Insulin Receptor Substrate-1, p70S6K and Cell Size in Transformation and Differentiation of Hemopoietic Cells.

After an initial burst of cell proliferation, the type 1 insulin-like growth factor receptor (IGF-IR) induces granulocytic differentiation of 32D IGF-IR cells, an interleukin-3-dependent murine hemopoietic cell line devoid of insulin receptor substrate-1 (IRS-1). The combined expression of the IGF-IR and IRS-1 (32D IGF-IR/IRS-1 cells) inhibits IGF-I-mediated differentiation, and causes malignant transformation of 32D cells. Because of the role of IRS-1 in changing the fate of 32D IGF-IR cells from differentiation (and subsequent cell death) to malignant transformation, we have looked for differences in IGF-IR signaling between 32D IGF-IR and 32D IGF-IR/IRS-1 cells. In this report, we have focused on p70(S6K), which is activated by the IRS-1 pathway. We find that the ectopic expression of IRS-1 and the inhibition of differentiation correlated with a sustained activation of p70(S6K) and an increase in cell size. Phosphorylation in vivo of threonine 389 and, to a lesser extent, of threonine 421/serine 424 of p70(S6K) seemed to be a requirement for inhibition of differentiation. A role of IRS-1 and p70(S6K) in the alternative between transformation or differentiation of 32D IGF-IR cells was confirmed by findings that inhibition of p70(S6K) activation or IRS-1 signaling, by rapamycin or okadaic acid, induced differentiation of 32D IGF-IR/IRS-1 cells. We have also found that the expression of myeloperoxidase mRNA (a marker of differentiation, which sharply increases in 32D IGF-IR cells), does not increase in 32D IGF-IR/IRS-1 cells, suggesting that the expression of IRS-1 in 32D IGF-IR cells causes the extinction of the differentiation program initiated by the IGF-IR, while leaving intact its proliferation program

The baculovirus anti-apoptotic p35 protein promotes transformation of mouse embryo fibroblasts.

The baculovirus p35 protein is a potent inhibitor of programmed cell death induced by a variety of stimuli in insects, nematodes, and mammalian cell lines. The broad ability of p35 in preventing apoptosis has led us to investigate its effect on mouse embryo fibroblasts in vitro and in vivo. For this purpose, we have used R- cells (3T3-like fibroblasts derived from mouse embryos with a targeted disruption of the insulin-like growth factor I receptor (IGF-IR) genes) and R508 cells (derived from R- and with 15 x 10(3) IGF-IRs per cell). Both cell lines grow normally in monolayer, but they do not form colonies in soft agar, and they are non-tumorigenic in nude mice. We show here that, in addition to its anti-apoptotic effect, p35 causes transformation of R508 cells, as evidenced by the following: 1) decreased growth factor requirements, 2) ability to form foci in monolayer and colonies in soft agar, and 3) ability to form tumors in nude mice. Since R- cells stably transfected with p35 do not transform, our observations suggest that in addition to its effect as an inhibitor of apoptosis, the baculovirus p35 protein has transforming potential that requires the presence of the IGF-IR. The possibility that these two properties could be separated was confirmed by demonstrating that R508 cells expressing another anti-apoptotic protein, Bcl-2, could not form tumors in nude mice

Long-Term IGF-I Exposure Decreases Autophagy and Cell Viability

A reduction in IGF-I signaling has been found to increase lifespan in multiple organisms despite the fact that IGF-I is a trophic factor for many cell types and has been found to have protective effects against multiple forms of damage in acute settings. The increase in longevity seen in response to reduced IGF-I signaling suggests that there may be differences between the acute and chronic impact of IGF-I signaling. We have examined the possibility that long-term stimulation with IGF-I may have a negative impact at the cellular level using quiescent human fibroblasts. We find that fibroblast cells exposed to IGF-I for 14 days have reduced long-term viability as judged by colony forming assays, which is accompanied by an accumulation of senescent cells. In addition we observe an accumulation of cells with depolarized mitochondria and a reduction in autophagy in the long-term IGF-I treated cultures. An examination of mice with reduced IGF-I levels reveals evidence of enhanced autophagy and fibroblast cells derived from these mice have a larger mitochondrial mass relative to controls indicating that changes in mitochondrial turnover occurs in animals with reduced IGF-I. The results indicate that chronic IGF-I stimulation leads to mitochondrial dysfunction and reduced cell viability

Biomarkers and outcome after tamoxifen treatment in node-positive breast cancers from elderly women

The predictive role of tumour proliferative rate and expression of p53, bcl-2 and bax proteins, alone and in association with tumour size, nodal involvement and oestrogen receptors (ER), was analysed on 145 elderly patients (≥70 years of age) with histologically assessed node-positive breast cancers treated with radical or conservative surgery plus radiotherapy followed by adjuvant tamoxifen for at least 1 year. The 7-year probability of relapse was significantly higher for patients with tumours rapidly proliferating (hazard ratio (HR) = 2.0, P = 0.01), overexpressing p53 (HR = 4.4, P = 0.0001), weakly or not exhibiting bcl-2 (HR = 1.9, P = 0.02), without ERs (HR = 3.4, P = 0.0001) or with ≥ 4 positive lymph nodes (HR = 2.3, P = 0.003) than for patients with tumours expressing the opposite patho-biological profile. Conversely, tumour size and bax expression failed to influence relapse-free survival. Adjustment for the duration of tamoxifen treatment did not change these findings. Oestrogen receptors, cell proliferation, p53 accumulation and bcl-2 expression were also predictive for overall survival. Within ER-positive tumours, cell proliferation, p53 accumulation, bcl-2 expression and lymph node involvement provided significant and independent information for relapse and, in association, identified subgroups of patients with relapse probabilities of 20% (low-risk group, exhibiting only one unfavourable factor) to 90% (high-risk group, exhibiting three unfavourable factors). Such data could represent the initial framework for a biologically tailored therapy even for elderly patients and highlight the importance of a patho-biological characterization of their breast cancers. © 2000 Cancer Research Campaig

