Integrin Αvβ3 Research Articles

Treadmill Exercise Alleviates Cognition Disorder by Activating the FNDC5: Dual Role of Integrin αV/β5 in Parkinson's Disease.

Parkinson's disease with cognitive impairment (PD-CI) results in several clinical outcomes for which specific treatment is lacking. Although the pathogenesis of PD-CI has not yet been fully elucidated, it is related to neuronal plasticity decline in the hippocampus region. The dopaminergic projections from the substantia nigra to the hippocampus are critical in regulating hippocampal plasticity. Recently, aerobic exercise has been recognized as an effective therapeutic strategy for enhancing plasticity through the secretion of various muscle factors. The exact role of FNDC5-an upregulated, newly identified myokine produced after exercise-in mediating hippocampal plasticity and regional dopaminergic projections in PD-CI remains unclear. In this study, the effect of treadmill exercise on hippocampal synaptic plasticity was evaluated in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced chronic PD models. The results showed that treadmill exercise substantially alleviated the motor dysfunction, cognition disorder, and dopaminergic neuron degeneration induced by MPTP. Here, we discovered that the quadriceps, serum, and brain FNDC5 levels were lower in PD mice and that intervention with treadmill exercise restored FNDC5 levels. Moreover, treadmill exercise enhanced the synaptic plasticity of hippocampal pyramidal neurons via increased dopamine levels and BDNF in the PD mice. The direct protective effect of FNDC5 is achieved by promoting the secretion of BDNF in the hippocampal neurons via binding the integrin αVβ5 receptor, thereby improving synaptic plasticity. Regarding the indirect protection effect, FNDC5 promotes the dopaminergic connection from the substantia nigra to the hippocampus by mediating the interaction between the integrin αVβ5 of the hippocampal neurons and the CD90 molecules on the membrane of dopaminergic terminals. Our findings demonstrated that treadmill exercise could effectively alleviate cognitive disorders via the activation of the FNDC5-BDNF pathway and enhance the dopaminergic synaptic connection from SNpc to the hippocampus in the MPTP-induced chronic PD model.

Imaging angiogenesis in an intracerebrally induced model of brain macrometastasis using αv β3 -targeted iron oxide microparticles.

Brain metastasis is responsible for a large proportion of cancer mortality, and there are currently no effective treatments. Moreover, the impact of treatments, particularly antiangiogenic therapeutics, is difficult to ascertain using current magnetic resonance imaging (MRI) methods. Imaging of the angiogenic vasculature has been successfully carried out in solid tumours using microparticles of iron oxide (MPIO) conjugated to a Arg-Gly-Asp peptide (RGD) targeting integrin αv β3 . The aim of this study was to determine whether RGD-MPIO could be used to identify angiogenic blood vessels in brain metastases in vivo. A mouse model of intracerebrally implanted brain macrometastasis was established through intracerebral injection of 4T1-GFP cells. T2 *-weighted imaging was used to visualise MPIO-induced hypointense voxels in vivo, and Prussian blue staining was used to visualise MPIO and endogenous iron histologically ex vivo. The RGD-MPIO showed target-specific binding in vivo, but the sensitivity of the agent for visualising angiogenic vessels per se was reduced by the presence of endogenous iron-laden macrophages in larger metastases, resulting in pre-existing hypointense areas within the tumour. Further, our data suggest that peptide-targeted MPIO, but not antibody-targeted MPIO, are taken up by perivascular macrophages within the macrometastatic microenvironment, resulting in additional nonspecific contrast. While pre-MPIO imaging will circumvent the issues surrounding pre-existing hypointensities and enable detection of specific contrast, our preliminary findings suggest that the use of antibodies rather than peptides as the targeting ligand may represent a preferable route forward for new angiogenesis-targeted molecular MRI agents.

Validation of In-House Kit-Like Synthesis of 68Ga-Trivehexin and Its Biodistribution for Targeting the Integrin αvβ6 Expressing Tumors.

Background: Integrin αvβ6 has become an extremely promising theranostic target for precise delineation of fast-growing malignant cells in the recent years. The aim of the study was to validate the in-house kit-like synthesis of 68Ga-Trivehexin (integrin αvβ6) and to evaluate its uptake in patients with integrin αvβ6 expressing head and neck and pancreatic cancer. Materials and Methods: 68Ga-Trivehexin was synthesized by adding the variable amount of integrin αvβ6 (30-50 μg) to full volume (4-5 mL) Ga-68 in 0.05 M HCl and heating the reaction mixture at 90°C for 12 min at pH 3.5-4 to obtain the radiotracer with high radiochemical purity (RCP) and high yield. Quality control procedures were done to assess the RCP, stability, pyrogenicity and sterility of the radiotracer. 68Ga-Trivehexin was then administered in patients who met the eligibility criteria. Whole body PET/CT scans were done at variable time points post intravenous (i.v.) injection of 84-185 MBq of 68Ga-Trivehexin to assess its biodistribution and maximum uptake time. Results: 0.2 mCi of 68Ga/μg of Trivehexin at 90°C for 12 min was the optimal parameter to obtain 85%-88% of noncorrected yield and 99% of RCP. The 68Ga-Trivehexin showed in vitro stability upto 6 h and was also found to be sterile and pyrogen free. Intense radiotracer uptake was noticed in the tumor and no uptake was noticed in healthy tissues. PET/CT imaging at 60 min post injection was found to be the optimal time for imaging the tumors with 68Ga-Trivehexin. Conclusion: The protocol for in-house kit-like labeling of 68Ga-Trivehexin was safe, reproducible, and cost-effective. 68Ga-Trivehexin is an extremely promising agent for noninvasive molecular imaging of integrin αvβ6 expressing tumors.

