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ABSTRACT: The mitogenic and anti-apoptotic effects of insulin-like growth factor-I (IGF-I) are regulated by a family of insulin-like growth factor binding proteins (IGFBPs), particularly IGFBP-3. Little is known about the IGF-independent role of IGFBP-3 in breast cancer and the mechanisms regulating its production. The expression of IGFBP-3 in paired malignant and adjacent normal (n=53), and healthy normal (n=17) breast tissue samples was investigated using RT-PCR, immunohistochemistry and ELISA. We compared IGFBP-3 expression with other members of the IGF-I axis, other known tumorigenic genes and clinicopathological parameters. We also developed a novel tissue explant system using fresh normal and malignant breast tissue, with which we examined the in vitro effects of IGFBP-3 alone and in combination with known apoptotic agent, doxorubicin (n=6), on tissue viability and apoptosis. We demonstrated a high level of expression of IGFBP-3 mRNA in all samples. 96% of samples also expressed IGFBP-3 protein. No significant correlation was seen between IGFBP-3 expression and other clinicopathological parameters. The in vitro tissue explant system demonstrated that IGFBP-3 had little effect by itself on apoptosis. However, when used in combination with doxorubicin, increased apoptosis was seen in tumours. In contrast, less apoptosis was seen in normal tissue suggesting a protective effect. These divergent effects suggest a potential novel chemotherapeutic approach in the treatment of breast cancer. These findings suggest that IGFBP-3 may play a role in tumorigenesis, and that IGFBP-3 levels could be used in the future in cancer risk assessment/prevention or as markers of response to cancer treatments.
Anticancer research 10/2009; 29(10):3785-90. · 1.73 Impact Factor
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ABSTRACT: The purpose of this study was to evaluate COX-2 mRNA expression with known clinical prognostic features of breast cancer, oestrogen/progesterone receptor status, tumour size and grade.
Total RNA was extracted from 45 frozen breast tumour (invasive) and 22 normal breast tissue samples. COX-2 mRNA transcription was quantified using a real time RT-PCR assay and expressed as copy number/microg total RNA. All specimens were assessed for tumour grade, size, nodal status and presence of vascular invasion and oestrogen and progesterone receptor status.
COX-2 mRNA was detected in all samples with a median copy number of 1.15 x 10(7) for tumours and 6.5 x 10(6) for normal samples. Expression was significantly higher in oestrogen receptor negative tumours compared to the receptor positive group. There was no correlation between COX-2 mRNA levels and tumour size, grade, nodal status and presence of vascular invasion.
COX-2 mRNA expression is increased in oestrogen and progesterone receptor negative breast cancers.
European Journal of Surgical Oncology 10/2006; 32(7):707-9. · 2.50 Impact Factor
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ABSTRACT: IGF-binding protein-3 (IGFBP-3) has been reported to exert a protective influence on the pathogenesis of colorectal cancer. This may reflect its modulation of IGF-I bioactivity as well as IGF-I-independent effects on cell proliferation and apoptosis. Although local expression of IGF-I in the colon is increasingly recognised as having important regulatory consequences, the role of locally expressed IGFBP-3 remains unknown. The aims of the present study were: (i) to quantify and localise the expression of IGFBP-3 in human normal and malignant colon; (ii) to relate this expression to that of other components of the IGF-I axis; and (iii) to investigate the effects of IGFBP-3 on colonic epithelial cell proliferation and apoptosis. RNA was extracted from 46 paired samples of normal and malignant colonic tissue. IGFBP-3, IGF-I, IGF-I receptor and GH receptor mRNA levels were quantified using real-time RT-PCR. Laser-capture microdissection of the same samples was used to isolate mRNA from epithelium and stromal components and localise mRNA expression. Expression was confirmed at a protein level by immunohistochemistry. Human colorectal cancer HT-29 and CaCo-2 cells were cultured with IGFBP-3 (200 ng/ml), +/- IGF-I (20 ng/ml), +/- sodium butyrate (5 mM). Cell number was assessed by an MTS assay (a modification of the MTT assay), and apoptosis assessed by cell morphology and FACS analysis using both annexin and propidium iodide staining. UO146, a MAP kinase inhibitor, and wortmannin, an inhibitor of the phosphatidylinositol 3-kinase (PI-3K) pathway, were used to determine the contribution of these signalling pathways on the effects of IGFBP-3. IGFBP-3 mRNA was detected in all samples (mean copy number/mug total RNA in normal colon, 2.6 x 10(6) compared with 1.3 x 10(7) in the cancers; P < 0.0001). Immunohistochemistry confirmed the expression and showed it to be equally distributed between epithelial and stromal components in normal tissue, but to be mainly restricted to the stromal component of malignant tissue. This differential expression was confirmed by RT-PCR of RNA from laser-capture microdissected samples. IGF-I mRNA was detected in 31 samples of normal colon; mean IGFBP-3 copy number was higher in the IGF-I-positive samples compared with IGF-I-negative samples. IGFBP-3 on its own induced apoptosis in HT-29 cells (P < 0.001). Co-incubation of 200 ng/ml IGFBP-3 with butyrate (5 mM) resulted in the potentiation of its apoptosis (P < 0.0001), which was not rescued by co-incubation with IGF-I (P < 0.0001). The addition of UO126 caused a decrease in cell number and increased the effects of IGFBP-3. IGFBP-3 is differentially expressed between stromal and epithelial components of normal and malignant colon, which may reflect its pro-apoptotic, IGF-I-independent effect on colonic epithelial cells. These effects are mediated in part by the PI-3K pathway in contrast to the MAP kinase pathway used by IGF-I.
Endocrine Related Cancer 12/2005; 12(4):891-901. · 4.36 Impact Factor
