Effects of 9-cis-Retinoic Acid on the Insulin-Like Growth Factor Axis in Former Smokers
http://www.100md.com
《临床肿瘤学》
The University of Texas Graduate School of Biomedical Sciences at Houston and The University of Texas M.D. Anderson Cancer Center, Houston, TX
ABSTRACT
PURPOSE: Insulin-like growth factor (IGF) axis has been associated with the risk of lung cancer. 9-cis-retinoic acid (9-cis-RA) has shown potential chemopreventive activities in former smokers. This study was designed to evaluate the effects of 9-cis-RA on IGF axis in former smokers to identify any benefit the retinoid may have in preventing lung cancer.
PATIENTS AND METHODS: Serum concentrations of IGF-I, IGF binding protein (IGFBP)-3, and their molar ratio (IGF-I/IGFBP-3) were measured with radioimmunoassay kits in stored blood samples from the participants of an original chemoprevention trial. The participants had ceased smoking for at least 12 months and were randomly assigned to receive 3 months of daily oral 9-cis-RA (100 mg) or placebo. All statistical tests were two-sided.
RESULTS: A total of 111 samples from the study's baseline and 84 samples from the 3 months treatment were analyzed. The serum concentrations of IGF-I and IGF-I/IGFBP-3 at baseline were significantly lower in female than in male participants. After 3 months of treatment, the serum level of IGF-I and IGF-I/IGFBP-3 were significantly lower in the 9-cis-RA group than in the placebo group (P = .03 and P < .01, respectively), but the IGFBP-3 level was significantly higher (P = .03).
CONCLUSION: 9-cis-RA treatment modulated the IGF axis in former smokers, suggesting that the IGF axis is a potential target for the chemopreventive activities of 9-cis-RA and that the serum concentrations of IGF, IGFBP-3, and IGF-I/IGFBP-3 could serve as surrogate end point biomarkers of 9-cis-RA treatment.
INTRODUCTION
Lung cancer is the leading cause of cancer death in men and women in the United States,1,2 and cigarette smoking is the predominant risk factor for lung cancer. Therefore, smoking cessation campaigns have been a major focus of preventive effort.3,4 However, the risk for lung cancer does not diminish during the first 5 years after smoking cessation5,6: former smokers continue to have an increased risk compared with people who have never smoked.7 These findings indicate that additional preventive strategies for former smokers are needed. One effective strategy is the administration of agents that suppress the promotion or progression steps of lung carcinogenesis by inhibiting the proliferation and survival of preneoplastic cells that have acquired genomic DNA damage as a result of exposure to cigarette carcinogens.
An increasingly recognized mediator of cell proliferation and survival is insulin-like growth factor (IGF).8,9 IGFs can also inhibit apoptosis and play an important role in differentiation of many normal and cancer cell types and in neoplastic transformation and metastasis.9-11 The IGF system is regulated by IGF binding proteins (IGFBPs), especially IGFBP-3, which bind to IGFs in the extracellular milieu with high affinity and specificity, thus reducing the bioavailability of IGFs; more than 90% of circulating IGF-I is bound within a large complex containing IGFBP-3 and its acid-labile subunit.9 A growing number of epidemiologic studies have suggested that increased serum concentrations of IGFs, altered concentrations of IGFBP-3, or both, are associated with an increased risk for several types of cancer, including lung cancer, and that high IGFBP-3 concentrations can attenuate this risk.12-17 We have shown that loss of IGFBP-3 expression, due partly to hypermethylation of its promoter,18 is a marker of poor prognosis in patients with early-stage non–small-cell lung cancer (NSCLC).19,20 We have also demonstrated that overexpression of IGFBP-3 inhibits the growth of NSCLC cells in vitro and in vivo by inducing apoptosis.21 These data indicate that for high-risk patients, the IGF system is a potential target for preventive strategies, for novel antineoplastic therapies, or for both.
An increasing body of evidence has suggested that retinoids, the most frequently studied chemopreventive agents for lung cancer, modulate the IGF axis.22-24 Results of several in vivo studies in experimental animals have shown that retinoids suppress carcinogenesis in a variety of tissue types, including the lung.25,26 Of the naturally occurring retinoids, all-trans-retinoic acid (all-trans-RA) binds to RA receptors (RARs), and 9-cis-retinoic acid (9-cis-RA) binds to RARs, retinoid X receptors (RXRs), and other nuclear receptor complexes in which the RXR is a ligand-binding partner, such as the vitamin D receptor and the peroxisome proliferator–activated receptor.27 13-cis-RA binds to RAR and, after stereoisomerization, to either all-trans-RA or 9-cis-RA in a process that occurs intracellularly.28
We demonstrated in a clinical lung cancer chemoprevention trial that 9-cis-RA treatment can restore RAR expression in former smokers after 3 months of treatment.29 RAR expression has been implicated in the prevention of tumor development and has shown growth-inhibitory and apoptotic effects on the bronchial epithelia of former smokers.30 These previous findings thus raised the possibility that 9-cis-RA has potential chemopreventive properties in former smokers.
To shed more light on the beneficial effects of 9-cis-RA on former smokers, we analyzed the effects of 9-cis-RA on the IGF axis, especially on the serum concentrations of IGF-I and IGFBP-3 and on the molar ratio of IGF-I to IGFBP-3, which has been proposed to reflect tissue bioactivity, in a previously studied population of former smokers.31 We found that 9-cis-RA treatment modulated the IGF axis in these former smokers, suggesting that the serum IGF axis is a target of the potential chemopreventive activities of 9-cis-RA in former smokers.
PATIENTS AND METHODS
Patients
The original study from which we derived our data was a three-arm, randomized, double-blinded, placebo-controlled trial comparing the effects of 9-cis-RA (100 mg) with those of 13-cis-RA (1 mg/kg) plus -tocopherol (AT; 1,200 U) administered for 3 months. In this study, a significant increase in RAR expression and a reduction of metaplasia were observed in individuals treated with 9-cis-RA, but not with 13-cis-RA plus AT, compared with those treated with placebo. Because antioxidants such as AT can affect the concentrations of the components of the IGF axis,32 we decided not to include the 13-cis-RA plus AT treatment group in our analysis. The eligibility criteria were previously described.29 Briefly, the study population consisted of former heavy smokers clinically free of any cancer, who were registered in the Departments of Thoracic/Head and Neck Medical Oncology and of Thoracic Surgery at The University of Texas M.D. Anderson Cancer Center. To be eligible, subjects had to have adequate renal, hematologic, and hepatic function and must not have taken more than 25,000 U of vitamin A or other retinoids daily for at least 3 months before study entry. Subjects were allowed to have had a prior smoking-related cancer, but they had to have been tumor free for 6 months before enrollment in the study. Subjects were required to abstain from consuming dietary vitamin supplements while on the study. The treatment duration was 3 months, based on the toxicity data from a previous phase I trial that included 9-cis-RA treatment.33 Subjects were seen monthly and were evaluated for compliance with the trial protocol, drug-related toxic effects, and serum cotinine concentrations.
The clinical trial from which the samples analyzed in this study were derived indicated that 9-cis-RA had some side effects. Specifically, subjects in the 9-cis-RA group experienced grade 2 (46 subjects) and grade 3 (nine subjects) toxic effects typical of retinoid treatment, including skin rash, hypertriglyceridemia, headache, cheilitis, conjunctivitis, arthralgia, and myalgia.29 Drug-related toxicity was graded according to the National Cancer Institute's Common Toxicity Criteria.34
The original study had been approved by the institutional review board of The M.D. Anderson Cancer Center and by the US Department of Health and Human Services. Our current study was also approved by the institutional review board at M.D. Anderson.
IGF-I and IGFBP-3 Measurements
For analysis of serum concentrations of IGF-I and IGFBP-3, blood samples were drawn from nonfasting subjects and collected in heparinized tubes that were transported immediately to the laboratory, where the samples were immediately centrifuged for 10 minutes at 4,000 x g and then stored at –80°C until the assays were performed. The serum concentration of IGF-I was measured by a specific radioimmunoassay (Diagnostic Systems Laboratories Inc, Webster, TX) with intra-assay and interassay coefficients of variation of less than 4% and 8%, respectively. To separate IGFs from their binding proteins, we mixed serum specimens with an acid-ethanol extraction buffer before measurement. The extraction procedure had been previously evaluated, and the efficiency of the extraction was identical to that for acid-column chromatography.35 The IGFBP-3 concentration was also measured by a specific radioimmunoassay (Diagnostic Systems Laboratories Inc) with intra-assay and interassay coefficients of variation of less than 3.5% and 7.5%, respectively, and no cross-reaction with other members of the IGFBP family. The assays were performed according to the instructions of the manufacturer and without knowledge of who the subject was. The molar ratio of IGF-I to IGFBP-3 was calculated as (0.130 x IGF-I concentration [ng/mL])/(0.036 x IGFBP-3 concentration [ng/mL]).
Statistical Analysis
The characteristics of the study subjects were compared pre–random assignment, according to sex or study group, by using Fisher's exact test for dichotomous characteristics and the Kruskal-Wallis test for quantitative characteristics. Because the distributions of serum concentrations of IGF-I and IGFBP-3 were skewed, the differences between groups were tested by using the Wilcoxon rank sum test. An overall treatment effect over time was determined by comparing the modulation (ie, the value at the subsequent visit minus the value at the baseline evaluation) in serum concentrations of IGF-I and IGFBP-3 and in the molar ratio of IGF-I to IGFBP-3 between the 9-cis-RA and placebo groups. All P values were determined by two-sided tests. Associations were considered statistically significant at P values less than .05. In the multivariate analysis, the variables of sex, age, smoking status, and body mass index (body weight in kg/height in m2) were included in the model.
RESULTS
Subject Characteristics
The characteristics of eligible subjects were described in detail previously.29 Of the 226 former smokers enrolled in the original chemoprevention trial, 149 were randomly assigned to placebo or 9-cis-RA treatments, and 113 of them completed 3 months of treatment. The characteristics of subjects assessable for our study are detailed in Table 1. Each treatment group was well balanced for sex, race, age, body mass index, and history of smoking. Blood samples from 111 of these subjects (52 women and 59 men; 56 from the 9-cis-RA group and 55 from the placebo group) at the baseline of the study, and from 84 (40 women and 44 men; 41 from the 9-cis-RA group and 43 from the placebo group) after 3 months of treatment were assessable for serum concentrations of IGF-I and IGFBP-3. The two treatment groups had comparable baseline mean serum concentrations of IGF-I and IGFBP-3 and similar molar ratios of IGF-I to IGFBP-3 (Table 2).
