Depletion of insulin-like growth factor 1 receptor increases radiosensitivity in colorectal cancer

BackgroundAlthough radiation therapy for advanced colorectal cancer (CRC) is very effective in some patients, treatment resistance limits its efficacy. Insulin-like growth factor 1 receptor (IGF1R) can affect tumor responsiveness and sensitivity to radiation in several cancer types. Herein, we studied the underlying function of IGF1R in the resistance of advanced CRC to radiation therapy and the possible use of drugs targeting IGF1R to overcome this resistance in patients with CRC.MethodsDifferences in the expression levels of the IGF1R were assessed in CRC samples from patients who were radiosensitive or radioresistant. Two radio-resistant colorectal cancer cell lines, SW480 and HT29, were selected for in vitro studies, and the involvement of the IGF1R in their radiation resistance was elucidated by suppressing its expression through a targeted siRNA and through the use of a specific IGF1R inhibitor, BMS-754807. We assessed radiosensitivity in these human CRC cells lines by examining their proliferation and colony formation, as well as cell cycle analysis. Activation of the Akt pathway was assessed using western blotting.ResultsCompared with tissues from radiosensitive patients, higher IGF1R expression levels were found in patients with radiation-resistant colorectal cancer, while BMS-754807 had improved radiosensitivity and reversed radiation tolerance in both colorectal cancer cell lines. Pre-treatment with BMS-754807 prior to irradiation inhibited Akt phosphorylation, induced cell cycle arrest, and increased DNA damage. Therefore, the IGF1R contributes to radiation resistance of CRC cells in vitro.ConclusionsThis study supports the notion that the radiosensitivity of radiation-resistant colorectal cancer cells can be enhanced by directly targeting IGF1R expression or activity. Ultimately, the combination of radiotherapy with IGF1R targeted inhibitors could potentially increase its effectiveness in the treatment of advanced colorectal cancer.     

The type 1 insulin-like growth factor receptor pathway

Research conducted over the past two decades has shown the importance of the type 1 insulin-like growth factor receptor (IGF1R) in tumorigenesis, metastasis, and resistance to existing forms of cancer therapy. The IGF1R itself has only recently been accepted as a credible treatment target, however, perhaps reflecting the potential problems for drug design posed by normal tissue IGF1R expression, and close homology with the insulin receptor. Currently approximately 12 anti-IGF1R therapeutics are undergoing clinical evaluation, including blocking antibodies and tyrosine kinase inhibitors. This review will summarize the principal signaling pathways activated by IGF1R and the preclinical data that validated this receptor as a treatment target. We will review clinical progress in the testing of IGF1R inhibitory drug candidates, the relative benefits and potential toxicities of coinhibition of the insulin receptor, and the rationale for combining IGF1R blockade with other cancer treatments. An understanding of IGF1R signaling is important because it will guide the incorporation of appropriate molecular markers into clinical trial design. This will be key to the identification of patients most likely to benefit, and so will influence the ability of IGF1R inhibition to make the transition from experimental intervention to clinical therapy.     

Targeting insulin-like growth factor type 1 receptor in cancer therapy

It is believed that the insulin-like growth factor receptor type 1 (IGF-1R) signaling pathway plays a pivotal role in cancer growth, progression, and resistance to anticancer therapies. Strategies are being developed to block IGF-1R as an anticancer treatment. We reviewed several potential strategies for disrupting the IGF axis. We also reviewed the effects of two drugs that target the IGF-1R: monoclonal antibodies and tyrosine kinase inhibitors. Preliminary results of studies involving these agents provided a foundation for ongoing clinical trials, whose results in the near future will help us understand how to incorporate anti IGF-1R strategies into the current anticancer armamentarium.     

胰岛素样生长因子l及其受体对子宫肌瘤生长的影响

目的 测定胰岛素样生长因子1(IGF 1)及其受体(IGF 1R)在子宫肌瘤患者的子宫平滑肌瘤组织、血清中的表达,旨在探讨IGF 1、IGF 1R在子宫肌瘤发病机制中的作用。方法 采用酶联免疫吸附实验(ELISA)方法,测定26例子宫肌瘤患者子宫平滑肌瘤、正常子宫平滑肌组织及血清中IGF1、IGF 1R的表达,每例患者充当组织测定的自身对照,选择30例健康妇女作为血清测定的对照组。结果 ①子宫平滑肌瘤组织中IGF 1、IGF 1R的表达较正常子宫平滑肌增强,进行统计学分析,两组差异有显著性(P<0.01)。②子宫肌瘤患者血清中IGF-1表达与对照组差异无显著性(P>0.05);IGF1R表达较对照组增强,两组差异有显著性(P<0.01)。结论 由此推测子宫平滑肌瘤组织中IGF 1、IGF 1R过度表达可能是子宫肌瘤发生的机制之一,IGF 1主要以自分泌、旁分泌作用方式促进肌瘤的生长,IGF 1可能是子宫肌瘤生长的调节因子。     

Epidermal growth factor receptor and K-Ras mutations and resistance of lung cancer to insulin-like growth factor 1 receptor tyrosine kinase inhibitors

BACKGROUND:

Most patients with nonsmall cell lung cancer (NSCLC) have responded poorly to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs). The authors investigated the involvement of insulinlike growth factor 1 receptor (IGF‐1R) signaling in primary resistance to EGFR TKIs and the molecular determinants of resistance to IGF‐1R TKIs.

