Perfusable host microfluidic endothelial networks for quantitative assessment of the effects of interstitial flows, macrophage polarization and monoclonal‐based immunotherapy on immune cell infiltration into a tumor spheroid

Abstract

Monoclonal antibody-based immunotherapy is a promising treatment for several diseases such as cancer and virus infection. However, due to its highly selective target, drug testing using animal models or classical 2D human cell cultures are not sufficient to predict the drug effects on patients. New 3D organotypic models that employ 3D cell culture of human cells within a microfluidic device allow detailed characterization of critical biological interactions. In this project, we describe a new 3D cell culture model that allows the creation of in-vitro human perfusable vasculature containing hollow spaces for the insertion of tumor spheroids. A polydimethylsiloxane device with several vertical holes on top of the gel channel is used to create the host microvascular endothelial (HOME) network (Fig.1A). Endothelial cells and fibroblasts create perfusable vascular networks (Fig. 1B). Tumor spheroids are formed by coculturing MDA-MB-468 tumor cells (T), fibroblasts (F), and non-polarized (M0) or M2 macrophages, and denoted by TFM0 and TFM2 respectively (Fig. 1C). One day after inserting a tumor spheroid in collagen into the HOME networks through the hole, freshly-isolated monocytes are perfused into the vascular networks by luminal flows. We observe that after 2 days, the majority of monocytes remain either within the vasculature or extravasated but stay inside the fibrin gel outside of the hole containing the tumor spheroid (Fig. 1Di). On day 2, interstitial flows from the tumor spheroid cause monocyte migration from the vascular network side into the hole containing the tumor spheroid (figure 1Dii). When a TFM2 spheroid is inserted into the HOME networks, it recruits more monocytes than the TFM0 spheroid (Fig. Ei and Eii). Recruitment % is the percentage of monocytes that leave the blood vessel and migrate into the hole among the total number of monocytes in 3mmx3mm region of interest around the hole. Monocytes are recruited better by the TFM2 spheroid than the TFM0 spheroid (Fig. 1Gi). These results showed that interstitial flows from the tumor spheroid to the vascular networks and macrophage polarization are essential for monocyte recruitment. Multi-specific antibody UniTI102 developed by Elstar Therapeutics (MA, USA) neutralizes chemokine CCL2 to block monocyte recruitment. To characterize the response to the drug UniTI102, we compared devices having a TFM0 spheroid that are untreated (Fig. 1D) or treated with UniTI102 (Fig. 1F) and observed that less monocytes has migrated into the hole in the presence of UniTI102 at 100nM. Quantitative data also showed that less monocytes are recruited when devices are treated with 100nM UniTI102 than 100nM IgG (Fig. 1Gii).