Insulin-like Growth Factor-1 Receptor Overexpression Research Articles

Pathways to precision medicine: deciphering the secrets of physiological and pathological atrial enlargement.

Cardiac functional, morphological, and histological analysis, coupled with liquid chromatography and mass spectrometry, of two transgenic mouse models with cardiomyocyte-specific overexpression of insulin-like growth factor 1 receptor (IGF1R) or a dominant-negative PI3K mutant (DCM-dnPI3K) revealed distinctive functional and molecular profiles during physiological (driven by IGF1R overexpression) and pathological (driven by dn-PI3K overexpression) atrial remodeling. The current study confirmed previously reported findings, including ventricular dilatation and enhanced systolic function with no evidence of arrhythmia in IGF1R model, as well as ventricular hypertrophy and decreased systolic function with intermittent atrial fibrillation in DCM-dnPI3K model. Novel findings obtained from the left atrial (LA) characterization of female mice revealed that physiological atrial enlargement resulted from increased atrial myocyte size and was associated with preserved atrial function, as determined by maintained LA ejection fraction (EF). The proteomic profile of IGF1R transgenic (Tg) mice was enriched for metabolic remodeling and showed a protein expression pattern similar to that of healthy human atria; on the other hand, pathological atrial enlargement resulted from increased atrial fibrosis with normal myocyte size and was associated with impaired atrial function due to a reduced LA EF. The proteomic profile of DCM-dnPI3K mice was enriched to both metabolic and structural remodeling and showed a protein expression pattern similar to that of human AF atria.

Functional Investigation of IGF1R Mutations in Multiple Myeloma.

High expression of the receptor tyrosine kinase (RTK) insulin-like growth factor-1 receptor (IGF1R) and RTK mutations are associated with high-risk/worse prognosis in multiple myeloma (MM). Combining the pIGF1R/pINSR inhibitor linsitinib with the proteasome inhibitor (PI) bortezomib seemed promising in a clinical trial, but IGF1R expression was not associated with therapy response. Because the oncogenic impact of IGF1R mutations is so far unknown, we investigated the functional impact of IGF1R mutations on survival signaling, viability/proliferation and survival response to therapy. We transfected four human myeloma cell lines (HMCLs) with IGF1RWT, IGF1RD1146N and IGF1RN1129S (Sleeping Beauty), generated CRISPR-Cas9 IGF1R knockouts in the HMCLs U-266 (IGF1RWT) and L-363 (IGF1RD1146N) and tested the anti-MM activity of linsitinib alone and in combination with the second-generation PI carfilzomib in seven HMCLs. IGF1R knockout entailed reduced proliferation. Upon IGF1R overexpression, survival signaling was moderately increased in all HCMLs and slightly affected by IGF1RN1129S in one HMCL, whereby the viability remained unaffected. Expression of IGF1RD1146N reduced pIGF1R-Y1135, especially under serum reduction, but did not impact downstream signaling. Linsitinib and carfilzomib showed enhanced anti-myeloma activity in six out of seven HMCL irrespective of the IGF1R mutation status. In conclusion, IGF1R mutations can impact IGF1R activation and/or downstream signaling, and a combination of linsitinib with carfilzomib might be a suitable therapeutic approach for MM patients potentially responsive to IGF1R blockade.

From Insight to Eyesight: Unveiling the Secrets of the Insulin-Like Growth Factor Axis in Retinal Health.

Insulin-like growth factor-1 (IGF-1) plays a diverse role in the retina, exerting its effects in both normal and diseased conditions. Deficiency of IGF-1 in humans leads to issues such as microcephaly, mental retardation, deafness, and postnatal growth failure. IGF-1 is produced in the retinal pigment epithelium (RPE) and activates the IGF-1 receptor (IGF-1R) in photoreceptor cells. When IGF-1R is absent in rod cells, it results in the degeneration of photoreceptors, emphasizing the neuroprotective function of IGF signaling in these cells. Contrastingly, in neovascular age-related macular degeneration (AMD), there is an overexpression of both IGF-1 and IGF-1R in RPE. The mechanisms behind this altered regulation of IGF-1 in diseased states are currently unknown. This comprehensive review provides recent insights into the role of IGF-1 in the health and disease of the retina, raising several unanswered questions that still need further investigation.

Synergic Action of Insulin-like Growth Factor-2 and miRNA-483 in Pterygium Pathogenesis.