Tumor‐Targeted Poly(ArgGlyAsp) Nanocapsules for Personalized Cancer Therapy – In Vivo Study

AbstractThe arginine‐glycine‐glutamic acid (RGD) sequence, an αvβ3 integrin recognition site, is overexpressed in malignancies and neovasculature, making it a potential therapeutic target. Herein, efficacy/safety of tumor‐targeted RGD‐based proteinoid nanocapsules (NCs) entrapping a synergistic combination of two drugs—palbociclib (Pal), a CDK4/6 inhibitor, and alpelisib (Alp), a P13K inhibitor, as a cancer treatment, is assessed. P(RGD) proteinoid polymers are produced by thermal step‐growth polymerization of R, G, and D under an inert atmosphere. P(RGD) NCs, hollow and encapsulating 25 w% each of Pal and Alp, are formed by self‐assembly of the proteinoid polymer. The encapsulation yields of Pal and Alp are 72% and 95%, respectively. Long‐term stability, controlled release, cellular uptake,synergistic cytotoxicity, and induced cell death are evident from in vitro experiments. Findings from in vivo breast, colon and gastric patient‐derived xenograft (PDX) mice experiments are consistent with the in vitro studies showing that the response to treatment with drug‐loaded NCs is similar to that elicited by free drugs, with reduced side effects. The study demonstrates the potential clinical utility of P(RGD) NCs for cancer treatment.

Contact Guidance of Connective Tissue Fibroblasts on Submicrometer Anisotropic Topographical Cues Is Dependent on Tissue of Origin, β1 Integrins, and Tensin-1 Recruitment.

The substratum topography of both natural and synthetic materials is a prominent regulator of cell behaviors including adhesion, migration, matrix fibrillogenesis, and cell phenotype. Connective tissue fibroblasts are known to respond to repeating groove topographical modifications by aligning and exhibiting directed migration, a phenomenon termed contact guidance. Although both reside in collagen rich connective tissues, dermal and gingival fibroblasts are known to exhibit differences in phenotype during wound healing, with gingival tissue showing a fetal-like scarless response. Differences in adhesion formation and maturation are known to underlie both a scarring phenotype and cell response to topographical features. Utilizing repeating groove substrates with periodicities of 600, 900, and 1200 nm (depth, 100 nm), we investigated the roles of integrins αvβ3 and β1 associated adhesions on contact guidance of human gingival (HGFs) and dermal fibroblasts (HDFs). HGFs showed a higher degree of orientation with the groove long axis than HDFs, with alignment of both vinculin and tensin-1 evident on 600 and 900 nm periodicities in both cell types. Orientation with grooves of any periodicity in HGFs and HDFs did not alter the adhesion number or area compared to smooth control surfaces. Growth of both cell types on all periodicities reduced fibronectin fibrillogenesis compared to control surfaces. Independent inhibition of integrin αvβ3 and β1 in both cell types induced changes in spreading up to 6 h and reduced alignment with the groove long axis. At 24 h post-seeding with blocking antibodies, HGFs recovered orientation, but in HDFs, blocking of β1, but not αvβ3 integrins, inhibited alignment. Blocking of β1 and αvβ3 in HDFs, but not HGFs, inhibited tensin-1-associated fibrillar adhesion formation. Furthermore, inhibition of β1 integrins in HDFs, but not HGFs, resulted in recruitment of tensin-1 to αvβ3 focal adhesions, preventing HDFs from aligning with the groove long axis. Our work demonstrates that tensin-1 localization with specific integrins in adhesion sites is an important determinant of contact guidance. This work emphasizes further the need for tissue-specific biomaterials, when integration into host tissues is required.

Scandium-44: Diagnostic Feasibility in Tumor-Related Angiogenesis.