To determine whether subject characteristics affected the baseline serum concentrations of IGF-I or IGFBP-3, we evaluated the correlation between certain variables (eg, sex, race, age, body mass index, pack-years of smoking, and number of years since stopping smoking) and the baseline serum concentrations of IGF-I and IGFBP-3 and the molar ratio of IGF-I to IGFBP-3 (Table 3). The baseline serum concentration of IGF-I and the molar ratio of IGF-I to IGFBP-3 were significantly lower in women, whereas the IGFBP-3 concentration was slightly higher, though the difference did not reach statistical significance. Moreover, the serum concentration of IGF-I and the molar ratio of IGF-I to IGFBP-3 in women significantly decreased with increasing age (Fig 1A). Because serum concentrations of IGF-I are reduced in women treated with hormone replacement therapy (HRT),36,37 we analyzed whether the changes in the baseline serum concentration of IGF-I and the baseline molar ratio of IGF-I to IGFBP-3 were associated with HRT use among women. Self-reported data for HRT use were available for 38 of the 52 women (Table 3). Overall, baseline serum concentrations IGF-I and molar ratios of IGF-I to IGFBP-3 concentrations were significantly lower in the HRT users than the non–HRT users (Fig 1B). Among HRT users, differences in the IGFBP-3 concentrations were not significantly different. The mean baseline serum concentrations of IGF-I and IGFBP-3, and the molar ratio of IGF-I to IGFBP-3 in the HRT users are summarized in Table 3. Race, body mass index, number of pack-years, and years since stopping smoking did not affect the serum concentrations of IGF-I and IGFBP-3 or the molar ratio of IGF-I to IGFBP-3 in the study population.
Effect of 9-cis-RA on IGF-I and IGFBP-3 Serum Concentrations and on the IGF-I to IGFBP-3 Molar Ratio
The primary end point of the original lung cancer chemoprevention trial was restoration of RAR expression in the bronchial epithelium.29 In the previous study, we demonstrated that, compared with the effect of placebo, the median change in receptor index was significantly different from placebo for 9-cis-RA but not for 13-cis-RA plus AT,29 raising the possibility that 9-cis-RA has potential chemopreventive properties in former smokers.
In the current study, we evaluated the serum concentrations of the IGF axis in former smokers during treatment with 9-cis-RA. The modulations in mean serum concentrations of IGF-I and IGFBP-3 and in the molar ratio of IGF-I to IGFBP-3 in the two treatment groups during 3 months of treatment are illustrated in Figure 2. The mean changes in the placebo and 9-cis-RA groups are summarized in Table 4. Compared with the placebo group, the 9-cis-RA group exhibited a statistically significant modulation in the IGF axis. The mean changes from the study's baseline to the end of 3 months of treatment in the placebo and 9-cis-RA groups were as follows: IGF-I, 14.3 and –19.2, respectively; IGFBP-3, –175.1 and 196.6, respectively; and molar ratio of IGF-I to IGFBP-3, 0.05 and –0.06, respectively. These findings suggested that 9-cis-RA increases serum concentrations of IGFBP-3 and decreases serum concentrations of IGF-I, thus reducing the molar ratio of IGF-I to IGFBP-3.
To determine whether the differences in the serum concentrations of IGFBP3 between untreated and treated groups reflected differences at the tissue level, we also performed immunohistochemical analysis of IGFBP3, using the methodology described previously for lung cancer samples,20 in more than 90% of the cases. The intensity of IGFBP-3 expression in the normal and hyperplastic epithelial tissue samples was high, and differences between placebo–and 9-cis-RA–treated groups were not significant (data not shown). We suggest that the reason for the apparent discrepancy between the results of the plasma concentration analysis reported in this article and the tissue analysis is the higher sensitivity and dynamic range of the enzyme-linked immunosorbent assay for the plasma IGFBP3 and IGF1 compared with the limited dynamic range of the immunohistochemical analysis. In addition, the sources of IGF1 and IGFBP-3 in the blood include the liver and other tissues and is not expected to be directly related to expression in bronchial epithelial cells.
Correlation Between Changes in Tissue Level of RAR- and Modulation of the IGF Axis Induced by 9-cis-RA
We further evaluated the association between the changes in serum concentrations of the IGF axis peptides and changes in the tissue expression level of RAR during 9-cis-RA treatment. To determine the effect of treatment on loss of RAR expression at any biopsy site, the subjects were grouped according to whether their biopsy samples were positive (ie, RAR was detected in all six biopsy sites) or negative (ie, RAR was not detected in at least one biopsy site) for RAR expression, and the effects of treatment on RAR expression was determined as a binary variable (ie, loss of RAR expression at any biopsy site). The modulation of serum concentrations of IGF-I and IGFBP-3 and the molar ratio of IGF-I to IGFBP-3 were not significantly correlated with baseline RAR expression (Fig 3A) or with the changes in tissue RAR expression induced by 9-cis-RA (Fig 3B).
DISCUSSION
Our main finding in this analysis of data collected from a completed chemoprevention trial assessing the benefits of 9-cis-RA for former smokers was that the IGF axis can be modulated by 9-cis-RA in a population of former smokers. To our knowledge, this is the first report showing modulation of the IGF axis by 9-cis-RA in the setting of a chemoprevention trial.
Growing evidence supports an association between the IGF axis and the risk for lung cancer8 and suggests that the development of agents targeting the IGF axis could be an effective strategy in chemoprevention of the disease. Therefore, retinoids, shown to regulate the IGF axis in vitro22,38,39 have a potential to exert chemopreventive activities. Indeed, several findings have demonstrated the potential use of retinoids as chemopreventive agents. For example, 9-cis-RA has shown antiproliferative activity against a broad range of neoplastic cells, including those from prostate cancer,40 breast cancer,41,42 leukemia and lymphoma,43 lung cancer,44 and head and neck cancer.45 In vivo, 9-cis-RA has substantial anticarcinogenic activity in rat mammary glands46,47 and rat colons.48 More recently, the synthetic retinoid fenretinide has been shown to modulate circulating IGF-I and IGFBP-3 concentrations in breast cancer patients,49,50 and the relative risk of a second breast cancer was 35% lower in premenopausal women treated with fenretinide than in those who received no treatment.51 Thus, from the standpoint of directing future chemoprevention trials, our findings are important because we have shown both that clinical trials targeting the IGF axis in former smokers are feasible and that 9-cis-RA can modulate the IGF axis in this population.
To determine whether the characteristics of subjects affected the baseline IGF axis, we evaluated many variables, such as sex, age, nutritional status, and growth hormone secretion level, which have all been shown to affect the serum concentrations of IGF-I.52 The baseline serum level of IGF-I and the molar ratio of IGF-I to IGFBP-3 were significantly lower in women than in men. Estrogen is most likely responsible for the sex-related difference in the serum concentration of IGF-I. Endogenous estrogens have been shown to directly regulate circulating IGF-I synthesis53 and oral administration of estrogen decreases IGF-I serum concentrations.54,55 In our study, 38 of the 52 women took HRT during the treatment period, and the serum concentrations of IGF-I were significantly lower for HRT users than for non-HRT users. Therefore, the significant difference in serum concentrations of IGF-I and the molar ratio of IGF-I to IGFBP-3 might have resulted from the HRT. The role of estrogen in regulating the IGF-I level is further supported by the recent finding that the IGF-I concentrations of women using HRT were significantly lower than those of women not using HRT.37 Studies have suggested that HRT use is associated with a decreased risk for several types of cancer, including lung cancer, through decreases in the production of IGF-I.56,57 Therefore, if a high level of IGF-I is a putative risk factor for lung cancer, HRT use appears to lower IGF-I concentrations, thereby decreasing lung cancer risk.
We also observed that baseline serum concentrations of IGF-I and the molar ratio of IGF-I to IGFBP-3 were significantly decreased with increasing age in women. Duration of HRT use is also likely responsible for the age-related difference in the serum concentration of IGF-I in a female population because older women may conceivably have used HRT for a longer time. The decrease in IGF-I concentrations with aging has been described many times,49,58,59 and may be due in part to decreased growth hormone levels.60 In normal subjects, the interaction between growth hormone and its specific hepatic receptor stimulates expression of the IGF-I gene and the release of the IGF-I polypeptide.52 Therefore, it is likely that the concentration of IGF-I declines with age, as do secretions of growth hormone. However, we have not observed a significant correlation between the age of the study population in men and serum concentrations of IGF-I and IGFBP-3 or the molar ratio of IGF-I to IGFBP-3 in our study.
Growth hormone deficiency is also associated with a decreased muscle mass and increased body fat,61 and it has been hypothesized that low IGF-I concentrations are associated with high body fat, weight gain over time, and high body mass index.62 This hypothesis was supported by a study of Swedish men and women, which showed an inverse association between IGF-I concentrations and body mass index.53 However, we have not observed such correlation between IGF-I and body mass index, similar to the Rancho Bernardo study.59
The effects of smoking on serum concentrations of IGF-I are unclear. A positive association between IGF-I concentrations and pack-years of smoking and a negative association between pack-years or the number of cigarettes/d and concentrations of IGFBP-3 have been shown.63 However, an inverse association between IGF-I concentrations and smoking has been also reported among men, although not among women.53 Recently, Holmes et al58 showed that lower serum concentrations of IGF-I are significantly associated with smoking. In our study subjects, we observed a very modest decrease in IGF-I concentrations and an increase in IGFBP-3 concentrations associated with pack-years. However, smoking history generally did not affect the serum concentrations of IGF-I and IGFBP-3 or the molar ratio of IGF-I to IGFBP-3.
