Progress in the development of modulators targeting Frizzleds.

The Frizzleds (FZDs) receptors on the cell surface belong to the class F of G protein-coupled receptors (GPCRs) which are the major receptors of WNT protein that mediates the classical WNT signaling pathway and other non-classical pathways. Besides, the FZDs also play a core role in tissue regeneration and tumor occurrence. With the structure and mechanism of FZDs activation becoming clearer, a series of FZDs modulators (inhibitors and agonists) have been developed, with the hope of bringing benefits to the treatment of cancer and degenerative diseases. Most of the FZDs inhibitors (small molecules, antibodies or designed protein inhibitors) block WNT signaling through binding to the cysteine-rich domain (CRD) of FZDs. Several small molecules impede FZDs activation by targeting to the third intracellular domain or the transmembrane domain of FZDs. However, three small molecules (FZM1.8, SAG1.3 and purmorphamine) activate the FZDs through direct interaction with the transmembrane domain. Another type of FZDs agonists are bivalent or tetravalent antibodies which activate the WNT signaling via inducing FZD-LRP5/6 heterodimerization. In this article, we reviewed the FZDs modulators reported in recent years, summarized the critical molecules' discovery processes and the elucidated relevant structural and pharmacological mechanisms. We believe the summaried molecular mechanisms of the relevant modulators could provide important guidance and reference for the future development of FZD modulators.

Frizzled receptors (FZDs) in Wnt signaling: potential therapeutic targets for human cancers.

Frizzled receptors (FZDs) are key contributors intrinsic to the Wnt signaling pathway, activation of FZDs triggering the Wnt signaling cascade is frequently observed in human tumors and intimately associated with an aggressive carcinoma phenotype. It has been shown that the abnormal expression of FZD receptors contributes to the manifestation of malignant characteristics in human tumors such as enhanced cell proliferation, metastasis, chemotherapy resistance as well as the acquisition of cancer stemness. Given the essential roles of FZD receptors in the Wnt signaling in human tumors, this review aims to consolidate the prevailing knowledge on the specific status of FZD receptors (FZD1-10) and elucidate their respective functions in tumor progression. Furthermore, we delineate the structural basis for binding of FZD and its co-receptors to Wnt, and provide a better theoretical foundation for subsequent studies on related mechanisms. Finally, we describe the existing biological classes of small molecule-based FZD inhibitors in detail in the hope that they can provide useful assistance for design and development of novel drug candidates targeted FZDs.

FZD10-targeted α-radioimmunotherapy with 225 Ac-labeled OTSA101 achieves complete remission in a synovial sarcoma model.

Synovial sarcomas are rare tumors arising in adolescents and young adults. The prognosis for advanced disease is poor, with an overall survival of 12-18 months. Frizzled homolog 10 (FZD10) is overexpressed in most synovial sarcomas, making it a promising therapeutic target. The results of a phase 1 trial of ß-radioimmunotherapy (RIT) with the 90 Y-labeled anti-FZD10 antibody OTSA101 revealed a need for improved efficacy. The present study evaluated the potential of α-RIT with OTSA101 labeled with the α-emitter 225 Ac. Competitive inhibition and cell binding assays showed that specific binding of 225 Ac-labeled OTSA101 to SYO-1 synovial sarcoma cells was comparable to that of the imaging agent 111 In-labeled OTSA101. Biodistribution studies showed high uptake in SYO-1 tumors and low uptake in normal organs, except for blood. Dosimetric studies showed that the biologically effective dose (BED) of 225 Ac-labeled OTSA101 for tumors was 7.8 Bd higher than that of 90 Y-labeled OTSA101. 90 Y- and 225 Ac-labeled OTSA101 decreased tumor volume and prolonged survival. 225 Ac-labeled OTSA101 achieved a complete response in 60% of mice, and no recurrence was observed. 225 Ac-labeled OTSA101 induced a larger amount of necrosis and apoptosis than 90 Y-labeled OTSA101, although the cell proliferation decrease was comparable. The BED for normal organs and tissues was tolerable; no treatment-related mortality or obvious toxicity, except for temporary body weight loss, was observed. 225 Ac-labeled OTSA101 provided a high BED for tumors and achieved a 60% complete response in the synovial sarcoma mouse model SYO-1. RIT with 225 Ac-labeled OTSA101 is a promising therapeutic option for synovial sarcoma.