METHODS:

Phosphorylated IGF‐1R/insulin receptor (pIGF‐1R/IR) was immunohistochemically evaluated in an NSCLC tissue microarray. The authors analyzed the antitumor effects of an IGF‐1R TKI (PQIP or OSI‐906), either alone or in combination with a small‐molecular inhibitor (PD98059 or U0126) or with siRNA targeting K‐Ras or mitogen‐activated protein kinase/extracellular signal‐regulated kinase kinase (MEK), in vitro and in vivo in NSCLC cells with variable histologic features and EGFR or K‐Ras mutations.

RESULTS:

pIGF‐1R/IR expression in NSCLC specimens was associated with a history of tobacco smoking, squamous cell carcinoma histology, mutant K‐Ras, and wild‐type (WT) EGFR, all of which have been strongly associated with poor response to EGFR TKIs. IGF‐1R TKIs exhibited significant antitumor activity in NSCLC cells with WT EGFR and WT K‐Ras but not in those with mutations in these genes. Introduction of mutant K‐Ras attenuated the effects of IGF‐1R TKIs on NSCLC cells expressing WT K‐Ras. Conversely, inactivation of MEK restored sensitivity to IGF‐TKIs in cells carrying mutant K‐Ras.

CONCLUSIONS:

The mutation status of both EGFR and K‐Ras could be a predictive marker of response to IGF‐1R TKIs. Also, MEK antagonism can abrogate primary resistance of NSCLC cells to IGF‐1R TKIs. Cancer 2012. © 2012 American Cancer Society.     

Production and characterization of monoclonal antibodies against insulin-like growth factor type 1 receptor

The type 1 insulin-like growth factor receptor (IGF-1R) has been extensively reported to play an important role in cancer. Activation of the IGF-1R by IGF-I and IGF-II binding to the extracellular domains of the receptor induces mitogenic and anti-apoptotic effects, which are important events in tumor growth and survival. Several cancer cell types overexpress IGF-1R, suggesting a possible use of monoclonal antibodies (MAbs) against IGF-1R as diagnostic reagents. Here, we report the production and characterization of two independent MAbs, namely 7C2 and 9E11, generated by immunizing mice with the soluble extracellular part of this receptor (amino acids 1-906). Both MAbs bind to membrane bound IGF-1R and do not cross-react with insulin receptor isoforms, IR-A and IR-B expressed on IGF-1R() cells. MAbs 7C2 and 9E11 stained the IGF- 1R on frozen or paraffin-embedded tissue sections or frozen cells. The MAbs 7C2 and 9E11 immunoprecipitated the IGF-1R from P6 cell lysates (cells overexpressing human IGF-1R) and could detect non-reduced intact IGF-1R on immunoblots. However, the MAbs were not able to detect reduced and denatured receptor alpha and beta chains. Sequencing of the heavy- and light-chain variable regions revealed that the 7C2 and 9E11 CDR amino acid sequences are different but result in antibodies with similar properties. MAbs 7C2 and 9E11 are therefore potentially useful diagnostic tools and could be of therapeutic use for humans in the future.     

Heterogeneity of receptor function in colon carcinoma cells determined by cross-talk between type I insulin-like growth factor receptor and epidermal growth factor receptor

This study identifies a novel cross-talk paradigm between the type I insulin-like growth factor receptor (IGF1R) and epidermal growth factor receptor (EGFR) in colon cancer cells. IGF1R activation by ligand exposure in growth factor-deprived cells induces Akt activation in the FET, CBS, and GEO colon cancer cell lines. Investigation of IGF1R-mediated signaling pathways using small interfering RNA approaches indicated that, as expected, phosphatidylinositol 3'-kinase (PI3K) was activated by IGF1R. Mitogen-activated protein kinase (MAPK) activity as reflected by phospho-extracellular signal-regulated kinase (ERK) induction was not significantly activated until later times following release of these cells from growth factor deprivation stress. The appearance of phospho-ERK was proximal to EGFR activation. Treatment of cells with the PI3K inhibitor LY294002 before release from stress resulted in a concentration-dependent loss of EGFR activation, whereas treatment with the MAPK inhibitor PD98059 did not block EGFR activation, indicating that EGFR activation was downstream of the IGF1R/PI3K pathway. PD98059 inhibition of MAPK was associated with a concentration-dependent reduction in EGFR-mediated phospho-ERK. EGFR inhibitor blocked induction of phospho-ERK, showing that MAPK activity was a consequence of EGFR-mediated signaling. On the other hand, a small-molecule IGF1R inhibitor, PQIP, blocked Akt phosphorylation. The divergent signaling functions of IGF1R and EGFR suggested the potential for synergism by a combination of therapy directed at the two receptors. Combination treatment with PQIP and EGFR inhibitor Tarceva resulted in synergistic effects as indicated by combination index analysis in all three cell lines tested.     