We observed that subjects in the 9-cis-RA group had a significant decrease in their IGF-I serum concentration, an increase in their IGFBP-3 serum concentration, and a decrease in the molar ratio of IGF-I to IGFBP-3 compared with subjects in the placebo group. The magnitude of the 9-cis-RA–induced changes in the IGF axis was moderate. This may be a consequence of the substantial interindividual variability in concentrations of 9-cis-RA and IGFs, which is at least in part genetically driven.33 Given the potential therapeutic activity of both exogenous growth hormone and IGF-I against heart failure64 and the long-term positive association between the decline in serum IGF-I concentrations and aging,65 a moderate yet durable effect of 9-cis-RA on the IGF axis may be desirable in a preventive context.
Although the exact mechanisms underlying the modulation of the IGF axis by 9-cis-RA are unclear, several in vitro results suggest the ability of retinoids to regulate the IGF axis; retinoids have been shown to regulate the expression of IGFBPs22,23,66; likewise, IGFs have been shown to modulate the cellular response of RAs, and vice versa.24 The induction of IGFBP-3 expression is activated by retinoids. We previously demonstrated that all-trans-RA increases IGFBP-3 expression at a transcriptional level through a RAR--dependent signaling pathway.22 It is well known that the cellular effects of RAs are mediated by RXRs and RARs. 9-cis-RA is a ligand for both RXRs and RARs, but its affinity for RXRs is 40-fold higher than for RARs. On the other hand, RA is primarily a ligand for RARs and only activates RXRs at high concentrations.67 Recently, the vitamin D receptor has been identified in the IGFBP-3 promoter, and RXR has been shown to be required for 1,25-dihydroxyvitamin D3–induced gene transcription.68 Therefore, treatment with 9-cis-RA could lead to there being available ligand for activation of both the RAR:RXR and the VDR:RXR heterodimeric complex, so that IGFBP-3 gene transcription could be efficiently activated.
Given the inverse association between estrogen and serum IGF-I concentrations, the ability of 9-cis-RA to regulate type 1 17-hydroxysteroid dehydrogenases,69 which catalyzes the conversion of estrone and 17-estradiol, could increase the level of 17-estradiol, the physiologically significant molecule of estrogen, and thereby decrease IGF-I concentrations. The ability of estrogens to increase the expression of vitamin D receptors in vitro70 may also contribute to decreased concentrations of IGF-I by inducing IGFBP-3 expression.71 Additional work will be necessary to investigate the mechanism that mediates the regulation of the IGF axis by 9-cis-RA in former smokers.
Because 9-cis-RA treatment increased the tissue expression of RAR, we also explored the correlation between tissue levels of RAR and serum concentrations of the IGF axis. However, we did not find any significant correlation between the modulation of the serum concentrations of IGF-I or IGFBP-3 and RAR expression in the tissue. These findings provide evidence that the mechanisms involved in 9-cis-RA–mediated gene expression are diverse and complex.
In conclusion, we are the first to show that 3 months of treatment with 9-cis-RA decreased the serum level of IGF-I and the molar ratio of IGF-I to IGFBP-3 and increased the serum level of IGFBP-3 in former smokers. These effects may contribute to the chemopreventive benefit of 9-cis-RA to former smokers. Despite these promising findings, enthusiasm for the use of 9-cis-RA as a chemopreventive agent for lung cancer could be tempered by the toxic effects of the agent.29 Although there were no serious side effects such as cardiovascular problems, pancreatitis, or death in the subjects treated with 9-cis-RA in our previous prevention study, and only one patient with grade 4 hypertriglyceridemia stopped 9-cis-RA treatment, it would be better to develop newer agents that are related to 9-cis-RA but that do not have the toxic effects of 9-cis-RA. Because 9-cis-RA can activate both RARs and RXRs and because the toxicities are thought to be mediated by RAR-RXR heterodimers, it had been suggested that "pure" RXR selective synthetic retinoids may exert the beneficial effects of retinoids without the toxicities. Indeed, a group of RXR-selective retinoids ("rexinoids") has demonstrated efficacy with fewer adverse effects in patients with NSCLC in early clinical trials.72 Therefore, whether rexinoids regulate the IGF axis and thereby reduce lung cancer risk would be a worthwhile topic for future chemoprevention trials in former smokers. In addition, because Ras-mediated signaling pathways may participate in the development of resistance to IGFBP-3,73 the combination of RXR-selective agonists and inhibitors of the Ras-mediated signaling pathway could also be considered. Clearly, additional work will be necessary to determine whether the modulation of the IGF axis by 9-cis-RA correlates with the ability to reduce lung cancer risk in former smokers. In addition, further investigation into the role of serum concentrations of IGF-I and IGFBP-3 as surrogate biomarkers in determining the chemopreventive effects of retinoids are warranted.
Authors' Disclosures of Potential Conflicts of Interest
The authors indicated no potential conflicts of interest.
NOTES
Supported in part by National Institutes of Health Grants U19 CA68437 (W.K.H.), R01 CA109520-01 (H.-Y.L.), CA-100816-01A1 (H.-Y.L), American Cancer Society grant RSG-04-082-01-TBE 01(H.-Y.L), and W81XWH-04-1-0142-01-VITAL from the Department of Defense (W.K.H.). W.K.H. is an American Cancer Society Clinical Research Professor.
Presented in part at the 40th Annual Meeting of the American Society of Clinical Oncology, New Orleans, LA, June 5-8, 2004.
Authors' disclosures of potential conflicts of interest are found at the end of this article.
REFERENCES
Khuri FR, Herbst RS, Fossella FV: Emerging therapies in non-small-cell lung cancer. Ann Oncol 12:739-744, 2001
Parkin DM, Pisani P, Ferlay J: Estimates of the worldwide incidence of 25 major cancers in 1990. Int J Cancer 80:827-841, 1999
Ginsberg RJ, Vokes EE, Kenneth R: Non-small cell lung cancer, in DeVita VT Jr, Hellman S, Rosenberg SA (eds): Cancer: Principles and Practice of Oncology (ed 6). Philadelphia, PA, Lippincott Williams & Wilkins, 2001, pp 925-983
Mattson ME, Pollack ES, Cullen JW: What are the odds that smoking will kill you Am J Public Health 77:425-431, 1987
Wistuba II, Lam S, Behrens C, et al: Molecular damage in the bronchial epithelium of current and former smokers. J Natl Cancer Inst 89:1366-1373, 1997
Sekido Y, Fong KM, Minna JD: Progress in understanding the molecular pathogenesis of human lung cancer. Biochim Biochim Biophys Acta 1378:21-59, 1998
Burns DM: Primary prevention, smoking, and smoking cessation: Implications for future trends in lung cancer prevention. Cancer 89:2506-2509, 2000
Ibrahim YH, Yee D: Insulin-like growth factor-I and cancer risk. Growth Horm IGF Res 14:261-269, 2004
Butt AJ, Firth SM, Baxter RC: The IGF axis and programmed cell death. Immunol Cell Biol 77:256-262, 1999
Lopez T, Hanahan D: Elevated levels of IGF-1 receptor convey invasive and metastatic capability in a mouse model of pancreatic islet tumorigenesis. Cancer Cell 1:339-353, 2002
Samani AA, Chevet E, Fallavollita L, et al: Loss of tumorigenicity and metastatic potential in carcinoma cells expressing the extracellular domain of the type 1 insulin-like growth factor receptor. Cancer Res 64:3380-3385, 2004
Yu H, Spitz MR, Mistry J, et al: Plasma levels of insulin-like growth factor-I and lung cancer risk: A case-control analysis. J Natl Cancer Inst 91:151-156, 1999
Wu X, Tortolero-Luna G, Zhao H, et al: Serum levels of insulin-like growth factor I and risk of squamous intraepithelial lesions of the cervix. Clin Cancer Res 9:3356-3361, 2003
Pollak M: Insulin-like growth factors (IGFs) and prostate cancer. Epidemiol Rev 23:59-66, 2001
Lukanova A, Lundin E, Toniolo P, et al: Circulating levels of insulin-like growth factor-1 and risk of ovarian cancer. Int J Cancer 101:549-554, 2002
Zhao H, Grossman HB, Spitz MR, et al: Plasma levels of insulin-like growth factor-1 and binding protein-3, and their association with bladder cancer risk. J Urol 169:714-717, 2003
Kaaks R, Toniolo P, Akhmedkhanov A, et al: Serum C-peptide, insulin-like growth factor (IGF)-I, IGF binding proteins, and colorectal cancer risk in women. J Natl Cancer Inst 92:1592-1600, 2000
Chang YS, Wang L, Mao L, et al: Mechanisms underlying lack of insulin like-growth factor binding protein-3 expression in NSCLC cells. Oncogene 23:6569-6580, 2004
Chang YS, Wang L, Liu D, et al: Correlation between IGFBP-3 promoter methylation and prognosis of patients with stage I non-small cell lung cancer. Clin Cancer Res 8:3669-3675, 2002
Chang YS, Gong K, Sun S, et al: Clinical significance of IGFBP-3 expression in stage I non-small cell lung cancer. Clin Cancer Res 8:3796-3802, 2002
Lee HY, Chun KH, Liu B, et al: Insulin-like growth factor binding protein-3 inhibits the growth of non-small cell lung cancer. Cancer Res 62:3530-3537, 2002
Han G-R, Dohi DF, Lee H-Y, et al: All-trans-retinoic acid increases transforming growth factor-2 and insulin-like growth factor binding protein-3 expression through a retinoic acid receptor--dependent signaling pathway. J Biol Chem 272:13711-13716, 1997
Fontana JA, Burrows-Mezu A, Clemmons DR, et al: Retinoid modulation of insulin-like growth factor-binding proteins and inhibition of breast carcinoma proliferation. Endocrinology 128:1115-1122, 1991
Bentel JM, Lebwohl DE, Cullen KJ, et al: Insulin-like growth factors modulate the growth inhibitory effects of retinoic acid on MCF-7 breast cancer cells. J Cell Physiol 165:212-221, 1995
