Obesity: a perfect storm for carcinogenesis.

Obesity-related cancers account for 40% of the cancer cases observed in the USA and obesity is overtaking smoking as the most widespread modifiable risk factor for carcinogenesis. Here, we use the hallmarks of cancer framework to delineate how obesity might influence the carcinogenic hallmarks in somatic cells. We discuss the effects of obesity on (a) sustaining proliferative signaling; (b) evading growth suppressors; (c) resisting cell death; (d) enabling replicative immortality; (e) inducing angiogenesis; (f) activating invasion and metastasis; (g) reprogramming energy metabolism; and (h) avoiding immune destruction, together with its effects on genome instability and tumour-promoting inflammation. We present the current understanding and controversies in this evolving field, and highlight some areas in need of further cross-disciplinary focus. For instance, the relative importance of the many potentially causative obesity-related factors is unclear for each type of malignancy. Even within a single tumour type, it is currently unknown whether one obesity-related factor consistently plays a predominant role, or if this varies between patients or, even in a single patient with time. Clarifying how the hallmarks are affected by obesity may lead to novel prevention and treatment strategies for the increasingly obese population.

A Phase Ib/II study of IGF-neutralising antibody xentuzumab with enzalutamide in metastatic castration-resistant prostate cancer.

BACKGROUND: This multicentre, open-label, Phase Ib/II trial evaluated the insulin-like growth factor (IGF) 1/2 neutralising antibody xentuzumab plus enzalutamide in metastatic castrate-resistant prostate cancer (mCRPC). METHODS: The trial included Phase Ib escalation and expansion parts and a randomised Phase II part versus enzalutamide alone. Primary endpoints in the Phase Ib escalation, Phase Ib expansion and Phase II parts were maximum tolerated dose (MTD), prostate-specific antigen response and investigator-assessed progression-free survival (PFS), respectively. Patients in the Phase Ib escalation and Phase II parts had progressed on/after docetaxel/abiraterone. RESULTS: In the Phase Ib escalation (n = 10), no dose-limiting toxicities were reported, and xentuzumab 1000 mg weekly plus enzalutamide 160 mg daily (Xe1000 + En160) was defined as the MTD and recommended Phase 2 dose. In the Phase Ib expansion (n = 24), median PFS was 8.2 months, and one patient had a confirmed, long-term response. In Phase II (n = 86), median PFS for the Xe1000 + En160 and En160 arms was 7.4 and 6.2 months, respectively. Subgroup analysis suggested trends towards benefit with Xe1000 + En160 in patients whose tumours had high levels of IGF1 mRNA or PTEN protein. Overall, the combination was well tolerated. CONCLUSIONS: Xentuzumab plus enzalutamide was tolerable but lacked antitumour activity in unselected patients with mCRPC. CLINICAL TRIAL REGISTRATION: EudraCT number 2013-004011-41.

Type 1 IGF receptor associates with adverse outcome and cellular radioresistance in paediatric high-grade glioma.

High-grade glioma (HGG) is highly resistant to therapy, prompting us to investigate the contribution of insulin-like growth factor receptor (IGF-1R), linked with radioresistance in other cancers. IGF-1R immunohistochemistry in 305 adult HGG (aHGG) and 103 paediatric/young adult HGG (pHGG) cases revealed significant association with adverse survival in pHGG, with median survival of 13.5 vs 29 months for pHGGs with moderate/strong vs negative/weak IGF-1R (p = 0.011). Secondly, we tested IGF-1R inhibitor BMS-754807 in HGG cells, finding minimal radiosensitisation of 2/3 aHGG cell lines (dose enhancement ratios DERs < 1.60 at 2-8 Gy), and greater radiosensitisation of 2/2 pHGG cell lines (DERs ≤ 4.16). BMS-754807 did not influence radiation-induced apoptosis but perturbed the DNA damage response with altered induction/resolution of γH2AX, 53BP1 and RAD51 foci. These data indicate that IGF-1R promotes radioresistance in pHGG, potentially contributing to the association of IGF-1R with adverse outcome and suggesting IGF-1R as a candidate treatment target in pHGG.

Radioresistant laryngeal cancers upregulate type 1 IGF receptor and exhibit increased cellular dependence on IGF and EGF signalling.