Pterygium is a multifactorial disease in which UV-B is speculated to play a key role by inducing oxidative stress and phototoxic DNA damage. In search for candidate molecules that are useful for justifying the intense epithelial proliferation observed in pterygium, our attention has been focused on Insulin-like Growth Factor 2 (IGF-2), mainly detected in embryonic and fetal somatic tissues, which regulate metabolic and mitogenic functions. The binding between IGF-2 and its receptor Insulin-like Growth Factor 1 Receptor (IGF-1R) activates the PI3K-AKT pathway, which leads to the regulation of cell growth, differentiation, and the expression of specific genes. Since IGF2 is regulated by parental imprinting, in different human tumors, the IGF2 Loss of Imprinting (LOI) results in IGF-2- and IGF2-derived intronic miR-483 overexpression. Based on these activities, the purpose of this study was to investigate the overexpression of IGF-2, IGF-1R, and miR-483. Using an immunohistochemical approach, we demonstrated an intense colocalized epithelial overexpression of IGF-2 and IGF-1R in most pterygium samples (Fisher's exact test, p = 0.021). RT-qPCR gene expression analysis confirmed IGF2 upregulation and demonstrated miR-483 expression in pterygium compared to normal conjunctiva (253.2-fold and 12.47-fold, respectively). Therefore, IGF-2/IGF-1R co-expression could suggest their interplay through the two different paracrine/autocrine IGF-2 routes for signaling transfer, which would activate the PI3K/AKT signaling pathway. In this scenario, miR-483 gene family transcription might synergically reinforce IGF-2 oncogenic function through its boosting pro-proliferative and antiapoptotic activity.

Role of let-7 family in the invasion and metastasis of osteosarcoma.

Role of let-7 family in the invasion and metastasis of osteosarcoma.

IGF1R and HIF-1α Gene Silencing Inhibits Cancer Cell Growth

Background: According to several studies, hypoxia-inducible factor-1α (HIF-1α) and insulin-like growth factor 1 receptor (IGF1R) promote cancer progression and drug resistance. The overexpression of IGF1R and HIF-1 α is observed in various cancers, including breast and colon cancers. Thus, we tried to inhibit the progression of tumor cells by blocking IGF1R and HIF-1α. Methods: This study used Lipofectamine to transfect small interfering RNAs (siRNAs) into tumor cells. We treated murine mammary carcinoma (4T1) and murine colon carcinoma (CT26) cells with anti-IGF1R and anti-HIF-1 siRNAs. The real-time polymerase chain reaction (PCR) assay studied the impact of siRNA transfection on the mRNA levels of affected factors. Moreover, the viability of cancer cells was investigated by the MTT assay. Results: The investigations demonstrated that the mRNA levels of IGF1R and HIF-1α strongly reduced in tumor cell lines following siRNA transfection. Moreover, silencing IGF1R and HIF-1 additively downregulated cell viability in cancer cell lines. Conclusion: These findings imply that cancer combination therapy based on IGF1R and HIF-1α targeting may be a novel promising approach for the immunotherapy of breast and colorectal cancers; however, more study is required.

Protective effects of endothelial progenitor cell microvesicles carrying miR‑98‑5p on angiotensin II‑induced rat kidney cell injury

With the increasing number of patients with hypertensive nephropathy worldwide, it has posed a major threat to health and studies on its treatment and pathogenesis are imminent. The present study investigated the mechanism through which microRNA (miR)-98-5p in microvesicles (MVs) secreted by endothelial progenitor cells (EPCs) is involved in the repair of angiotensin II (Ang II)-induced injury of rat primary renal kidney cells (PRKs). After isolation of rat renal cortical sections, PRKs were isolated by density gradient centrifugation and identified by immunofluorescence staining. Transmission electron microscopy identifies successful separation of Mvs. An in vitro cell injury model was constructed using Ang II. The Gene Expression Omnibus was used to analyze the differentially expressed genes between diabetic rats and normal rats, and the Kyoto Encyclopedia of Genes and Genomes was used to analyze the signaling pathways involved in these differentially expressed genes. Reverse transcription-quantitative PCR was used to analyze the effect of EPC-MVs on the expression of miRNA induced by Ang II, and the levels of target genes and signaling pathway-related proteins involved were analyzed by western blot. luciferase was used to detect the targeted binding of miR-98-5p to insulin-like growth factor 1 receptor (IGF1R). Enzyme-linked immunosorbent assay was used to analyze the effect of EPC-MVs on Ang II-induced oxidative stress and inflammation levels on PRKs. Cell Counting Kit-8 was used to analyze the effect of EPC-MVs on the cell viability of PRKs induced by Ang II. The results showed that treatment of PRKs with Ang II decreased cell viability, whereas oxidative stress and inflammation were increased. However, EPC-MVs alleviated Ang II-induced damage of the PRKs. During this process, the Ang-II-induced downregulation of miR-98-5p was reversed by EPC-MVs, so miR-98-5p may be a key factor regulating the action of EPC-MVs. Dual-luciferase assay confirmed that miR-98-5p targets IGF1R. It was subsequently demonstrated that EPC-MVs overexpressing miR-98-5p promoted phosphorylation of PI3K/Akt/endothelial nitric oxide synthase (eNOS), and inhibited the oxidative stress and inflammation in PRKs, which were reversed by the overexpression of IGF1R. In conclusion, the results of the present study demonstrated that EPC-MVs with high expression of miR-98-5p can activate the PI3K/Akt/eNOS pathway by regulating IGF1R, as well as protect PRKs from Ang II-induced oxidative stress, inflammation and inhibition of cell viability.