Nagy L, Thomazy VA, Heyman RA, et al: Retinoid-induced apoptosis in normal and neoplastic tissues. Cell Death Differ 5:11-19, 1998
Hansen LA, Sigman CC, Andreola F, et al: Retinoids in chemoprevention and differentiation therapy. Carcinogenesis 21:1271-1279, 2000
Mangelsdorf DJ, Evans RM: The RXR heterodimers and orphan receptors. Cell 83:841-850, 1995
Marchetti MN, Samplo E, Bun H, et al: In vitro metabolism of three major isomers of retinoic acid in rats: Intersex and interstrain comparison. Drug Metab Dispos 25:637-646, 1997
Kurie JM, Lotan R, Lee JJ, et al: Treatment of former smokers with 9-cis-retinoic acid reverses loss of retinoic acid receptor-expression in the bronchial epithelium: Results from a randomized placebo-controlled trial. J Natl Cancer Inst 95:206-214, 2003
Seewaldt VL, Johnson BS, Parker MB, et al: Expression of retinoic acid receptor beta mediates retinoic acid-induced growth arrest and apoptosis in breast cancer cells. Cell Growth Differ 6:1077-1088, 1995
Juul A, Dalgaard P, Blum W, et al: Serum levels of insulin-like growth factor (IGF)-binding protein-3 (IGFBP-3) in healthy infants, children, and adolescents: The relation to IGF-I, IGF-II, IGFBP-1, IGFBP-2, age, sex, body mass index, and pubertal maturation. J Clin Endocrinol Metab 80:2534-2542, 1995
Zi X, Zhang J, Agarwal R, et al: Silibinin up-regulates insulin-like growth factor-binding protein 3 expression and inhibits proliferation of androgen-independent prostate cancer cells. Cancer Res 60:5617-5620, 2000
Miller VA, Rigas JR, Benedetti FM, et al: Initial clinical trial of the retinoid receptor pan agonist 9-cis retinoic acid. Clin Cancer Res 6:471-475, 1996
National Cancer Institute: Common toxicity criteria, version 2.0. http://ctep.cancer.gov/forms/CTCv20_4-30-992.pdf
Baxter RC, Martin JL: Radioimmunoassay of growth hormone-dependent insulin-like growth factor binding protein in human plasma. J Clin Invest 78:1504-1512, 1986
Campagnoli C, Biglia N, Altare F, et al: Differential effects of oral conjugated estrogens and transdermal estradiol on insulinlike growth factor 1, growth hormone and sex hormone binding globulin serum levels. Gynecol Endocrinol 7:251-258, 1993
Schabath MB, Wu X, Sellin RV, et al: Hormone replacement therapy and lung cancer risk: A case-control analysis. Clin Cancer Res 10:113-123, 2004
Zhou Y, Mohan S, Linkhart TA, et al: Retinoic acid regulates insulin-like growth factor-binding protein expression in human osteoblast cells. Endocrinology 137:975-983, 1996
Gabbitas B, Canalis E: Retinoic acid regulates the expression of insulin-like growth factors I and II in osteoblasts. J Cell Physiology 172:253-264, 1997
Blutt SE, Allegretto EA, Pike JW, et al: 1,25-Dihydroxyvitamin D3 and 9-cis-retinoic acid act synergistically to inhibit the growth of LNCaP prostate cells and cause accumulation of cells in G1. Endocrinology 138:1491-1497, 1997
Rubin M, Fenig E, Rosenauer A, et al: 9-cis-Retinoic acid inhibits growth of breast cancer cells and down-regulates estrogen receptor RNA and protein. Cancer Res 54:6549-6556, 1994
Gottardis MM, Lamph WW, Shalinsky DR, et al: The efficacy of 9-cis-retinoic acid in experimental models of cancer. Breast Cancer Res Treat 38:85-96, 1996
Lutzky J, Vujicic M, Yamanishi DT, et al: Antiproliferative effects of all-trans-retinoic acid (tRA) and 9-cis-retinoic acid (9-cisRA) on human lymphoid cell lines. Proc Am Assoc Cancer Res 34:292, 1993 (abstr 1738)
Guzey M, Demirpence E, Criss W, et al: Effects of retinoic acid (all-trans and 9-cis) on tumor progression in small-cell lung carcinoma. Biochem Biophys Res Commun 242:369-375, 1998
Giannini F, Maestro R, Vukosavljevic T, et al: All-trans, 13-cis, and 9-cis retinoic acids induce a fully reversible growth inhibition in HNSCC cell lines: Implications for in vivo retinoic acid use. Int J Cancer 70:194-200, 1997
Anzano MA, Byers SW, Smith JM, et al: Prevention of breast cancer in the rat with 9-cis-retinoic acid as a single agent and in combination with tamoxifen. Cancer Res 54:4614-4617, 1994
Anzano MA, Peer CW, Smith JM, et al: Chemoprevention of mammary carcinogenesis in the rat: Combined use of raloxifene and 9-cis-retinoic acid. J Natl Cancer Inst 88:123-125, 1996
Zheng Y, Kramer PM, Olson G, et al: Prevention by retinoids of azoxymethane-induced tumors and aberrant crypt foci and their modulation of cell proliferation in the colon of rats. Carcinogenesis 18:2119-2125, 1997
Decensi A, Johansson H, Miceli R, et al: Long-term effects of fenretinide, a retinoic acid derivative, on the insulin-like growth factor system in women with early breast cancer. Cancer Epidemiol Biomarkers Prev 10:1047-1053, 2001
Torrisi R, Parodi S, Fontana V, et al: Effect of fenretinide on plasma IGF-I and IGFBP-3 in early breast cancer patients. Int J Cancer 76:787-790, 1998
Veronesi U, De Palo G, Marubini E, et al: Randomized trial of fenretinide to prevent second breast malignancy in women with early breast cancer. J Natl Cancer Inst 91:1847-1856, 1999
Le Roith D: Seminars in medicine of the Beth Israel Deaconess Medical Center: Insulin-like growth factors. N Engl J Med 336:633-640, 1997
Landin-Wilhelmsen K, Wilhelmsen L, Lappas G, et al: Serum insulin-like growth factor I in a random population sample of men and women: Relation to age, sex, smoking habits, coffee consumption and physical activity, blood pressure and concentrations of plasma lipids, fibrinogen, parathyroid hormone and osteocalcin. Clin Endocrinol (Oxf) 41:351-357, 1994
Helle SI, Omsjo IH, Hughes SC, et al: Effects of oral and transdermal oestrogen replacement therapy on plasma levels of insulin-like growth factors and IGF binding proteins 1 and 3: a cross-over study. Clin Endocrinol (Oxf) 45:727-732, 1996
Raudaskoski T, Knip M, Laatikainen T: Plasma insulin-like growth factor-I and its binding proteins 1 and 3 during continuous non-oral and oral combined hormone replacement therapy. Menopause 5:217-222, 1998
Janne PA, Mayer RJ: Chemoprevention of colorectal cancer. N Engl J Med 342:1960-1968, 2000
Grodstein F, Newcomb PA, Stampfer MJ: Postmenopausal hormone therapy and the risk of colorectal cancer: A review and meta-analysis. Am J Med 106:574-578, 1999
Holmes MD, Pollak MN, Hankinson SE: Lifestyle correlates of plasma insulin-like growth factor I and insulin-like growth factor binding protein 3 concentrations. Cancer Epidemiol Biomarkers Prev 11:862-867, 2002
Goodman-Gruen D, Barrett-Connor E: Epidemiology of insulin-like growth factor-I in elderly men and women: The Rancho Bernardo Study. Am J Epidemiol 146:970-976, 1997
Bondy CA, Underwood LE, Clemmons DR, et al: Clinical uses of insulin-like growth factor I. Ann Intern Med 120:593-601, 1994
Sonksen PH, Salomon F, Cuneo R: Metabolic effects of hypopituitarism and acromegaly. Horm Res 36:27-31, 1991
Harris T, Kiel D, Roubenoff R, et al: Association of insulin-like growth factor-I with body composition, weight history, and past health behaviors in the very old: The Framingham Heart Study. J Am Geriatr Soc 45:133-139, 1997
Kaklamani VG, Linos A, Kaklamani E, et al: Age, sex, and smoking are predictors of circulating insulin-like growth factor 1 and insulin-like growth factor-binding protein 3. J Clin Oncol 17:813-817, 1999
Fazio S, Sabatini D, Capaldo B, et al: A preliminary study of growth hormone in the treatment of dilated cardiomyopathy. N Engl J Med 334:809-814, 1996
Janssen JA, Stolk RP, Pols HA, et al: Serum free IGF-I, total IGF-I, IGFBP-1 and IGFBP-3 levels in an elderly population: Relation to age and sex steroid levels. Clin Endocrinol (Oxf) 48:471-478, 1998
Woodward TL, Turner JD, Hung HT, et al: Inhibition of cellular proliferation and modulation of insulin-like growth factor binding proteins by retinoids in a bovine mammary epithelial cell line. J Cell Physiol 167:488-499, 1996
Heyman RA, Mangelsdorf DJ, Dyck JA, et al: 9-cis Retinoic acid is a high affinity ligand for the retinoid X receptor. Cell 68:397-406, 1992
Pathrose P, Barmina O, Chang CY, et al: Inhibition of 1,25-dihydroxyvitamin D3-dependent transcription by synthetic LXXLL peptide antagonists that target the activation domains of the vitamin D and retinoid X receptors. J Bone Miner Res 17:2196-2205, 2002
Piao Y-S, Peltoketo H, Jouppila A, et al: Retinoic acids increase 17-hydroxysteroid dehydrogenase type 1 expression in JEG-3 and T47D cells, but the stimulation is potentiated by epidermal growth factor, 12-O-tetradecanoulphorbol-13-acetate, and cyclic adenosine 3', 5'-monophosphate only in JEG-3 cells. Endocrinology 138:898-904, 1997
Smirnoff P, Liel Y, Gnainsky J, et al: The protective effect of estrogen against chemically induced murine colon carcinogenesis is associated with decreased CpG island methylation and increased mRNA and protein expression of the colonic vitamin D receptor. Oncol Res 11:255-264, 1999
Boyle BJ, Zhao XY, Cohen P, et al: Insulin-like growth factor binding protein-3 mediates 1,25-dihydroxyvitamin D3 growth inhibition in the LNCaP prostate cancer cell line through p21/WAF1. J Urol 165:1319-1324, 2001
Rizvi NA, Marshall JL, Dahut W, et al: A phase I study of LGD1069 in adults with advanced cancer. Clin Cancer Res 5:1658-1664, 1999
Lee H-Y, Moon HJ, Chun K-H, et al: Effects of insulin-like growth factor binding protein-3 and farnesyltransferase inhibitor SCH66336 on Akt expression and apoptosis in non–small-cell lung cancer cells. J Natl Cancer Inst 96:1536-1548, 2004(Ho-Young Lee, Yoon Soo Ch)
ABSTRACT
PURPOSE: Insulin-like growth factor (IGF) axis has been associated with the risk of lung cancer. 9-cis-retinoic acid (9-cis-RA) has shown potential chemopreventive activities in former smokers. This study was designed to evaluate the effects of 9-cis-RA on IGF axis in former smokers to identify any benefit the retinoid may have in preventing lung cancer.
