An Acquired Vulnerability of Drug-Resistant Melanoma with Therapeutic Potential.

BRAF(V600E) mutant melanomas treated with inhibitors of the BRAF and MEK kinases almost invariably develop resistance that is frequently caused by reactivation of the mitogen activated protein kinase (MAPK) pathway. To identify novel treatment options for such patients, we searched for acquired vulnerabilities of MAPK inhibitor-resistant melanomas. We find that resistance to BRAF+MEK inhibitors is associated with increased levels of reactive oxygen species (ROS). Subsequent treatment with the histone deacetylase inhibitor vorinostat suppresses SLC7A11, leading to a lethal increase in the already-elevated levels of ROS in drug-resistant cells. This causes selective apoptotic death of only the drug-resistant tumor cells. Consistently, treatment of BRAF inhibitor-resistant melanoma with vorinostat in mice results in dramatic tumor regression. In a study in patients with advanced BRAF+MEK inhibitor-resistant melanoma, we find that vorinostat can selectively ablate drug-resistant tumor cells, providing clinical proof of concept for the novel therapy identified here.

EGFR activation limits the response of liver cancer to lenvatinib.

Hepatocellular carcinoma (HCC)-the most common form of liver cancer-is an aggressive malignancy with few effective treatment options1. Lenvatinib is a small-molecule inhibitor of multiple receptor tyrosine kinases that is used for the treatment of patients with advanced HCC, but this drug has only limited clinical benefit2. Here, using a kinome-centred CRISPR-Cas9 genetic screen, we show that inhibition of epidermal growth factor receptor (EGFR) is synthetic lethal with lenvatinib in liver cancer. The combination of the EGFR inhibitor gefitinib and lenvatinib displays potent anti-proliferative effects in vitro in liver cancer cell lines that express EGFR and in vivo in xenografted liver cancer cell lines, immunocompetent mouse models and patient-derived HCC tumours in mice. Mechanistically, inhibition of fibroblast growth factor receptor (FGFR)  by lenvatinib treatment leads to feedback activation of the EGFR-PAK2-ERK5 signalling axis, which is blocked by EGFR inhibition. Treatment of 12 patients with advanced HCC who were unresponsive to lenvatinib treatment with the combination of lenvatinib plus gefitinib (trial identifier NCT04642547) resulted in meaningful clinical responses. The combination therapy identified here may represent a promising strategy for the approximately 50% of patients with advanced HCC who have high levels of EGFR.

Small-molecule inhibition of MAP2K4 is synergistic with RAS inhibitors in KRAS-mutant cancers.

The Kirsten rat sarcoma viral oncogene homologue KRAS is among the most commonly mutated oncogenes in human cancers, thus representing an attractive target for precision oncology. The approval for clinical use of the first selective inhibitors of G12C mutant KRAS therefore holds great promise for cancer treatment. However, despite initial encouraging clinical results, the overall survival benefit that patients experience following treatment with these inhibitors has been disappointing to date, pointing toward the need to develop more powerful combination therapies. Here, we show that responsiveness to KRASG12C and pan-RAS inhibitors in KRAS-mutant lung and colon cancer cells is limited by feedback activation of the parallel MAP2K4-JNK-JUN pathway. Activation of this pathway leads to elevated expression of receptor tyrosine kinases that reactivate KRAS and its downstream effectors in the presence of drug. We find that the combination of sotorasib, a drug targeting KRASG12C, and the MAP2K4 inhibitor HRX-0233 prevents this feedback activation and is highly synergistic in a panel of KRASG12C-mutant lung and colon cancer cells. Moreover, combining HRX-0233 and sotorasib is well-tolerated and resulted in durable tumor shrinkage in mouse xenografts of human lung cancer cells, suggesting a therapeutic strategy for KRAS-driven cancers.

Trajectories of sickness absence and disability pension days among people with multiple sclerosis by type of occupation.