IGFBP-rP1, a potential molecule associated with colon cancer differentiation

<p>Abstract</p> <p>Background</p> <p>In our previous studies, we have demonstrated that insulin-like growth factor binding protein-related protein1 (IGFBP-rP1) played its potential tumor suppressor role in colon cancer cells through apoptosis and senescence induction. In this study, we will further uncover the role of IGFBP-rP1 in colon cancer differentiation and a possible mechanism by revealing responsible genes.</p> <p>Results</p> <p>In normal colon epithelium, immunohistochemistry staining detected a gradient IGFBP-rP1 expression along the axis of the crypt. IGFBP-rP1 strongly expressed in the differentiated cells at the surface of the colon epithelium, while weakly expressed at the crypt base. In colon cancer tissues, the expression of IGFBP-rP1 correlated positively with the differentiation status. IGFBP-rP1 strongly expressed in low grade colorectal carcinoma and weakly expressed in high grade colorectal carcinoma. In vitro, transfection of PcDNA3.1(IGFBP-rP1) into RKO, SW620 and CW2 cells induced a more pronounced anterior-posterior polarity morphology, accompanied by upregulation with alkaline phosphatase (AKP) activity. Upregulation of carcino-embryonic antigen (CEA) was also observed in SW620 and CW2 transfectants. The addition of IGFBP-rP1 protein into the medium could mimic most but not all effects of IGFBP-rP1 cDNA transfection. Seventy-eight reproducibly differentially expressed genes were detected in PcDNA3.1(IGFBP-rP1)-RKO transfectants, using Affymetrix 133 plus 2.0 expression chip platform. Directed Acyclic Graph (DAG) of the enriched GO categories demonstrated that differential expression of the enzyme regulator activity genes together with cytoskeleton and actin binding genes were significant. IGFBP-rP1 could upreguate Transgelin (TAGLN), downregulate SRY (sex determining region Y)-box 9(campomelic dysplasia, autosomal sex-reversal) (SOX9), insulin receptor substrate 1(IRS1), cyclin-dependent kinase inhibitor 2B (p15, inhibits CDK4) (CDKN2B), amphiregulin(schwannoma-derived growth factor) (AREG) and immediate early response 5-like(IER5L) in RKO, SW620 and CW2 colon cancer cells, verified by Real time Reverse Transcription Polymerase Chain Reaction (rtRT-PCR). During sodium butyrate-induced Caco2 cell differentiation, IGFBP-rP1 was upregulated and the expression showed significant correlation with the AKP activity. The downregulation of IRS1 and SOX9 were also induced by sodium butyrate.</p> <p>Conclusion</p> <p>IGFBP-rP1 was a potential key molecule associated with colon cancer differentiation. Downregulation of IRS1 and SOX9 may the possible key downstream genes involved in the process.</p

IGF-I induced genes in stromal fibroblasts predict the clinical outcome of breast and lung cancer patients

<p>Abstract</p> <p>Background</p> <p>Insulin-like growth factor-1 (IGF-I) signalling is important for cancer initiation and progression. Given the emerging evidence for the role of the stroma in these processes, we aimed to characterize the effects of IGF-I on cancer cells and stromal cells separately.</p> <p>Methods</p> <p>We used an <it>ex vivo </it>culture model and measured gene expression changes after IGF-I stimulation with cDNA microarrays. <it>In vitro </it>data were correlated with <it>in vivo </it>findings by comparing the results with published expression datasets on human cancer biopsies.</p> <p>Results</p> <p>Upon stimulation with IGF-I, breast cancer cells and stromal fibroblasts show some common and other distinct response patterns. Among the up-regulated genes in the stromal fibroblasts we observed a significant enrichment in proliferation associated genes. The expression of the IGF-I induced genes was coherent and it provided a basis for the segregation of the patients into two groups. Patients with tumours with highly expressed IGF-I induced genes had a significantly lower survival rate than patients whose tumours showed lower levels of IGF-I induced gene expression (<it>P </it>= 0.029 - Norway/Stanford and <it>P </it>= 7.96e-09 - NKI dataset). Furthermore, based on an IGF-I induced gene expression signature derived from primary lung fibroblasts, a separation of prognostically different lung cancers was possible (<it>P </it>= 0.007 - Bhattacharjee and <it>P </it>= 0.008 - Garber dataset).</p> <p>Conclusion</p> <p>Expression patterns of genes induced by IGF-I in primary breast and lung fibroblasts accurately predict outcomes in breast and lung cancer patients. Furthermore, these IGF-I induced gene signatures derived from stromal fibroblasts might be promising predictors for the response to IGF-I targeted therapies.</p> <p>See the related commentary by Werner and Bruchim: <url>http://www.biomedcentral.com/1741-7015/8/2</url></p