An Engineered Patient‐Derived Tumor Organoid Model That Can Be Disassembled to Study Cellular Responses in a Graded 3D Microenvironment

Landon-Brace, Natalie; Cadavid, Jose L.; Latour, Simon; Co, Ileana L.; Rodenhizer, Darren; Li, Nancy T; Wu, Nila C.; Bugbee, Eryn; Chebotarev, Aleks; Zhang, Ji; Wouters, Bradly G.; McGuigan, Alison P.

doi:10.1002/adfm.202105349

Cited by 20 publications

(76 citation statements)

References 68 publications

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“…Lastly, the flow culture model exhibits increased sensitivity to Docetaxel, with a uniform cell death distribution and higher differential transcriptional response. This finding is consistent with previous studies which suggest that removing hypoxia-induced cellular drug resistance could increase the cellular response to chemotherapy [55,56,57], and which show flow culturederived spheroids can show reduced sensitivity compared to 2D models [58].…”

Section: Discussionsupporting

confidence: 93%

Microwell-Based Flow Culture Increases Viability and Restores Drug Response in Prostate Cancer Spheroids

Payne

Hashimoto

et al. 2022

Preprint

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3D cancer spheroids represent a highly promising model for study of cancer progression and therapeutic development. Wide-scale adoption of cancer spheroids, however, remains a challenge due to the lack of control over hypoxic gradients that may cloud the assessment of cell morphology and drug response. Here, we present a Microwell Flow Device (MFD) that generates in-well laminar flow around 3D tissues via repetitive tissue sedimentation. Using a prostate cancer cell line, we demonstrate the spheroids in the MFD exhibit improved cell growth, reduced necrotic core formation, enhanced cellular structure, and down-regulated expression of cell stress genes. The flow-cultured spheroids also exhibit an improved sensitivity to chemotherapy with greater transcriptional response. These results demonstrate how fluidic stimuli reveal the cellular phenotype previously impacted by severe hypoxia. Our platform advances 3D cellular models and enables study into hypoxia modulation, cancer metabolism, and drug screening within pathophysiologically relevant conditions.

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Section: Discussionsupporting

confidence: 93%

Microwell-Based Flow Culture Increases Viability and Restores Drug Response in Prostate Cancer Spheroids

Payne

Hashimoto

et al. 2022

Preprint

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“…Alternatives to preclinical and monolayer cell culture models such as tissue-engineered in vitro tumor models , have been introduced, including 3D hydrogel culture systems, ,, spheroids, organoid cultures, − and organ-on-a-chip ,, platforms. Among the fabrication tools, extrusion bioprinting offers the potential to create culture systems with user-defined settings to control the geometry of the construct, placement of cell types, cell density, and scaffold materials or bioinks. , Several bioinks have been developed for this technique using synthetic hydrogels, which have excellent mechanical properties and have been proven to be reliable and consistent, with reproducible mechanical characteristics. , Nevertheless, these materials often fail to recapitulate the biochemical and biomechanical complexity found in native ECM, resulting in the absence of intrinsic physiological function and aberrant cell behavior .…”

Section: Introductionmentioning

confidence: 99%

Decellularized Extracellular Matrix Composite Hydrogel Bioinks for the Development of 3D Bioprinted Head and Neck in Vitro Tumor Models

Kort-Mascort

Bao

Elkashty

et al. 2021

ACS Biomater. Sci. Eng.

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Reinforced extracellular matrix (ECM)-based hydrogels recapitulate several mechanical and biochemical features found in the tumor microenvironment (TME) in vivo. While these gels retain several critical structural and bioactive molecules that promote cell–matrix interactivity, their mechanical properties tend toward the viscous regime limiting their ability to retain ordered structural characteristics when considered as architectured scaffolds. To overcome this limitation characteristic of pure ECM hydrogels, we present a composite material containing alginate, a seaweed-derived polysaccharide, and gelatin, denatured collagen, as rheological modifiers which impart mechanical integrity to the biologically active decellularized ECM (dECM). After an optimization process, the reinforced gel proposed is mechanically stable and bioprintable and has a stiffness within the expected physiological values. Our hydrogel’s elastic modulus has no significant difference when compared to tumors induced in preclinical xenograft head and neck squamous cell carcinoma (HNSCC) mouse models. The bioprinted cell-laden model is highly reproducible and allows proliferation and reorganization of HNSCC cells while maintaining cell viability above 90% for periods of nearly 3 weeks. Cells encapsulated in our bioink produce spheroids of at least 3000 μm2 of cross-sectional area by day 15 of culture and are positive for cytokeratin in immunofluorescence quantification, a common marker of HNSCC model validation in 2D and 3D models. We use this in vitro model system to evaluate the standard-of-care small molecule therapeutics used to treat HNSCC clinically and report a 4-fold increase in the IC50 of cisplatin and an 80-fold increase for 5-fluorouracil compared to monolayer cultures. Our work suggests that fabricating in vitro models using reinforced dECM provides a physiologically relevant system to evaluate malignant neoplastic phenomena in vitro due to the physical and biological features replicated from the source tissue microenvironment.