OBJECTIVES: Patients failing radiotherapy for laryngeal squamous cell carcinoma (LSCC) often require salvage total laryngectomy which has major functional consequences, highlighting a need for biomarkers of radiotherapy resistance. In other tumour types, radioresistance has been linked to epidermal growth factor receptor (EGFR) and type 1 insulin-like growth factor receptor (IGF-1R). Here, we evaluated IGF-1R and EGFR as predictors and mediators of LSCC radioresistance. DESIGN: We compared IGF-1R and EGFR immunohistochemical scores in patients with LSCC achieving long-term remission post-radiotherapy (n = 23), patients treated with primary laryngectomy (n = 22) or salvage laryngectomy following radiotherapy recurrence (n = 18). To model radioresistance in vitro, two LSCC cell lines underwent clinically relevant irradiation to 55 Gy in 2.75 Gy fractions. RESULTS: Type 1 insulin-like growth factor receptor expression was higher in pre-treatment biopsies of radiotherapy failures compared with those in long-term remission and was upregulated post-radiotherapy. Patients undergoing primary laryngectomy had more advanced T/N stage and greater tumour IGF-1R content than those achieving long-term remission. Pre-treatment EGFR did not associate with radiotherapy outcomes but showed a trend to upregulation post-irradiation. In vitro, radiosensitivity was enhanced by inhibition of EGFR but not IGF. Repeated irradiation upregulated IGF-1R in BICR18 and SQ20B cells and EGFR in SQ20B, and enhanced SQ20B radioresistance. Repeatedly irradiated SQ20B_55 cells were not radiosensitised by inhibition of IGF and/or EGFR, but IGF-1R:EGFR co-inhibition suppressed baseline cell survival more effectively than blockade of either pathway alone, and more effectively than in parental cells. CONCLUSIONS: Radiation upregulates IGF-1R and may enhance IGF/EGFR dependence, suggesting that IGF/EGFR blockade may have activity in LSCCs that recur post-radiotherapy.

IGF-1R associates with adverse outcomes after radical radiotherapy for prostate cancer.

BACKGROUND: Activated type 1 insulin-like growth factor receptors (IGF-1Rs) undergo internalisation and nuclear translocation, promoting cell survival. We previously reported that IGF-1R inhibition delays DNA damage repair, sensitising prostate cancer cells to ionising radiation. Here we tested the clinical relevance of these findings. METHODS: We assessed associations between IGF-1R and clinical outcomes by immunohistochemistry in diagnostic biopsies of 136 men treated with 55-70 Gy external beam radiotherapy for prostate cancer, comparing results with publicly available transcriptional data in surgically treated patients. RESULTS: Following radiotherapy, overall recurrence-free survival was shorter in patients whose tumours contained high total, cytoplasmic and internalised (nuclear/cytoplasmic) IGF-1R. High total IGF-1R associated with high primary Gleason grade and risk of metastasis, and cytoplasmic and internalised IGF-1R with biochemical recurrence, which includes patients experiencing local recurrence within the radiation field indicating radioresistance. In multivariate analysis, cytoplasmic, internalised and total IGF-1R were independently associated with risk of overall recurrence, and cytoplasmic IGF-1R was an independent predictor of biochemical recurrence post radiotherapy. Insulin-like growth factor receptors expression did not associate with biochemical recurrence after radical prostatectomy. CONCLUSIONS: These data reveal increased risk of post-radiotherapy recurrence in men whose prostate cancers contain high levels of total or cytoplasmic IGF-1R.

Synthesis and bioactivity of fused- and spiro-ß-lactone-lactam systems.

An investigation of the formation of fused- and spiro-ß-lactone annulate to γ-lactams has shown that the fused systems are formed preferentially, under standard conditions, but that spiro systems are accessible only when the formation of the fused system is blocked and require careful optimisation of reaction conditions. These systems display both weak antibacterial activity and proteasome inhibition.

IGF-1R expression is associated with HPV-negative status and adverse survival in head and neck squamous cell cancer.