Angiogenesis-related cell-surface molecules, including integrins, aminopeptidase N, vascular endothelial growth factor, and gastrin-releasing peptide receptor (GRPR), play a crucial role in tumour formation. Radiolabelled imaging probes targeting angiogenic biomarkers serve as valuable vectors in tumour identification. Nowadays, there is a growing interest in novel radionuclides other than gallium-68 (68Ga) or copper-64 (64Cu) to establish selective radiotracers for the imaging of tumour-associated neo-angiogenesis. Given its ideal decay characteristics (Eβ+average: 632 KeV) and a half-life (T1/2 = 3.97 h) that is well matched to the pharmacokinetic profile of small molecules targeting angiogenesis, scandium-44 (44Sc) has gained meaningful attention as a promising radiometal for positron emission tomography (PET) imaging. More recently, intensive research has been centered around the investigation of 44Sc-labelled angiogenesis-directed radiopharmaceuticals. Previous studies dealt with the evaluation of 44Sc-appended avb3 integrin-affine Arg-Gly-Asp (RGD) tripeptides, GRPR-selective aminobenzoyl-bombesin analogue (AMBA), and hypoxia-associated nitroimidazole derivatives in the identification of various cancers using experimental tumour models. Given the tumour-related hypoxia- and angiogenesis-targeting capability of these PET probes, 44Sc seems to be a strong competitor of the currently used positron emitters in radiotracer development. In this review, we summarize the preliminary preclinical achievements with 44Sc-labelled angiogenesis-specific molecular probes.

Vascular endothelial-derived Von Willebrand factor inhibits lung cancer progression through the αvβ3/ERK1/2 axis

Lung cancer remains a common malignant tumor causing death due to the rapid industrialization and serious pollution of the environment. The Von Willebrand Factor (vWF) protein is an endothelial marker and is widely used to diagnose cancer and other inflammations, however its exact mechanism of action remains largely unexplored. In particular, how it plays two opposing roles in tumor development is not clear. Our study aimed to the impact of endothelial-derived vWF on tumor development by co-culturing human umbilical vein endothelial cells (HUVECs) with lung cancer cells (95D and A549). A knockdown of endothelial-derived vWF assisted lung cancer cell in proliferation, migration and inhibited apoptosis in vitro, while overexpression of endothelial-derived vWF inhibited the proliferation, migration and induced apoptosis of lung cancer cells. The results of further experiments indicated that the vWF secreted by endothelial cells could affect lung cancer cell migration and apoptosis via its binding to integrin αvβ3 on the surface of lung cancer cells. Furthermore, a novel finding was the fact that endothelial-derived vWF inhibited lung cancer cell apoptosis by phosphorylating ERK1/2. At the same time, we established experimental lung metastasis model and xenograft model in normal mice and vWF−/− mice, and found that knockout of vWF in mice significantly promoted lung cancer growth and metastasis. In conclusion, our research found that endothelial-derived vWF could directly combine to αvβ3 on the exterior of A549 and 95D, thereby mediating lung cancer proliferation, migration and apoptosis and inhibiting the development of lung cancer.

Targeted Nanodrugs to Destroy the Tumor Extracellular Matrix Barrier for Improving Drug Delivery and Cancer Therapeutic Efficacy.

One of the main reasons why most cancer patients do not respond well to chemotherapy is that drugs cannot accumulate in tumors at an optimal dose, eventually resulting in failure to prevent cancer cell growth. To improve drug delivery efficiency, we engineered a highly efficient tumor-targeted and stroma-breaking nanocarrier by the modification of iron oxide nanoparticles (IONPs) with a tumor-targeting peptide c(RGDyK) and a hyaluronidase (HAase) on the surface. The yielding nanocomplex, c(RGDyK)-HAase-IONP, targeted the tumor by binding integrin αvβ3 and went deeply into the tumors by the degradation of hyaluronic acid (HA), which was highly expressed in the tumor extracellular matrix (ECM). Good biostability and a low pH preferred drug release profile were characterized for c(RGDyK)-HAase-IONP carrying DOX in vitro. c(RGDyK)-HAase-IONP showed an improved tumor-targeting (2.5 times higher) effect after intravenous injection in the MC38 tumor-bearing mice model, as determined by whole-body fluorescence imaging compared to the non-targeted IONPs without HAase. After 5 systemic treatments, c(RGDyK)-HAase-IONP/DOX (5 mg/kg of equivalent dose of DOX) significantly inhibited MC38 tumor growth (22.1 ± 7.4 times relative to the non-treated group). Elevated apoptosis and reduced proliferation in the tumor cell were detected in the c(RGDyK)-HAase-IONP/DOX treated tumors compared to the control groups. Overall, the highly efficient targeted nanocarrier c(RGDyK)-HAase-IONP demonstrated tremendous potency for improving drug delivery and tumor therapy efficacy by targeted degradation of the dense HA barrier in the tumor ECM. We determined that such a tumor stroma-degrading nanosystem was capable of reducing tumor recurrence and drug resistance and could ultimately improve clinical tumor treatment responses.