Spermidine overrides INSR (insulin receptor)-IGF1R (insulin-like growth factor 1 receptor)-mediated inhibition of autophagy in the aging heart

ABSTRACT Although attenuated IGF1R (insulin-like growth factor 1 receptor) signaling has long been viewed to promote longevity in model organisms, adverse effects on the heart have been the subject of major concern. We observed that IGF1R is overexpressed in cardiac tissues from patients with end-stage non-ischemic heart failure, coupled to the activation of the IGF1R downstream effector AKT/protein kinase B and inhibition of ULK1 (unc-51 like autophagy activating kinase 1). Transgenic overexpression of human IGF1R in cardiomyocytes from mice initially induces physiological cardiac hypertrophy and superior function, but later in life confers a negative impact on cardiac health, causing macroautophagy/autophagy inhibition as well as impaired oxidative phosphorylation, thus reducing life expectancy. Treatment with the autophagy inducer and caloric restriction mimetic spermidine ameliorates most of these IGF1R-induced cardiotoxic effects in vivo. Moreover, inhibition of IGF1R signaling by means of a dominant-negative phosphoinositide 3-kinase (PI3K) mutant induces cardioprotective autophagy, restores myocardial bioenergetics and improves late-life survival. Hence, our results demonstrate that IGF1R exerts a dual biphasic impact on cardiac health, and that autophagy mediates the late-life geroprotective effects of IGF1R inhibition in the heart.

IGF-1 receptor regulates upward firing rate homeostasis via the mitochondrial calcium uniporter

Regulation of firing rate homeostasis constitutes a fundamental property of central neural circuits. While intracellular Ca2+ has long been hypothesized to be a feedback control signal, the molecular machinery enabling a network-wide homeostatic response remains largely unknown. We show that deletion of insulin-like growth factor-1 receptor (IGF-1R) limits firing rate homeostasis in response to inactivity, without altering the distribution of baseline firing rates. The deficient firing rate homeostatic response was due to disruption of both postsynaptic and intrinsic plasticity. At the cellular level, we detected a fraction of IGF-1Rs in mitochondria, colocalized with the mitochondrial calcium uniporter complex (MCUc). IGF-1R deletion suppressed transcription of the MCUc members and burst-evoked mitochondrial Ca2+ (mitoCa2+) by weakening mitochondria-to-cytosol Ca2+ coupling. Overexpression of either mitochondria-targeted IGF-1R or MCUc in IGF-1R-deficient neurons was sufficient to rescue the deficits in burst-to-mitoCa2+ coupling and firing rate homeostasis. Our findings indicate that mitochondrial IGF-1R is a key regulator of the integrated homeostatic response by tuning the reliability of burst transfer by MCUc. Based on these results, we propose that MCUc acts as a homeostatic Ca2+ sensor. Faulty activation of MCUc may drive dysregulation of firing rate homeostasis in aging and in brain disorders associated with aberrant IGF-1R/MCUc signaling.

Fine-Tuning Cardiac Insulin-Like Growth Factor 1 Receptor Signaling to Promote Health and Longevity.

