In recent years, the application of conditioned medium (CM) in regenerative medicine has received much attention. However, the incidence of cancer in Japan is continuing to rise, with one in two people having cancer, among those who may be candidates for CM treatment and may be affected by or have a risk of cancer. We investigated the effects of human dental pulp stem cell (DPSC)-derived CM on human oral squamous cell carcinoma (OSCC) cell lines. CM was extracted from DPSC isolated from human dental pulp, and cell proliferation rates upon contact with OSCC cell lines were compared. Furthermore, chemosensitivity was evaluated with the collagen gel droplet embedded culture drug sensitivity test and was compared using Dulbecco's modified Eagle's medium (DMEM). To test the effect of DPSC-CM on xenograft tumors, an in vivo comparative study was conducted to investigate the tumor proliferation rate upon DPSC-CM and DMEM administration in tumor-bearing nude mice. The tumor growth factor production in culture medium over time was measured with enzyme-linked immunosorbent assay (ELISA) and was compared using DMEM. Vascular endothelial growth factor (VEGF) level significantly increased in the DPSC-CM contact group with ELISA. Therefore, VEGF-A-mRNA expression in the OSCC cell line was studied with real-time polymerase chain reaction for comparison. No significant differences were observed in the cell proliferation rate or drug sensitivity between different culture media in each cell line or in in vivo tumor proliferation rate. However, VEGF level contained in the cultured medium was significantly higher than that in the DP-SC-CM group. VEGF-A-mRNA level in OSCC cell lines was significantly higher in the DPSC-CM group, which increased over time. These results suggested that exposure to DPSC-CM does not immediately affect tumor growth or drug resistance, but induces VEGF overexpression in tumor cells.

Abstract. Using trace three-dimensional culture, the collagen gel droplet-embedded culture drug sensitivity test (CD-DST) can be tested even in cases with a small number of cells, including oral squamous cell carcinoma (OSCC), and evaluation of the antitumor effect with a drug concentration close to the in vivo level is possible. The present report aimed to evaluate the utility of the CD-DST in the assessment of the in vitro efficacy of single-agent and multidrug combination chemotherapy for OSCC in comparison with the clinical response rates and to examine the possible clinical application of CD-DST for such cases. A total of 33 OSCC patients from whom 33 samples were obtained from January 2010 to September 2015 were included. CD-DST was performed, individually and in combination, on the three drugs [i.e., cisplatin (CDDP), 5-fluorouracil (5-FU), and docetaxel (DOC)] and on super selective intra-arterial infusion chemoradiotherapy (IACRT). The overall evaluable rate of the CD-DST in OSCC was 81.8% (27 of 33 cases) and the sensitivity to each anticancer drug was evaluated. The in vitro efficacy rates of IACRT, cisplatin + 5-fluorouracil, and docetaxel + cisplatin + 5-fluorouracil (TPF) confirmed the estimated clinical response rates. In 14 of 33 patients, the results of CD-DST were compared with clinical efficacy, which was judged based on measurable lesions on imaging. For TPF therapy, the sensitivity test of the IACRT had a positive predictive value of 90.9% (10 of 11 cases) and a negative predictive value of 100% (3 of 3 cases); the accuracy of the susceptibility test for the anticancer agents was 92.8% (13 of 14 cases). The CD-DST may be useful in selecting multidrug combination chemotherapy and IACRT for OSCC, however, accumulation of further clinical data is required in the future. IntroductionThe treatment of advanced or unresectable oral squamous cell carcinoma (OSCC) is multidisciplinary and entails the use of both multidrug combination chemotherapy and radiotherapy (1-3). The usefulness of super selective intraarterial infusion chemoradiotherapy (IACRT) to deliver high concentrations of anticancer drugs to tumors has been reported (4). The selection of such anticancer drugs for IACRT had been based on statistical information and clinical reports on a number of cases (1-3); however, the expected therapeutic effects of these anticancer drugs have not been consistent and adverse events were common. Therefore, drug sensitivity testing prior to administration of an anticancer agent is ideal to avoid the serious side effects of less effective anticancer drugs. Collagen gel droplet-embedded culture drug sensitivity test (CD-DST) uses an image colorimetric method to assess the combination of an anticancer drug and microcollagen gel embedded in a three-dimensional serum-free culture medium (3 to 10x10 3 cells/30 µg/drop) (5,6). It is a susceptibility test that is currently widely applied in various clinical fields, including gastrointestinal cancer (7-9). However, reports on the use of CD-DST in OSCC were few a...

Anticancer drug sensitivity testing using the collagen gel droplet embedded culture drug sensitivity test (CD-DST) on oral squamous cell carcinoma (OSCC) samples beginning from 2010 has been conducted. The present study investigated the effect of adding cetuximab (Erbitux ® ), a molecularly targeted drug, on anticancer drug activity against clinical OSCC specimens. A total of 25 specimens were obtained from 25 patients with OSCC between October 2013 and December 2017. The present study conducted anticancer drug sensitivity testing for cisplatin (CDDP), 5-fluorouracil (5-FU), cetuximab, three-drug combination, single agent and multi drug combinations, and cetuximab addition to the aforementioned regimens using CD-DST. In addition, the optimum concentration of each drug was evaluated. The overall evaluation success rate of the CD-DST method for OSCC specimens was 84.0% (21 of 25 cases); sensitivity to anticancer drugs and cetuximab could be evaluated. The in vitro efficacy rate of a cetuximab single agent and CDDP + 5-FU (PF) at a cut-off value of 50% was similar to the known clinical response rate. However, at a cut-off value of 50%, the in vitro efficacy of PF + cetuximab was calculated to be 40%, which was higher than the clinical response rate. The CD-DST method could be used to evaluate cetuximab, a molecularly targeted drug. Furthermore, its additive effect on conventional chemotherapy could be evaluated.The CD-DST method is suitable for evaluating and selecting chemotherapy regimens, including molecularly targeted drugs. Future studies are required to generate and evaluate relevant clinical data. Identification of the optimal cetuximab concentration that is effective against oral squamous cell carcinoma in collagen gel droplet embedded culture drug sensitivity testingAbbreviations: ADCC, antibody-dependent cell-mediated cytotoxicity; CG, collagen gel-coated; EGFR, epidermal growth factor receptor; OSCC, oral squamous cell carcinoma; SCCHN, squamous cell carcinoma of the head and neck Key words: anticancer drug sensitivity test, oral squamous cell carcinoma, collagen gel droplet embedded culture drug sensitivity test, cetuximab, anticancer drugs