Analysis of the role of irisin receptor signaling in regulating osteogenic/adipogenic differentiation of bone marrow mesenchymal stem cells

ABSTRACT The study aimed to explore the role of the irisin receptor (integrin αVβ5) signaling pathway in obesity-induced osteoporosis and its potential mechanism. The integrin αVβ5 gene of bone marrow mesenchymal stem cells (BMSCs) was silenced and overexpressed, and the cells were exposed to irisin treatment and mechanical stretch. Mouse models of obesity were established by feeding mice a high-fat diet, and 8-week caloric restriction/aerobic exercise regimens were implemented. The results showed that after silencing the integrin αVβ5, the osteogenic differentiation of BMSCs was significantly reduced. While overexpression of the integrin αVβ5 increased the osteogenic differentiation of BMSCs. Besides, mechanical stretch promoted the osteogenic differentiation of BMSCs. Obesity did not affect integrin αVβ5 expression in the bone, but it downregulated the expression of irisin and osteogenic factors, upregulated the expression of adipogenic factors, increased bone marrow fat, reduced bone formation, and destroyed the bone microstructure. Caloric restriction, exercise, and a combined regimen reversed these effects and improved obesity-induced osteoporosis, with the combined treatment exhibiting the most potent effect. This study confirms that the irisin receptor signaling pathway has a significant part in transmitting ‘mechanical stress’ and regulating ‘osteogenic/adipogenic differentiation’ of BMSCs via recombinant irisin, mechanical stretch, and overexpression/silencing of the integrin αVβ5 gene.

Recent advances in the tumor-penetrating peptide internalizing RGDfor cancer treatment and diagnosis.

Cancer has become a prominent disease that seriously endangers human health. The complexity of the biological characteristics of the tumor makes it challenging for traditional therapeutic drugs to penetrate tumor tissues and exert their antitumor effects.Internalizing RGD peptide (iRGD) is a novel tumor-homing peptide that binds to αvβ3 and αvβ5 integrins on the surface of tumor vessels through the C-end rule (CendR) motif. The CendR motif binds to the neuropilin-1 (NRP-1) receptor on tumor cells, initiating NRP-1-mediated transcytosis to facilitate drug entry into the tumor tissue. Multiple studies demonstrated that iRGD improved the penetration and targeting of antitumor drugs, thereby enhancing their antitumor efficacy. In this review, we initially described the origins of iRGD and its penetration mechanism. Furthermore, we presented updates on the application of iRGD in cancer chemotherapy, photodynamic therapy, gene therapy, immunotherapy, treatment with antibodiesor protein-based biologics, and tumor imaging.

Can targeted nanoparticles distinguish cancer metastasis from inflammation?

Targeting ligands have been widely used to increase the intratumoral accumulation of nanoparticles and their uptake by cancer cells. However, these ligands aim at targets that are often also upregulated in inflamed tissues. Here, we assessed the ability of targeted nanoparticles to distinguish metastatic cancer from sites of inflammation. Using common targeting ligands and a 60-nm liposome as a representative nanoparticle, we generated three targeted nanoparticle (NP) variants that targeted either fibronectin, folate, or αvβ3 integrin, whose deposition was compared against that of standard untargeted NP. Using fluorescently labeled NPs and ex vivo fluorescence imaging of organs, we assessed the deposition of the NPs into the lungs of mice modeling 4 different biological landscapes, including healthy lungs, aggressive metastasis in lungs, dormant/latent metastasis in lungs, and lungs with general pulmonary inflammation. Among the four NP variants, fibronectin-targeting NP and untargeted NP exhibited the highest deposition in lungs harboring aggressive metastases. However, the deposition of all targeted NP variants in lungs with metastasis was similar to the deposition in lungs with inflammation. Only the untargeted NP was able to exhibit higher deposition in metastasis than inflammation. Moreover, flow-cytometry analysis showed all NP variants accumulated predominantly in immune cells rather than cancer cells. For example, the number of NP+ macrophages and dendritic cells was 16-fold greater than NP+ cancer cells in the case of fibronectin-targeting NP. Overall, targeted NPs were unable to distinguish cancer metastasis from general inflammation, which may have clinical implications to the nanoparticle-mediated delivery of cancer drugs.

Abstract 3073: Action of thyroid hormones in sorafenib treatment of thyroid cancer