PATIENTS AND METHODS: Serum concentrations of IGF-I, IGF binding protein (IGFBP)-3, and their molar ratio (IGF-I/IGFBP-3) were measured with radioimmunoassay kits in stored blood samples from the participants of an original chemoprevention trial. The participants had ceased smoking for at least 12 months and were randomly assigned to receive 3 months of daily oral 9-cis-RA (100 mg) or placebo. All statistical tests were two-sided.
RESULTS: A total of 111 samples from the study's baseline and 84 samples from the 3 months treatment were analyzed. The serum concentrations of IGF-I and IGF-I/IGFBP-3 at baseline were significantly lower in female than in male participants. After 3 months of treatment, the serum level of IGF-I and IGF-I/IGFBP-3 were significantly lower in the 9-cis-RA group than in the placebo group (P = .03 and P < .01, respectively), but the IGFBP-3 level was significantly higher (P = .03).
CONCLUSION: 9-cis-RA treatment modulated the IGF axis in former smokers, suggesting that the IGF axis is a potential target for the chemopreventive activities of 9-cis-RA and that the serum concentrations of IGF, IGFBP-3, and IGF-I/IGFBP-3 could serve as surrogate end point biomarkers of 9-cis-RA treatment.
INTRODUCTION
Lung cancer is the leading cause of cancer death in men and women in the United States,1,2 and cigarette smoking is the predominant risk factor for lung cancer. Therefore, smoking cessation campaigns have been a major focus of preventive effort.3,4 However, the risk for lung cancer does not diminish during the first 5 years after smoking cessation5,6: former smokers continue to have an increased risk compared with people who have never smoked.7 These findings indicate that additional preventive strategies for former smokers are needed. One effective strategy is the administration of agents that suppress the promotion or progression steps of lung carcinogenesis by inhibiting the proliferation and survival of preneoplastic cells that have acquired genomic DNA damage as a result of exposure to cigarette carcinogens.
An increasingly recognized mediator of cell proliferation and survival is insulin-like growth factor (IGF).8,9 IGFs can also inhibit apoptosis and play an important role in differentiation of many normal and cancer cell types and in neoplastic transformation and metastasis.9-11 The IGF system is regulated by IGF binding proteins (IGFBPs), especially IGFBP-3, which bind to IGFs in the extracellular milieu with high affinity and specificity, thus reducing the bioavailability of IGFs; more than 90% of circulating IGF-I is bound within a large complex containing IGFBP-3 and its acid-labile subunit.9 A growing number of epidemiologic studies have suggested that increased serum concentrations of IGFs, altered concentrations of IGFBP-3, or both, are associated with an increased risk for several types of cancer, including lung cancer, and that high IGFBP-3 concentrations can attenuate this risk.12-17 We have shown that loss of IGFBP-3 expression, due partly to hypermethylation of its promoter,18 is a marker of poor prognosis in patients with early-stage non–small-cell lung cancer (NSCLC).19,20 We have also demonstrated that overexpression of IGFBP-3 inhibits the growth of NSCLC cells in vitro and in vivo by inducing apoptosis.21 These data indicate that for high-risk patients, the IGF system is a potential target for preventive strategies, for novel antineoplastic therapies, or for both.
An increasing body of evidence has suggested that retinoids, the most frequently studied chemopreventive agents for lung cancer, modulate the IGF axis.22-24 Results of several in vivo studies in experimental animals have shown that retinoids suppress carcinogenesis in a variety of tissue types, including the lung.25,26 Of the naturally occurring retinoids, all-trans-retinoic acid (all-trans-RA) binds to RA receptors (RARs), and 9-cis-retinoic acid (9-cis-RA) binds to RARs, retinoid X receptors (RXRs), and other nuclear receptor complexes in which the RXR is a ligand-binding partner, such as the vitamin D receptor and the peroxisome proliferator–activated receptor.27 13-cis-RA binds to RAR and, after stereoisomerization, to either all-trans-RA or 9-cis-RA in a process that occurs intracellularly.28
We demonstrated in a clinical lung cancer chemoprevention trial that 9-cis-RA treatment can restore RAR expression in former smokers after 3 months of treatment.29 RAR expression has been implicated in the prevention of tumor development and has shown growth-inhibitory and apoptotic effects on the bronchial epithelia of former smokers.30 These previous findings thus raised the possibility that 9-cis-RA has potential chemopreventive properties in former smokers.
To shed more light on the beneficial effects of 9-cis-RA on former smokers, we analyzed the effects of 9-cis-RA on the IGF axis, especially on the serum concentrations of IGF-I and IGFBP-3 and on the molar ratio of IGF-I to IGFBP-3, which has been proposed to reflect tissue bioactivity, in a previously studied population of former smokers.31 We found that 9-cis-RA treatment modulated the IGF axis in these former smokers, suggesting that the serum IGF axis is a target of the potential chemopreventive activities of 9-cis-RA in former smokers.
PATIENTS AND METHODS
Patients
The original study from which we derived our data was a three-arm, randomized, double-blinded, placebo-controlled trial comparing the effects of 9-cis-RA (100 mg) with those of 13-cis-RA (1 mg/kg) plus -tocopherol (AT; 1,200 U) administered for 3 months. In this study, a significant increase in RAR expression and a reduction of metaplasia were observed in individuals treated with 9-cis-RA, but not with 13-cis-RA plus AT, compared with those treated with placebo. Because antioxidants such as AT can affect the concentrations of the components of the IGF axis,32 we decided not to include the 13-cis-RA plus AT treatment group in our analysis. The eligibility criteria were previously described.29 Briefly, the study population consisted of former heavy smokers clinically free of any cancer, who were registered in the Departments of Thoracic/Head and Neck Medical Oncology and of Thoracic Surgery at The University of Texas M.D. Anderson Cancer Center. To be eligible, subjects had to have adequate renal, hematologic, and hepatic function and must not have taken more than 25,000 U of vitamin A or other retinoids daily for at least 3 months before study entry. Subjects were allowed to have had a prior smoking-related cancer, but they had to have been tumor free for 6 months before enrollment in the study. Subjects were required to abstain from consuming dietary vitamin supplements while on the study. The treatment duration was 3 months, based on the toxicity data from a previous phase I trial that included 9-cis-RA treatment.33 Subjects were seen monthly and were evaluated for compliance with the trial protocol, drug-related toxic effects, and serum cotinine concentrations.
The clinical trial from which the samples analyzed in this study were derived indicated that 9-cis-RA had some side effects. Specifically, subjects in the 9-cis-RA group experienced grade 2 (46 subjects) and grade 3 (nine subjects) toxic effects typical of retinoid treatment, including skin rash, hypertriglyceridemia, headache, cheilitis, conjunctivitis, arthralgia, and myalgia.29 Drug-related toxicity was graded according to the National Cancer Institute's Common Toxicity Criteria.34
The original study had been approved by the institutional review board of The M.D. Anderson Cancer Center and by the US Department of Health and Human Services. Our current study was also approved by the institutional review board at M.D. Anderson.
IGF-I and IGFBP-3 Measurements
For analysis of serum concentrations of IGF-I and IGFBP-3, blood samples were drawn from nonfasting subjects and collected in heparinized tubes that were transported immediately to the laboratory, where the samples were immediately centrifuged for 10 minutes at 4,000 x g and then stored at –80°C until the assays were performed. The serum concentration of IGF-I was measured by a specific radioimmunoassay (Diagnostic Systems Laboratories Inc, Webster, TX) with intra-assay and interassay coefficients of variation of less than 4% and 8%, respectively. To separate IGFs from their binding proteins, we mixed serum specimens with an acid-ethanol extraction buffer before measurement. The extraction procedure had been previously evaluated, and the efficiency of the extraction was identical to that for acid-column chromatography.35 The IGFBP-3 concentration was also measured by a specific radioimmunoassay (Diagnostic Systems Laboratories Inc) with intra-assay and interassay coefficients of variation of less than 3.5% and 7.5%, respectively, and no cross-reaction with other members of the IGFBP family. The assays were performed according to the instructions of the manufacturer and without knowledge of who the subject was. The molar ratio of IGF-I to IGFBP-3 was calculated as (0.130 x IGF-I concentration [ng/mL])/(0.036 x IGFBP-3 concentration [ng/mL]).
Statistical Analysis
The characteristics of the study subjects were compared pre–random assignment, according to sex or study group, by using Fisher's exact test for dichotomous characteristics and the Kruskal-Wallis test for quantitative characteristics. Because the distributions of serum concentrations of IGF-I and IGFBP-3 were skewed, the differences between groups were tested by using the Wilcoxon rank sum test. An overall treatment effect over time was determined by comparing the modulation (ie, the value at the subsequent visit minus the value at the baseline evaluation) in serum concentrations of IGF-I and IGFBP-3 and in the molar ratio of IGF-I to IGFBP-3 between the 9-cis-RA and placebo groups. All P values were determined by two-sided tests. Associations were considered statistically significant at P values less than .05. In the multivariate analysis, the variables of sex, age, smoking status, and body mass index (body weight in kg/height in m2) were included in the model.
RESULTS
Subject Characteristics
The characteristics of eligible subjects were described in detail previously.29 Of the 226 former smokers enrolled in the original chemoprevention trial, 149 were randomly assigned to placebo or 9-cis-RA treatments, and 113 of them completed 3 months of treatment. The characteristics of subjects assessable for our study are detailed in Table 1. Each treatment group was well balanced for sex, race, age, body mass index, and history of smoking. Blood samples from 111 of these subjects (52 women and 59 men; 56 from the 9-cis-RA group and 55 from the placebo group) at the baseline of the study, and from 84 (40 women and 44 men; 41 from the 9-cis-RA group and 43 from the placebo group) after 3 months of treatment were assessable for serum concentrations of IGF-I and IGFBP-3. The two treatment groups had comparable baseline mean serum concentrations of IGF-I and IGFBP-3 and similar molar ratios of IGF-I to IGFBP-3 (Table 2).
