Continual depletion of circulating tumor cells using an automated device enriched with affinity glass bead substrates in breast and CRC patient’s whole blood.

Abstract

e12505 Background: Despite no evidence of disease radiologically or pathologically, about 25-50% colorectal (CRC) Stage II-III and early stage breast cancer (BC) cases are known to recur. Presence of circulating tumor cells (CTCs) having EMT traits represents aggressive disease systemically. Upon autonomous oncogenic activation, epithelial cells acquire invasiveness to form metastasis. High EMT score with immune checkpoints, like PD-L1 expression, could evade immune system. Following curative-intent surgery and treatments, CTCs represent minimal cellular residual disease (MCRD) with strong predictor for recurrence. We present automated extracorporeal device, to capture, analyze, and deplete CTC for further analysis. Methods: We retrospectively analyzed 66, CRC stage II-III (n=41) and BC (n=25) patients. Whole blood was processed to deplete CTCs using OncoMetastat device. In 41 CRC patients, 12 were female and 29 were male. The average patient age for BC and CRC patients was 53.6 and 58, respectively. Device consists of spiral channel (127 × 85 × 5 mm; spiral span: 66 mm and width: 4 mm), 3D-printed using biocompatible resin, filled with anti-EpCAM antibody conjugated glass beads (GB). Device possess controller, peristaltic pump for blood circulation in and out from spiral channels. Vibrational-energy imparted motion to GB. We analyzed hemolysis, protein, and leucocyte adsorption, and CTC capture in the device. CTC capture efficiency was compared with OncoDiscover CTC technology approved by CDSCO, India. Blood was pumped in device and incubated with affinity enriched GBs for 30 min, under constant vibration (200 Hz) to enhance CTC capture and prevent blood stagnancy. CTCs were confirmed for CK18+ve, DAPI+ve and CD45-ve and analyzed using an automated fluorescence microscope. Results: A total of 48 CTCs were detected in 58% (38/66) of patients. CTC +ve was higher in BC patients (60.0%) over CRC patients (56.1%). Total mean CTC distribution was 0.73, with CRC 0.73 and BC 0.72, respectively. The NPV was determined to be 0.86 (86%). Auto-scan imaging demonstrated 100% efficiency in detecting CTCs. We observed low leukocyte adhesion with anti-EpCAM GB. WBCs counts varied by cancer type (mean WBC count/ml of 4.9 × 10 6 ; Breast CA, 3.9 × 10 6 ; Colorectal CA), lower compared to healthy controls (6.9 × 10 6 ). Clinically insignificant hemolysis (<1%) and protein-binding (~1.5%) was noted in the spiral channel. GBs with vibrational-energy showed enhanced CTC sequestration (>90% cell capture efficiency) compared to vibration-free. Conclusions: We show efficiency of CTC depletion in 66 CRC and BC patients using an automated device. Early stage CRC and BC patients having CTCs possess high risk of progression of distant metastasis. Thus, post complete remission (CR), using an extracorporeal device to deplete CTCs could reduce the risk of metastasis.