Abstract Background: Thyroid carcinoma (TC) is the most common endocrine neoplasia. Its incidence has increased in the last 40 years worldwide. Sorafenib (Sor), a tyrosine kinase inhibitor (TKI), was approved for the treatment of TC, being hypothyroidism the most frequent consequence of Sor-induced endocrine dysfunction. We have described that thyroid hormones (TH) increase cell proliferation in TC cell lines. Thus, hormone replacement therapy to treat Sor-induced hypothyroidism could negatively affect its antitumor action. We have also shown that the selective inhibition of the TH membrane receptor, integrin αVβ3, diminishes proliferation. Objective: To study integrin αVβ3 inhibition to enhance Sor antineoplastic activity in TC and the molecular mechanisms involved. Experimental Procedures: 8505C human anaplastic TC cell viability was evaluated by MTS assay. Sor was the TKI used, and Cilengitide (Cile) was used to inhibit integrin αVβ3. Protein modulation was measured by Western blot. Apoptosis induction was determined by APC-Annexin V Propidium iodide staining, followed by flow cytometry. In silico analysis on several databases were performed using CBioPortal (https://cbioportal.org/; TCGA, Cell 2014, n=504) and R2 (http://r2.amc.nl; GSE126729, n=28). Results: We first performed a bioinformatic analysis by cBioPortal on the PanCancer Atlas data and found that thyroid tumors are those with the highest integrin αVβ3 subunits expression. We then studied the role of integrin αVβ3 in Sor inhibition of TH-induced proliferation in TC cells. Treatment of 8505C cells with integrin αVβ3 antagonist Cile inhibits TH-induced proliferation (p&lt0.0001), confirming the participation of the integrin. As expected, Sor treatment decreases proliferation (p&lt0.05) while Cile addition did not change the inhibition level. Also, we found that Sor and Cile treatment diminished PCNA and Cyclin D1 expression levels. Cells were preincubated with Sor and Cile and then treated or not with TH for 48h to analyze apoptosis. Sor treatment increases the number of apoptotic cells relative to untreated control (p&lt0.0001) and the presence of TH reduces the rate of apoptosis (p&lt0.01). We then studied Sor target genes expression by R2 on an anaplastic TC patients’ database. FGFR is highly expressed among Sor targets, and it is correlated with integrin αVβ3 expression. Thus, FGFR expression modulation was studied by Western blot on cells treated as previously described. Finally, we found that TH treatment increased FGFR expression, Sor diminishes expression and in presence of TH, FGFR expression was rescued. Interestingly, Cile addition resensitized cells to Sor in presence of TH. Conclusion: Our results establish that the effective dual Sor and Cile treatment can significantly drive tumor proliferation inhibition and apoptosis, and it could provide alternatives to the treatments currently used for this disease. Citation Format: Mateo N. Campos Haedo, Maria C. Diaz Flaque, Helena A. Sterle, Johanna A. Diaz Albuja, Maria F. Cayrol, Maria M. Debernardi, Marina Perona, Guillermo J. Juvenal, Graciela A. Cremaschi, Cinthia Rosemblit. Action of thyroid hormones in sorafenib treatment of thyroid cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3073.

Abstract 2001: Engineering extracellular vesicle like liposomes with integrin αVβ5 to study its role in cancer metastasis

Abstract Background: Extracellular vesicles (EVs) are key mediators of cancer metastasis. EV membrane-associated integrins show organotrophic properties by interacting with microenvironmental stromal cells and extracellular matrix. Specifically, integrin αVβ5 is associated with the metastasis of cancers with remarkable liver tropism, including pancreatic cancer, uveal melanoma, and colorectal cancer. Despite the importance of understanding the pathogenic roles of EVs, EV heterogeneity, lengthy isolation, and low yields make it difficult to study the function of EVs at the molecular level. Therefore, we explored engineering EV-like lipid nanoparticles with selected cargo, such as integrin αVβ5, as a tool to study EV biology. Methods: EV-like liposomes (in terms of size and charge) have been produced by our group by nanoprecipitation. Aqueous solvent (proteins in Milli-Q water) and organic solvent (cholesterol and DMPC in ethanol) were mixed in a 3D-printed microfluidic chip. Recombinant integrin αVβ5 was purchased from R&D System and encapsulated into the liposomes in one step. Liposomes with recombinant GFP protein (Thermo Fisher) and liposomes with cellular proteins isolated from 92.1 and A431 cells were produced as proof of concept. Liposomes were produced with dye SP-DilC18 or labelled by PKH after production. Unencapsulated proteins and unbound dye was removed by ultracentrifugation or size exclusion chromatography (qEV, Izon). Stain-free gel (Biorad), flow cytometry, and Tecan Reader were used to detect encapsulated proteins. Hepatocyte uptake of liposomes was analyzed by Incucyte. The liposomes were characterized by transmission emission microscopy (TEM), nanoparticle tracking analysis, and dynamic light scattering. Results: Liposomes with and without protein cargo presented a bi-layer lipid membrane by cryoTEM. 92.1 cellular protein, A431 cellular protein, GFP, and integrin αVβ5 encapsulated by liposomes were detected by stain-free gel. Positive signals from GFP encapsulated by liposomes were detected by flow cytometry and Tecan Reader, indicating encapsulated proteins can remain functional. By Incucyte fluorescence microscopy, we analyzed the uptake of liposomes by hepatocytes and found that different sizes and charges affect the uptake efficiency. Conclusion: In this study, we produced EV-like liposomes with lipid bilayers in a controllable way in terms of size, charges, and functional cargo. We showed that EV-like liposomes could be a novel and easy-to-use model to study the role of specific EV cargo in cancer metastasis. Citation Format: Yunxi Chen, Rubén R. López, Chaymaa Zouggari Ben El Khyat, Thupten Tsering, Vahé Nerguizian, Julia V. Burnier. Engineering extracellular vesicle like liposomes with integrin αVβ5 to study its role in cancer metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2001.

