Immersion Bioprinting of Tumor Organoids in Multi-Well Plates for Increasing Chemotherapy Screening Throughput

Abstract

The current drug development pipeline takes approximately fifteen years and $2.6 billion to get a new drug to market. Typically, drugs are tested on two-dimensional (2D) cell cultures and animal models to estimate their efficacy before reaching human trials. However, these models are often not representative of the human body. The 2D culture changes the morphology and physiology of cells, and animal models often have a vastly different anatomy and physiology than humans. The use of bioengineered human cell-based organoids may increase the probability of success during human trials by providing human-specific preclinical data. They could also be deployed for personalized medicine diagnostics to optimize therapies in diseases such as cancer. However, one limitation in employing organoids in drug screening has been the difficulty in creating large numbers of homogeneous organoids in form factors compatible with high-throughput screening (e.g., 96-and 384-well plates). Bioprinting can be used to scale up deposition of such organoids and tissue constructs. Unfortunately, it has been challenging to 3D print hydrogel bioinks into small-sized wells due to well-bioink interactions that can result in bioinks spreading out and wetting the well surface instead of maintaining a spherical form. Here, we demonstrate an immersion printing technique to bioprint tissue organoids in 96-well plates to increase the throughput of 3D drug screening. A hydrogel bioink comprised of hyaluronic acid and collagen is bioprinted into a viscous gelatin bath, which blocks the bioink from interacting with the well walls and provides support to maintain a spherical form. This method was validated using several cancerous cell lines, and then applied to patient-derived glioblastoma (GBM) and sarcoma biospecimens for drug screening.

Engineering of tissue constructs using coaxial bioprinting

“…Organoids have been used in a staggering number of applications, including intestinal, cerebral, pancreatic, kidney, hepatic, retinal, lung, colonic, gastric, thyroid, prostate, salivary, mammary, lingual, placental, and spinal tissues, which are reviewed elsewhere [ 88 ]. Bioprinting has only recently seen application in organoid literature [ [89] , [90] , [91] , [92] , [93] , [94] , [95] , [96] ]. In coaxial systems, developing organoid tissues might be printed within a sacrificial sheath hydrogel, allowing for mechanical strength and hierarchal arrangement, without compromising cell viability ( Fig.…”

“…Importantly, bioprinting within soft material preserves cell-cell contacts. A few studies have demonstrated spheroid or organoid bioprinting into pharmaceutical well-plate assays [ 89 , 90 , 97 ]. Coaxial bioprinting with a sacrificial sheath and soft core could augment these approaches, bringing the same benefits of enhanced cell-cell contact and self-assembly.…”

Bioprinting for combating infectious diseases

“…Recent advances in additive manufacturing technologies allow for the automated bioprinting of 3D constructs in assay format, providing constructs for testing that exhibit responses more similar to natural physiological responses compared to 2D assays ( Fig. 4 ) [ 34 , 61 ].

Fig.

…”

“…In order to apply 3D bioprinting to high-throughput screening, cells of interest are selected and loaded into a bioink [ 34 , 61 ]. The bioink can then be printed into a multi-well plate containing a matrix material, or printed as a structurally stable construct in order to form a 3D cellular model [ 34 , 61 ]. Drug compounds can then be added and their effect can be assessed in a more physiologically relevant format [ 34 , 61 ].…”

“…The bioink can then be printed into a multi-well plate containing a matrix material, or printed as a structurally stable construct in order to form a 3D cellular model [ 34 , 61 ]. Drug compounds can then be added and their effect can be assessed in a more physiologically relevant format [ 34 , 61 ]. However, technologies used to screen 3D constructs used in assay format are not currently available or integrated into pharmaceutical screening systems [ 34 ].…”