BACKGROUND: Multiple sclerosis (MS) can impact working life, sickness absence (SA) and disability pension (DP). Different types of occupations involve different demands, which may be associated with trajectories of SA/DP among people with MS (PwMS). OBJECTIVES: To explore, among PwMS and references, if SA/DP differ according to type of occupation. Furthermore, to examine how trajectories of SA/DP days are associated with type of occupation among PwMS. METHODS: A longitudinal nationwide Swedish register-based cohort study was conducted, including 6100 individuals with prevalent MS and 38,641 matched references from the population. Trajectories of SA/DP were identified with group-based trajectory modelling. Multinomial logistic regressions were estimated for associations between identified trajectories and occupations. RESULTS: Increase of SA/DP over time was observed in all occupational groups, in both PwMS and references, with higher levels of SA/DP among PwMS. The lowest levels of SA/DP were observed among managers. Three trajectory groups of SA/DP were identified: Persistently Low (55.2%), Moderate Increasing (31.9%) and High Increasing (12.8%). Managers and those working in Science & Technology, and Economics, Social & Cultural were more likely to belong to the Persistently Low group. CONCLUSION: Results suggest that type of occupation plays a role in the level and course of SA/DP.

Comparative Network Reconstruction using mixed integer programming.

Motivation: Signal-transduction networks are often aberrated in cancer cells, and new anti-cancer drugs that specifically target oncogenes involved in signaling show great clinical promise. However, the effectiveness of such targeted treatments is often hampered by innate or acquired resistance due to feedbacks, crosstalks or network adaptations in response to drug treatment. A quantitative understanding of these signaling networks and how they differ between cells with different oncogenic mutations or between sensitive and resistant cells can help in addressing this problem. Results: Here, we present Comparative Network Reconstruction (CNR), a computational method to reconstruct signaling networks based on possibly incomplete perturbation data, and to identify which edges differ quantitatively between two or more signaling networks. Prior knowledge about network topology is not required but can straightforwardly be incorporated. We extensively tested our approach using simulated data and applied it to perturbation data from a BRAF mutant, PTPN11 KO cell line that developed resistance to BRAF inhibition. Comparing the reconstructed networks of sensitive and resistant cells suggests that the resistance mechanism involves re-establishing wild-type MAPK signaling, possibly through an alternative RAF-isoform. Availability and implementation: CNR is available as a python module at https://github.com/NKI-CCB/cnr. Additionally, code to reproduce all figures is available at https://github.com/NKI-CCB/CNR-analyses. Supplementary information: Supplementary data are available at Bioinformatics online.

The BRCA1ness signature is associated significantly with response to PARP inhibitor treatment versus control in the I-SPY 2 randomized neoadjuvant setting.

BACKGROUND: Patients with BRCA1-like tumors correlate with improved response to DNA double-strand break-inducing therapy. A gene expression-based classifier was developed to distinguish between BRCA1-like and non-BRCA1-like tumors. We hypothesized that these tumors may also be more sensitive to PARP inhibitors than standard treatments. METHODS: A diagnostic gene expression signature (BRCA1ness) was developed using a centroid model with 128 triple-negative breast cancer samples from the EU FP7 RATHER project. This BRCA1ness signature was then tested in HER2-negative patients (n = 116) from the I-SPY 2 TRIAL who received an oral PARP inhibitor veliparib in combination with carboplatin (V-C), or standard chemotherapy alone. We assessed the association between BRCA1ness and pathologic complete response in the V-C and control arms alone using Fisher's exact test, and the relative performance between arms (biomarker × treatment interaction, likelihood ratio p < 0.05) using a logistic model and adjusting for hormone receptor status (HR). RESULTS: We developed a gene expression signature to identify BRCA1-like status. In the I-SPY 2 neoadjuvant setting the BRCA1ness signature associated significantly with response to V-C (p = 0.03), but not in the control arm (p = 0.45). We identified a significant interaction between BRCA1ness and V-C (p = 0.023) after correcting for HR. CONCLUSIONS: A genomic-based BRCA1-like signature was successfully translated to an expression-based signature (BRC1Aness). In the I-SPY 2 neoadjuvant setting, we determined that the BRCA1ness signature is capable of predicting benefit of V-C added to standard chemotherapy compared to standard chemotherapy alone. TRIAL REGISTRATION: I-SPY 2 TRIAL beginning December 31, 2009: Neoadjuvant and Personalized Adaptive Novel Agents to Treat Breast Cancer (I-SPY 2), NCT01042379 .