Abstract 3946: Radiolabeled αvβ3 analog 177Lu-EB-RGD is an effective therapeutic agent in thyroid cancer xenograft mouse model

Abstract Background: Integrins are cell adhesion receptors consisting of 18α and 8β subunits. A subset of integrins (αvβ3) recognizes Arg-Gly-Asp (RGD) peptide motifs which are involved in the neovascularization and progression of various cancers. The aim of the study was to investigate if αvβ3 can serve as a molecular target for the treatment of thyroid cancer (TC) with a novel radiolabeled RGD analog 177Lu-EB-RGD. Methods: Integrin αvβ3 mRNA and/or protein expression was evaluated in 496 TC included in The Cancer Genome Atlas, tissue microarray including 70 TC and 10 normal thyroid samples, and 14 TC cell lines. BRAF-like or RAS-like expression profile was determined through standard BRS scores ranging from -1 to 0 for BRAF-like and 0 to 1 for RAS-like TC. The association between BRS and the αvβ3 expression was tested by the Spearman correlation (r). Nude mice xenografts developing αvβ3 expressing TC after subcutaneous injection of 5*10^6 TC cells were subjected to monotherapy with 0.5 mCi 177Lu-EB-RGD (177Lu) or in combination with Lenvatinib (177Lu+L). The therapeutic efficacy of 177Lu-EB-RGD was compared with standard-of-care Lenvatinib alone (L) and placebo (P). The continuous data were presented as medians with [25-75% interquartile ranges] and compared using Kruskal-Wallis test. The mixed-effects models were used for longitudinal data analysis with adjusted p≤ 0.05 considered statistically significant. Results: We found a moderate negative correlation between BRS and αv (r=-0.5, p<0.001), and β3 (r=-0.27, p<0.001), revealing that BRAF-like tumors have a higher mRNA expression of αvβ3 integrins. Consistently, the highest αvβ3 mRNA and/or protein expression was found in the BRAF-like TC cell lines OCUT2 (BRS=-0.56), TPC1 (BRS=-0.4), K1 (BRS=-0.29), and Hurthle TC cell line XTC1 (BRS=-0.46). The immunostaining revealed a higher αvβ3 expression in papillary TC compared with follicular TC (p=0.002), and in normal thyroid (p<0.001). Poorly differentiated TC had a similar αvβ3 expression to papillary TC (p=0.14). In the thyroid cancer xenograft mouse model, all treatment modalities were more effective than the placebo in decreasing tumor size as early as 5 days after therapy initiation. A significant difference in growth curve and tumor volume was maintained at the study endpoint (L 0.584 cm3 [0.196-0.984] vs P 0.911 cm3 [0.183-1.68], p=0.001; 177Lu 0.259 cm3 [0.103-0.376] vs P, p<0.001; and 177Lu+L 0.274 cm3 [0.108-0.406] vs P, p<0.001). The combination therapy (177Lu+L) resulted in decreased tumor volume as compared with monotherapy with Lenvatinib (p=0.05) but had a similar effect as compared with 177Lu-EB-RGD monotherapy (p=0.99). Conclusions: The radiolabeled αvβ3 analog 177Lu-EB-RGD has potent growth inhibitory effects in TC characterized by a high integrins expression. The αvβ3 integrin could potentially serve as a molecular target for therapy with radiolabeled RGD analogs in TC. Citation Format: Sonam Kumari, Zhantong Wang, Shilpa Thakur, Laura Abaandou, Oksana Gavrilova, Huiyan Lu, Lixin Lang, Dale Kiesewetter, Vasyl Vasko, Joanna Klubo-Gwiezdzinska. Radiolabeled αvβ3 analog 177Lu-EB-RGD is an effective therapeutic agent in thyroid cancer xenograft mouse model. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3946.

Abstract 3949: CYR61, a member of the CCN protein family, regulates IGF-1 in metastatic prostate cancer

