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Background: In 2007, the International Agency for Research on Cancer (IARC) classified long-term shift work as a probable human carcinogen; however, the mechanism through which shift work potentially increases cancer risk is not known. One hypothesis is that diminished melatonin production may be involved, possibly as a result of exposure to light during night work. Experimental studies suggest a link between melatonin and peptides in the insulin-like growth factor (IGF) family, also implicated in carcinogenesis. This research aimed to describe the distributions of circulating concentrations of insulin-like growth factor-I (IGF-I) and insulin-like growth factor binding protein-3 (IGFBP-3) and their associations with urinary melatonin as possible intermediates in the pathway between work at night and breast cancer. Methods: A cross-sectional study was conducted among 85 premenopausal nurses working a rotating shift pattern of two 12-hour days, two 12-hour nights, and five days off. Once during both the summer and winter seasons, melatonin metabolites were measured in urine samples and circulating concentrations of IGF-I and IGFBP-3 were determined from serum samples. Weight and height were measured by the study coordinator, while a questionnaire and study diaries were used to collect all other covariate information. Predictors of IGF levels were identified using multivariate mixed effects modeling and relationships between melatonin and the IGFs were investigated using Spearman’s rank correlation and multivariate mixed effects modeling. Results: Both age (β = -3.6, p < 0.0001) and current OC use (β = -40.8, p = 0.003) were associated with decreases in circulating IGF-I, while levels of IGF-I were increased in the winter months (β = 26.3, p = 0.02). A positive relationship between recent alcohol consumption and serum IGFBP-3 was also suggested (β = 197.8, p = 0.05). Neither Spearman’s rank correlations nor mixed effects modeling indicated that urinary melatonin was a determinant of serum IGFs. Conclusions: Age, season, and current OC use were observed to predict circulating IGF-I, while recent alcohol consumption was a determinant of IGFBP-3 levels. A relationship between melatonin and IGFs, theorized as a component of the mechanism linking shift work and cancer, was not supported by the results of this project. / Thesis (Master, Community Health & Epidemiology) -- Queen's University, 2012-05-30 15:29:31.253

Recent observations have demonstrated that Insulin-like Growth Factors (IGFs) are able to form complexes with the extracellular matrix protein Vitronectin (VN). These complexes of VN:IGFBP:IGF-I significantly enhance the proliferation and migration of various cell lines including skin and corneal epithelial cells, as well as primary cells derived from human skin and corneal tissue. These enhanced effects arise from co- activation of the IGF-binding type-1 IGF receptor (IGF-1R) as well as activation of the VN-binding αv-integrins. Further studies suggest that these complexes can replace the requirement for serum in the ex vivo expansion of cells. In order to translate the VN:IGFBP:IGF-I technology into techniques for the improved culture of cells, we have designed, expressed and purified synthetic chimeric molecules, consisting of various domains of VN and mature IGF-I, using a baculovirus based expression system. The recombinant VN:IGF-I (rVN:IGF-I) chimeras were secreted into conditioned media of transfected Sf9 insect cells. Purification of the chimeras was achieved via methods including heparin-sepharose chromatography, Q-sepharose ion-exchange chromatography and Ni2+-NTA affinity chromatography. The rVN:IGF-I chimeras were detectable by Western blot analysis using a poly-clonal anti-VN antibody. Functional characterisation studies indicate that the chimeras promote cellular growth and migration to a similar extent as the VN:IGFBP:IGF-I complexes at 10x and 30x molar ratios. Additionally, function blocking antibodies directed to the IGF-1R and the VN binding αv-integrin were able to abolish this effect indicating that co-activation of these receptors is critical to the migratory effect of the chimeras. A functional chimera may lead to the development of cell culture techniques and methodologies that are devoid of xenogeneic or allogeneic support systems, thus paving the way to approved tissue engineering therapeutics that incorporate ex vivo expanded adult stem and progenitor cells.