Background:The insulin-like growth factor 1 (IGF1) pathway is a key regulator of cellular metabolism and aging. Although its inhibition promotes longevity across species, the effect of attenuated IGF1 signaling on cardiac aging remains controversial.Methods:We performed a lifelong study to assess cardiac health and lifespan in 2 cardiomyocyte-specific transgenic mouse models with enhanced versus reduced IGF1 receptor (IGF1R) signaling. Male mice with human IGF1R overexpression or dominant negative phosphoinositide 3-kinase mutation were examined at different life stages by echocardiography, invasive hemodynamics, and treadmill coupled to indirect calorimetry. In vitro assays included cardiac histology, mitochondrial respiration, ATP synthesis, autophagic flux, and targeted metabolome profiling, and immunoblots of key IGF1R downstream targets in mouse and human explanted failing and nonfailing hearts, as well.Results:Young mice with increased IGF1R signaling exhibited superior cardiac function that progressively declined with aging in an accelerated fashion compared with wild-type animals, resulting in heart failure and a reduced lifespan. In contrast, mice with low cardiac IGF1R signaling exhibited inferior cardiac function early in life, but superior cardiac performance during aging, and increased maximum lifespan, as well. Mechanistically, the late-life detrimental effects of IGF1R activation correlated with suppressed autophagic flux and impaired oxidative phosphorylation in the heart. Low IGF1R activity consistently improved myocardial bioenergetics and function of the aging heart in an autophagy-dependent manner. In humans, failing hearts, but not those with compensated hypertrophy, displayed exaggerated IGF1R expression and signaling activity.Conclusions:Our findings indicate that the relationship between IGF1R signaling and cardiac health is not linear, but rather biphasic. Hence, pharmacological inhibitors of the IGF1 pathway, albeit unsuitable for young individuals, might be worth considering in older adults.

Role of IGF-1R in epithelial-mesenchymal transdifferentiation of human peritoneal mesothelial cells.

Peritoneal fibrosis (PF) is caused by epithelial-mesenchymal transdifferentiation (EMT) in the peritoneum under high glucose (HG) conditions. The study aimed to explored the role of Insulin-like growth factor 1 receptor (IGF-1R) in the regulation of EMT in human peritoneal mesothelial cells (HPMCs). We used HG peritoneal dialysis fluid (PDF) to induce in vivo PF in mice, and treated HPMCs with HG in vitro to stimulate EMT. In the mice, the higher the glucose concentration in the dialysate, the more obvious the peritoneal tissue thickening and the more that collagen was deposited. The in vitro study indicated that the expression of IGF-1R, α-SMA, vimentin was upregulated, while the expression of occludin, ZO-1, and E-cadherin was downregulated in HPMCs under HG and IGF-1R overexpression conditions. Conversely, the expression of IGF-1R, α-SMA, and vimentin was downregulated, while the expression of occludin, ZO-1, and E-cadherin was upregulated in IGF-1R-underexpressed HPMCs under HG conditions. The cell migration abilities were increased, while the cell adhesion abilities were reduced in HPMCs under HG and IGF-1R overexpression conditions. In contrast, cell migration abilities were reduced, while cell adhesion abilities were increased in IGF-1Runderexpressed HPMCs under HG conditions. Targeting at IGF-1R may provide novel insights into the prevention and treatment of PF.

CircRASSF2 promotes IGF1R and osteosarcoma metastasis via sponging miR-6838-5p.

BackgroundOsteosarcoma (OS) often occurs in children and adolescents and is highly malignant. Analyzing the pathogenesis of OS has great significance for prognosis and the discovery of new treatment strategies.MethodsThe effects and mechanism of circular RNA (circRNA) on OS were analyzed, as was the correlation between circRASSF2 and insulin-like growth factor 1 receptor (IGF1R) in data from The Cancer Genome Atlas (TCGA). The expression levels of microRNA (miR)-6838-5p and circRASSF2 in OS cells and osteoblasts were detected. The dual luciferase report was used to verify the targeting relationship. OS cells overexpressing circRASSF2, miR-6838-5p and/or IGF1R were constructed. The expression level of IGF1R and the biological behavior of the cells were detected. Eighty-two pairs of OS tissue and adjacent normal tissue samples were collected, and the levels of circRASSF2, miR-6838-5p, and IGF1R mRNA were detected by reverse transcription-quantitative PCR (RT-qPCR).ResultsCompared with osteoblasts, OS cells showed lower expression of miR-6838-5p and higher expression of circRASSF2. The dual luciferase report confirmed that miR-6838-5p targeted IGF1R. Overexpression of IGF1R significantly blocked the anticancer effects of miR-6838-5p. The dual luciferase report verified that circRASSF2 targeted miR-6838-5p, and promoted the expression of IGF1R. Overexpression of circRASSF2 not only promoted the malignant biological behavior of OS cells, but also blocked the anticancer effects of miR-6838-5p. In OS tissue, circRASSF2 and IGF1R were upregulated, and the two were positively correlated. MiR-6838-5p was downregulated, which negatively correlated with both circRASSF2 and IGF1R. High levels of circRASSF2 were associated with higher stage and metastasis of OS.ConclusionsIn conclusion, the promoting effects of IGF1R on OS are targeted by miR-6838-5p. CircRASSF2 restored the expression of IGF1R by sponging miR-6838-5p, thereby promoting the progression of OS.