MiR-340 regulates the growth and metabolism of renal cell carcinoma cells by targeting frizzled class receptor 3.

MicroRNA(miR)-340 is known as a multifunctional miRNA related to various types of cancer while its role in renal cell carcinoma (RCC) remains to be further investigated. In the present study, an apparent increase in miR-340 expression was observed in both clear cell RCC tissues and RCC cell line 786-O and Caki-1. Functionally, the overexpression of miR-340 promoted cell proliferation, migration, invasion, extracellular alanine (Ala) level, and glycolysis level in 786-O cells. Then, frizzled class receptor 3 (FZD3) was determined as the target gene of miR-340 and its expression level was negatively regulated by miR-340. The FZD3 silencing abrogated the inhibitory effect of miR-340 knockdown on cell proliferation, migration, invasion, Ala level, and glycolysis level in 786-O cells. In conclusion, miR-340 promotes proliferation, migration, and invasion of RCC cells via suppressing FZD3 expression, and the promotion effect of miR-340 on RCC progression may be due to its regulatory effect on glycolysis and Ala level.

The flavonoid luteolin suppresses infantile hemangioma by targeting FZD6 in the Wnt pathway.

Infantile hemangioma is the most common vascular tumor of childhood. It is characterized by clinical expansion of endothelial cells and promoted by angiogenic factors. Luteolin is a flavonoid compound that carries anti-cancer and anti-angiogenesis properties. The study aimed to investigate the effect of luteolin in treating infantile hemangioma. We first tested the effect of luteolin on cell proliferative potential and VEGFA expression in hemangioma-derived stem cells (HemSCs). We then examined the efficacy of systemic application of luteolin in a murine hemangioma model. We then identified the downstream factor regulated by luteolin in HemSCs and validated its causative relationship with knock-down method in both in vitro and in vivo models. We also investigated the protein expression change of this targeting factor in proliferating hemangiomas. Luteolin inhibited HemSC growth and suppressed VEGF-A expression in a dose-dependent manner. Luteolin inhibited microvessel formation and de novo vasculogenesis in the murine model. FZD6 was induced by luteolin and exerted the anti-angiogenesis effect in our tumor models. Lastly, FZD6 level was repressed in the clinical tissues of human proliferating hemangiomas. Luteolin is a promising new agent to treat infantile hemangioma. Targeting the Wnt pathway may represent a potential therapeutic strategic to inhibit angiogenesis in proliferating hemangiomas.

Inhibition of Frizzled-2 by small interfering RNA protects rat hepatic BRL-3A cells against cytotoxicity and apoptosis induced by Hypoxia/Reoxygenation / La inhibición de Frizzled-2 por ARN interferente pequeño protege a los hepatocitos BRL-3A de rata frente a la citotoxicidad y la apoptosis inducidas por hipoxia/reoxigenación

Frizzled-2 plays an important role in maintaining normal hepatic cell functionality. This study aimed to investigate the role of inhibition of Frizzled-2 in protecting rat liver BRL-3A cells from Hypoxia/Reoxygenation (H/R). In vitro H/R hepatic cell model was established by culturing BRL-3A cells under H/R condition. Frizzled-2 siRNA was transfected into BRL-3A cells to inhibit Frizzled-2 signaling. Wnt5a and Frizzled-2 were significantly increased in BRL-3A cells upon H/R treatment. H/R treatment induced cell cytotoxicity, the early apoptosis rate and the intracellular Ca2+ level in BRL-3A cells while silencing frizzled-2 gene decreased the H/R induced cell cytotoxicity, apoptosis and intracellular Ca2+ level. In vivo mice study further showed the up-regulation of Frizzled-2/Wnt 5 pathway and cleaved Caspase-3 expression in liver tissues under ischemia and reperfusion injury (IRI). In summary, inhibition of Frizzled-2 by its siRNA may protects BRL-3A cells by attenuating the H/R induced cell cytotoxicity and apoptosis