Luteolin inhibits insulin-like growth factor 1 receptor signaling in prostate cancer cells

Insulin-like growth factor 1 receptor (IGF-1R) activation is required for prostate cell proliferation. Prostate cancer is one of the most commonly diagnosed malignant tumors in Western countries. Overexpression of IGF-1R in prostate cancer is associated with tumor growth. These suggest that IGF-1R inhibitory agents may be of preventive and/or therapeutic value. With evidence accumulating for a chemopreventive role of flavonoids, the effects of luteolin, a bioactive flavonoid, on IGF-1R signaling in prostate cancer cells were examined. Luteolin inhibited insulin-like growth factor 1 (IGF-1) induced activation of IGF-1R and AKT in prostate cancer PC-3 and DU145 cells. Inhibition of AKT by luteolin resulted in decreased phosphorylation of its downstream targets, including p70S6K1, GSK-3beta and FKHR/FKHRL1. Luteolin also inhibited the IGF-1-induced activation of EGFR and MAPK/ERK signaling. Luteolin inhibited expression of cyclin D1 and increased expression of p21. As a result, luteolin suppressed proliferation and induced apoptosis of prostate cancer cells. Knockdown of IGF-1R by siRNA led to inhibition of proliferation of prostate cancer cells. Results of in vivo tumor growth assay indicated that luteolin inhibited PC-3 tumor growth. Immunoblotting of the extracts of tumor tissues showed that luteolin inhibited IGF-1R/AKT signaling. Our results provide a new insight into the mechanisms that luteolin is against cancer cells.     

Ras-related protein 1 and the insulin-like growth factor type I receptor are associated with risk of progression in patients diagnosed with carcinoma in situ

Currently, there are no applied molecular markers to aid in predicting risk of carcinoma in situ (CIS) progression to invasive cancer, and therefore, all women diagnosed with CIS undergo surgery. Standard assessment of protein expression in fixed tissue by immunohistochemistry (IHC) is not quantitative and hence is not well suited for measuring biomarkers. In this study, we developed an original analytical method for IHC quantification. Using our novel image-based uniplex (IBU) method, quantitative protein profiling was performed on 90 samples of the breast (17 histologically normal tissues, 16 benign lesions, 15 CIS, and 42 invasive carcinomas). Differences between groups were assessed using analysis of variance (ANOVA) and mixed effects models. Measuring protein expression on a continuous scale revealed a significant increase in Ras-related protein 1 (Rap1) and the insulin-like growth factor type I receptor (IGF-IR) in conjunction with the presence of cancer invasion. Women with invasive cancers were four times more likely to have increased levels of Rap1 [odds ratio (OR) = 3.91; P = 0.0002] and IGF-IR (OR = 4.33; P < 0.0001) than women with non-invasive lesions. Furthermore, expression of both proteins was also increased significantly in CIS adjacent to invasive tumors compared with non-cancerous tissue. These novel findings of a significant up-regulation of Rap1 and IGF-IR in CIS progressing to invasive cancers warrant further investigation of Rap1 and IGF-IR together as a dual biomarker to aid in predicting risk of progression and ultimately providing non-surgical treatment options to those at lower risk.     

The contradictions of the insulin-like growth factor 1 receptor

In recent years, the type 1 insulin-like growth factor receptor (IGF-IR) has emerged as a receptor that plays a very important role in the growth of cells, both in vivo and in vitro. The ability of the IGF-IR to induce mitogenesis and to promote survival of cells against a variety of apoptotic agents is well documented. Somewhat less known are other functions of the IGF-IR, like its ability to induce differentiation, to regulate cell size and to affect the organization of the cytoskeleton of cells. This review will focus on these lesser known functions of the IGF-IR. At the same time, we will emphasize how the IGF-IR can send contradictory signals, which depend on different domains of the receptor and the availability of downstream transducing molecules.     