LncRNA SNHG11 facilitates prostate cancer progression through the upregulation of IGF‑1R expression and by sponging miR‑184.

Long non-coding RNA (lncRNA) small nucleolar RNA host gene 11 (SNHG11) has been shown to play an important role in the development and progression of numerous types of cancer. However, to the best of our knowledge, the role of SNHG11 in prostate cancer (PCa) development and metastasis remains unclear. Thus, the aim of the present study was to investigate the functional role and molecular mechanisms of SNHG11 in PCa progression. It was revealed that the SNHG11 expression levels were significantly upregulated in PCa tissues, in comparison with those in adjacent normal tissues. Functionally, SNHG11 knockdown significantly suppressed PCa cell proliferation, migration, invasion and metastasis in vitro and in vivo. Furthermore, SNHG11 was found to positively regulate insulin-like growth factor 1 receptor (IGF-1R) expression by sponging microRNA (miRNA/miR)-184 in PCa cells. The results of rescue experiments demonstrated that IGF-1R overexpression reversed the suppressive effects of SNHG11 knockdown on the proliferation, migration and invasion of PCa cells. On the whole, the findings of the present study suggest that SNHG11 expression is upregulated in PCa and that it facilitates PCa progression, at least in part, via the modulation of the miR-184/IGF-1R signaling axis.

Long non-coding RNA DUXAP8 promotes tumorigenesis by regulating IGF1R via miR-9-3p in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related death worldwide with a low 5-year survival rate. Long non-coding RNA (lncRNA) double homeobox A pseudogene 8 (DUXAP8) is an oncogene and a potential biomarker in various tumors, such as ovarian, colorectal and non-small-cell lung cancer. However, the function and molecular mechanism underlying DUXAP8 in HCC progression is not completely understood. The expression of DUXAP8, microRNA (miR)-9-3p and insulin-like growth factor 1 receptor (IGF1R) in HCC tissues and cells was detected via reverse transcription-quantitative PCR. The expression levels of IGF1R and epithelial-mesenchymal transition-associated proteins (Snail, Slug, E-cadherin, N-cadherin and vimentin) were assessed via western blotting. The effects of DUXAP8, miR-9-3p and IGF1R on proliferation, migration and invasion were examined by conducting Cell Counting Kit-8 and Transwell assays, respectively. The interaction between miR-9-3p and DUXAP8 or IGF1R was predicted using StarBase or TargetScan, and further assessed using dual luciferase reporter and RNA immunoprecipitation assays. DUXAP8 and IGF1R were upregulated and miR-9-3p was downregulated in HCC tissues and cells compared with adjacent healthy tissues and a normal liver cell line, respectively. miR-9-3p overexpression decreased the protein expression level of IGF1R, and miR-9-3p knockdown enhanced the protein expression level of IGF1R in HCC cells compared with the corresponding control groups. Moreover, compared with the corresponding control groups, DUXAP8 knockdown and miR-9-3p overexpression increased E-cadherin protein expression levels, and decreased Snail, Slug, N-cadherin and vimentin protein expression levels. However, miR-9-3p inhibitor and IGF1R overexpression reversed DUXAP8 knockdown- and miR-9-3p overexpression-induced effects, respectively. In addition, compared with the corresponding control groups, DUXAP8 knockdown and miR-9-3p overexpression suppressed proliferation, migration and invasion, which was reversed by miR-9-3p inhibitor and IGF1R overexpression, respectively. Moreover, miR-9-3p as the target of DUXAP8 and IGF1R as the target of miR-9-3p were verified in HCC cells. lncRNA DUXAP8 contributed to HCC tumorigenesis via the miR-9-3p/IGF1R axis, providing a novel therapeutic approach for HCC diagnosis and treatment.

Ginkgoic acid impedes gastric cancer cell proliferation, migration and EMT through inhibiting the SUMOylation of IGF-1R

