3D stem-like spheroids-on-a-chip for personalized combinatorial drug testing in oral cancer.

BACKGROUND: Functional drug testing (FDT) with patient-derived tumor cells in microfluidic devices is gaining popularity. However, the majority of previously reported microfluidic devices for FDT were limited by at least one of these factors: lengthy fabrication procedures, absence of tumor progenitor cells, lack of clinical correlation, and mono-drug therapy testing. Furthermore, personalized microfluidic models based on spheroids derived from oral cancer patients remain to be thoroughly validated. Overcoming the limitations, we develop 3D printed mold-based, dynamic, and personalized oral stem-like spheroids-on-a-chip, featuring unique serpentine loops and flat-bottom microwells arrangement. RESULTS: This unique arrangement enables the screening of seven combinations of three drugs on chemoresistive cancer stem-like cells. Oral cancer patients-derived stem-like spheroids (CD 44+) remains highly viable (> 90%) for 5 days. Treatment with a well-known oral cancer chemotherapy regimen (paclitaxel, 5 fluorouracil, and cisplatin) at clinically relevant dosages results in heterogeneous drug responses in spheroids. These spheroids are derived from three oral cancer patients, each diagnosed with either well-differentiated or moderately-differentiated squamous cell carcinoma. Oral spheroids exhibit dissimilar morphology, size, and oral tumor-relevant oxygen levels (< 5% O2). These features correlate with the drug responses and clinical diagnosis from each patient's histopathological report. CONCLUSIONS: Overall, we demonstrate the influence of tumor differentiation status on treatment responses, which has been rarely carried out in the previous reports. To the best of our knowledge, this is the first report demonstrating extensive work on development of microfluidic based oral cancer spheroid model for personalized combinatorial drug screening. Furthermore, the obtained clinical correlation of drug screening data represents a significant advancement over previously reported personalized spheroid-based microfluidic devices. Finally, the maintenance of patient-derived spheroids with high viability under oral cancer relevant oxygen levels of less than 5% O2 is a more realistic representation of solid tumor microenvironment in our developed device.

Osteomatrix as a personalized 3D tissue-specific invasion test-bed for oral carcinoma.

The clinical challenge in the successful management of oral cancer malignancy remains in the inaccuracy of detecting regional invasion potential and inefficient treatment of recurrent tumors. The presence and extent of bone invasion by the oral tumor are of critical importance as they can influence the preoperative strategy altering the prognosis outcome. Here, we are examining the patient-specific osteotropism of oral carcinoma using a bone derived extracellular matrix. The extracellular matrix (ECM) was obtained from caprine bone by a combination of demineralization, delipidation and decellularization (D3) techniques. The bone D3-derived ECM (BdECM) tissue was characterized for analyzing the effective removal of cells, minerals, and lipids with an intact structure and chemical composition. The human adipose-derived stem cells (ADSCs) on the osteomatrix (BdECM derived hydrogel) exhibited excellent cell viability and early osteogenic differentiation capacity in vitro. Furthermore, the osteomatrix polarized monocytes towards an anti-inflammatory phenotype (M2 macrophage) indicating its low immunogenicity. In the second phase of this study, we isolated and established primary cancer cell cultures from patient-derived tissue exhibiting the cancer stem cell marker phenotype (EpCAM+/CD44high/CD24-). Moreover, the presence of side population (SP) cells confirmed a contributing factor for resistance to cancer therapy. The spheroid formed from primary cells embedded in the osteomatrix was used as a test-bed to monitor the invasion profile and screening of anti-cancer drugs. Our 3D test platform captured the inter-patient heterogeneity by displaying variation in the degree of invasion and response towards tested doses of anticancer drugs. Altogether, our data emphasize the necessity of a tissue-specific in vitro preclinical model for the evaluation of oral carcinogenesis and drug sensitivity.