Abstract 7495: Validation of microfluidic isolation of circulating pancreatic tumor cells from blood
Authors: Macaraniag Celine , Khan Ifra , Zhou Jian , Giulianotti Pier Cristoforo , Borgeat Alain , Votta-Velis Gina , Papautsky Ian 
Published: March 2024

Abstract
Background. Pancreatic ductal adenocarcinoma (PDAC) has a poor prognosis and is the fourth leading cause of cancer-related deaths in the US. PDAC is diagnosed in the late stages, which limits treatment options and yields poor clinical outcomes. Recurrence is a significant issue for patients undergoing surgery for PDAC. Circulating tumor cells (CTCs) are thought to be a potential mediator of recurrence, facilitating the formation of secondary tumor sites after surgery. Thus, there is a need to isolate CTCs to improve understanding of disease recurrence, to ultimately improve outcomes. Herein, we aim to validate our inertial microfluidic (iMF) system for isolating pancreatic CTCs from blood. We also benchmark our iMF system to a commercially available immunomagnetic negative selection system, EasySep™, and quantify target cell recovery and cell enrichment.
Methods. Our iMF system employs the differential cell sizes of CTCs from blood cells and inertial migration of such cells in a straight microchannel (150 µm × 50 µm × 24 mm) to enrich CTCs from blood. The iMF system sample flow rate used is 100 µL/min, with a 200 µL/min PBS buffer flow rate. The EasySep™ Direct Human CTC Enrichment kit targets CD2, CD14, CD16, CD19, CD45, CD61, CD66b, and Glycophorin to remove blood cells from the sample using magnetic particles, thereby separating them from CTCs. We spiked low cell concentrations of 50, 100, and 500 Hoechst-stained pancreatic carcinoma cells, PANC1, in 1 mL of healthy blood then captured using either EasySep™ or iMF. Cell enumeration was performed after centrifugation or Cytospin.
Results. PANC1 cells and WBCs have average sizes of 18 µm and 8.4 µm, respectively. We characterized our iMF device to capture cells ≥ 12 µm, which yields capture of most PANC1 cells. Indeed, our iMF system achieved 82.8±19.2%, 72.5±16.3%, and 59.3±3.8% recovery for 50, 100, and 500 cell-spikes in lysed blood, respectively. For the same 50, 100, and 500 cell-spikes, the total iMF CTC yields after Cytospin were 46.3±11.8, 44.3±9.1, and 32.7±2.1%, while the total CTC yields from EasySep™ were 4.6±3, 5.3±2.1, 8.3±1%, respectively. When we processed whole blood directly, EasySep™ had lower cell recovery, 3.8±0.2, 4.6±1, and 3.3±0.4%, respectively. Our system also demonstrated 8 × higher target cell enrichment compared to EasySep™. Blood lysis before iMF separation did not drastically affect the recovery of target cells.
Conclusions. We determined that our system has improved performance over the commercial isolation system, EasySep™, thereby validating the effectiveness of microfluidic isolation for PDAC. In future studies, we can then use our iMF system in CTC isolation from routine blood draws from PDAC patients undergoing surgical resection. We may then isolate real patient samples and study relevant phenotypes of PDAC CTCs which may therefore be used for identifying certain mechanisms of PDAC progression.
Citation Format: Celine Macaraniag, Ifra Khan, Jian Zhou, Pier Cristoforo Giulianotti, Alain Borgeat, Gina Votta-Velis, Ian Papautsky. Validation of microfluidic isolation of circulating pancreatic tumor cells from blood [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7495.