High-Throughput Microfluidic 3D Cytotoxicity Assay for Cancer Immunotherapy (CACI-IMPACT Platform)

Abstract

Adoptive cell transfer against solid tumors faces challenges to overcome tumor microenvironment (TME), which plays as a physical barrier and provides immuno-suppressive conditions. Classical cytotoxicity assays are widely used to measure killing ability of the engineered cytotoxic lymphocytes as therapeutics, but the results cannot represent the performance in clinical application due to the absence of the TME. This paper describes a 3D cytotoxicity assay using an injection molded plastic array culture (CACI-IMPACT) device for 3D cytotoxicity assay to assess killing abilities of cytotoxic lymphocytes in 3D microenvironment through a spatiotemporal analysis of the lymphocytes and cancer cells embedded in 3D extra cellular matrix (ECM). Rail-based microfluidic design was integrated within a single 96-well and the wells were rectangularly arrayed in 2 × 6 to enhance the experimental throughput. The rail-based microstructures facilitate hydrogel patterning with simple pipetting so that hydrogel pre-solution aspirated with 10 μl pipette can be patterned in 10 wells within 30 s. To demonstrate 3D cytotoxicity assay, we patterned HeLa cells encapsulated by collagen gel and observed infiltration, migration and cytotoxic activity of NK-92 cells against HeLa cells in the collagen matrix. We found that 3D ECM significantly reduced migration of cytotoxic lymphocytes and access to cancer cells, resulting in lower cytotoxicity compared with 2D assays. In dense ECM, the physical barrier function of the 3D matrix was enhanced, but the cytotoxic lymphocytes effectively killed cancer cells once they contacted with cancer cells. The results implied ECM significantly influences migration and cytotoxicity of cytotoxic lymphocytes. Hence, the CACI-IMPACT platform, enabling high-throughput 3D co-culture of cytotoxic lymphocyte with cancer cells, has the potential to be used for pre-clinical evaluation of cytotoxic lymphocytes engineered for immunotherapy against solid tumors.

Novel 3‐aryl‐5‐substituted‐coumarin analogues: Synthesis and bioactivity profile

“…SK‐N‐SH human neuroblastoma cell line was selected to allow comparisons on the cytotoxic activity evaluation of hydroxy and oxyprenylated coumarin derivatives studied previously by our group (Kavetsou et al, ). Both SK‐N‐SH and HeLa cells have been widely used to examine the anticancer effect of bioactive agents (Akram et al, ; Dickens et al, ; Goswami et al, ; Park et al, ; Qi et al, ; Swift, Zhang, Trippett, & Narendran, ; Yen et al, ).…”

Human Ocular Angiogenesis‐Inspired Vascular Models on an Injection‐Molded Microfluidic Chip

“…The fluid guide structure allows the fluid to be spontaneously transported by capillary forces in a hydrophilic environment. Thus, the structure can be designed to engineer capillary flow patterns . We considered the height from the substrate and width of the concentric layer as key factors affecting the patterning conditions.…”

“…All images are then converted to a binary mask image after threshold adjustment. After careful verification, you can quantify vascular network coverage and other angiogenic parameters using the Fiji‐based AngioTool following automated protocols . If we input images that are well‐prepared for acquiring the morphological features, the tool can calculate the quantitative results automatically.…”

3D Immunocompetent Organ‐on‐a‐Chip Models

“…In an another study, a higher‐throughput microfluidic plastic array culture (CACI‐IMPACT) device was designed to study the cytotoxic efficacy of lymphocytes to ablate cancer cells in 3D microenvironments. [ 162 ] Rail‐based microstructures, comprised of the low‐rail (cancer cell) zone, the high‐rail (immune cell) zone, and the reservoir wall, were placed in microwells and integrated to form a 2 × 6 rectangular arrays of the microwells. HeLa cells encapsulated within the collagen hydrogel were patterned in a low‐rail zone, whereas NK‐92 cell suspension was loaded in the high‐rail zone and allowed to migrate toward the HeLa cells in the low‐rail zone ( Figure Aa,b).…”

“…Reproduced with permission under the Creative Commons Attribution 4.0. [ 162 ] Copyright 2019, the Authors. Published by Frontiers.…”