Insulin-like growth factors (IGF) are known paracrine/autocrine regulators of ovarian development in teleosts. Initial studies investigated the hormonal and intracellular signal cascades involved in regulating the expression of ovarian-derived IGFs in zebrafish (Danio rerio). Quantitative real-time PCR was used to quantify the expression of igf3, igf2a, and igf2b in full grown immature (FG; 0.57-0.65 mm) and mid-vitellogenic (MV; 0.45-0.56 mm) follicles. Addition of the gonadotropin analogue human chorionic gonadotropin (hCG) and the adenylate cyclase activator forskolin increased igf3 expression in FG and MV follicles, but had no effect on igf2a or igf2b expression. The effects of hCG were blocked by the addition of the protein kinase A inhibitor H-89. Pituitary adenylate cyclase activating peptide stimulated a small increase in igf3 expression in FG follicles, while growth hormone and salmon gonadotropin releasing hormone had no effect on igf3, igf2a, or igf2b expression. Treatment with melittin, prostaglandin F2α, and prostaglandin E2 inhibited igf3 and igf2b expression in FG follicles whereas the protein kinase C activators, PMA and A23187, significantly inhibited igf3, igf2a, igf2b expression in FG and MV follicles. Secondary studies investigated the involvement of ovarian-derived IGFs in mediating the ovarian actions of gonadotropins on cell survival and steroidogenesis. Treatment of FG follicles with recombinant human IGF-I, hCG, or forskolin inhibited the induction of caspase-3/7 activity, which was used as a measure of apoptosis. The effects of hCG and forskolin on caspase-3/7 were attenuated by co-treatment with NVP-AEW54, an IGF-I receptor antagonist. hCG increased production of the maturation-inducing steroid 17α, 20β-dihydroxy-4-pregnen-3-one and co-treatment with NVP-AEW541 had no effect. These results suggest there is a high degree of hormonal specificity in regulating IGFs in the zebrafish ovary and the ovarian-derived IGFs, presumably IGF-III, are downstream mediators of gonadotropin-dependent cell survival, but are not involved in gonadotropin-induced steroidogenesis.

The IGF system is a crucial regulator of normal growth and development, however dysregulation of the system on multiple levels is associated with the incidence of a wide variety of malignancies including the breast, thyroid, lung, and colon, making the IGF system an important anti-cancer therapeutic target. Due to its role in mediating cellular proliferation, protection from apoptosis, and metastasis, traditional focus has been set on examining the role of the type 1 IGF receptor [IGF1R] in cancer. However there is mounting evidence to suggest the insulin receptor [IR] may also be involved in the potentiation and pathogenesis of cancers. The observation that IGF-II is overexpressed, compared to normal tissues, by cancers suggests signaling via target receptors by this ligand has important implications on cancer pathogenesis. Indeed, both the IGF1R and IR have been demonstrated to be up-regulated in a variety of malignancies. In regards to IR isoform, the IGF-II binding IR-A is preferentially expressed by a number of cancer cell types. Together with the observation that an autocrine proliferative loop exists between IGF-II and the IR-A in malignant thyrocytes and cultured breast cancer cells, suggests signaling via the IR-A may play a role in cancer cell growth and survival. However, very few studies on the IR-A have been conducted in cells co-expressing the IGF1R. This is mainly due to the difficulties associated with discrimination between signaling arising from IGF1R homodimers, IR-A homodimers, and IGF1R/IR-A hybrid receptors. It is not known how the IR-A interacts, and functions in conjunction with the other receptors of the IGF system to signal biologically relevant outcomes, especially in terms of anti-cancer therapeutics that aim to block and down-regulate the IGF1R. Current anti-cancer therapies targeting the IGF system have concentrated on blocking IGF signaling via the IGF1R, due mostly to the functional properties of the receptor, but also in part due to the metabolic consequences associated with blockade and inhibition of the IR. This individual targeting of the IGF1R potentially leaves a pathway by which IGF-II secreted by the tumour can circumvent current IGF1R based therapies. Consequently, this thesis investigated whether the IR-A could compensate for the targeted loss of the IGF1R and how the IR-A interacts with the IGF1R in cells co-expressing these two receptors. In addition, the individual ability of the IR isoforms to signal biological outcomes in response to IGF stimulation was assessed. The main experimental techniques used throughout this body of work included; assessment of protein expression and activation by Western blot, siRNA mediated gene silencing, and measures of cell proliferation, survival, and migration. The key areas of investigation included: 1. Investigation of the individual ability of the IR isoforms to signal biological outcomes in response to IGF stimulation 2. Identification of an appropriate cell line model in which to investigate the interactions between the IR-A and IGF1R 3. Optimisation of siRNA mediated knock-down of the IR-A and IGF1R in SW480 colorectal adenocarcinoma cells 4. Determination of the biological role of the IR-A in SW480 cells co-expressing the IGF1R The key findings from this work included: 1. The IR-A could not compensate for IGF1R depletion in SW480 cells 2. Dual silencing of the IR-A and IGF1R indicated signaling via the IGF1R was dominant to signaling via the IR-A in SW480 cells 3. Signaling via IR-A/IGF1R hybrid receptors may not be as potent as signaling via IGF1R homodimers 4. IGF-I at physiological concentrations can stimulate biological responses via both isoforms of the IR. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1337339 / Thesis (Ph.D.) -- University of Adelaide, School of Molecular and Biomedical Science, 2008