To determine whether subject characteristics affected the baseline serum concentrations of IGF-I or IGFBP-3, we evaluated the correlation between certain variables (eg, sex, race, age, body mass index, pack-years of smoking, and number of years since stopping smoking) and the baseline serum concentrations of IGF-I and IGFBP-3 and the molar ratio of IGF-I to IGFBP-3 (Table 3). The baseline serum concentration of IGF-I and the molar ratio of IGF-I to IGFBP-3 were significantly lower in women, whereas the IGFBP-3 concentration was slightly higher, though the difference did not reach statistical significance. Moreover, the serum concentration of IGF-I and the molar ratio of IGF-I to IGFBP-3 in women significantly decreased with increasing age (Fig 1A). Because serum concentrations of IGF-I are reduced in women treated with hormone replacement therapy (HRT),36,37 we analyzed whether the changes in the baseline serum concentration of IGF-I and the baseline molar ratio of IGF-I to IGFBP-3 were associated with HRT use among women. Self-reported data for HRT use were available for 38 of the 52 women (Table 3). Overall, baseline serum concentrations IGF-I and molar ratios of IGF-I to IGFBP-3 concentrations were significantly lower in the HRT users than the non–HRT users (Fig 1B). Among HRT users, differences in the IGFBP-3 concentrations were not significantly different. The mean baseline serum concentrations of IGF-I and IGFBP-3, and the molar ratio of IGF-I to IGFBP-3 in the HRT users are summarized in Table 3. Race, body mass index, number of pack-years, and years since stopping smoking did not affect the serum concentrations of IGF-I and IGFBP-3 or the molar ratio of IGF-I to IGFBP-3 in the study population.
Effect of 9-cis-RA on IGF-I and IGFBP-3 Serum Concentrations and on the IGF-I to IGFBP-3 Molar Ratio
The primary end point of the original lung cancer chemoprevention trial was restoration of RAR expression in the bronchial epithelium.29 In the previous study, we demonstrated that, compared with the effect of placebo, the median change in receptor index was significantly different from placebo for 9-cis-RA but not for 13-cis-RA plus AT,29 raising the possibility that 9-cis-RA has potential chemopreventive properties in former smokers.
In the current study, we evaluated the serum concentrations of the IGF axis in former smokers during treatment with 9-cis-RA. The modulations in mean serum concentrations of IGF-I and IGFBP-3 and in the molar ratio of IGF-I to IGFBP-3 in the two treatment groups during 3 months of treatment are illustrated in Figure 2. The mean changes in the placebo and 9-cis-RA groups are summarized in Table 4. Compared with the placebo group, the 9-cis-RA group exhibited a statistically significant modulation in the IGF axis. The mean changes from the study's baseline to the end of 3 months of treatment in the placebo and 9-cis-RA groups were as follows: IGF-I, 14.3 and –19.2, respectively; IGFBP-3, –175.1 and 196.6, respectively; and molar ratio of IGF-I to IGFBP-3, 0.05 and –0.06, respectively. These findings suggested that 9-cis-RA increases serum concentrations of IGFBP-3 and decreases serum concentrations of IGF-I, thus reducing the molar ratio of IGF-I to IGFBP-3.
To determine whether the differences in the serum concentrations of IGFBP3 between untreated and treated groups reflected differences at the tissue level, we also performed immunohistochemical analysis of IGFBP3, using the methodology described previously for lung cancer samples,20 in more than 90% of the cases. The intensity of IGFBP-3 expression in the normal and hyperplastic epithelial tissue samples was high, and differences between placebo–and 9-cis-RA–treated groups were not significant (data not shown). We suggest that the reason for the apparent discrepancy between the results of the plasma concentration analysis reported in this article and the tissue analysis is the higher sensitivity and dynamic range of the enzyme-linked immunosorbent assay for the plasma IGFBP3 and IGF1 compared with the limited dynamic range of the immunohistochemical analysis. In addition, the sources of IGF1 and IGFBP-3 in the blood include the liver and other tissues and is not expected to be directly related to expression in bronchial epithelial cells.
Correlation Between Changes in Tissue Level of RAR- and Modulation of the IGF Axis Induced by 9-cis-RA
We further evaluated the association between the changes in serum concentrations of the IGF axis peptides and changes in the tissue expression level of RAR during 9-cis-RA treatment. To determine the effect of treatment on loss of RAR expression at any biopsy site, the subjects were grouped according to whether their biopsy samples were positive (ie, RAR was detected in all six biopsy sites) or negative (ie, RAR was not detected in at least one biopsy site) for RAR expression, and the effects of treatment on RAR expression was determined as a binary variable (ie, loss of RAR expression at any biopsy site). The modulation of serum concentrations of IGF-I and IGFBP-3 and the molar ratio of IGF-I to IGFBP-3 were not significantly correlated with baseline RAR expression (Fig 3A) or with the changes in tissue RAR expression induced by 9-cis-RA (Fig 3B).
DISCUSSION
Our main finding in this analysis of data collected from a completed chemoprevention trial assessing the benefits of 9-cis-RA for former smokers was that the IGF axis can be modulated by 9-cis-RA in a population of former smokers. To our knowledge, this is the first report showing modulation of the IGF axis by 9-cis-RA in the setting of a chemoprevention trial.
Growing evidence supports an association between the IGF axis and the risk for lung cancer8 and suggests that the development of agents targeting the IGF axis could be an effective strategy in chemoprevention of the disease. Therefore, retinoids, shown to regulate the IGF axis in vitro22,38,39 have a potential to exert chemopreventive activities. Indeed, several findings have demonstrated the potential use of retinoids as chemopreventive agents. For example, 9-cis-RA has shown antiproliferative activity against a broad range of neoplastic cells, including those from prostate cancer,40 breast cancer,41,42 leukemia and lymphoma,43 lung cancer,44 and head and neck cancer.45 In vivo, 9-cis-RA has substantial anticarcinogenic activity in rat mammary glands46,47 and rat colons.48 More recently, the synthetic retinoid fenretinide has been shown to modulate circulating IGF-I and IGFBP-3 concentrations in breast cancer patients,49,50 and the relative risk of a second breast cancer was 35% lower in premenopausal women treated with fenretinide than in those who received no treatment.51 Thus, from the standpoint of directing future chemoprevention trials, our findings are important because we have shown both that clinical trials targeting the IGF axis in former smokers are feasible and that 9-cis-RA can modulate the IGF axis in this population.
To determine whether the characteristics of subjects affected the baseline IGF axis, we evaluated many variables, such as sex, age, nutritional status, and growth hormone secretion level, which have all been shown to affect the serum concentrations of IGF-I.52 The baseline serum level of IGF-I and the molar ratio of IGF-I to IGFBP-3 were significantly lower in women than in men. Estrogen is most likely responsible for the sex-related difference in the serum concentration of IGF-I. Endogenous estrogens have been shown to directly regulate circulating IGF-I synthesis53 and oral administration of estrogen decreases IGF-I serum concentrations.54,55 In our study, 38 of the 52 women took HRT during the treatment period, and the serum concentrations of IGF-I were significantly lower for HRT users than for non-HRT users. Therefore, the significant difference in serum concentrations of IGF-I and the molar ratio of IGF-I to IGFBP-3 might have resulted from the HRT. The role of estrogen in regulating the IGF-I level is further supported by the recent finding that the IGF-I concentrations of women using HRT were significantly lower than those of women not using HRT.37 Studies have suggested that HRT use is associated with a decreased risk for several types of cancer, including lung cancer, through decreases in the production of IGF-I.56,57 Therefore, if a high level of IGF-I is a putative risk factor for lung cancer, HRT use appears to lower IGF-I concentrations, thereby decreasing lung cancer risk.
We also observed that baseline serum concentrations of IGF-I and the molar ratio of IGF-I to IGFBP-3 were significantly decreased with increasing age in women. Duration of HRT use is also likely responsible for the age-related difference in the serum concentration of IGF-I in a female population because older women may conceivably have used HRT for a longer time. The decrease in IGF-I concentrations with aging has been described many times,49,58,59 and may be due in part to decreased growth hormone levels.60 In normal subjects, the interaction between growth hormone and its specific hepatic receptor stimulates expression of the IGF-I gene and the release of the IGF-I polypeptide.52 Therefore, it is likely that the concentration of IGF-I declines with age, as do secretions of growth hormone. However, we have not observed a significant correlation between the age of the study population in men and serum concentrations of IGF-I and IGFBP-3 or the molar ratio of IGF-I to IGFBP-3 in our study.
Growth hormone deficiency is also associated with a decreased muscle mass and increased body fat,61 and it has been hypothesized that low IGF-I concentrations are associated with high body fat, weight gain over time, and high body mass index.62 This hypothesis was supported by a study of Swedish men and women, which showed an inverse association between IGF-I concentrations and body mass index.53 However, we have not observed such correlation between IGF-I and body mass index, similar to the Rancho Bernardo study.59
The effects of smoking on serum concentrations of IGF-I are unclear. A positive association between IGF-I concentrations and pack-years of smoking and a negative association between pack-years or the number of cigarettes/d and concentrations of IGFBP-3 have been shown.63 However, an inverse association between IGF-I concentrations and smoking has been also reported among men, although not among women.53 Recently, Holmes et al58 showed that lower serum concentrations of IGF-I are significantly associated with smoking. In our study subjects, we observed a very modest decrease in IGF-I concentrations and an increase in IGFBP-3 concentrations associated with pack-years. However, smoking history generally did not affect the serum concentrations of IGF-I and IGFBP-3 or the molar ratio of IGF-I to IGFBP-3.
We observed that subjects in the 9-cis-RA group had a significant decrease in their IGF-I serum concentration, an increase in their IGFBP-3 serum concentration, and a decrease in the molar ratio of IGF-I to IGFBP-3 compared with subjects in the placebo group. The magnitude of the 9-cis-RA–induced changes in the IGF axis was moderate. This may be a consequence of the substantial interindividual variability in concentrations of 9-cis-RA and IGFs, which is at least in part genetically driven.33 Given the potential therapeutic activity of both exogenous growth hormone and IGF-I against heart failure64 and the long-term positive association between the decline in serum IGF-I concentrations and aging,65 a moderate yet durable effect of 9-cis-RA on the IGF axis may be desirable in a preventive context.