Frizzled-2 desempeña un papel importante en el mantenimiento de la funcionalidad normal de los hepatocitos. Este estudio tiene como objetivo analizar el papel de la inhibición de Frizzled-2 en la protección de los hepatocitos BRL-3A de rata de la hipoxia/reoxigenación (H/R). El modelo de hepatocitos H/R in vitro se demostró con el cultivo de células BRL-3A en condiciones de H/R. El ARNip de Frizzled-2 se transinfectó en células BRL-3A para inhibir la señalización de Frizzled-2. Wnt5a y Frizzled-2 aumentaron considerablemente en las células BRL-3A tras el tratamiento con H/R. El tratamiento con H/R provocó citotoxicidad celular, una tasa de apoptosis temprana y el nivel de Ca2+ intracelular en células BRL-3A mientras que el gen frizzled-2 silenciado redujo la citotoxicidad celular inducida por H/R, la apoptosis y el nivel de Ca2+ intracelular. El estudio in vivo con ratones mostró, además, la regulación al alza de la vía de Frizzled-2/Wnt 5 y la expresión de caspasa 3 escindida en tejidos hepáticos con lesión por isquemia y reperfusión (LIR). En resumen, la inhibición de Frizzled-2 por su ARNip puede proteger a las células BRL-3A al atenuar la citotoxicidad celular y la apoptosis inducida por H/R

In Silico Identification of Potential Inhibitors of the Wnt Signaling Pathway in Human Breast Cancer.

Triple-negative breast cancer is the leading worldwide cause of cancer-related deaths in women. The prospection and development of new substances with antitumoral potential is of great importance for the treatment of this disease. The objective of this work was to identify a commercial drug or ligand that could potentially bind to the FZD7 transmembrane protein and inactivate the Wnt signaling pathway in triple-negative breast cancer cells. We aimed at computationally modeling the FZD7, Wnt3, and Wnt3a proteins, at making them available in protein model databases, and at conducting docking analysis to assess the binding free energy between FZD7 and the selected ligands. The Wnt3 and Wnt3a proteins were modeled by homology modeling, and the FZD7 protein was modeled by homology modeling and ab initio modeling. The ligands were selected based on their similarity to the palmitoleic acid and were gathered from the ZINC database. A total of 30 commercially available ligands were found in the ZINC database. The docking results show that the ligands zinc08221009, zinc13546050, zinc05260769, zinc04529321, and zinc05972969 are good candidates for novel drug development. The created models and conducted analysis by this work will most certainly help in future research on the Wnt signaling pathway and its components.

Lead Optimization Yields High Affinity Frizzled 7-Targeting Peptides That Modulate Clostridium difficile Toxin B Pathogenicity in Epithelial Cells.

Frizzled 7 (FZD7) receptors have been shown to play a central role in intestinal stem cell regeneration and, more recently, in Clostridium difficile pathogenesis. Yet, targeting FZD7 receptors with small ligands has not been explored as an approach to block C. difficile pathogenesis. Here, we report the discovery of high affinity peptides that selectively bind to FZD7 receptors. We describe an integrated approach for lead optimization, utilizing structure-based rational design and directed evolution, to enhance the peptide binding affinity while still maintaining FZD7 receptor selectivity. This work yielded new peptide leads with picomolar binding constants to FZD7 as measured by biophysical methods. The new peptides block the interaction between C. difficile toxin B (TcdB) and FZD receptors and perturb C. difficile pathogenesis in epithelial cells. As such, our findings provide a proof of concept that targeting FZD receptors could be a viable pharmacological approach to protect epithelial cells from TcdB pathogenicity.

Wnt Signaling in Cancer: Not a Binary ON:OFF Switch.

In the March 1 issue of Cancer Research, we identified the Wnt receptor Fzd7 as an attractive therapeutic target for the treatment of gastric cancer. In summary, we showed that pharmacological inhibition of Wnt receptors, or genetic deletion of Fzd7, blocks the initiation and growth of gastric tumors. Inhibiting Fzd receptors, specifically Fzd7, inhibits the growth of gastric cancer cells even in the presence of adenomatous polyposis coli (Apc) mutation. Apc is located in the cytoplasm downstream of Fzd7 in the Wnt signaling cascade and APC mutations activate Wnt/ß-catenin signaling, therefore, this result seems counterintuitive. Here, we analyze this result in greater detail in the context of current knowledge of Wnt signaling and discuss the wider implications of this aspect of Wnt signaling in other cancers.

Frizzled-7-targeted delivery of zinc oxide nanoparticles to drug-resistant breast cancer cells.