The IGF system is a crucial regulator of normal growth and development, however dysregulation of the system on multiple levels is associated with the incidence of a wide variety of malignancies including the breast, thyroid, lung, and colon, making the IGF system an important anti-cancer therapeutic target. Due to its role in mediating cellular proliferation, protection from apoptosis, and metastasis, traditional focus has been set on examining the role of the type 1 IGF receptor [IGF1R] in cancer. However there is mounting evidence to suggest the insulin receptor [IR] may also be involved in the potentiation and pathogenesis of cancers. The observation that IGF-II is overexpressed, compared to normal tissues, by cancers suggests signaling via target receptors by this ligand has important implications on cancer pathogenesis. Indeed, both the IGF1R and IR have been demonstrated to be up-regulated in a variety of malignancies. In regards to IR isoform, the IGF-II binding IR-A is preferentially expressed by a number of cancer cell types. Together with the observation that an autocrine proliferative loop exists between IGF-II and the IR-A in malignant thyrocytes and cultured breast cancer cells, suggests signaling via the IR-A may play a role in cancer cell growth and survival. However, very few studies on the IR-A have been conducted in cells co-expressing the IGF1R. This is mainly due to the difficulties associated with discrimination between signaling arising from IGF1R homodimers, IR-A homodimers, and IGF1R/IR-A hybrid receptors. It is not known how the IR-A interacts, and functions in conjunction with the other receptors of the IGF system to signal biologically relevant outcomes, especially in terms of anti-cancer therapeutics that aim to block and down-regulate the IGF1R. Current anti-cancer therapies targeting the IGF system have concentrated on blocking IGF signaling via the IGF1R, due mostly to the functional properties of the receptor, but also in part due to the metabolic consequences associated with blockade and inhibition of the IR. This individual targeting of the IGF1R potentially leaves a pathway by which IGF-II secreted by the tumour can circumvent current IGF1R based therapies. Consequently, this thesis investigated whether the IR-A could compensate for the targeted loss of the IGF1R and how the IR-A interacts with the IGF1R in cells co-expressing these two receptors. In addition, the individual ability of the IR isoforms to signal biological outcomes in response to IGF stimulation was assessed. The main experimental techniques used throughout this body of work included; assessment of protein expression and activation by Western blot, siRNA mediated gene silencing, and measures of cell proliferation, survival, and migration. The key areas of investigation included: 1. Investigation of the individual ability of the IR isoforms to signal biological outcomes in response to IGF stimulation 2. Identification of an appropriate cell line model in which to investigate the interactions between the IR-A and IGF1R 3. Optimisation of siRNA mediated knock-down of the IR-A and IGF1R in SW480 colorectal adenocarcinoma cells 4. Determination of the biological role of the IR-A in SW480 cells co-expressing the IGF1R The key findings from this work included: 1. The IR-A could not compensate for IGF1R depletion in SW480 cells 2. Dual silencing of the IR-A and IGF1R indicated signaling via the IGF1R was dominant to signaling via the IR-A in SW480 cells 3. Signaling via IR-A/IGF1R hybrid receptors may not be as potent as signaling via IGF1R homodimers 4. IGF-I at physiological concentrations can stimulate biological responses via both isoforms of the IR. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1337339 / Thesis (Ph.D.) -- University of Adelaide, School of Molecular and Biomedical Science, 2008

The IGF system is a crucial regulator of normal growth and development, however dysregulation of the system on multiple levels is associated with the incidence of a wide variety of malignancies including the breast, thyroid, lung, and colon, making the IGF system an important anti-cancer therapeutic target. Due to its role in mediating cellular proliferation, protection from apoptosis, and metastasis, traditional focus has been set on examining the role of the type 1 IGF receptor [IGF1R] in cancer. However there is mounting evidence to suggest the insulin receptor [IR] may also be involved in the potentiation and pathogenesis of cancers. The observation that IGF-II is overexpressed, compared to normal tissues, by cancers suggests signaling via target receptors by this ligand has important implications on cancer pathogenesis. Indeed, both the IGF1R and IR have been demonstrated to be up-regulated in a variety of malignancies. In regards to IR isoform, the IGF-II binding IR-A is preferentially expressed by a number of cancer cell types. Together with the observation that an autocrine proliferative loop exists between IGF-II and the IR-A in malignant thyrocytes and cultured breast cancer cells, suggests signaling via the IR-A may play a role in cancer cell growth and survival. However, very few studies on the IR-A have been conducted in cells co-expressing the IGF1R. This is mainly due to the difficulties associated with discrimination between signaling arising from IGF1R homodimers, IR-A homodimers, and IGF1R/IR-A hybrid receptors. It is not known how the IR-A interacts, and functions in conjunction with the other receptors of the IGF system to signal biologically relevant outcomes, especially in terms of anti-cancer therapeutics that aim to block and down-regulate the IGF1R. Current anti-cancer therapies targeting the IGF system have