Although the exact mechanisms underlying the modulation of the IGF axis by 9-cis-RA are unclear, several in vitro results suggest the ability of retinoids to regulate the IGF axis; retinoids have been shown to regulate the expression of IGFBPs22,23,66; likewise, IGFs have been shown to modulate the cellular response of RAs, and vice versa.24 The induction of IGFBP-3 expression is activated by retinoids. We previously demonstrated that all-trans-RA increases IGFBP-3 expression at a transcriptional level through a RAR--dependent signaling pathway.22 It is well known that the cellular effects of RAs are mediated by RXRs and RARs. 9-cis-RA is a ligand for both RXRs and RARs, but its affinity for RXRs is 40-fold higher than for RARs. On the other hand, RA is primarily a ligand for RARs and only activates RXRs at high concentrations.67 Recently, the vitamin D receptor has been identified in the IGFBP-3 promoter, and RXR has been shown to be required for 1,25-dihydroxyvitamin D3–induced gene transcription.68 Therefore, treatment with 9-cis-RA could lead to there being available ligand for activation of both the RAR:RXR and the VDR:RXR heterodimeric complex, so that IGFBP-3 gene transcription could be efficiently activated.
Given the inverse association between estrogen and serum IGF-I concentrations, the ability of 9-cis-RA to regulate type 1 17-hydroxysteroid dehydrogenases,69 which catalyzes the conversion of estrone and 17-estradiol, could increase the level of 17-estradiol, the physiologically significant molecule of estrogen, and thereby decrease IGF-I concentrations. The ability of estrogens to increase the expression of vitamin D receptors in vitro70 may also contribute to decreased concentrations of IGF-I by inducing IGFBP-3 expression.71 Additional work will be necessary to investigate the mechanism that mediates the regulation of the IGF axis by 9-cis-RA in former smokers.
Because 9-cis-RA treatment increased the tissue expression of RAR, we also explored the correlation between tissue levels of RAR and serum concentrations of the IGF axis. However, we did not find any significant correlation between the modulation of the serum concentrations of IGF-I or IGFBP-3 and RAR expression in the tissue. These findings provide evidence that the mechanisms involved in 9-cis-RA–mediated gene expression are diverse and complex.
In conclusion, we are the first to show that 3 months of treatment with 9-cis-RA decreased the serum level of IGF-I and the molar ratio of IGF-I to IGFBP-3 and increased the serum level of IGFBP-3 in former smokers. These effects may contribute to the chemopreventive benefit of 9-cis-RA to former smokers. Despite these promising findings, enthusiasm for the use of 9-cis-RA as a chemopreventive agent for lung cancer could be tempered by the toxic effects of the agent.29 Although there were no serious side effects such as cardiovascular problems, pancreatitis, or death in the subjects treated with 9-cis-RA in our previous prevention study, and only one patient with grade 4 hypertriglyceridemia stopped 9-cis-RA treatment, it would be better to develop newer agents that are related to 9-cis-RA but that do not have the toxic effects of 9-cis-RA. Because 9-cis-RA can activate both RARs and RXRs and because the toxicities are thought to be mediated by RAR-RXR heterodimers, it had been suggested that "pure" RXR selective synthetic retinoids may exert the beneficial effects of retinoids without the toxicities. Indeed, a group of RXR-selective retinoids ("rexinoids") has demonstrated efficacy with fewer adverse effects in patients with NSCLC in early clinical trials.72 Therefore, whether rexinoids regulate the IGF axis and thereby reduce lung cancer risk would be a worthwhile topic for future chemoprevention trials in former smokers. In addition, because Ras-mediated signaling pathways may participate in the development of resistance to IGFBP-3,73 the combination of RXR-selective agonists and inhibitors of the Ras-mediated signaling pathway could also be considered. Clearly, additional work will be necessary to determine whether the modulation of the IGF axis by 9-cis-RA correlates with the ability to reduce lung cancer risk in former smokers. In addition, further investigation into the role of serum concentrations of IGF-I and IGFBP-3 as surrogate biomarkers in determining the chemopreventive effects of retinoids are warranted.
Authors' Disclosures of Potential Conflicts of Interest
The authors indicated no potential conflicts of interest.
NOTES
Supported in part by National Institutes of Health Grants U19 CA68437 (W.K.H.), R01 CA109520-01 (H.-Y.L.), CA-100816-01A1 (H.-Y.L), American Cancer Society grant RSG-04-082-01-TBE 01(H.-Y.L), and W81XWH-04-1-0142-01-VITAL from the Department of Defense (W.K.H.). W.K.H. is an American Cancer Society Clinical Research Professor.
Presented in part at the 40th Annual Meeting of the American Society of Clinical Oncology, New Orleans, LA, June 5-8, 2004.
Authors' disclosures of potential conflicts of interest are found at the end of this article.
REFERENCES
Khuri FR, Herbst RS, Fossella FV: Emerging therapies in non-small-cell lung cancer. Ann Oncol 12:739-744, 2001
Parkin DM, Pisani P, Ferlay J: Estimates of the worldwide incidence of 25 major cancers in 1990. Int J Cancer 80:827-841, 1999
Ginsberg RJ, Vokes EE, Kenneth R: Non-small cell lung cancer, in DeVita VT Jr, Hellman S, Rosenberg SA (eds): Cancer: Principles and Practice of Oncology (ed 6). Philadelphia, PA, Lippincott Williams & Wilkins, 2001, pp 925-983
Mattson ME, Pollack ES, Cullen JW: What are the odds that smoking will kill you Am J Public Health 77:425-431, 1987
Wistuba II, Lam S, Behrens C, et al: Molecular damage in the bronchial epithelium of current and former smokers. J Natl Cancer Inst 89:1366-1373, 1997
Sekido Y, Fong KM, Minna JD: Progress in understanding the molecular pathogenesis of human lung cancer. Biochim Biochim Biophys Acta 1378:21-59, 1998
Burns DM: Primary prevention, smoking, and smoking cessation: Implications for future trends in lung cancer prevention. Cancer 89:2506-2509, 2000
Ibrahim YH, Yee D: Insulin-like growth factor-I and cancer risk. Growth Horm IGF Res 14:261-269, 2004
Butt AJ, Firth SM, Baxter RC: The IGF axis and programmed cell death. Immunol Cell Biol 77:256-262, 1999
Lopez T, Hanahan D: Elevated levels of IGF-1 receptor convey invasive and metastatic capability in a mouse model of pancreatic islet tumorigenesis. Cancer Cell 1:339-353, 2002
Samani AA, Chevet E, Fallavollita L, et al: Loss of tumorigenicity and metastatic potential in carcinoma cells expressing the extracellular domain of the type 1 insulin-like growth factor receptor. Cancer Res 64:3380-3385, 2004
Yu H, Spitz MR, Mistry J, et al: Plasma levels of insulin-like growth factor-I and lung cancer risk: A case-control analysis. J Natl Cancer Inst 91:151-156, 1999
Wu X, Tortolero-Luna G, Zhao H, et al: Serum levels of insulin-like growth factor I and risk of squamous intraepithelial lesions of the cervix. Clin Cancer Res 9:3356-3361, 2003
Pollak M: Insulin-like growth factors (IGFs) and prostate cancer. Epidemiol Rev 23:59-66, 2001
Lukanova A, Lundin E, Toniolo P, et al: Circulating levels of insulin-like growth factor-1 and risk of ovarian cancer. Int J Cancer 101:549-554, 2002
Zhao H, Grossman HB, Spitz MR, et al: Plasma levels of insulin-like growth factor-1 and binding protein-3, and their association with bladder cancer risk. J Urol 169:714-717, 2003
Kaaks R, Toniolo P, Akhmedkhanov A, et al: Serum C-peptide, insulin-like growth factor (IGF)-I, IGF binding proteins, and colorectal cancer risk in women. J Natl Cancer Inst 92:1592-1600, 2000
Chang YS, Wang L, Mao L, et al: Mechanisms underlying lack of insulin like-growth factor binding protein-3 expression in NSCLC cells. Oncogene 23:6569-6580, 2004
Chang YS, Wang L, Liu D, et al: Correlation between IGFBP-3 promoter methylation and prognosis of patients with stage I non-small cell lung cancer. Clin Cancer Res 8:3669-3675, 2002
Chang YS, Gong K, Sun S, et al: Clinical significance of IGFBP-3 expression in stage I non-small cell lung cancer. Clin Cancer Res 8:3796-3802, 2002
Lee HY, Chun KH, Liu B, et al: Insulin-like growth factor binding protein-3 inhibits the growth of non-small cell lung cancer. Cancer Res 62:3530-3537, 2002
Han G-R, Dohi DF, Lee H-Y, et al: All-trans-retinoic acid increases transforming growth factor-2 and insulin-like growth factor binding protein-3 expression through a retinoic acid receptor--dependent signaling pathway. J Biol Chem 272:13711-13716, 1997
Fontana JA, Burrows-Mezu A, Clemmons DR, et al: Retinoid modulation of insulin-like growth factor-binding proteins and inhibition of breast carcinoma proliferation. Endocrinology 128:1115-1122, 1991
Bentel JM, Lebwohl DE, Cullen KJ, et al: Insulin-like growth factors modulate the growth inhibitory effects of retinoic acid on MCF-7 breast cancer cells. J Cell Physiol 165:212-221, 1995
Nagy L, Thomazy VA, Heyman RA, et al: Retinoid-induced apoptosis in normal and neoplastic tissues. Cell Death Differ 5:11-19, 1998
Hansen LA, Sigman CC, Andreola F, et al: Retinoids in chemoprevention and differentiation therapy. Carcinogenesis 21:1271-1279, 2000