There is a need for novel strategies to treat aggressive breast cancer subtypes and overcome drug resistance. ZnO nanoparticles (NPs) have potential in cancer therapy due to their ability to potently and selectively induce cancer cell apoptosis. Here, we tested the in vitro chemotherapeutic efficacy of ZnONPs loaded via a mesoporous silica nanolayer (MSN) towards drug-sensitive breast cancer cells (MCF-7: estrogen receptor-positive, CAL51: triple-negative) and their drug-resistant counterparts (MCF-7TX, CALDOX). ZnO-MSNs were coated on to gold nanostars (AuNSs) for future imaging capabilities in the NIR-II range. Electron and confocal microscopy showed that MSN-ZnO-AuNSs accumulated close to the plasma membrane and were internalized by cells. High-resolution electron microscopy showed that MSN coating degraded outside the cells, releasing ZnONPs that interacted with cell membranes. MSN-ZnO-AuNSs efficiently reduced the viability of all cell lines, and CAL51/CALDOX cells were more susceptible than MCF7/MCF-7-TX cells. MSN-ZnO-AuNSs were then conjugated with the antibody to Frizzled-7 (FZD-7), the receptor upregulated by several breast cancer cells. We used the disulphide (S-S) linker that could be cleaved with a high concentration of glutathione normally observed within cancer cells, releasing Zn2+ into the cytoplasm. FZD-7 targeting resulted in approximately three-fold amplified toxicity of MSN-ZnO-AuNSs towards the MCF-7TX drug-resistant cell line with the highest FZD-7 expression. This study shows that ZnO-MSs are promising tools to treat triple-negative and drug-resistant breast cancers and highlights the potential clinical utility of FZD-7 for delivery of nanomedicines and imaging probes specifically to these cancer types.

Receptor subtype discrimination using extensive shape complementary designed interfaces.

To discriminate between closely related members of a protein family that differ at a limited number of spatially distant positions is a challenge for drug discovery. We describe a combined computational design and experimental selection approach for generating binders targeting functional sites with large, shape complementary interfaces to read out subtle sequence differences for subtype-specific antagonism. Repeat proteins are computationally docked against a functionally relevant region of the target protein surface that varies in the different subtypes, and the interface sequences are optimized for affinity and specificity first computationally and then experimentally. We used this approach to generate a series of human Frizzled (Fz) subtype-selective antagonists with extensive shape complementary interaction surfaces considerably larger than those of repeat proteins selected from random libraries. In vivo administration revealed that Wnt-dependent pericentral liver gene expression involves multiple Fz subtypes, while maintenance of the intestinal crypt stem cell compartment involves only a limited subset.

Structure-guided design fine-tunes pharmacokinetics, tolerability, and antitumor profile of multispecific frizzled antibodies.

Aberrant activation of Wnt/ß-catenin signaling occurs frequently in cancer. However, therapeutic targeting of this pathway is complicated by the role of Wnt in stem cell maintenance and tissue homeostasis. Here, we evaluated antibodies blocking 6 of the 10 human Wnt/Frizzled (FZD) receptors as potential therapeutics. Crystal structures revealed a common binding site for these monoclonal antibodies (mAbs) on FZD, blocking the interaction with the Wnt palmitoleic acid moiety. However, these mAbs displayed gastrointestinal toxicity or poor plasma exposure in vivo. Structure-guided engineering was used to refine the binding of each mAb for FZD receptors, resulting in antibody variants with improved in vivo tolerability and developability. Importantly, the lead variant mAb significantly inhibited tumor growth in the HPAF-II pancreatic tumor xenograft model. Taken together, our data demonstrate that anti-FZD cancer therapeutic antibodies with broad specificity can be fine-tuned to navigate in vivo exposure and tolerability while driving therapeutic efficacy.

Frizzled-7 Is Required for Wnt Signaling in Gastric Tumors with and Without Apc Mutations.