An α-E-catenin (CTNNA1) mutation in hereditary diffuse gastric cancer.

Diffuse gastric cancers typically present as late-stage tumours and, as a result, the 5 year survival rate is poor. Some gastric cancers are hereditary and these tend to be of the diffuse type; 30-40% of hereditary diffuse gastric cancers (HDGCs) can be explained by defective germline alleles of E-cadherin (CDH1), but for the remaining families the factors driving susceptibility remain unknown. We had access to a large HDGC pedigree with no obvious mutation in CDH1, and applied exome sequencing to identify new genes involved in gastric cancer. We identified a germline truncating allele of α-E-catenin (CTNNA1) that was present in two family members with invasive diffuse gastric cancer and four in which intramucosal signet ring cells were detected as part of endoscopic surveillance. The remaining CTNNA1 allele was silenced in the two diffuse gastric cancers from the family that were available for screening, and this was also true for signet ring cells identified in endoscopic biopsies. Since α-E-catenin functions in the same complex as E-cadherin, our results call attention to the broader signalling network surrounding these proteins in HDGC. We also detected somatic mutations in one tumour and found substantial overlap with genes mutated in sporadic gastric cancer, including PIK3CA, ARID1A, MED12 and MED23.

Identification of recurrent FGFR3 fusion genes in lung cancer through kinome-centred RNA sequencing.

Oncogenic fusion genes that involve kinases have proven to be effective targets for therapy in a wide range of cancers. Unfortunately, the diagnostic approaches required to identify these events are struggling to keep pace with the diverse array of genetic alterations that occur in cancer. Diagnostic screening in solid tumours is particularly challenging, as many fusion genes occur with a low frequency. To overcome these limitations, we developed a capture enrichment strategy to enable high-throughput transcript sequencing of the human kinome. This approach provides a global overview of kinase fusion events, irrespective of the identity of the fusion partner. To demonstrate the utility of this system, we profiled 100 non-small cell lung cancers and identified numerous genetic alterations impacting fibroblast growth factor receptor 3 (FGFR3) in lung squamous cell carcinoma and a novel ALK fusion partner in lung adenocarcinoma.

Support needs for return to work among self-employed workers: A focus group study.

OBJECTIVE: The aim of this study is to gain insight into the facilitators, barriers, and support needs of Dutch self-employed workers when returning to work (RTW) after sick leave. METHODS: Three focus groups were conducted, involving 15 Dutch self-employed workers who were on sick leave due to health problems. The transcripts were analysed through thematic content analysis. RESULTS: Five main themes regarding barriers, facilitators and needs of self-employed workers to RTW were identified: autonomy, social support, client management, financial security and information on sick leave. Having autonomy was considered a facilitator for RTW. However, the participants expressed a need for more financial support, additional guidance from occupational health professionals, and tailored information regarding RTW. CONCLUSION: The results of this study emphasize a need for optimizing national policy and support for self-employed workers regarding sick leave and RTW.

Barriers and Facilitators to Participation and Key Components of Sleep Health Programs: Perspectives for the Corporate Work Environment.