Head and neck squamous cell carcinomas (HNSCC) are treated with surgery, radiotherapy and cisplatin-based chemotherapy, but survival from locally-advanced disease remains poor, particularly in patients whose tumors are negative for Human papillomavirus (HPV). Type 1 IGF receptor (IGF-1R) is known to promote tumorigenesis and resistance to cancer therapeutics. Here, we assessed IGF-1R immunohistochemistry on tissue microarrays containing 852 cores from 346 HNSCC patients with primary tumors in the oropharynx (n = 231), larynx (85), hypopharynx (28), oral cavity (2). Of these, 236 (68%) were HPV-negative, 110 (32%) positive. IGF-1R was detected in the cell membrane of 36% and cytoplasm of 92% of HNSCCs; in 64 cases with matched normal tonsillar epithelium, IGF-1R was overexpressed in the HNSCCs (P < 0.001). Overall survival (OS) and disease-specific survival (DSS) were reduced in patients whose tumors contained high membrane IGF-1R [OS: hazard ratio (HR) = 1.63, P = 0.006; DSS: HR = 1.63, P = 0.016], cytoplasmic IGF-1R (OS: HR = 1.58, P = 0.009; DSS: HR = 1.58, P = 0.024) and total IGF-1R (OS: HR = 2.02, P < 0.001; DSS: HR = 2.2, P < 0.001). High tumor IGF-1R showed significant association with high-tumor T-stage (P < 0.001) and HPV-negativity (P < 0.001), and was associated with shorter OS when considering patients with HPV-positive (P = 0.01) and negative (P = 0.006) tumors separately. IGF-1R was independently associated with survival in multivariate analysis including HPV, but not when lymphovascular invasion, perineural spread and T-stage were included. Of these factors, only IGF-1R can be manipulated; the association of IGF-1R with aggressive disease supports experimental incorporation of anti-IGF-1R agents into multimodality treatment programs for HPV-negative and high IGF-1R HPV-positive HNSCC.

Suppression of homologous recombination sensitizes human tumor cells to IGF-1R inhibition.

Inhibition of type 1 IGF receptor (IGF-1R) sensitizes to DNA-damaging cancer treatments, and delays repair of DNA double strand breaks (DSBs) by non-homologous end-joining and homologous recombination (HR). In a recent screen for mediators of resistance to IGF-1R inhibitor AZ12253801, we identified RAD51, required for the strand invasion step of HR. These findings prompted us to test the hypothesis that IGF-1R-inhibited cells accumulate DSBs formed at endogenous DNA lesions, and depend on residual HR for their repair. Indeed, initial experiments showed time-dependent accumulation of γH2AX foci in IGF-1R -inhibited or -depleted prostate cancer cells. We then tested effects of suppressing HR, and found that RAD51 depletion enhanced AZ12253801 sensitivity in PTEN wild-type prostate cancer cells but not in cells lacking functional PTEN. Similar sensitization was induced in prostate cancer cells by depletion of BRCA2, required for RAD51 loading onto DNA, and in BRCA2(-/-) colorectal cancer cells, compared with isogenic BRCA2(+/-) cells. We also assessed chemical HR inhibitors, finding that RAD51 inhibitor BO2 blocked RAD51 focus formation and sensitized to AZ12253801. Finally, we tested CDK1 inhibitor RO-3306, which impairs HR by inhibiting CDK1-mediated BRCA1 phosphorylation. R0-3306 suppressed RAD51 focus formation consistent with HR attenuation, and sensitized prostate cancer cells to IGF-1R inhibition, with 2.4-fold reduction in AZ12253801 GI50 and 13-fold reduction in GI80. These data suggest that responses to IGF-1R inhibition are enhanced by genetic and chemical approaches to suppress HR, defining a population of cancers (PTEN wild-type, BRCA mutant) that may be intrinsically sensitive to IGF-1R inhibitory drugs.

WNT5A-ROR2 axis mediates VEGF dependence of BRAF mutant melanoma.

PURPOSE: Despite recent advances, approximately 50% of patient with metastatic melanoma eventually succumb to the disease. Patients with melanomas harboring a BRAF mutation (BRAFMut) have a worse prognosis than those with wildtype (BRAFWT) tumors. Unexpectedly, interim AVAST-M Phase III trial data reported benefit from adjuvant anti-VEGF bevacizumab only in the BRAFMut group. We sought to find mechanisms underpinning this sensitivity. METHODS: We investigated this finding in vitro and in vivo using melanoma cell lines and clones generated by BRAFV600E knock-in on a BRAFWT background. RESULTS: Compared with BRAFWT cells, isogenic BRAFV600E clones secreted more VEGF and exhibited accelerated growth rates as spheroids and xenografts, which were more vascular and proliferative. Recapitulating AVAST-M findings, bevacizumab affected only BRAFV600E xenografts, inducing significant tumor growth delay, reduced vascularity and increased necrosis. We identified 814 differentially expressed genes in isogenic BRAFV600E/BRAFWT clones. Of 61 genes concordantly deregulated in clinical melanomas ROR2 was one of the most upregulated by BRAFV600E. ROR2 was shown to be RAF-MEK regulated in BRAFV600E cells and its depletion suppressed VEGF secretion down to BRAFWT levels. The ROR2 ligand WNT5A was also overexpressed in BRAFMut melanomas, and in ROR2-overexpressing BRAFV600E cells MEK inhibition downregulated WNT5A and VEGF secretion. CONCLUSIONS: These data implicate WNT5A-ROR2 in VEGF secretion, vascularity, adverse outcomes and bevacizumab sensitivity of BRAFMut melanomas, suggesting that this axis has potential therapeutic relevance.