The imbalance of SUMOylation is related to different cancers, including gastric cancer (GC). Ginkgolic acid (GA) inhibits the growth and invasion of many cancer cells, and it has been reported to restrain SUMOylation. However, the role of GA in GC and whether it functions through SUMOylation remains to be clarified. Our research revealed that GA (15:1) inhibited cell proliferation, migration, epithelial-mesenchymal transition (EMT) and overall protein SUMOylation in BGC823 and HGC27 cells. In addition, knockdown of SUMO1 (small ubiquitin-like modifier) instead of SUMO2/3 played a similar role to GA in cell behaviors. Besides, nuclear IGF-1R (insulin-like growth factor 1 receptor) expression was markedly upregulated in GC cells compared to normal gastric epithelial cells. GA prevented IGF-1R from binding to SUMO1, thereby suppressing its nuclear accumulation. Further research found that IGF-1R directly bound to SNAI2 (snail family zinc finger 2) promoter. The interference of IGF-1R downregulated the mRNA and protein levels of SNAI2, while the overexpression of SUMO1, IGF-1R and UBC9 (SUMO-conjugating enzyme) played the opposite role. Furthermore, the co-transfection of SUMO1, UBC9 and IGF-1R vectors or the overexpression of SNAI2 reversed the inhibitory effects of GA on cell proliferation, migration and EMT. Finally, GA impeded the growth of GC xenografts and decreased the expression of nuclear IGF-1R and SNAI2 in vivo. In conclusion, these findings demonstrated that GA hindered the progression of GC by inhibiting the SUMOylation of IGF-1R. Thus, GA might be a promising therapeutic for GC.

MiR-520b Inhibits IGF-1R to Increase Doxorubicin Sensitivity and Promote Cell Apoptosis in Breast Cancer.

Doxorubicin (DOX) is currently one of the most widely used and effective drugs for the treatment of breast cancer, but drug resistance in breast cancer often leads to poor efficacy. MicroRNAs (miRNAs) are involved in the development and progression of various tumors and increasing number of studies have confirmed that abnormal miR-520b expression is closely associated breast cancer. We analyzed the clinical features, including miR-520b, of 30 patients with breast cancer. Further, we analyzed the interaction between miR-520b and insulin-like growth factor 1 receptor (IGF-1R) in breast cancer cell. miR-520b expression was significantly increased in chemotherapy-sensitive patients and was positively correlated with the chemotherapeutic efficacy in breast cancer. Cell proliferation assay confirmed that miR-520b promotes DOX-induced breast cancer cell apoptosis by regulating the PI3K/AKT signaling pathway. Moreover, bioinformatics method and dual luciferase reporter assay demonstrated that miR-520b negatively regulates IGF-1R, and IGF-1R overexpression and enhanced activity are closely associated with tumor development, progression, metastasis, and chemotherapy resistance. Similarly, cell proliferation assay showed that IGF-1R is negatively correlated with the efficacy of DOX chemotherapy and affects cell apoptosis mediated by the PI3K/AKT signaling pathway. On the contrary, miR-520b can downregulate the expression of IGF-1R. miR-520b increases DOX sensitivity and promotes cell apoptosis in breast cancer by inhibiting IGF-1R expression by the PI3K/AKT signaling pathway.

SOX4 Mediates BRAF Inhibitor Resistance in Melanoma through Regulation of IGF-1R Signaling: In Vitro Study

