Use of dynamic blood flow device with conjugated affinity ligands on glass substrate to capture circulating tumor cells in cancer patients.

Abstract

e15029 Background: Primary tumors are known to shed circulating tumor cells (CTCs), promoting disease in circulation and thus rendering the patient highly prone to the progression of metastasis to distant organs. 90% of cancer-related deaths, are unswervingly correlated with metastasis. Few studies show the concept of enhanced overall survival in animal models, attributable to reduction in the number of CTCs in blood circulation. We demonstrate capture of CTCs in a continual blood flow device with 3-dimensional glass substrates (3D GS) conjugated to affinity ligands anti-epithelial cell adhesion molecule (EpCAM) antibody and transferrin (Tf) in cancer patients. Methods: A bi-spiral, plano-horizontal, optically transparent device having biocompatible resin having multiple channels for continuous blood flow was designed using 3D printer. The circulation device consisted 14 loops for holding 17.5ml of blood sample bearing 680 glass substrate (2 mm dia) with conjugated ligands, anti-EpCAM antibody and Tf protein. Device was mechanised for functional circulation with 3 pumps. To elicit pyrogenicity, arising from passage of blood through the device, was evaluated in 3 New Zealand White rabbits by ISO:10993-11, for systemic toxicity. 27 blood samples from early stage and late cancers were processed using OncoDialysis assay across 9 cancer types including colorectal, lung, breast, ovarian, etc. 48.15% of the patients were male and 51.85% of the patients were female. CTCs were captured using 5 GSs in a blood volume 1.5-5 mL. CTCs were validated with CK 18 and CD45 and verified using fluorescence microscopy. Blood samples were also processed using Drug Controller, India approved OncoDiscover technology for comparative analysis. Results: Vitally, no blood hemolysis was observed as arising from the device, and further CTC capture in 5 ml of continually circulated blood in the flow system was demonstrated. All rabbits appeared healthy during the test and none of the tested rabbits exhibited an individual increase in temperature of .5 °C or more, when compared to control test. OncoDialysis assay captured CTCs in 48.15% of patients (n = 13/27). A total of 14 CTCs (12 CTCs and 2 CTC clusters) with a mean CTC distribution of 0.51 in blood volume 1-5 ml. 23 CTCs (18 CTCs and 5 CTC clusters) were isolated using the OncoDiscover platform (mean CTC distribution of 0.8). Concurrence of 74.07% was observed for CTC capture across the two platforms. In 40.74% of cases (n = 11/27), CTCs were detected by both platforms. Conclusions: We designed a dynamic in vitro blood circulation device, with evidenced safety in animal studies to capture CTCs thereby demonstrating an ability to selectively separate CTCs from a patients blood. CTC load reduction has an immense therapeutic utility in stanching metastatic invasion to improve overall survival of epithelial-origin cancer patients, both with and without therapy.