Single-Cell Analysis of Rohon-Beard Neurons Implicates Fgf Signaling in Axon Maintenance and Cell Survival.

Peripheral sensory neurons are a critical part of the nervous system that transmit a multitude of sensory stimuli to the central nervous system. During larval and juvenile stages in zebrafish, this function is mediated by Rohon-Beard somatosensory neurons (RBs). RBs are optically accessible and amenable to experimental manipulation, making them a powerful system for mechanistic investigation of sensory neurons. Previous studies provided evidence that RBs fall into multiple subclasses; however, the number and molecular makeup of these potential RB subtypes have not been well defined. Using a single-cell RNA sequencing (scRNA-seq) approach, we demonstrate that larval RBs in zebrafish fall into three, largely nonoverlapping classes of neurons. We also show that RBs are molecularly distinct from trigeminal neurons in zebrafish. Cross-species transcriptional analysis indicates that one RB subclass is similar to a mammalian group of A-fiber sensory neurons. Another RB subclass is predicted to sense multiple modalities, including mechanical stimulation and chemical irritants. We leveraged our scRNA-seq data to determine that the fibroblast growth factor (Fgf) pathway is active in RBs. Pharmacological and genetic inhibition of this pathway led to defects in axon maintenance and RB cell death. Moreover, this can be phenocopied by treatment with dovitinib, an FDA-approved Fgf inhibitor with a common side effect of peripheral neuropathy. Importantly, dovitinib-mediated axon loss can be suppressed by loss of Sarm1, a positive regulator of neuronal cell death and axonal injury. This offers a molecular target for future clinical intervention to fight neurotoxic effects of this drug.

New frontiers against sorafenib resistance in renal cell carcinoma: From molecular mechanisms to predictive biomarkers.

Renal cell carcinoma (RCC) is a highly vascularized tumor and prone to distant metastasis. Sorafenib is the first targeted multikinase inhibitor and first-line chemical drug approved for RCC therapy. In fact, only a small number of RCC patients benefit significantly from sorafenib treatment, while the growing prevalence of sorafenib resistance has become a major obstacle for drug therapy effectivity of sorafenib. The molecular mechanisms of sorafenib resistance in RCC are not completely understood by now. Herein, we comprehensively summarize the underlying mechanisms of sorafenib resistance and molecular biomarkers for predicting sorafenib responsiveness. Moreover, we outline strategies suitable for overcoming sorafenib resistance and prospect potential approaches for identifying biomarkers associated with sorafenib resistance in RCC, which contributes to guide individualized and precision drug therapy.

In Vitro Effect of Dovitinib (TKI258), a Multi-Target Angiokinase Inhibitor on Aggressive Meningioma Cells.

Background: Meningiomas represent ∼30% of primary central nervous system (CNS) tumors. Although advances in surgery and radiotherapy have significantly improved survival, there remains an important subset of patients whose tumors have more aggressive behavior and are refractory to conventional therapy. Recent advances in molecular genetics and epigenetics suggest that this aggressive behavior may be due to the deletion of the DNA repair and tumor suppressor gene, CHEK2, neurofibromatosis Type 2 (NF2) mutation on chromosome 22q12, and genetic abnormalities in multiple RTKs including FGFRs. Management of higher-grade meningiomas, such as anaplastic meningiomas (AM: WHO grade III), is truly challenging and there isn't an established chemotherapy option. We investigate the effect of active multi tyrosine receptor kinase inhibitor Dovitinib at stopping AM cell growth in in vitro with either frequent codeletion or mutated CHEK2 and NF2 gene.Methods: Treatment effects were assessed using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay, western blot analysis, caspases assay, and DNA fragmentation assay.Results: Treatment of CH157MN and IOMM-Lee cells with Dovitinib suppressed multiple angiokinases-mainly FGFRs, leading to suppression of downstream signaling by RAS-RAF-MAPK molecules and PI3K-AKT molecules which are involved in cell proliferation, cell survival, and tumor invasion. Furthermore, Dovitinib induced apoptosis via downregulation of survival proteins (Bcl-XL), and over-expression of apoptotic factors (Bax and caspase-3) regardless of CHEK2 and NF2 mutation status.Conclusions: This study establishes the groundwork for the development of Dovitinib as a therapeutic agent for high-grade AM with either frequent codeletion or mutated CHEK2 and NF2, an avenue with high translational potential.