OBJECTIVE: The aim of the study is to explore the barriers and facilitators of participation and key components for sleep health programs designed for corporate work environments. METHODS: Semistructured interviews with corporate executives and occupational medicine specialists in the decision making and management of workplace health promotion programs (WHPP) within their companies were held before and during COVID-19. Interviews were transcribed verbatim and analyzed using thematic content analysis to identify themes. RESULTS: Barrier and facilitator themes emerging from the data include sleep health awareness, work culture, work-family balance, and confidentiality. Key components for sleep health programs included the following: identifying the need for a program, incorporating sleep health risk screening to WHPP, and promoting sleep health by raising awareness thereof. CONCLUSIONS: The identified barriers and facilitators to employee participation and key components of an ideal sleep health program provide guidance for further WHPP.

A roadmap for sustainable implementation of vocational rehabilitation for people with mental disorders and its outcomes: a qualitative evaluation.

BACKGROUND: People suffering from mental health disorders have lower work participation compared to people without mental challenges. To increase work participation within this group vocational rehabilitation interventions are often offered. Collaboration between the mental health care and social security sectors is needed to enable professionals to perform optimally when carrying out these interventions. Yet, regulatory and financial barriers often hinder sustainable implementation. To overcome these barriers an experimental roadmap for sustainable funding based on a shared savings strategy was piloted in four regions. The aim of the present qualitative study was to gain understanding of the uses of this roadmap and the factors that were important in the experiment's process. METHOD: The roadmap consisted of five steps based upon insights from shared savings strategies and implementation science knowledge, and was initiated by a national steering board. The roadmap aimed to make sustainable funding agreements (based on shared savings) for the implementation of a vocational rehabilitation intervention. In four regions, stakeholders from the mental health care and social security services sector followed the roadmap. We conducted interviews (n = 16) with involved participants and project leaders of the experiment and collected 54 sets of field notes and documents to evaluate the roadmap process. A thematic analysis was used to analyse the data. RESULTS: Regions perceived improved stakeholder collaboration around vocational rehabilitation after they were guided by the roadmap. Three regions made, or intended to make, agreements on collaboration and funding, yet not based on shared savings. Moreover, going through the roadmap took more time than anticipated. Stakeholder collaboration depended on factors like personal and organizational interests and collaboration conditions and values. Financial legislation and politics were regarded as barriers and personal motives were mentioned as a facilitator in this process. CONCLUSIONS: Our study showed that the roadmap supported stakeholders to establish a more sustainable collaboration, even though no sustainable financial agreements were made yet. Although participants acknowledged the function of financial insights and the need for financial resources, the driver for collaboration was found to be more on improving clients' perspectives than on solving unfair financial distribution issues. This suggests modifying the focus of the roadmap from financial benefits to improving clients' perspectives.

Rational combination of SHP2 and mTOR inhibition for the treatment of hepatocellular carcinoma.

Liver cancer is the fourth most common cause of cancer-related death worldwide, with hepatocellular carcinoma (HCC) being the main primary malignancy affecting the liver. Unfortunately, there are still limited therapeutic options for HCC, and even the latest advances have only increased the overall survival modestly. Thus, new treatment strategies and rational drug combinations are urgently needed. Reactivation of receptor tyrosine kinases (RTK) has been described as a mechanism of intrinsic resistance to targeted therapies in a variety of cancers, including inhibitors of mTOR. The design of rational combination therapies to overcome this type of resistance is complicated by the notion that multiple RTK can be upregulated during the acquisition of resistance. SHP2, encoded by the gene PTPN11, acts downstream of virtually all RTK, and has proven to be a good target for small molecule inhibitors. Here, we report activation of multiple RTK upon mTOR inhibition in HCC which, through SHP2, leads to reactivation of the mTOR pathway. We show that co-inhibition of both mTOR and SHP2 is highly synergistic in vitro by triggering apoptosis. More importantly, the combination is well-tolerated and outperforms the monotherapies in impairing tumor growth in multiple HCC mouse models. Our findings suggest a novel rational combination therapy for the treatment of HCC.