Abstract The cysteine-rich angiogenic inducer 61 (CYR61) is a member of the cellular communication network (CCN) protein family that can be induced by growth factors. CYR61-specific functions are dependent on cellular context. In prostate cancer (PCa), CYR61 contributes to cell growth and survival through its interactions with extracellular ligands, such as integrins avb3 and a6b4, in the extracellular matrix space. Interestingly, CYR61 contains an insulin-like growth factor-binding protein domain suggesting that it may interact with insulin-like growth factor-I (IGF-1). High serum levels of IGF-1 are directly linked to PCa development and recently have been studied as a predictor of metastatic disease. Given the important roles that IGF-1 and CYR61 play in PCa and their potential interaction, it is critical to investigate their molecular interplay. This study is designated to specifically determine and characterize the molecular interaction between CYR61 and IGF-1 in metastatic PCa and to determine if this interaction contributes to aggressive tumor properties (e.g., proliferation, tumorsphere formation). Using immunoblotting, we demonstrated that CYR61 is upregulated in the metastatic cell line PC3 and downregulated in the docetaxel-resistant cell line PC3-DR. Furthermore, optimized knockdown of CYR61 using siRNA significantly reduced PC3 cell proliferation, viability, and prostasphere formation. To examine the underlying mechanisms associated with IGF-1 signaling, we assessed the activation of either the PI3/Akt and MAPK pathways in PC3 cells with CYR61 silencing. CYR61 siRNA-mediated knockdown decreased activation of the PI3/Akt pathway but did not affect the MAPK pathway. We also initiated studies to determine the co-localization of CYR61 and IGF-1 using immunofluorescence microscopy (IF). Based on the initial IF observations, we will proceed to examine if their interaction is primarily intra- or extra-cellular. For intracellular interactions, we will co-immunoprecipitate CYR61 with IGF-1 and for extracellular interactions, we will use the AVidity-based EXtracellular Interaction Screen (AVEXIS) system. Our goal is to establish the contribution of CYR61-IGF1 protein-protein interaction to PCa cell proliferation. Specifically, the correlation between PCa aggressiveness and circulating levels of CYR61 and IGF-1 is poorly studied. Their combined role may explain a proportion of aggressive PCa. Citation Format: Greisha L. Ortiz-Hernandez, Mya Walker, Leanne Woods-Burnham, Rick A. Kittles, Carlos A. Casiano, Susan L. Neuhausen. CYR61, a member of the CCN protein family, regulates IGF-1 in metastatic prostate cancer. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3949.

Abstract 484: VIP236: A small molecule drug conjugate with an optimized camptothecin payload has significant activity in patient-derived and metastatic cancer models

Abstract Introduction: We previously presented the design and preliminary characterization of VIP236—an SMDC consisting of an αvβ3 integrin binder linked to an optimized camptothecin topoisomerase I (TOP1) inhibitor payload released by neutrophil elastase in the tumor microenvironment (Lerchen et al, Cancers 2022). Herein we evaluate VIP236 in patient-derived and metastatic models of cancer. Methods: Subcutaneous patient-derived xenograft mouse models in non-small cell lung cancer (NSCLC), renal cell carcinoma (RCC), colorectal cancer (CRC), and triple negative breast cancer (TNBC) were treated with VIP236. Metastasis formation was measured by human DNA PCR in an orthotopic TNBC mouse model. Gastric cancer models included patient-derived and cell line xenograft mouse models. Tumor/control (T/C) ratios were calculated, and body weight was measured. Pharmacodynamic (PD) effect of VIP236 in a gastric cancer SNU16 xenograft mouse model was investigated. Tumor samples from vehicle- and VIP236-treated groups collected up to 144 hours postdose were analyzed for % positive γH2AX cells by a fit-for-purpose qualified immunohistochemistry assay. Results: VIP236 is efficacious in vivo patient-derived models of NSCLC, RCC, CRC and TNBC with T/C ratios of 0.0, 8.9,14.5 and 5.6 compared with vehicle control (p<0.0001, CRC p<0.01). VIP236 showed statistically improved in vivo efficacy in gastric cancer models versus trastuzumab deruxtecan (a commercial antibody-drug conjugate [ADC] with a camptothecin payload) in NCIH87 (HER2high, p<0.05), SNU16 (HER2neg, p<0.001), and GXA3040 (patient-derived, HER2low, p<0.001) xenografts. VIP236 induced statistically significant tumor growth inhibition in a patient-derived CRC liver metastasis model (p<0.01) and statistically significant reduction in lung (p<0.001) and brain (p<0.01) metastasis from an orthotopic TNBC model was observed. Across all in vivo models no clinically significant weight loss or treatment-related mortality were observed. Furthermore, a time-dependent increase of γH2AX % positive cells in SNU16 in vivo VIP236-treated tumors confirms that VIP236 drives robust TOP1 inhibition and subsequent DNA damage. Conclusions: In summary, VIP236 monotherapy efficacy in NSCLC, gastric, TNBC, RCC, CRC, and metastatic TNBC in vivo cancer models provides a rationale for clinical investigation in advanced metastatic solid tumors. In gastric cancer, VIP236 showed improved efficacy compared with the approved ADC, trastuzumab deruxtecan, regardless of HER2 expression. These results warrant further evaluation in clinical trials. Citation Format: Beatrix Stelte-Ludwig, Melanie M. Frigault, Hans-Georg Lerchen, Tibor Schomber, Raquel Izumi, Amy J. Johnson, Ahmed Hamdy. VIP236: A small molecule drug conjugate with an optimized camptothecin payload has significant activity in patient-derived and metastatic cancer models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 484.

3,5,3'-Triiodo-L-Thyronine Regulates Actin Cytoskeleton Dynamic in The Differentiated PC-12 Cells during Hypoxia through An αvβ3 Integrin.