Abstract Objective: SOX4, a transcription factor, has been found to contribute to tumorigenesis in several cancers. This study was performed to determine whether SOX4 mediates BRAF inhibitor resistance in melanoma. Methods: Melanoma cell lines with acquired resistance to BRAF inhibitor (SK-MEL-5R, SK-MEL-28R, and A375R) were generated by adding escalating concentrations of PLX4032 into parental SK-MEL-5, SK-MEL-28, and A375 cells for >6 months. The expression of SOX4 and insulin-like growth factor 1 receptor (IGF-1R) was measured by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. The downstream signaling of IGF-1R was detected by Western blotting. SOX4 and IGF-1R overexpression or knockdown was conducted by lentivirus transfection. Cell viability and apoptosis were demonstrated by MTT and flow cytometry, respectively. The binding ability of SOX4 to IGF-1R promoter was determined by chromatin immunoprecipitation quantitative PCR assay. Results: SOX4 was upregulated in BRAF inhibitor-resistant melanoma cells as compared with parental cells (SK-MEL-5 group, 1.02 vs. 6.33; SK-MEL-28 group, 1.03 vs. 3.22; A375 group, 1.00 vs. 1.86; t =°7.069, 29.26, and 5.291, respectively; all P < 0.01), and PLX4032 treatment could not alter the expression of SOX4 in resistant cells. SOX4 overexpression attenuated the response of parental cells to PLX4032 (for cell viability, SK-MEL-5 group: 77.76% vs. 104.28%, F = 91.50; SK-MEL-28 group: 60.59% vs. 93.13%, F = 171.8; A375 group: 62.50% vs. 80.87%, F = 47.15. For apoptosis rates, SK-MEL-5 group: 34.90% vs. 14.31%, F = 4.781; SK-MEL-28 group, 40.8% vs. 29.4%, F = 13.32, P = 0.063; A375 group: 40.20% vs. 17.09%, F = 11.39; all P < 0.05, otherwise indicated). While SOX4 knockdown enhanced the response of resistant cells to PLX4032 (for cell viability, SK-MEL-5R group: 93.75% vs. 69.53%, F = 94.45, SK-MEL-28R group: 95.60% vs. 66.79%, F = 30.41, A375R group: 95.51% vs. 59.98%, F = 111.6; for apoptosis rates, SK-MEL-5R group: 16.2% vs. 44.4%, F = 25.67, SK-MEL-28R group: 26.59% vs. 44.20%, F = 158.0, A375R group: 5.98% vs. 31.51%, F = 14.35, and all P < 0.01). Chromatin immunoprecipitation quantitative PCR assay demonstrated that SOX4 binded to the promoter of IGF-1R (1.04 vs. 1.94 [−1044 to −920 bp] and 0.110 vs. 0.139 [GAPDH], F = 534.5, P < 0.01). In addition, SOX4 overexpression increased IGF-1R and its downstream phosphorylated ERK, phosphorylated AKT, and phosphorylated STAT3 expression, while SOX4 knockdown exerted the opposite effects. Moreover, IGF-1R knockdown overcame SOX4 overexpression-induced PLX4032 resistance (cell viability: 35.85% vs. 52.79% vs. 37.84% [A375 group, negative control group vs. SOX4 overexpressing group vs. SOX4 overexpressing + sh-IGF-1R group]; apoptosis rates: 25.30% vs. 9.56% vs. 22.26 [A375 group, negative control group vs. SOX4 overexpressing group vs. SOX4 overexpressing + sh-IGF-1R group]; F = 13.01 and 41.87, respectively; all P < 0.01), while IGF-1R overexpression abrogated SOX4 knockdown-induced response enhancement to PLX4032 for comparison of negative control group, sh-SOX4 group and sh-SOX4 + IGF-1R overexpressing group (cell viability: 96.62% vs. 86.86% vs. 97.26% (A375R), 98.15% vs. 81.63% vs. 98.49% [SK-MEL-5R]; apoptosis rates: 13.81% vs. 32.00% vs. 12.16 [A375R], 29.70% vs. 41.40% vs. 26.10% [SK-MEL-5R]; F = 13.56, 12.86, 38.81, and 39.85, respectively; all P < 0.01). Conclusion: SOX4 mediates BRAF inhibitor resistance in melanoma through regulation of IGF-1R signaling. SOX4 might serve as a potential target for the treatment of BRAF inhibitor-resistant melanoma.

Activation of \u03b21 integrins and caveolin-1 by TF/FVIIa promotes IGF-1R signaling and cell survival

The tissue factor/coagulation factor VIIa (TF/FVIIa) complex induces transactivation of the IGF-1 receptor (IGF-1R) in a number of different cell types. The mechanism is largely unknown. The transactivation leads to protection from apoptosis and nuclear translocation of the IGF-1R. The aim of this study was to clarify the signaling pathway between TF and IGF-1R after FVIIa treatment with PC3 and DU145 prostate or MDA-MB-231 breast cancer cells as model systems. Protein interactions, levels, and phosphorylations were assessed by proximity ligation assay or flow cytometry in intact cells and by western blot on cell lysates. The transactivation of the IGF-1R was found dependent on TF/FVIIa-induced activation of β1-integrins. A series of experiments led to the conclusion that the caveolae protein caveolin-1 prevented IGF-1R activation in resting cells via its scaffolding domain. TF/FVIIa/β1-integrins terminated this inhibition by activation of Src family kinases and subsequent phosphorylation of caveolin-1 on tyrosine 14. This phosphorylation was not seen after treatment with PAR1 or PAR2 agonists. Consequently, the protective effect of FVIIa against apoptosis induced by the death receptor agonist TRAIL and the de novo synthesis of cyclin D1 induced by nuclear IGF-1R accumulation were both significantly reduced by down-regulation of β1-integrins or overexpression of the caveolin-1 scaffolding domain. In conclusion, we present a plausible mechanism for the interplay between TF and IGF-1R involving FVIIa, β1-integrins, Src family proteins, and caveolin-1. Our results increase the knowledge of diseases associated with TF and IGF-1R overexpression in general but specifically of TF-mediated signaling with focus on cell survival.