Participatory Approach to Create a Supportive Work Environment for Employees With Chronic Conditions: A Pilot Implementation Study.

OBJECTIVE: To evaluate a pilot implementation of an organizational-level intervention. The participatory approach (PA) was used to create a supportive work environment for employees with chronic conditions, with a key role for occupational physicians (OPs). METHODS: Twenty-eight semi-structured interviews were conducted with OPs and stakeholders within their organizations. Furthermore, observational data and research notes were gathered. Data analysis occurred through content analysis. RESULTS: Recruitment of organizations was challenging, with a reach of 25%. Dose delivered, dose received, and fidelity differed across the three organizations. Organizations were positive about the PA as a method to improve support for employees with chronic conditions. CONCLUSIONS: The PA could be of added value for creating a supportive work environment. However, research is needed on activating organizations to improve support for employees with chronic conditions.

Workers' views on involving significant others in occupational health care: a focus group study among workers with a chronic disease.

PURPOSE: To explore workers' views and considerations on involving their significant others (SOs) in occupational health care. METHODS: Four focus group interviews in the Netherlands, with 21 workers who had visited an occupational health physician (OHP) due to work absence caused by a chronic disease. Data was analyzed using thematic analysis. RESULTS: We distinguished four main themes: (i) attitudes towards involving SOs, (ii) preferences on how to involve SOs, (iii) benefits of involving SOs, and (iv) concerns with regard to involving SOs. Workers expressed both positive and critical opinions about involving SOs in occupational health care. Benefits mentioned included provision of emotional and informational support by SOs before, during, and after consultations. According to workers, support from SOs can be enhanced by informing SOs about re-integration plans and involving them in decision making. However, workers were concerned about overburdening SOs, and receiving unwanted support from them. CONCLUSIONS: According to interviewed workers, engagement of SOs in occupational health care can help workers with a chronic disease in their recovery and return to work. However, they felt it is important to take SO characteristics and the worker's circumstances and preferences into account, and to balance the potential benefits and drawbacks of involving SOs.Implications for rehabilitationThis study suggests that the worker's re-integration process could benefit from informing significant others about the return to work plans, involving them in decision-making, and explicitly discussing how the significant other can support the worker.Occupational health physicians have an important role in informing workers about the possibility and potential benefits of involving their significant others in the re-integration process.The involvement of a significant other in the re-integration process needs to be tailored to the specific situation of the individual worker, taking into account the preferences of both the worker and significant other.Findings suggest that it is important that occupational health physicians, workers and significant others are not only aware of the possible benefits of significant other involvement, but also of potential drawbacks such as interference during consultations, overburdening significant others, and significant others providing unwanted support.

Allelic expression imbalance of PIK3CA mutations is frequent in breast cancer and prognostically significant.

PIK3CA mutations are the most common in breast cancer, particularly in the estrogen receptor-positive cohort, but the benefit of PI3K inhibitors has had limited success compared with approaches targeting other less common mutations. We found a frequent allelic expression imbalance between the missense mutant and wild-type PIK3CA alleles in breast tumors from the METABRIC (70.2%) and the TCGA (60.1%) projects. When considering the mechanisms controlling allelic expression, 27.7% and 11.8% of tumors showed imbalance due to regulatory variants in cis, in the two studies respectively. Furthermore, preferential expression of the mutant allele due to cis-regulatory variation is associated with poor prognosis in the METABRIC tumors (P = 0.031). Interestingly, ER-, PR-, and HER2+ tumors showed significant preferential expression of the mutated allele in both datasets. Our work provides compelling evidence to support the clinical utility of PIK3CA allelic expression in breast cancer in identifying patients of poorer prognosis, and those with low expression of the mutated allele, who will unlikely benefit from PI3K inhibitors. Furthermore, our work proposes a model of differential regulation of a critical cancer-promoting gene in breast cancer.