Acquired resistance to tamoxifen is associated with loss of the type I insulin-like growth factor receptor: implications for breast cancer treatment

The role of the insulin-like growth factor (IGF) system in breast cancer is well defined, and inhibitors of this pathway are currently in clinical trials. The majority of anti-IGF1R clinical trials are in estrogen receptor-positive patients who have progressed on prior endocrine therapy; early reports show no benefit for addition of IGF1R inhibitors to endocrine therapy in this setting. In this study, we examined the effectiveness of IGF1R inhibitors in vitro by generating tamoxifen-resistant (TamR) cells. We found that TamR cells had diminished levels of IGF1R with unchanged levels of insulin receptor (IR), and failed to respond to IGF-I-induced Akt activation, proliferation, and anchorage-independent growth while retaining responsiveness to both insulin and IGF-II. The IGF1R antibody dalotuzumab inhibited IGF-I-mediated Akt phosphorylation, proliferation, and anchorage-independent growth in parental cells, but had no effect on TamR cells. An IGF1R tyrosine kinase inhibitor, AEW541, with equal potency for the IGF1R and IR, inhibited IGF-I-, IGF-II-, and insulin-stimulated Akt phosphorylation, proliferation, and anchorage-independent growth in parental cells. Interestingly, AEW541 also inhibited insulin- and IGF-II-stimulated effects in TamR cells. Tamoxifen-treated xenografts also had reduced levels of IGF1R, and dalotuzumab did not enhance the effect of tamoxifen. We conclude that cells selected for tamoxifen resistance in vitro have downregulated IGF1R making antibodies directed against this receptor ineffective. Inhibition of IR may be necessary to manage tamoxifen-resistant breast cancer.     

Increase in hepatocyte growth factor receptor tyrosine kinase activity in renal carcinoma cells is associated with increased motility partly through phosphoinositide 3-kinase activation.

Dysregulated cell motility is one of the major characteristics of invasion and metastatic potentials of malignant tumor cells. Here, we examined the hepatocyte growth factor (HGF)-induced cell motility of two human renal carcinoma cell lines, ACHN and VMRC-RCW. Scattering and migration was induced in ACHN in an HGF-dependent manner, whereas they were maintained in VMRC-RCW even in the absence of HGF. In VMRC-RCW, HGF receptor (HGFR) tyrosine kinase was constitutively active, and sequence analysis showed N375S, A1209G and V1290L mutations. However, transfection experiments using porcine aortic endothelial (PAE) cells demonstrated that no single mutation or combination of two or three mutations caused HGF-independent constitutive activation. Conversely, the expressed amount of receptor protein had a pivotal role in the basal kinase activity. With respect to downstream signaling molecules of HGFR in ACHN or VMRC-RCW, the Ras-MAPK pathway was downregulated, whereas phosphoinositide 3-kinase (PI3-kinase) was not further activated by HGF-treatment in VMRC-RCW cells. The PI3-kinase inhibitors, wortmannin and LY294002 strongly inhibited spontaneous migration of VMRC-RCW. One transfected PAE cell line with massive overexpression of HGFR demonstrated scattered morphology and increased PI3-kinase activity in association with increased motility, which was partially inhibited by LY294002. Taken together, our results indicate that the overexpression of HGFR causes increase in cellular motility and PI3-kinase shows the important contribution on the increased motility of renal carcinoma cells.     

Expression of the type 1 insulin-like growth factor receptor is up-regulated in primary prostate cancer and commonly persists in metastatic disease

The type 1 insulin-like growth factor receptor (IGF1R) mediates tumor cell growth, adhesion, and protection from apoptosis. High plasma IGF-I levels predispose to prostate cancer, but there is no consensus regarding IGF1R expression in primary and metastatic prostate cancer. Recent studies in a human cell line and a mouse model suggest that metastatic prostate cancer cell detachment may be favored by impairing cadherin function via loss of expression of insulin receptor substrate-1 (IRS-1), the principal IGF1R docking molecule. This may be accompanied by PTEN mutation, reactivating a key antiapoptotic pathway, and by IGF1R down-regulation to prevent Shc-mediated differentiation. We studied IGF1R expression in 54 samples of primary prostate tissue including 44 archival and 10 prospectively collected biopsies. We performed semiquantitative immunostaining for the IGF1R, IRS-1, and PTEN, and in situ hybridization for IGF1R. The IGF1R was significantly up-regulated at the protein and mRNA level in primary prostate cancer compared with benign prostatic epithelium. There was a trend toward increased expression of IRS-1 in the malignant biopsies. We also measured IGF1R, IRS-1, and PTEN expression in 12 paired biopsies of primary prostate cancer and subsequent bone metastases. In four cases, IGF1R and IRS-1 levels were lower in the metastases than in the primary tumors. Three of these metastases also lacked significant PTEN staining, compatible with findings in the model systems described above. However, this pattern was relatively uncommon, and 8 of 12 cases expressed detectable IGF1R and IRS-1 in both primary and metastatic biopsies. These findings challenge earlier reports of IGF1R down-regulation in metastatic disease and reinforce the importance of the IGF1R in prostate cancer biology.