ABSTRACT
BACKGROUND: Cancer chemotherapeutic treatment is a complex scientific task. The ChemoFx Drug Response Marker (DRM) assists physicians in identifying treatment protocols likely to be effective for specific patients. MATERIALS AND METHODS: The ChemoFx DRM was used to study drug response in vitro. Established human cancer cell lines and primary cultures of patient tumor specimens were challenged with chemotherapeutic agents to observe response of multiple tumor samples and determine whether drugs with similar mechanisms of action elicit similar response. RESULTS: These studies demonstrated heterogeneous response among patient tumor samples and clustering of drug response with similar mechanisms of action. Also highlighted was the reproducibility of ChemoFx DRM and its utility in characterizing tumor response to chemotherapy. CONCLUSION: Heterogeneous drug responses observed in vitro were similar to those observed clinically. Response characteristics were similar for drugs with similar mechanisms of action, suggesting response heterogeneity is determined at a cellular and molecular level.
Subject(s)
Antineoplastic Agents/pharmacology , Drug Screening Assays, Antitumor/methods , Neoplasms/drug therapy , Anthracyclines/pharmacology , Antineoplastic Agents/classification , Breast Neoplasms/drug therapy , Breast Neoplasms/pathology , Bridged-Ring Compounds/pharmacology , Cell Line, Tumor , Colonic Neoplasms/drug therapy , Colonic Neoplasms/pathology , Female , Humans , Lung Neoplasms/drug therapy , Lung Neoplasms/pathology , Neoplasms/pathology , Ovarian Neoplasms/drug therapy , Ovarian Neoplasms/pathology , Platinum Compounds/pharmacology , Reproducibility of Results , Taxoids/pharmacologyABSTRACT
BACKGROUND: A localized hypoxic environment occurs during tumor growth necessitating an angiogenic response or tumor necrosis results. Novel cancer treatment strategies take advantage of tumor-induced vascularisation by combining standard chemotherapeutic agents with angiogenesis-inhibiting agents. This has extended the progression-free interval and prolonged survival in patients with various types of cancer. We postulated that the expression levels of angiogenesis-related proteins from various primary tumor cultures would be greater under hypoxic conditions than under normoxia. METHODS: Fifty cell sources, including both immortalized cell lines and primary carcinoma cells, were incubated under normoxic conditions for 48 hours. Then, cells were either transferred to a hypoxic environment (1% O2) or maintained at normoxic conditions for an additional 48 hours. Cell culture media from both conditions was collected and analyzed via an ELISA-based assay to determine expression levels of 11 angiogenesis-related factors: VEGF, PDGF-AA, PDGF-AA/BB, IL-8, bFGF/FGF-2, EGF, IP-10/CXCL10, Flt-3 ligand, TGF-beta1, TGF-beta2, and TGF-beta3. RESULTS: A linear correlation between normoxic and hypoxic growth conditions exists for expression levels of eight of eleven angiogenesis-related proteins tested including: VEGF, IL-8, PDGF-AA, PDGF-AA/BB, TGF-beta1, TGF-beta2, EGF, and IP-10. For VEGF, the target of current therapies, this correlation between hypoxia and higher cytokine levels was greater in primary breast and lung carcinoma cells than in ovarian carcinoma cells or tumor cell lines. Of interest, patient cell isolates differed in the precise pattern of elevated cytokines. CONCLUSION: As linear correlations exist between expression levels of angiogenic factors under normoxic and hypoxic conditions in vitro, we propose that explanted primary cells may be used to probe the in vivo hypoxic environment. Furthermore, differential expression levels for each sample across all proteins examined suggests it may be possible to build a predictor for angiogenesis-related anticancer agents, as each sample has a unique expression profile. Further studies should be performed to correlate in vitro protein expression levels of angiogenesis-related factors with in vivo patient response.
ABSTRACT
AIMS: Establishing and maintaining human tumours in primary culture can be challenging. In this application, a short-term primary culture process is desired to ensure cells maintained in culture are representative of the in vivo tumour for the purpose of chemoresponse testing. To ensure the appropriate cells are being grown, the cultures must be evaluated for malignancy. The clinical gold standard determination of malignancy is cytological evaluation by a cytopathologist. METHODS: Fifty human tumour specimens (breast, colon, lung, ovary) were established and maintained in primary culture. Cytospins were prepared upon initiation of culture and again at completion of the culture process. Cytospins were stained (Diff-Quik, Papanicolaou) and evaluated by a cytopathologist for the percentage of malignant cells at both times. RESULTS: An increase in the percentage of malignant cells was noted in 86% (43/50) of the cultures evaluated; 8% (4/50) of the cultures maintained the same percentage of malignant cells throughout the culture period, and 6% (3/50) displayed a decrease in malignant cells. On average, the percentage of malignant cells increased by 37% and was not associated with the length of culture (range 5-28 days). CONCLUSIONS: The described primary culture process enriches for malignant cells, which is desirable for further evaluation such as chemoresponse testing.