Extensive preclinical validation of combined RMC-4550 and LY3214996 supports clinical investigation for KRAS mutant pancreatic cancer.

Over 90% of pancreatic cancers present mutations in KRAS, one of the most common oncogenic drivers overall. Currently, most KRAS mutant isoforms cannot be targeted directly. Moreover, targeting single RAS downstream effectors induces adaptive resistance mechanisms. We report here on the combined inhibition of SHP2, upstream of KRAS, using the allosteric inhibitor RMC-4550 and of ERK, downstream of KRAS, using LY3214996. This combination shows synergistic anti-cancer activity in vitro, superior disruption of the MAPK pathway, and increased apoptosis induction compared with single-agent treatments. In vivo, we demonstrate good tolerability and efficacy of the combination, with significant tumor regression in multiple pancreatic ductal adenocarcinoma (PDAC) mouse models. Finally, we show evidence that 18F-fluorodeoxyglucose (FDG) positron emission tomography (PET) can be used to assess early drug responses in animal models. Based on these results, we will investigate this drug combination in the SHP2 and ERK inhibition in pancreatic cancer (SHERPA; ClinicalTrials.gov: NCT04916236) clinical trial, enrolling patients with KRAS-mutant PDAC.

The prognostic significance of tumour cell detection in the peripheral blood versus the bone marrow in 733 early-stage breast cancer patients.

INTRODUCTION: The detection of circulating tumour cells (CTCs) in the peripheral blood and disseminated tumour cells (DTCs) in the bone marrow are promising prognostic tools for risk stratification in early breast cancer. There is, however, a need for further validation of these techniques in larger patient cohorts with adequate follow-up periods. METHODS: We assayed CTCs and DTCs at primary surgery in 733 stage I or II breast cancer patients with a median follow-up time of 7.6 years. CTCs were detected in samples of peripheral blood mononuclear cells previously stored in liquid-nitrogen using a previously-developed multi-marker quantitative PCR (QPCR)-based assay. DTCs were detected in bone marrow samples by immunocytochemical analysis using anti-cytokeratin antibodies. RESULTS: CTCs were detected in 7.9% of patients, while DTCs were found in 11.7%. Both CTC and DTC positivity predicted poor metastasis-free survival (MFS) and breast cancer-specific survival (BCSS); MFS hazard ratio (HR) = 2.4 (P < 0.001)/1.9 (P = 0.006), and BCSS HR = 2.5 (P < 0.001)/2.3 (P = 0.01), for CTC/DTC status, respectively). Multivariate analyses demonstrated that CTC status was an independent prognostic variable for both MFS and BCSS. CTC status also identified a subset of patients with significantly poorer outcome among low-risk node negative patients that did not receive adjuvant systemic therapy (MFS HR 2.3 (P = 0.039), BCSS HR 2.9 (P = 0.017)). Using both tests provided increased prognostic information and indicated different relevance within biologically dissimilar breast cancer subtypes. CONCLUSIONS: These results support the use of CTC analysis in early breast cancer to generate clinically useful prognostic information.

A powerful drug combination strategy targeting glutamine addiction for the treatment of human liver cancer.

The dependency of cancer cells on glutamine may be exploited therapeutically as a new strategy for treating cancers that lack druggable driver genes. Here we found that human liver cancer was dependent on extracellular glutamine. However, targeting glutamine addiction using the glutaminase inhibitor CB-839 as monotherapy had a very limited anticancer effect, even against the most glutamine addicted human liver cancer cells. Using a chemical library, we identified V-9302, a novel inhibitor of glutamine transporter ASCT2, as sensitizing glutamine dependent (GD) cells to CB-839 treatment. Mechanically, a combination of CB-839 and V-9302 depleted glutathione and induced reactive oxygen species (ROS), resulting in apoptosis of GD cells. Moreover, this combination also showed tumor inhibition in HCC xenograft mouse models in vivo. Our findings indicate that dual inhibition of glutamine metabolism by targeting both glutaminase and glutamine transporter ASCT2 represents a potential novel treatment strategy for glutamine addicted liver cancers.