The role of fibroblast growth factor receptor (FGFR) protein-tyrosine kinase inhibitors in the treatment of cancers including those of the urinary bladder.

The human fibroblast growth factor family consists of 22 factors and five transmembrane receptors. Of the 22 factors, eighteen are secreted while four of them function exclusively within the cell. Four of the fibroblast growth factor receptors (FGFRs) possess intracellular protein-tyrosine kinase activity while the fifth (FGFRL1) has a short 105-residue intracellular non-enzymatic component. The FGFR protein kinase domain consists of a bi-lobed structure that is similar to that of all other protein kinases. FGFR gene alterations occur in a wide variety of cancers including those of the urinary bladder, breast, ovary, prostate, endometrium, lung, and stomach. The majority (66 %) of FGFR gene alterations involve gene amplifications, followed by mutations (26 %), and rearrangements that produce fusion proteins (8 %). Erdafitinib was the first orally effective FGFR antagonist approved by the FDA (2019) for the treatment of advanced cancer, that of the urinary bladder. FGF23 suppresses phosphate reabsorption in the proximal tubules of the kidney; FGF23 blockade allows phosphate reabsorption to occur and leads to elevated serum phosphate levels. Erdafitinib and several other, but not all, FGFR antagonists produce hyperphosphatemia. Erdafitinib binds to an inactive DGF-Din conformation of FGFR1 and is classified as a type I½ inhibitor. Similarly, dovitinib, AZD4547, CH5183284, infigratinib, lenvatinib, LY2874455, and lucitanib are type I½ inhibitors. The inactive conformations contain an autoinhibitory brake that is made up of three main residues: an asparagine (N) within the αC-ß4 back loop, a glutamate (E) corresponding to the second hinge residue, and a lysine (K) in the ß8-strand (the NEK triad). PDGFRα/ß, Kit, CSF1R, VEGFR1/2/3, Flt3, Tek, and Tie protein kinases are also regulated by a similar autoinhibitory brake mechanism. Ponatinib binds to FGFR4 in a DFG-Dout conformation and is classified as a type II inhibitor. Futibatinib, roblitinib, H3B-6527, fisogatinib, and PRN1371 bind covalently to their FGFR target and are classified as type VI inhibitors. Nintedanib, pazopanib, pemigatinib, rogaratinib, fisogatinib, and PRN1371 are FGFR inhibitors lacking drug-enzyme crystal structures. All of the aforementioned FGFR antagonists are orally effective. The development of FGFR inhibitors has lagged behind those of other receptor protein-tyrosine kinases. However, the FDA approval of erdafitinib for the treatment of urinary bladder cancers may stimulate additional work targeting the many other FGFR-driven neoplasms.

Anti-Proliferative Effect of Allium senescens L. Extract in Human T-Cell Acute Lymphocytic Leukemia Cells.

Allium species are well known plants distributed throughout the world, and they contain various bioactive components with different biological activities including anti-cancer effects. In this study, we investigated the inhibitory effect of Allium senescens L. (A.S.) extract on cell survival and IL-2-mediated inflammation in human T cell acute lymphocytic leukemia (T-ALL) Jurkat cells. Our results showed that A.S. extract induced caspase-dependent apoptosis of Jurkat cells with no significant cytotoxicity in the normal peripheral blood mononuclear cells. A.S. extract induced ROS generation through the activation of MAPK p38 phosphorylation. It also inhibited IL-2 mRNA expression and NF-κB signaling mediated by phorbol 12-myristate 13-acetate, and phytohemagglutinin. Combined treatment with A.S. extract and axitinib/dovitinib exerted enhanced inhibitory effects on T-ALL cell growth and IL-2 production. These results provide novel information on the potential use of A.S. extract as a therapeutic herbal agent for the treatment and prevention of T-ALL.