Mangelsdorf DJ, Evans RM: The RXR heterodimers and orphan receptors. Cell 83:841-850, 1995
Marchetti MN, Samplo E, Bun H, et al: In vitro metabolism of three major isomers of retinoic acid in rats: Intersex and interstrain comparison. Drug Metab Dispos 25:637-646, 1997
Kurie JM, Lotan R, Lee JJ, et al: Treatment of former smokers with 9-cis-retinoic acid reverses loss of retinoic acid receptor-expression in the bronchial epithelium: Results from a randomized placebo-controlled trial. J Natl Cancer Inst 95:206-214, 2003
Seewaldt VL, Johnson BS, Parker MB, et al: Expression of retinoic acid receptor beta mediates retinoic acid-induced growth arrest and apoptosis in breast cancer cells. Cell Growth Differ 6:1077-1088, 1995
Juul A, Dalgaard P, Blum W, et al: Serum levels of insulin-like growth factor (IGF)-binding protein-3 (IGFBP-3) in healthy infants, children, and adolescents: The relation to IGF-I, IGF-II, IGFBP-1, IGFBP-2, age, sex, body mass index, and pubertal maturation. J Clin Endocrinol Metab 80:2534-2542, 1995
Zi X, Zhang J, Agarwal R, et al: Silibinin up-regulates insulin-like growth factor-binding protein 3 expression and inhibits proliferation of androgen-independent prostate cancer cells. Cancer Res 60:5617-5620, 2000
Miller VA, Rigas JR, Benedetti FM, et al: Initial clinical trial of the retinoid receptor pan agonist 9-cis retinoic acid. Clin Cancer Res 6:471-475, 1996
National Cancer Institute: Common toxicity criteria, version 2.0. http://ctep.cancer.gov/forms/CTCv20_4-30-992.pdf
Baxter RC, Martin JL: Radioimmunoassay of growth hormone-dependent insulin-like growth factor binding protein in human plasma. J Clin Invest 78:1504-1512, 1986
Campagnoli C, Biglia N, Altare F, et al: Differential effects of oral conjugated estrogens and transdermal estradiol on insulinlike growth factor 1, growth hormone and sex hormone binding globulin serum levels. Gynecol Endocrinol 7:251-258, 1993
Schabath MB, Wu X, Sellin RV, et al: Hormone replacement therapy and lung cancer risk: A case-control analysis. Clin Cancer Res 10:113-123, 2004
Zhou Y, Mohan S, Linkhart TA, et al: Retinoic acid regulates insulin-like growth factor-binding protein expression in human osteoblast cells. Endocrinology 137:975-983, 1996
Gabbitas B, Canalis E: Retinoic acid regulates the expression of insulin-like growth factors I and II in osteoblasts. J Cell Physiology 172:253-264, 1997
Blutt SE, Allegretto EA, Pike JW, et al: 1,25-Dihydroxyvitamin D3 and 9-cis-retinoic acid act synergistically to inhibit the growth of LNCaP prostate cells and cause accumulation of cells in G1. Endocrinology 138:1491-1497, 1997
Rubin M, Fenig E, Rosenauer A, et al: 9-cis-Retinoic acid inhibits growth of breast cancer cells and down-regulates estrogen receptor RNA and protein. Cancer Res 54:6549-6556, 1994
Gottardis MM, Lamph WW, Shalinsky DR, et al: The efficacy of 9-cis-retinoic acid in experimental models of cancer. Breast Cancer Res Treat 38:85-96, 1996
Lutzky J, Vujicic M, Yamanishi DT, et al: Antiproliferative effects of all-trans-retinoic acid (tRA) and 9-cis-retinoic acid (9-cisRA) on human lymphoid cell lines. Proc Am Assoc Cancer Res 34:292, 1993 (abstr 1738)
Guzey M, Demirpence E, Criss W, et al: Effects of retinoic acid (all-trans and 9-cis) on tumor progression in small-cell lung carcinoma. Biochem Biophys Res Commun 242:369-375, 1998
Giannini F, Maestro R, Vukosavljevic T, et al: All-trans, 13-cis, and 9-cis retinoic acids induce a fully reversible growth inhibition in HNSCC cell lines: Implications for in vivo retinoic acid use. Int J Cancer 70:194-200, 1997
Anzano MA, Byers SW, Smith JM, et al: Prevention of breast cancer in the rat with 9-cis-retinoic acid as a single agent and in combination with tamoxifen. Cancer Res 54:4614-4617, 1994
Anzano MA, Peer CW, Smith JM, et al: Chemoprevention of mammary carcinogenesis in the rat: Combined use of raloxifene and 9-cis-retinoic acid. J Natl Cancer Inst 88:123-125, 1996
Zheng Y, Kramer PM, Olson G, et al: Prevention by retinoids of azoxymethane-induced tumors and aberrant crypt foci and their modulation of cell proliferation in the colon of rats. Carcinogenesis 18:2119-2125, 1997
Decensi A, Johansson H, Miceli R, et al: Long-term effects of fenretinide, a retinoic acid derivative, on the insulin-like growth factor system in women with early breast cancer. Cancer Epidemiol Biomarkers Prev 10:1047-1053, 2001
Torrisi R, Parodi S, Fontana V, et al: Effect of fenretinide on plasma IGF-I and IGFBP-3 in early breast cancer patients. Int J Cancer 76:787-790, 1998
Veronesi U, De Palo G, Marubini E, et al: Randomized trial of fenretinide to prevent second breast malignancy in women with early breast cancer. J Natl Cancer Inst 91:1847-1856, 1999
Le Roith D: Seminars in medicine of the Beth Israel Deaconess Medical Center: Insulin-like growth factors. N Engl J Med 336:633-640, 1997
Landin-Wilhelmsen K, Wilhelmsen L, Lappas G, et al: Serum insulin-like growth factor I in a random population sample of men and women: Relation to age, sex, smoking habits, coffee consumption and physical activity, blood pressure and concentrations of plasma lipids, fibrinogen, parathyroid hormone and osteocalcin. Clin Endocrinol (Oxf) 41:351-357, 1994
Helle SI, Omsjo IH, Hughes SC, et al: Effects of oral and transdermal oestrogen replacement therapy on plasma levels of insulin-like growth factors and IGF binding proteins 1 and 3: a cross-over study. Clin Endocrinol (Oxf) 45:727-732, 1996
Raudaskoski T, Knip M, Laatikainen T: Plasma insulin-like growth factor-I and its binding proteins 1 and 3 during continuous non-oral and oral combined hormone replacement therapy. Menopause 5:217-222, 1998
Janne PA, Mayer RJ: Chemoprevention of colorectal cancer. N Engl J Med 342:1960-1968, 2000
Grodstein F, Newcomb PA, Stampfer MJ: Postmenopausal hormone therapy and the risk of colorectal cancer: A review and meta-analysis. Am J Med 106:574-578, 1999
Holmes MD, Pollak MN, Hankinson SE: Lifestyle correlates of plasma insulin-like growth factor I and insulin-like growth factor binding protein 3 concentrations. Cancer Epidemiol Biomarkers Prev 11:862-867, 2002
Goodman-Gruen D, Barrett-Connor E: Epidemiology of insulin-like growth factor-I in elderly men and women: The Rancho Bernardo Study. Am J Epidemiol 146:970-976, 1997
Bondy CA, Underwood LE, Clemmons DR, et al: Clinical uses of insulin-like growth factor I. Ann Intern Med 120:593-601, 1994
Sonksen PH, Salomon F, Cuneo R: Metabolic effects of hypopituitarism and acromegaly. Horm Res 36:27-31, 1991
Harris T, Kiel D, Roubenoff R, et al: Association of insulin-like growth factor-I with body composition, weight history, and past health behaviors in the very old: The Framingham Heart Study. J Am Geriatr Soc 45:133-139, 1997
Kaklamani VG, Linos A, Kaklamani E, et al: Age, sex, and smoking are predictors of circulating insulin-like growth factor 1 and insulin-like growth factor-binding protein 3. J Clin Oncol 17:813-817, 1999
Fazio S, Sabatini D, Capaldo B, et al: A preliminary study of growth hormone in the treatment of dilated cardiomyopathy. N Engl J Med 334:809-814, 1996
Janssen JA, Stolk RP, Pols HA, et al: Serum free IGF-I, total IGF-I, IGFBP-1 and IGFBP-3 levels in an elderly population: Relation to age and sex steroid levels. Clin Endocrinol (Oxf) 48:471-478, 1998
Woodward TL, Turner JD, Hung HT, et al: Inhibition of cellular proliferation and modulation of insulin-like growth factor binding proteins by retinoids in a bovine mammary epithelial cell line. J Cell Physiol 167:488-499, 1996
Heyman RA, Mangelsdorf DJ, Dyck JA, et al: 9-cis Retinoic acid is a high affinity ligand for the retinoid X receptor. Cell 68:397-406, 1992
Pathrose P, Barmina O, Chang CY, et al: Inhibition of 1,25-dihydroxyvitamin D3-dependent transcription by synthetic LXXLL peptide antagonists that target the activation domains of the vitamin D and retinoid X receptors. J Bone Miner Res 17:2196-2205, 2002
Piao Y-S, Peltoketo H, Jouppila A, et al: Retinoic acids increase 17-hydroxysteroid dehydrogenase type 1 expression in JEG-3 and T47D cells, but the stimulation is potentiated by epidermal growth factor, 12-O-tetradecanoulphorbol-13-acetate, and cyclic adenosine 3', 5'-monophosphate only in JEG-3 cells. Endocrinology 138:898-904, 1997
Smirnoff P, Liel Y, Gnainsky J, et al: The protective effect of estrogen against chemically induced murine colon carcinogenesis is associated with decreased CpG island methylation and increased mRNA and protein expression of the colonic vitamin D receptor. Oncol Res 11:255-264, 1999
Boyle BJ, Zhao XY, Cohen P, et al: Insulin-like growth factor binding protein-3 mediates 1,25-dihydroxyvitamin D3 growth inhibition in the LNCaP prostate cancer cell line through p21/WAF1. J Urol 165:1319-1324, 2001
Rizvi NA, Marshall JL, Dahut W, et al: A phase I study of LGD1069 in adults with advanced cancer. Clin Cancer Res 5:1658-1664, 1999
Lee H-Y, Moon HJ, Chun K-H, et al: Effects of insulin-like growth factor binding protein-3 and farnesyltransferase inhibitor SCH66336 on Akt expression and apoptosis in non–small-cell lung cancer cells. J Natl Cancer Inst 96:1536-1548, 2004(Ho-Young Lee, Yoon Soo Ch)