Aberrant succination of proteins in fumarate hydratase-deficient mice and HLRCC patients is a robust biomarker of mutation status.

Germline mutations in the FH gene encoding the Krebs cycle enzyme fumarate hydratase predispose to hereditary leiomyomatosis and renal cell cancer (HLRCC) syndrome. FH-deficient cells and tissues accumulate high levels of fumarate, which may act as an oncometabolite and contribute to tumourigenesis. A recently proposed role for fumarate in the covalent modification of cysteine residues to S-(2-succinyl) cysteine (2SC) (termed protein succination) prompted us to assess 2SC levels in our existing models of HLRCC. Herein, using a previously characterized antibody against 2SC, we show that genetic ablation of FH causes high levels of protein succination. We next hypothesized that immunohistochemistry for 2SC would serve as a metabolic biomarker for the in situ detection of FH-deficient tissues. Robust detection of 2SC was observed in Fh1 (murine FH)-deficient renal cysts and in a retrospective series of HLRCC tumours (n = 16) with established FH mutations. Importantly, 2SC was undetectable in normal tissues (n = 200) and tumour types not associated with HLRCC (n = 1342). In a prospective evaluation of cases referred for genetic testing for HLRCC, the presence of 2SC-modified proteins (2SCP) correctly predicted genetic alterations in FH in every case. In two series of unselected type II papillary renal cancer (PRCC), prospectively analysed by 2SCP staining followed by genetic analysis, the biomarker accurately identified previously unsuspected FH mutations (2/33 and 1/36). The investigation of whether metabolites in other tumour types produce protein modification signature(s) that can be assayed using similar strategies will be of interest in future studies of cancer.

CHK1 inhibition exacerbates replication stress induced by IGF blockade.

We recently reported that genetic or pharmacological inhibition of insulin-like growth factor receptor (IGF-1R) slows DNA replication and induces replication stress by downregulating the regulatory subunit RRM2 of ribonucleotide reductase, perturbing deoxynucleotide triphosphate (dNTP) supply. Aiming to exploit this effect in therapy we performed a compound screen in five breast cancer cell lines with IGF neutralising antibody xentuzumab. Inhibitor of checkpoint kinase CHK1 was identified as a top screen hit. Co-inhibition of IGF and CHK1 caused synergistic suppression of cell viability, cell survival and tumour growth in 2D cell culture, 3D spheroid cultures and in vivo. Investigating the mechanism of synthetic lethality, we reveal that CHK1 inhibition in IGF-1R depleted or inhibited cells further downregulated RRM2, reduced dNTP supply and profoundly delayed replication fork progression. These effects resulted in significant accumulation of unreplicated single-stranded DNA and increased cell death, indicative of replication catastrophe. Similar phenotypes were induced by IGF:WEE1 co-inhibition, also via exacerbation of RRM2 downregulation. Exogenous RRM2 expression rescued hallmarks of replication stress induced by co-inhibiting IGF with CHK1 or WEE1, identifying RRM2 as a critical target of the functional IGF:CHK1 and IGF:WEE1 interactions. These data identify novel therapeutic vulnerabilities and may inform future trials of IGF inhibitory drugs.

Serial analysis of resected prostate cancer suggests up-regulation of type 1 IGF receptor with disease progression.