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“…The PMMA-acetone solution (1.75g PMMA to 7.84mL acetone) was mixed with blue nail polish (Essie) for visibility. A modification of the contact wicking printing technique developed in our group [39], [40] was used for PMMA infiltration. Briefly, the scaffold templates were taped to a sheet of parchment paper to prevent sticking and placed on the AxiDraw V3 (Evil Mad Scientists, USA) plotter support.…”

Section: Methodsmentioning

confidence: 99%

“…The remaining cells were pelleted by centrifugation (300 g , 5 min, 4 °C). In accordance with [39] the PPTO.46 cell pellet was re-suspended in an appropriate volume of a hydrogel blend to achieve the desired cell concentration. The hydrogel blend used for PPTO.46 cells was comprised of 25% Matrigel™ and 75% collagen hydrogel as reported previously [39].…”

Section: Methodsmentioning

confidence: 99%

“…Sixteen or four sites were acquired per 96- or 384-SPOT, respectively, to capture a full well bottom, which was subsequently stitched using MetaXpress software (Molecular Devices, USA). A custom script (adapted from [39]) was written in ImageJ to measure the mean grey value (MGV) of GFP signal of 100 randomly selected squares to obtain the standard deviation within 1 well. Standard deviations were used to calculate the coefficient of variation associated to each well using the following: …”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

An off-the-shelf multi-well scaffold-supported platform for tumour organoid-based tissues

Li¹,

Wu²,

Cao³

et al. 2022

Preprint

Self Cite

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Complex 3D bioengineered tumour models provide the opportunity to better capture the heterogeneity of patient tumours. Patient-derived organoids are emerging as a useful tool to study tumour heterogeneity and variation in patient responses. Organoid cultures typically require a 3D microenvironment that can be manufactured easily to facilitate screening. Here we set out to create a high-throughput, "off-the-shelf" platform which permits the generation of organoid- containing microtissues for standard phenotypic bioassays and image-based readings. To achieve this, we developed the Scaffold-supported Platform for Organoid-based Tissues (SPOT) platform. SPOT is a 3D gel-embedded in vitro platform that can be produced in a 96- or 384-well plate format and enables the generation of flat, thin and dimensionally-defined microgels. SPOT has high potential for adoption due to its reproducible manufacturing methodology, compatibility with existing instrumentation, and reduced within-sample and between-sample variation, which can pose challenges to both data analysis and interpretation. Using SPOT we generate cultures from patient derived pancreatic ductal adenocarcinoma organoids and assess the cellular response to standard-of-care chemotherapeutic compounds, demonstrating our platform's usability for drug screening. We envision 96/384-SPOT will provide a useful tool to assess drug sensitivity of patient-derived organoids and easily integrate into the drug discovery pipeline.

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An Engineered Patient‐Derived Tumor Organoid Model That Can Be Disassembled to Study Cellular Responses in a Graded 3D Microenvironment

Cited by 20 publications

References 68 publications

Microwell-Based Flow Culture Increases Viability and Restores Drug Response in Prostate Cancer Spheroids

Microwell-Based Flow Culture Increases Viability and Restores Drug Response in Prostate Cancer Spheroids

Decellularized Extracellular Matrix Composite Hydrogel Bioinks for the Development of 3D Bioprinted Head and Neck in Vitro Tumor Models

An off-the-shelf multi-well scaffold-supported platform for tumour organoid-based tissues

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