A subset of patients with gastric cancer have mutations in genes that participate in or regulate Wnt signaling at the level of ligand (Wnt) receptor (Fzd) binding. Moreover, increased Fzd expression is associated with poor clinical outcome. Despite these findings, there are no in vivo studies investigating the potential of targeting Wnt receptors for treating gastric cancer, and the specific Wnt receptor transmitting oncogenic Wnt signaling in gastric cancer is unknown. Here, we use inhibitors of Wnt/Fzd (OMP-18R5/vantictumab) and conditional gene deletion to test the therapeutic potential of targeting Wnt signaling in preclinical models of intestinal-type gastric cancer and ex vivo organoid cultures. Pharmacologic targeting of Fzd inhibited the growth of gastric adenomas in vivo. We identified Fzd7 to be the predominant Wnt receptor responsible for transmitting Wnt signaling in human gastric cancer cells and mouse models of gastric cancer, whereby Fzd7-deficient cells were retained in gastric adenomas but were unable to respond to Wnt signals and consequently failed to proliferate. Genetic deletion of Fzd7 or treatment with vantictumab was sufficient to inhibit the growth of gastric adenomas with or without mutations to Apc. Vantictumab is currently in phase Ib clinical trials for advanced pancreatic, lung, and breast cancer. Our data extend the scope of patients that may benefit from this therapeutic approach as we demonstrate that this drug will be effective in treating patients with gastric cancer regardless of APC mutation status. SIGNIFICANCE: The Wnt receptor Fzd7 plays an essential role in gastric tumorigenesis irrespective of Apc mutation status, therefore targeting Wnt/Fzd7 may be of therapeutic benefit to patients with gastric cancer.

A synthetic anti-Frizzled antibody engineered for broadened specificity exhibits enhanced anti-tumor properties.

Secreted Wnt ligands play a major role in the development and progression of many cancers by modulating signaling through cell-surface Frizzled receptors (FZDs). In order to achieve maximal effect on Wnt signaling by targeting the cell surface, we developed a synthetic antibody targeting six of the 10 human FZDs. We first identified an anti-FZD antagonist antibody (F2) with a specificity profile matching that of OMP-18R5, a monoclonal antibody that inhibits growth of many cancers by targeting FZD7, FZD1, FZD2, FZD5 and FZD8. We then used combinatorial antibody engineering by phage display to develop a variant antibody F2.A with specificity broadened to include FZD4. We confirmed that F2.A blocked binding of Wnt ligands, but not binding of Norrin, a ligand that also activates FZD4. Importantly, F2.A proved to be much more efficacious than either OMP-18R5 or F2 in inhibiting the growth of multiple RNF43-mutant pancreatic ductal adenocarcinoma cell lines, including patient-derived cells.

Wnt5a/FZD4 Mediates the Mechanical Stretch-Induced Osteogenic Differentiation of Bone Mesenchymal Stem Cells.

BACKGROUND/AIMS: Mechanical stimulation and WNT signalling have essential roles in regulating the osteogenic differentiation of bone marrow stromal cells (BMSCs) and bone formation. However, little is known regarding the regulation of WNT signalling molecule expression and therefore the osteogenic differentiation of BMSCs during osteogenesis. METHODS: Microarrays of BMSCs from elderly individuals or patients with osteoporosis (GSE35959) from the GEO database were analysed using GeneSight-Lite 4.1.6 (BioDiscovery) and C2 curated gene sets downloaded from Molecular Signatures Database (MSigDB). Realtime PCR and western blotting were used to measure the expression of the indicated genes. ALP and Alizarin red staining were used to evaluate the osteogenesis of BMSCs. RESULTS: In this study, we investigated whether mechanical loading directly regulates the expression of WNT signalling molecules and examined the role of WNT signalling in mechanical loading-triggered osteogenic differentiation and bone formation. We first studied the microarrays of samples from patients with osteoporosis and found downregulation of the GPCR ligand binding gene set in the BMSCs of patients with osteoporosis. Then, we demonstrated that mechanical stimuli can regulate osteogenesis and bone formation both in vivo and in vitro. FZD4 was upregulated during cyclic mechanical stretch (CMS)-induced osteogenic differentiation, and the JNK signalling pathway was activated. FZD4 knockdown inhibited the mechanical stimuli-induced osteogenesis and JNK activity. More importantly, we found an activating effect of WNT5A and FZD4 that regulated bone formation in response to hindlimb unloading in mice, and pretreatment with WNT5A or activation of the expression of FZD4 partly rescued the osteoporosis caused by mechanical unloading. CONCLUSIONS: Our results demonstrate, for the first time, that mechanical stimulation alters the expression of genes involved in the osteogenic differentiation of BMSCs via the direct regulation of FZD4 and that therapeutic WNT5A and FZD saRNA may be an efficient strategy for enhancing bone formation under mechanical stimulation.

hsa_circ_0000177-miR-638-FZD7-Wnt Signaling Cascade Contributes to the Malignant Behaviors in Glioma.