OBJECTIVE: • To compare immunostaining protocols using different antibodies for the type 1 insulin-like growth factor receptor (IGF-1R) in channel transurethal resection of the prostate (chTURP) chips, and to investigate how IGF-1R expression varies with time in serial prostate cancer specimens from individual patients. METHODS: • We studied IGF-1R expression in 44 prostate cancer specimens from 18 patients who had undergone serial chTURP at least 3 months apart. • Retrospective analysis of the hospital notes was undertaken to obtain clinical information, including age, Gleason score, prostate-specific antigen (PSA) level, hormone treatment and metastatic disease status at the time of each operation. • After an optimization process using three commercially-available IGF-1R antibodies, we used two antibodies for semiquantititve immunostaining of serial chTURP chips. RESULTS: • Santa Cruz antibody sc713 gave positive staining in IGF-1R null R- cells, and was not used further. Antibodies from Cell Signaling Technology (Beverly, MA, USA) (CS) and NeoMarkers Inc. (Fremont, CA, USA) (NM) did not stain R- cells and, in prostate tissue, showed staining of the glandular epithelium, with negligible stromal staining. All 44 chTURP samples contained identifiable malignant tissue and, of these, 73% and 64% scored moderately or strongly (score 3 or 4) with the CS and NM antibodies respectively. • There was significant correlation of IGF-1R scores of malignant tissue between the two antibodies (P < 0.001). By contrast, staining of benign glands showed poor correlation between antibodies: CS gave significantly weaker staining than malignant epithelium in the same sections (P < 0.001), whereas NM showed poor discrimination between malignant and benign glands. IGF-1R staining scores generated by the CS antibody were used to analyze the clinical data. • Most patients (six of seven) with falling IGF-1R staining scores were responding to androgen deprivation therapy (confirmed by PSA response) between operations. Conversely, in seven of eight patients who had progression to androgen-independence between procedures, IGF-1R levels increased or remained high. Finally, seven of 11 patients who developed radiologically confirmed metastases between procedures showed stable or increasing IGF-1R staining scores. CONCLUSION: • The present study is the first to assess changes in IGF-1R expression in serial prostate cancer samples. The results obtained indicate that IGF-1R expression usually remains high throughout the course of histologically-proven disease progression in serial specimens, suggesting that the IGF-1R remains a valid treatment target for advanced prostate cancer.

IGF-1R nuclear import and recruitment to chromatin involves both alpha and beta subunits.

Mature type 1 insulin-like growth factor receptors (IGF-1Rs) are heterotetrameric structures comprising two extracellular α-subunits disulphide-bonded to two transmembrane ß-subunits with tyrosine kinase activity. IGF-1R is a well-known cell surface mediator of malignant growth, with an incompletely understood role upon nuclear import as a transcriptional regulator. Previous characterisation of nuclear IGF-1R focused on IGF-1Rß. Here, we aimed to clarify the source of nuclear IGF-1R and investigate whether α-subunits contribute to nuclear IGF-1R function. Using prostate cancer cell lines DU145 and 22Rv1 we detected nuclear α- and ß-subunits, with increase in nuclear signal upon IGF-treatment and reduction in response to IGF-1R inhibitor BMS-754807. Following biotinylation of cell surface proteins, biotinylated α- and ß-subunits were detected in nuclear extracts of both cell lines. Furthermore, α- and ß-subunits reciprocally co-precipitated from nuclear extract. Finally, we detected recruitment of both subunits to regulatory regions of chromatin, including the promoter of the oncogene JUN, that we previously identified in ChIP-seq as sites of IGF-1Rß enrichment. These data confirm the cell surface origin of nuclear IGF-1R, suggest the presence of nuclear αß complexes and reveal that both IGF-1Rα- and ß-subunits contribute to pro-tumorigenic functions of nuclear IGF-1R. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12672-021-00407-8.

Targeting IGF Perturbs Global Replication through Ribonucleotide Reductase Dysfunction.