Abstract A20: Persistent high-risk HPV infection in the development of uterine cervical cancer

Abstract The early detection and treatment of preneoplastic uterine cervical lesions is certainly a major challenge for developing countries. Preneoplastic lesions result from human papilloma virus (HPV) infection in cells located in the transformation zone of the cervical epithelial tissue, causing low-grade or high-grade lesions. HPV infections are transient and most of the time cleared within a couple of years following exposure. However, 10–20% of infections persist latently, leading to disease progression and, ultimately, various forms of invasive cancer. The transformation of preneoplastic lesions in invasive cancer cells is provoked at least partly by HPV, especially through gene and protein expression modulation. This modulation involves tissue oxygenation, oncogene activation, loss of tumor suppressor genes and aberrant molecular signaling pathways (such as Insulin-like growth factor-1 receptor (IGF-1R). In recent studies, it was reported that IGF1R is overexpressed by effect of E6 AA-a and E-G350 HPV-16 variants. The aim of the present study was to prospectively and retrospectively report the detection of HPV-16 variants and IGF1R expression in patients with cervical cancer and assess the relationship with development of uterine cervical cancer. Sixty consecutive patients with invasive cervical cancer were assessed for HPV-16. A total of 35 patients (58.3%) were infected with HPV-16, with a mean age of 51 years (range: 30–76 years). Thirty patients (85.7%) were diagnosed with European HPV-16 variant and five patients (14.3%) were diagnosed with non-European HPV-16 variants. European HPV-16 variants (E-HPV-16) were mainly E-G350 (n = 14; 41.1%) and E-T350 (n = 12; 35.2%) subclasses. Of these 35 patients, 15 received exclusive radiotherapy and four radiochemotherapy, and the remaining 16 did not complete the treatment (NCT). Of the patients who presented an incomplete response, overexpression of IGF1R was detected in all. Of patients NCT, we retrospectively studied one case in IIIB stage FIGO. Samples of preneoplastic lesions and invasive cervical cancer were analyzed to confirm histopathologic diagnosis and detect the presence of HPV variants and overexpression of IGF-1R. Of the results found in this case, in which the patient was diagnosed in 1986 with a high-grade squamous intraepithelial cervical lesion, HPV-16 E-G350 and HPV-33 coinfection as low expression of IGF1R were detected and for the invasive cancer diagnosed in 2002, the presence of HPV-16 AA-c and overexpression of IGF1R was detected. The detection of high-risk HPV in samples of preneoplastic lesions and invasive cancer in a 60-year-old patient at the time of diagnosis showed that HPV33 infection was eliminated. Results such as these suggest that persistent infection with HPV-16 variants, overexpression of IGF1R, such as viral load and age, could be necessary factors for disease progression, and thus contribute to the identification of patients with a high risk of viral persistence and development of cervical neoplasia. Note: This abstract was not presented at the conference. Citation Format: Pablo Moreno Acosta, Diana Mayorga, Nicolas Magné. Persistent high-risk HPV infection in the development of uterine cervical cancer [abstract]. In: Proceedings of the AACR Special Conference on the Microbiome, Viruses, and Cancer; 2020 Feb 21-24; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2020;80(8 Suppl):Abstract nr A20.

IGF1R as druggable target mediating PI3K-δ inhibitor resistance in a murine model of chronic lymphocytic leukemia

AbstractTargeted therapy is revolutionizing the treatment of cancers, but resistance evolves against these therapies and derogates their success. The phosphatidylinositol 3-kinase delta (PI3K-δ) inhibitor idelalisib has been approved for treatment of chronic lymphocytic leukemia (CLL) and non-Hodgkin lymphoma, but the mechanisms conferring resistance in a subset of patients are unknown. Here, we modeled resistance to PI3K-δ inhibitor in vivo using a serial tumor transfer and treatment scheme in mice. Whole-exome sequencing did not identify any recurrent mutation explaining resistance to PI3K-δ inhibitor. In the murine model, resistance to PI3K-δ inhibitor occurred as a result of a signaling switch mediated by consistent and functionally relevant activation of insulin-like growth factor 1 receptor (IGF1R), resulting in enhanced MAPK signaling in the resistant tumors. Overexpression of IGF1R in vitro demonstrated its prominent role in PI3K-δ inhibitor resistance. IGF1R upregulation in PI3K-δ inhibitor–resistant tumors was mediated by functional activation and enhanced nuclear localization of forkhead box protein O1 transcription factors and glycogen synthase kinase 3β. In human CLL, high IGF1R expression was associated with trisomy 12. CLL cells from an idelalisib-treated patient showed decreased sensitivity to idelalisib in vitro concomitant with enhanced MAPK signaling and strong upregulation of IGF1R upon idelalisib exposure. Thus, our results highlight that alternative signaling cascades play a predominant role in the resistance and survival of cancer cells under PI3K-δ inhibition. We also demonstrate that these pathway alterations can serve as therapeutic targets, because inhibition of IGF1R offered efficacious salvage treatment of PI3K-δ inhibitor–resistant tumors in vitro and in vivo.