Antifolate pseudo-resistance due to elevated levels of thymidine and hypoxanthine in a commercial serum preparation.

BACKGROUND: Batch variability of sera used for cell culture is of considerable experimental concern. A novel fetal calf serum product, FCS Gold, was claimed to be the first defined fetal calf serum free of batch variation. MATERIALS AND METHODS: The efficacy of methotrexate (MTX) and LY231514 (multitargeted antifolate, MTA) in CCRF-CEM cells and KB cells was compared using media supplemented with FCS Gold or conventional fetal bovine serum. RESULTS: IC50 values from tests using conventional serum corresponded to published data. FCS Gold fully protected the cells from antifolate drug cytotoxicity. Dialysis of FCS Gold restored responsiveness to antifolate drugs. Elevated levels of hypoxanthine and thymidine were present in FCS Gold. They were approximately 10-fold greater than the concentrations required to overcome growth arrest mediated by 2 microM MTX. CONCLUSION: FCS Gold or identical products, e.g. FBS Gold, should not be used in studies on antifolate drug action.

Ex vivo responsiveness of head and neck squamous cell carcinoma to vinorelbine.

BACKGROUND: Vinorelbine has been identified as an effective cytostatic drug in head and neck squamous cell carcinoma (HNSCC). To predict the individual chemoresponse, the application of an ex vivo assay to quantify the response of HNSCC to vinorelbine is presented. PATIENTS AND METHODS: Twelve biopsies from primary HNSCC sites and five biopsies from metastatic lesions were analyzed (n = 17). The specimens were investigated ex vivo for the overall, epithelial and stromal chemoresponse to vinorelbine. RESULTS: By selective evaluation of the epithelial chemoresponse, the applied assay identified three specimens as vinorelbine-sensitive (18%), including one de novo sensitivity in a metastatic lesion of a vinorelbine-resistant hypopharyngeal carcinoma. CONCLUSION: Applying flavin-protecting culture methods and with careful correction for the stromal cell effect, the assay generated reliable data for the assessment of the individual chemoresponse of HNSCC specimens to vinorelbine. The assay may, therefore, facilitate the implementation of individualized chemotherapy protocols.

Ex vivo chemosensitivity of head and neck carcinoma to cytostatic drug combinations.

Previous studies focusing on response prediction to chemotherapy by chemosensitivity testing of tumor explants has focused on the response determination of single cytostatic drugs, in contrast to the common clinical application of cytostatic drug combinations. Therefore, the present study was aimed at determining the quantitative ex vivo chemoreactivity of epithelial cells from head and neck squamous cell carcinoma (HNSCC) specimens to cytostatic drug combinations. Specimens from 12 histologically-confirmed HNSCC were investigated. According to a previously established ex vivo colony formation assay, the individual cellular chemoreactivity was determined quantitatively for combinations of 4 cytostatic drugs: cis-platinum (cis-DDP), carboplatin (CBDCA), 5-Fluorouracil (5-FU) and docetaxel (DTX). The tests were performed using drug combinations according to recent clinical therapy regimens in the treatment of solid tumors: i) cis-DDP + 5FU, ii) CBDCA + 5FU, iii) cis-DDP + DTX and iv) CBDCA + DTX. The approach provides individual drug response patterns of epithelial as well as of stromal cells. Individual, selective sensitivities were found for each drug combination tested. The stromal and epithelial chemoreactivity profiles differed in most of the specimens. Moreover, stromal cell chemoresistance dominated selective epithelial chemosensitivities in the majority of cases. The determination of the epithelial ex vivo chemoreactivity identified individual chemosensitivities which were verified by the clinical history of the patient. Therefore, the described protocol to determine the ex vivo chemoresponse of HNSCC specimens to cytostatic drug combinations may help to provide clinicaly useful information concerning the individual chemoresponse of HNSCC with regard to individualization of oncological decision making.

Cytotoxicity determination without photochemical artifacts.

