and H1299) collected from buffy coat were shown to be viable, as demonstrated by infection with the telomerase-selective replicating adenovirus (TelomeScan, OBP-401). Cryopreservation of the buffy coat samples showed a slight reduction following cryopreservation. Discussion: The optimal performance of the Vortex chip on buffy coat samples has previously been shown at 10X dilution. This capability of the VTX-1 was further demonstrated through infection with a telomerase-selective replicating adenovirus. This combined workflow has shown seamless integration from whole blood. Noteworthy recovery and sensitivity were also observed with cryopreserved buffy coat. Patient biobanked samples are currently being pursued. The overall advantages are: a) viable cell detection, b) direct collection of cells into culture media ready for in vitro testing, c) no requirements for fixation, labeling, backflushing, d) a very low WBC background eliminating false positives, and e) the potential to retrospectively analyze clinical biobanked samples at a centralized laboratory.