Inhibition of IGF receptor (IGF1R) delays repair of radiation-induced DNA double-strand breaks (DSB), prompting us to investigate whether IGF1R influences endogenous DNA damage. Here we demonstrate that IGF1R inhibition generates endogenous DNA lesions protected by 53BP1 bodies, indicating under-replicated DNA. In cancer cells, inhibition or depletion of IGF1R delayed replication fork progression accompanied by activation of ATR-CHK1 signaling and the intra-S-phase checkpoint. This phenotype reflected unanticipated regulation of global replication by IGF1 mediated via AKT, MEK/ERK, and JUN to influence expression of ribonucleotide reductase (RNR) subunit RRM2. Consequently, inhibition or depletion of IGF1R downregulated RRM2, compromising RNR function and perturbing dNTP supply. The resulting delay in fork progression and hallmarks of replication stress were rescued by RRM2 overexpression, confirming RRM2 as the critical factor through which IGF1 regulates replication. Suspecting existence of a backup pathway protecting from toxic sequelae of replication stress, targeted compound screens in breast cancer cells identified synergy between IGF inhibition and ATM loss. Reciprocal screens of ATM-proficient/deficient fibroblasts identified an IGF1R inhibitor as the top hit. IGF inhibition selectively compromised growth of ATM-null cells and spheroids and caused regression of ATM-null xenografts. This synthetic-lethal effect reflected conversion of single-stranded lesions in IGF-inhibited cells into toxic DSBs upon ATM inhibition. Overall, these data implicate IGF1R in alleviating replication stress, and the reciprocal IGF:ATM codependence we identify provides an approach to exploit this effect in ATM-deficient cancers. SIGNIFICANCE: This study identifies regulation of ribonucleotide reductase function and dNTP supply by IGFs and demonstrates that IGF axis blockade induces replication stress and reciprocal codependence on ATM. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/81/8/2128/F1.large.jpg.

The type 1 insulin-like growth factor receptor pathway.

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

Therapeutic Targeting of the IGF Axis.

The insulin like growth factor (IGF) axis plays a fundamental role in normal growth and development, and when deregulated makes an important contribution to disease. Here, we review the functions mediated by ligand-induced IGF axis activation, and discuss the evidence for the involvement of IGF signaling in the pathogenesis of cancer, endocrine disorders including acromegaly, diabetes and thyroid eye disease, skin diseases such as acne and psoriasis, and the frailty that accompanies aging. We discuss the use of IGF axis inhibitors, focusing on the different approaches that have been taken to develop effective and tolerable ways to block this important signaling pathway. We outline the advantages and disadvantages of each approach, and discuss progress in evaluating these agents, including factors that contributed to the failure of many of these novel therapeutics in early phase cancer trials. Finally, we summarize grounds for cautious optimism for ongoing and future studies of IGF blockade in cancer and non-malignant disorders including thyroid eye disease and aging.

Bad to the Bone: The Role of the Insulin-Like Growth Factor Axis in Osseous Metastasis.

Bone metastases are a frequent complication of cancer that are associated with considerable morbidity. Current treatments may temporarily palliate the symptoms of bone metastases but often fail to delay their progression. Bones provide a permissive environment because they are characterized by dynamic turnover, secreting factors required for bone maintenance but also stimulating the establishment and growth of metastases. Insulin-like growth factors (IGF) are the most abundant growth factors in bone and are required for normal skeletal development and function. Via activation of the IGF-1 receptors (IGF-1R) and variant insulin receptors, IGFs promote cancer progression, aggressiveness, and treatment resistance. Of specific relevance to bone biology, IGFs contribute to the homing, dormancy, colonization, and expansion of bone metastases. Furthermore, preclinical evidence suggests that tumor cells can be primed to metastasize to bone by a high IGF-1 environment in the primary tumor, suggesting that bone metastases may reflect IGF dependency. Therapeutic targeting of the IGF axis may therefore provide an effective method for treating bone metastases. Indeed, anti-IGF-1R antibodies, IGF-1R tyrosine kinase inhibitors, and anti-IGF-1/2 antibodies have demonstrated antitumor activity in preclinical models of prostate and breast cancer metastases, either alone or in combination with other agents. Several studies suggest that such treatments can inhibit bone metastases without affecting growth of the primary tumor. Although previous trials of anti-IGF-1R drugs have generated negative results in unselected patients, these considerations suggest that future clinical trials of IGF-targeted agents may be warranted in patients with bone metastases.

RAPTOR up-regulation contributes to resistance of renal cancer cells to PI3K-mTOR inhibition.