As a new member of the noncoding RNA family, circular RNAs (circRNAs) have been demonstrated as critical regulators in various physiological and pathological processes, such as tumorigenesis. However, the role of circRNAs has not been well understood until now. In our study, we found that circRNA hsa_circ_0000177 was upregulated in glioma tissues and cell lines. Also, hsa_circ_0000177 overexpression was associated with poor prognosis in glioma patients. Through functional experiments, we found that hsa_circ_0000177 knockdown dramatically inhibited glioma cell proliferation and invasion in vitro. Consistently, hsa_circ_0000177 knockdown significantly repressed glioma growth in vivo. In terms of mechanism, we used bioinformatics analysis and identified hsa_circ_0000177 as a miR-638 sponge. We showed that miR-638 inhibition could restore the proliferation and invasion of glioma cells transfected with hsa_circ_0000177 small interfering RNA. Furthermore, we demonstrated that frizzled class receptor 7 (FZD7) was targeted by miR-638 and upregulated by hsa_circ_0000177. Through upregulating FZD7 expression, hsa_circ_0000177 activated Wnt signaling and facilitated glioma growth. Taken together, our study revealed a novel signaling pathway involved in glioma progression.

α-particle therapy for synovial sarcoma in the mouse using an astatine-211-labeled antibody against frizzled homolog 10.

Synovial sarcoma (SS) is a rare yet refractory soft-tissue sarcoma that predominantly affects young adults. We show in a mouse model that radioimmunotherapy (RIT) with an α-particle emitting anti-Frizzled homolog 10 (FZD10) antibody, synthesized using the α-emitter radionuclide astatine-211 (211 At-OTSA101), suppresses the growth of SS xenografts more efficiently than the corresponding ß-particle emitting anti-FZD10 antibody conjugated with the ß-emitter yettrium-90 (90 Y-OTSA101). In biodistribution analysis, 211 At was increased in the SS xenografts but decreased in other tissues up to 1 day after injection as time proceeded, albeit with a relatively higher uptake in the stomach. Single 211 At-OTSA101 doses of 25 and 50 µCi significantly suppressed SS tumor growth in vivo, whereas a 50-µCi dose of 90 Y-OTSA101 was needed to achieve this. Importantly, 50 µCi of 211 At-OTSA101 suppressed tumor growth immediately after injection, whereas this effect required several days in the case of 90 Y-OTSA101. Both radiolabeled antibodies at the 50-µCi dosage level significantly prolonged survival. Histopathologically, severe cellular damage accompanied by massive cell death was evident in the SS xenografts at even 1 day after the 211 At-OTSA101 injection, but these effects were relatively milder with 90 Y-OTSA101 at the same timepoint, even though the absorbed doses were comparable (3.3 and 3.0 Gy, respectively). We conclude that α-particle RIT with 211 At-OTSA101 is a potential new therapeutic option for SS.

A first-in-human study investigating biodistribution, safety and recommended dose of a new radiolabeled MAb targeting FZD10 in metastatic synovial sarcoma patients.

BACKGROUND: Synovial Sarcomas (SS) are rare tumors occurring predominantly in adolescent and young adults with a dismal prognosis in advanced phases. We report a first-in-human phase I of monoclonal antibody (OTSA-101) targeting FZD10, overexpressed in most SS but not present in normal tissues, labelled with radioisotopes and used as a molecular vehicle to specifically deliver radiation to FZD10 expressing SS lesions. METHODS: Patients with progressive advanced SS were included. In the first step of this trial, OTSA-101 in vivo bio-distribution and lesions uptake were evaluated by repeated whole body planar and SPECT-CT scintigraphies from H1 till H144 after IV injection of 187 MBq of 111In-OTSA-101. A 2D dosimetry study also evaluated the liver absorbed dose when using 90Y-OTSA-101. In the second step, those patients with significant tumor uptake were randomized between 370 MBq (Arm A) and 1110 MBq (Arm B) of 90Y-OTSA-101 for radionuclide therapy. RESULTS: From January 2012 to June 2015, 20 pts. (median age 43 years [21-67]) with advanced SS were enrolled. Even though 111In-OTSA-101 liver uptake appeared to be intense, estimated absorbed liver dose was less than 20 Gy for each patient. Tracer intensity was greater than mediastinum in 10 patients consistent with sufficient tumor uptake to proceed to treatment with 90Y-OTSA-101: 8 were randomized (Arm A: 3 patients and Arm B: 5 patients) and 2 were not randomized due to worsening PS. The most common Grade ≥ 3 AEs were reversible hematological disorders, which were more frequent in Arm B. No objective response was observed. Best response was stable disease in 3/8 patients lasting up to 21 weeks for 1 patient. CONCLUSIONS: Radioimmunotherapy targeting FZD10 is feasible in SS patients as all patients presented at least one lesion with 111In-OTSA-101 uptake. Tumor uptake was heterogeneous but sufficient to select 50% of pts. for 90Y-OTSA-101 treatment. The recommended activity for further clinical investigations is 1110 MBq of 90Y-OTSA-101. However, because of hematological toxicity, less energetic particle emitter radioisopotes such as Lutetium 177 may be a better option to wider the therapeutic index. TRIAL REGISTRATION: The study was registered on the NCT01469975 website with a registration code NCT01469975 on November the third, 2011.