A study was performed to improve cytotoxicity determinations by eliminating flavin-mediated photosensitization from tests with KB cells, NCI-H69 cells, P-glycoprotein expressing KBC5-8 cells, MRP1-expressing H69AR cells, and A240286S human lung adenocarcinoma cells. Growth inhibition by cis-platin, doxorubicin, etoposide, gemcitabine, taxol, vincristine, vinblastine, and vinorelbine was determined under flavin-protecting conditions using flavin-free culture media with fetal bovine serum as the only source of flavins. As compared to conventional tests, the IC50 values determined under flavin-protecting conditions reflected increased apparent drug cytotoxicities, and were flawlessly reproducible. Flavin-mediated photosensitization should, therefore, be strictly eliminated from in vitro experiments involving cytotoxic and other drugs.

Ex vivo responsiveness of head and neck squamous cell carcinoma to glufosfamide, a novel alkylating agent.

BACKGROUND: Glufosfamide is a novel alkylating agent in which the active metabolite of isophosphoramide mustard is glycosidically linked to beta-D-glucose. Targeting the elevated glucose uptake of tumor cells expressing the SAAT1 glucose transporter, glufosfamide represents an attractive new drug for cancer chemotherapy. The present study investigates the ex vivo responsiveness of Head and Neck Squamous Cell Carcinoma (HNSCC) specimens to glufosfamide. PATIENTS AND METHODS: Twenty-one unselected HNSCC specimens were investigated using a novel ex vivo colony formation assay to determine the epithelial drug response. The individual responsiveness to glufosfamide and to cis-platinum was determined. RESULTS: Five out of 21 evaluable HNSCC specimens were sensitive to glufosfamide. There was a tendency for glufosfamide sensitivity in platinum-resistant specimens and vice versa. CONCLUSION: The effectiveness of glufosfamide observed in the present ex vivo study suggests at least an equipotentiality of glufosfamide in comparison to cis-platinum. The potential clinical usefulness of glufosfamide in HNSCC warrants further evaluation.

Synthesis of new boron-rich building blocks for boron neutron capture therapy or energy-filtering transmission electron microscopy.

The synthesis of a new ortho-carborane derivative, tetracarboranylketone 4, is reported here. Ketone 4 was prepared from a tetraalkynylated ketone by the addition of decaborane. The keto group was then easily modified to yield the glycosides 17alpha and 18beta, which contain glucose or galactose, respectively, and the nucleotide 13b. In addition to ketone 4, which is acyclic, cyclic ketone 8 was also synthesised. X-ray diffraction analysis of compound 4 indicated the presence of two toluene guest molecules per molecule of the host compound. Furthermore, compound 4 displays a rather low cytotoxicity. These novel products can be used as building blocks to create a new class of biomolecules containing high-density carborane clusters. Such molecules may constitute powerful tools for applications like Boron Neutron Capture Therapy or Energy-Filtering Transmission Electron Microscopy.

The impact of stromal cell contamination on chemosensitivity testing of head and neck carcinoma.

BACKGROUND: Reliable chemosensitivity testing of head and neck squamous cell carcinoma (HNSCC) still faces methodical limitations. Since stromal cell contamination has been found to preclude reliable radiosensitivity testing of HNSCC as well as chemosensitivity testing of lung tumors, the present study investigates the impact of stromal cell contamination on chemosensitivity testing of HNSCC. PATIENTS AND METHODS: Seventeen biopsies from HNSCC were analyzed. The specimens were investigated using an ex vivo colony formation assay which allows for the quantitative and separate determination of the overall, as well as the epithelial, and stromal response to carboplatin, 5-fluorouracil and docetaxel. RESULTS: The overall chemoresponse was dominated by stromal cell multidrug resistance. However, by selective evaluation of the epithelial chemoresponse, individual chemosensitivity patterns could be identified. CONCLUSION: Multidrug-resistant stromal cells preclude the reliable assessment of the chemoresponse of HNSCC specimens. Carefiul correction for stromal cell effects is a prerequisite for the generation of therapeutically useful information.