The outlook for patients with advanced renal cell cancer (RCC) has been improved by targeted agents including inhibitors of the PI3 kinase (PI3K)-AKT-mTOR axis, although treatment resistance is a major problem. Here, we aimed to understand how RCC cells acquire resistance to PI3K-mTOR inhibition. We used the RCC4 cell line to generate a model of in vitro resistance by continuous culture in PI3K-mTOR kinase inhibitor NVP-BEZ235 (BEZ235, Dactolisib). Resistant cells were cross-resistant to mTOR inhibitor AZD2014. Sensitivity was regained after 4 months drug withdrawal, and resistance was partially suppressed by HDAC inhibition, supporting an epigenetic mechanism. BEZ235-resistant cells up-regulated and/or activated numerous proteins including MET, ABL, Notch, IGF-1R, INSR and MEK/ERK. However, resistance was not reversed by inhibiting or depleting these pathways, suggesting that many induced changes were passengers not drivers of resistance. BEZ235 blocked phosphorylation of mTOR targets S6 and 4E-BP1 in parental cells, but 4E-BP1 remained phosphorylated in resistant cells, suggesting BEZ235-refractory mTORC1 activity. Consistent with this, resistant cells over-expressed mTORC1 component RAPTOR at the mRNA and protein level. Furthermore, BEZ235 resistance was suppressed by RAPTOR depletion, or allosteric mTORC1 inhibitor rapamycin. These data reveal that RAPTOR up-regulation contributes to PI3K-mTOR inhibitor resistance, and suggest that RAPTOR expression should be included in the pharmacodynamic assessment of mTOR kinase inhibitor trials.

Nuclear IGF1R Interacts with Regulatory Regions of Chromatin to Promote RNA Polymerase II Recruitment and Gene Expression Associated with Advanced Tumor Stage.

Internalization of ligand-activated type I IGF receptor (IGF1R) is followed by recycling to the plasma membrane, degradation or nuclear translocation. Nuclear IGF1R reportedly associates with clinical response to IGF1R inhibitory drugs, yet its role in the nucleus is poorly characterized. Here, we investigated the significance of nuclear IGF1R in clinical cancers and cell line models. In prostate cancers, IGF1R was predominantly membrane localized in benign glands, while malignant epithelium contained prominent internalized (nuclear/cytoplasmic) IGF1R, and nuclear IGF1R associated significantly with advanced tumor stage. Using ChIP-seq to assess global chromatin occupancy, we identified IGF1R-binding sites at or near transcription start sites of genes including JUN and FAM21, most sites coinciding with occupancy by RNA polymerase II (RNAPol2) and histone marks of active enhancers/promoters. IGF1R was inducibly recruited to chromatin, directly binding DNA and interacting with RNAPol2 to upregulate expression of JUN and FAM21, shown to mediate tumor cell survival and IGF-induced migration. IGF1 also enriched RNAPol2 on promoters containing IGF1R-binding sites. These functions were inhibited by IGF1/II-neutralizing antibody xentuzumab (BI 836845), or by blocking receptor internalization. We detected IGF1R on JUN and FAM21 promoters in fresh prostate cancers that contained abundant nuclear IGF1R, with evidence of correlation between nuclear IGF1R content and JUN expression in malignant prostatic epithelium. Taken together, these data reveal previously unrecognized molecular mechanisms through which IGFs promote tumorigenesis, with implications for therapeutic evaluation of anti-IGF drugs.Significance: These findings reveal a noncanonical nuclear role for IGF1R in tumorigenesis, with implications for therapeutic evaluation of IGF inhibitory drugs. Cancer Res; 78(13); 3497-509. ©2018 AACR.

The type 1 insulin-like growth factor receptor is over-expressed in bladder cancer.

OBJECTIVE: To analyse bladder cancer biopsies and investigate the pattern of expression of the type 1 insulin-like growth factor receptor (IGF1R), a receptor tyrosine kinase that mediates tumour cell proliferation, motility and protection from apoptosis. MATERIALS AND METHODS: Formalin-fixed specimens of bladder cancer (40 whole-mount, 80 cores on a tumour microarray) and normal bladder (15 samples) were stained immunohistochemically for the IGF1R. The IGF1R expression was also measured by quantitative reverse transcription-polymerase chain reaction (Q-RT-PCR) on RNA extracted from fresh frozen bladder cancers (61) and benign bladder (12). RESULTS: Of the 15 samples of normal bladder, 14 showed negligible (1+) or light (2+) IGF1R immunostaining. By contrast moderate (3+) or heavy (4+) staining for IGF1R was detected in 89 (74%) of the 120 samples of malignant urothelium. Q-RT-PCR showed significantly higher levels of steady-state IGF1R mRNA in tumours (all cases, Ta-T4) than in normal bladder (P < 0.05), indicating up-regulation at the transcriptional level. This difference was particularly evident when comparing normal urothelium with superficial (Ta-T1) or invasive (T2-4) tumours; only the latter showed significant IGF1R over-expression at the RNA level (P < 0.05 vs normal bladder). CONCLUSION: The IGF1R is up-regulated in bladder cancer compared with non-malignant bladder, and might contribute to a propensity for invasion.