Comparison of angiogenesis-related factor expression in primary tumor cultures under normal and hypoxic growth conditions

Abstract

BackgroundA 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.MethodsFifty 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-β1, TGF-β2, and TGF-β3.ResultsA 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-β1, TGF-β2, 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.ConclusionAs 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.

Angiogenesis and ovarian cancer

“…A linear correlation between normoxic and hypoxic growth conditions exists for expression levels of angiogenesis-related proteins: VEGF, IL-8, PDGF-AA, PDGF-AA/ BB, TGF-ß1, TGF-ß2, EGF and IP-10 [64]. As is known, VEGF-A may be stimulated by tumour-associated hypoxic conditions via HIF1-α.…”

Molecular targets and gene therapy of lung cancer

“…Specific CD44 isoforms may also be associated with lung cancer metastasis. A correlation between upregulated VEGF and progression of disease in NSCLC has been shown (22,23). A number of agents attacking either the protein or the receptor of VEGF have been developed, including bevacizumab (Avastin ® ), which is a human monoclonal antibody against VEGF that has shown increased time-to-progression in NSCLC when combined with chemotherapy in phase II trials (24).…”

Inhibition of aldose reductase prevents angiogenesis in vitro and in vivo

“…Angiogenesis in tumors allows the growth and invasiveness of tumor cells leading to metastasis to distant organs (2–4). Angiogenesis, especially in a solid tumor progression plays a critical role because when tumor mass increases beyond 2–3 mm, formation of new blood vessels is presumably essential in supplying nutrients and oxygen and removing waste products (1–3).…”

“…Angiogenesis, especially in a solid tumor progression plays a critical role because when tumor mass increases beyond 2–3 mm, formation of new blood vessels is presumably essential in supplying nutrients and oxygen and removing waste products (1–3). In angiogenesis the interaction of tumor cells with vascular endothelial cells leads to the release of various growth factors, cytokines, chemokines, etc (4,5). These molecules in turn by autocrine and paracrine fashion activate inflammatory signaling leading to differentiation of vascular endothelial cells.…”

“…These molecules in turn by autocrine and paracrine fashion activate inflammatory signaling leading to differentiation of vascular endothelial cells. In addition, the important concept of tumor angiogenesis is that in the tumor microenvironment tumor-associated endothelial cells are genetically unstable and show enhanced responsiveness to growth factors such as VEGF, FGF and EGF (4–6). Therefore, antiangiogenic therapy represents one of the most promising approaches to regulate tumor growth and metastasis.…”