Thyroid hormones are involved in the pathogenesis of various neurological disorders. Ischemia/hypoxia that induces rigidity of the actin filaments, which initiates neurodegeneration and reduces synaptic plasticity. We hypothesized that thyroid hormones via alpha-v-beta-3 (αvβ3) integrin could regulate the actin filament rearrangement during hypoxia and increase neuronal cell viability. In this experimental study, we analysed the dynamics of actin cytoskeleton according to the G/F actin ratio, cofilin-1/p-cofilin-1 ratio, and p-Fyn/Fyn ratio in differentiated PC-12 cells with/without T3 hormone (3,5,3'-triiodo-L-thyronine) treatment and blocking αvβ3-integrin-antibody under hypoxic conditions using electrophoresis and western blotting methods. We assessed NADPH oxidase activity under the hypoxic condition by the luminometric method and Rac1 activity using the ELISA-based (G-LISA) activation assay kit. The T3 hormone induces the αvβ3 integrin-dependent dephosphorylation of the Fyn kinase (P=0.0010), modulates the G/F actin ratio (P=0.0010) and activates the Rac1/NADPH oxidase/cofilin-1 (P=0.0069, P=0.0010, P=0.0045) pathway. T3 increases PC-12 cell viability (P=0.0050) during hypoxia via αvβ3 integrin-dependent downstream regulation systems. The T3 thyroid hormone may modulate the G/F actin ratio via the Rac1 GTPase/NADPH oxidase/ cofilin1signaling pathway and αvβ3-integrin-dependent suppression of Fyn kinase phosphorylation.

Modeling the Interaction of Low Molecular Weight Targeting Ligands and Synthesis of Lipotripeptides with Potential Inhibitory Ability Against Integrin αVβ3

Modeling the Interaction of Low Molecular Weight Targeting Ligands and Synthesis of Lipotripeptides with Potential Inhibitory Ability Against Integrin αVβ3

Dual-Ligand-Functionalized Liposomes Based on Glycyrrhetinic Acid and cRGD for Hepatocellular Carcinoma Targeting and Therapy.

Hepatocellular carcinoma (HCC) is one of the most common cancers, with high mortality. Chemotherapy is one of the main treatment options for HCC. However, the high toxicity and poor specificity of chemotherapeutic drugs have limited their clinical application. In this study, dual-ligand liposomes modified with glycyrrhetinic acid (GA) and cyclic arginine-glycine-aspartic acid (cRGD) (GA/cRGD-LP) were designed to target the GA receptor and αvβ3 integrin, respectively. The aim was to develop a highly selective targeted drug delivery system and further enhance the antitumor efficiency of drugs by targeting both hepatic tumor cells and vasculature. A novel lipid conjugate (mGA-DOPE) by coupling dioleoylphosphatidyl ethanolamine (DOPE) with methyl glycyrrhetinic acid (mGA) was synthesized, and its structure was confirmed. The targeting efficiency of GA/cRGD-LP by in vitro cellular uptake and ex vivo imaging was assessed. GA- and cRGD-modified doxorubicin-loaded liposomes (GA/cRGD-LP-DOX) were prepared, and their cytotoxicity in HepG2 and antitumor activity were evaluated. The results showed that the average particle size of the GA/cRGD-LP-DOX was 114 ± 4.3 nm, and the zeta potential was -32.9 ± 2.0 mV. The transmission electron microscopy images showed that the shapes of our liposomes were spherical. cGA/cRGD-LP-DOX displayed an excellent cellular uptake in both HepG2 and human umbilical vein endothelial cells. In the in vivo study, pharmacokinetic parameters indicated that cGA/cRGD-LP can prolong the circulation time of DOX in the blood. GA/cRGD-LP-DOX showed greater inhibition of tumor growth for HepG2-bearing mice than either the single-ligand-modified liposomes or nontargeted liposomes. GA/cRGD-LP-DOX displayed higher liver tumor localization than that of single-ligand-modified liposomes or free DOX. GA/cRGD-LP is a promising drug delivery system for liver cancer targeting and therapy and is worthy of further study.

Inhibition of Integrin αvβ3-FAK-MAPK signaling constrains the invasion of T-ALL cells

ABSTRACT The role of adhesion receptor integrin αvβ3 in T-ALL was unclear. Firstly, we performed quantitative real-time PCR to assess medullary expression of integrin β3(ITGB3) in T-ALL patients and high ITGB3 expression was relevant with the central nervous system leukemia(CNSL) incidence. Decreasing of cell invasion was observed in Jurkat and Molt4 treated with integrin αvβ3 specific antibody and inhibitor as well as cells with ITGB3 interference. Further, phosphorylation of FAK, cRAF, MEK and ERK decreased in cells with integrin αvβ3 inhibition or interference. Invasion decreased in T-ALL cells treated with FAK and ERK inhibitors. In conclusion, inhibition of integrin αvβ3 signals significantly limits the cell invasion of T-ALL cells.