Reversal effect of quercetin on multidrug resistance via FZD7/ß-catenin pathway in hepatocellular carcinoma cells.

BACKGROUND: Chemotherapy has been widely used to treat cancer, but the appearance of multidrug resistance (MDR) is the biggest obstacle to successful chemotherapy. One of the conventional mechanisms of MDR is overexpression of ATP-binding cassette (ABC) transporters such as P-glycoprotein (P-gp/ABCB1) and multidrug resistance-associated proteins (MRPs/ABCCs) that limits the prolonged and efficient use of chemotherapeutic drugs. To enhance the chemosensitivity of tumor cells, attentions have been focused on effective MDR modulators. PURPOSE: This study aimed to investigate the reversal effect of quercetin on MDR, and explored its mechanism of action in vitro. STUDY DESIGN/METHODS: The effect and mechanism of quercetin on MDR was examined by using MTT assay, flow cytometry, real-time PCR and western blot analysis in human hepatocellular carcinoma cells. RESULTS: Our data found that the intracellular accumulation of rhodamine-123 (Rh123) and doxorubicin (ADR) were increased, the sensitivity of BEL/5-FU cells to chemotherapeutic drugs were increased, and the expressions of ABCB1, ABCC1 and ABCC2 were all down-regulated, which indicated that the functions and expressions of ABCB1, ABCC1 and ABCC2 efflux pump were inhibited by quercetin treatment. Moreover, the suppression of ABCB1, ABCC1 and ABCC2 by quercetin was dependent on the FZD7 through the Wnt/ß-catenin pathway. Further research revealed that reduction of FZD7 by RNA interference (siFZD7) enhanced the sensitivity to chemotherapeutic drugs, increased the cellular accumulation of Rh123 and ADR, and induced inhibitory effects on the expression of FZD7, ABCB1, ABCC1, ABCC2 and ß-catenin, similar to quercetin. In the meanwhile, overexpression of FZD7 showed the inversely effect on the expressions. Interesting, it was confirmed that quercetin could inhibit the expression levels of FZD7, ABCB1, ABCC1, ABCC2 and ß-catenin in BEL-7402 cells; furthermore, treatment by quercetin combined with siFZD7 in BEL/5-FU cells, the expressions of these genes were effectively decreased in comparison to quercetin combined with siRNA negative control (sncRNA). CONCLUSION: Overall, these data suggested the effectiveness of using quercetin, at least in part, via inhibiting FZD7 to combat chemoresistance and showed that quercetin could be developed into an efficient natural sensitizer for resistant human hepatocellular carcinoma.

Frizzled Receptors as Potential Therapeutic Targets in Human Cancers.

Frizzled receptors (FZDs) are a family of seven-span transmembrane receptors with hallmarks of G protein-coupled receptors (GPCRs) that serve as receptors for secreted Wingless-type (WNT) ligands in the WNT signaling pathway. Functionally, FZDs play crucial roles in regulating cell polarity, embryonic development, cell proliferation, formation of neural synapses, and many other processes in developing and adult organisms. In this review, we will introduce the basic structural features and review the biological function and mechanism of FZDs in the progression of human cancers, followed by an analysis of clinical relevance and therapeutic potential of FZDs. We will focus on the development of antibody-based and small molecule inhibitor-based therapeutic strategies by targeting FZDs for human cancers.