Kinome capture sequencing of high-grade serous ovarian carcinoma reveals novel mutations in the JAK3 gene.

High-grade serous ovarian carcinoma (HGSOC) remains the deadliest form of epithelial ovarian cancer and despite major efforts little improvement in overall survival has been achieved. Identification of recurring "driver" genetic lesions has the potential to enable design of novel therapies for cancer. Here, we report on a study to find such new therapeutic targets for HGSOC using exome-capture sequencing approach targeting all kinase genes in 127 patient samples. Consistent with previous reports, the most frequently mutated gene was TP53 (97% mutation frequency) followed by BRCA1 (10% mutation frequency). The average mutation frequency of the kinase genes mutated from our panel was 1.5%. Intriguingly, after BRCA1, JAK3 was the most frequently mutated gene (4% mutation frequency). We tested the transforming properties of JAK3 mutants using the Ba/F3 cell-based in vitro functional assay and identified a novel gain-of-function mutation in the kinase domain of JAK3 (p.T1022I). Importantly, p.T1022I JAK3 mutants displayed higher sensitivity to the JAK3-selective inhibitor Tofacitinib compared to controls. For independent validation, we re-sequenced the entire JAK3 coding sequence using tagged amplicon sequencing (TAm-Seq) in 463 HGSOCs resulting in an overall somatic mutation frequency of 1%. TAm-Seq screening of CDK12 in the same population revealed a 7% mutation frequency. Our data confirms that the frequency of mutations in kinase genes in HGSOC is low and provides accurate estimates for the frequency of JAK3 and CDK12 mutations in a large well characterized cohort. Although p.T1022I JAK3 mutations are rare, our functional validation shows that if detected they should be considered as potentially actionable for therapy. The observation of CDK12 mutations in 7% of HGSOC cases provides a strong rationale for routine somatic testing, although more functional and clinical characterization is required to understand which nonsynonymous mutations alterations are associated with homologous recombination deficiency.

No effect of red clover-derived isoflavone intervention on the insulin-like growth factor system in women at increased risk of colorectal cancer.

BACKGROUND: Increased insulin-like growth factor (IGF)-I and IGF-II concentrations are related to increased colorectal cancer risk. Isoflavones have been associated with reduced colorectal cancer risk, and may affect the IGF system because of their weak estrogenic activity. The aim of the study was to investigate the effect of isolated isoflavones on serum concentrations of IGF system components. MATERIALS AND METHODS: We conducted a randomized, placebo-controlled, double-blinded, crossover trial in four hospitals in the Netherlands to investigate the effect of an 8-week supplementation with red clover-derived isoflavones (84 mg/d) on serum IGF-I concentrations. In addition, serum concentrations of IGF-II and IGF binding proteins (IGFBP)-1, IGFBP-2, and IGFBP-3 were assessed. Normal colorectal tissue biopsies were obtained after the first intervention period and mRNA expression of IGF-I, IGF-II, IGFBP-3, and IGF-IR was evaluated. Our study population consisted of 34 postmenopausal women with a family history of colorectal cancer or a personal history of colorectal adenomas. RESULTS: Isoflavone supplementation did not significantly affect serum concentrations of total IGF-I (mean relative within-person difference; IGF-I, -2.0%; 95% confidence interval, -8.0% to 3.9%). IGF-II and IGFBPs were also not significantly altered after isoflavone supplementation. Colorectal tissue mRNA expression of IGF system components did not significantly differ between individuals on isoflavone supplementation and those who received placebo. CONCLUSIONS: The results of our trial, supported by a qualitative review of soy trials published to date, suggest that isoflavones do not significantly affect circulating levels of IGF system components. Increased levels of IGF-I, as observed in most of these trials, are likely due to simultaneous protein supplementation.