concentrated on blocking IGF signaling via the IGF1R, due mostly to the functional properties of the receptor, but also in part due to the metabolic consequences associated with blockade and inhibition of the IR. This individual targeting of the IGF1R potentially leaves a pathway by which IGF-II secreted by the tumour can circumvent current IGF1R based therapies. Consequently, this thesis investigated whether the IR-A could compensate for the targeted loss of the IGF1R and how the IR-A interacts with the IGF1R in cells co-expressing these two receptors. In addition, the individual ability of the IR isoforms to signal biological outcomes in response to IGF stimulation was assessed. The main experimental techniques used throughout this body of work included; assessment of protein expression and activation by Western blot, siRNA mediated gene silencing, and measures of cell proliferation, survival, and migration. The key areas of investigation included: 1. Investigation of the individual ability of the IR isoforms to signal biological outcomes in response to IGF stimulation 2. Identification of an appropriate cell line model in which to investigate the interactions between the IR-A and IGF1R 3. Optimisation of siRNA mediated knock-down of the IR-A and IGF1R in SW480 colorectal adenocarcinoma cells 4. Determination of the biological role of the IR-A in SW480 cells co-expressing the IGF1R The key findings from this work included: 1. The IR-A could not compensate for IGF1R depletion in SW480 cells 2. Dual silencing of the IR-A and IGF1R indicated signaling via the IGF1R was dominant to signaling via the IR-A in SW480 cells 3. Signaling via IR-A/IGF1R hybrid receptors may not be as potent as signaling via IGF1R homodimers 4. IGF-I at physiological concentrations can stimulate biological responses via both isoforms of the IR. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1337339 / Thesis (Ph.D.) -- University of Adelaide, School of Molecular and Biomedical Science, 2008

The IGF system is a crucial regulator of normal growth and development, however dysregulation of the system on multiple levels is associated with the incidence of a wide variety of malignancies including the breast, thyroid, lung, and colon, making the IGF system an important anti-cancer therapeutic target. Due to its role in mediating cellular proliferation, protection from apoptosis, and metastasis, traditional focus has been set on examining the role of the type 1 IGF receptor [IGF1R] in cancer. However there is mounting evidence to suggest the insulin receptor [IR] may also be involved in the potentiation and pathogenesis of cancers. The observation that IGF-II is overexpressed, compared to normal tissues, by cancers suggests signaling via target receptors by this ligand has important implications on cancer pathogenesis. Indeed, both the IGF1R and IR have been demonstrated to be up-regulated in a variety of malignancies. In regards to IR isoform, the IGF-II binding IR-A is preferentially expressed by a number of cancer cell types. Together with the observation that an autocrine proliferative loop exists between IGF-II and the IR-A in malignant thyrocytes and cultured breast cancer cells, suggests signaling via the IR-A may play a role in cancer cell growth and survival. However, very few studies on the IR-A have been conducted in cells co-expressing the IGF1R. This is mainly due to the difficulties associated with discrimination between signaling arising from IGF1R homodimers, IR-A homodimers, and IGF1R/IR-A hybrid receptors. It is not known how the IR-A interacts, and functions in conjunction with the other receptors of the IGF system to signal biologically relevant outcomes, especially in terms of anti-cancer therapeutics that aim to block and down-regulate the IGF1R. Current anti-cancer therapies targeting the IGF system have concentrated on blocking IGF signaling via the IGF1R, due mostly to the functional properties of the receptor, but also in part due to the metabolic consequences associated with blockade and inhibition of the IR. This individual targeting of the IGF1R potentially leaves a pathway by which IGF-II secreted by the tumour can circumvent current IGF1R based therapies. Consequently, this thesis investigated whether the IR-A could compensate for the targeted loss of the IGF1R and how the IR-A interacts with the IGF1R in cells co-expressing these two receptors. In addition, the individual ability of the IR isoforms to signal biological outcomes in response to IGF stimulation was assessed. The main experimental techniques used throughout this body of work included; assessment of protein expression and activation by Western blot, siRNA mediated gene silencing, and measures of cell proliferation, survival, and migration. The key areas of investigation included: 1. Investigation of the individual ability of the IR isoforms to signal biological outcomes in response to IGF stimulation 2. Identification of an appropriate cell line model in which to investigate the interactions between the IR-A and IGF1R 3. Optimisation of siRNA mediated knock-down of the IR-A and IGF1R in SW480 colorectal adenocarcinoma cells 4. Determination of the biological role of the IR-A in SW480 cells co-expressing the IGF1R The key findings from this work included: 1. The IR-A could not compensate for IGF1R depletion in SW480 cells 2. Dual silencing of the IR-A and IGF1R indicated signaling via the IGF1R was dominant to signaling via the IR-A in SW480 cells 3. Signaling via IR-A/IGF1R hybrid receptors may not be as potent as signaling via IGF1R homodimers 4. IGF-I at physiological concentrations can stimulate biological responses via both isoforms of the IR. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1337339 / Thesis (Ph.D.) -- University of Adelaide, School of Molecular and Biomedical Science, 2008