Phase II study of Dovitinib in recurrent glioblastoma.

INTRODUCTION: Dovitinib is an oral, potent inhibitor of FGFR and VEGFR, and can be a promising strategy in patients with recurrent or progressive glioblastoma (GBM). METHODS: This was an open label phase II study of two arms: Arm 1 included anti-angiogenic naïve patients with recurrent GBM and Arm 2 included patients with recurrent GBM that had progressed on prior anti-angiogenic therapy. Nineteen subjects were enrolled in Arm 1 and 14 subjects in Arm 2. The primary endpoint was 6-month progression-free survival (PFS-6) in Arm 1 and time to progression (TTP) in Arm 2. The secondary endpoints were toxicity, objective response rate (ORR) and overall survival. RESULTS: Patients in Arm 2 (compared to Arm 1) tended to have longer intervals from diagnosis to study entry (median 26.9 vs. 8.9 months, p = 0.002), experienced more recurrences (64%, had 3-4 prior recurrences compared to 0, p < 0.0001) and tended to be heavily pretreated (71% vs. 26-32% p = 0.04 or 0.02). 6-month PFS was 12% ± 6% for the Arm 1 and 0% for Arm 2. TTP was similar in both treatment arms (median 1.8 months Arm 1 and 0.7-1.8 months Arm 2, p = 0.36). Five patients (15%) had grade 4 toxicities and 22 patients (67%) had grade 3 toxicities. There were no significant differences between the two arms with respect to the amount of change in the levels of biomarkers from baseline. CONCLUSION: Dovitinib was not efficacious in prolonging the PFS in patients with recurrent GBM irrespective of prior treatment with anti-angiogenic therapy (including bevacizumab).

Phase II, randomized, placebo-controlled study of dovitinib in combination with fulvestrant in postmenopausal patients with HR+, HER2- breast cancer that had progressed during or after prior endocrine therapy.

BACKGROUND: Overexpression of fibroblast growth factor receptor 1 (FGFR1), found in ≤8% of hormone receptor-positive (HR+), human epidermal growth factor receptor 2-negative (HER2-) breast cancer cases, is correlated with decreased overall survival and resistance to endocrine therapy (ET). Dovitinib, a potent FGFR inhibitor, has demonstrated antitumor activity in heavily pretreated patients with FGFR pathway-amplified breast cancer. METHODS: In this randomized, placebo-controlled phase II trial, we evaluated whether the addition of dovitinib to fulvestrant would improve outcomes in postmenopausal patients with HR+, HER2- advanced breast cancer that had progressed during or after prior ET. Patients were stratified by FGF pathway amplification and presence of visceral disease, and they were randomized 1:1 to receive fulvestrant plus dovitinib or placebo. The primary endpoint was progression-free survival (PFS). RESULTS: From 15 May 2012 to 26 November 2014, 97 patients from 36 centers were enrolled. The frequency of FGF pathway amplification was lower than anticipated, and the study was terminated early owing to slow accrual of patients with FGF pathway amplification. The median PFS (95% CI) was 5.5 (3.8-14.0) months vs 5.5 (3.5-10.7) months in the dovitinib vs placebo arms, respectively (HR, 0.68; did not meet predefined efficacy criteria). For the FGF pathway-amplified subgroup (n = 31), the median PFS (95% CI) was 10.9 (3.5-16.5) months vs 5.5 (3.5-16.4) months in the dovitinib vs placebo arms, respectively (HR, 0.64; met the predefined superiority criteria). Frequently reported adverse events in the dovitinib (diarrhea, nausea, vomiting, asthenia, and headache) and placebo (diarrhea, fatigue, nausea, and asthenia) arms were mostly low grade. CONCLUSIONS: The safety profile of dovitinib plus fulvestrant was consistent with the known safety profile of single-agent dovitinib. Dovitinib in combination with fulvestrant showed promising clinical activity in the FGF pathway-amplified subgroup. However, the data reported herein should be interpreted with caution, given that fewer PFS events occurred in the FGF pathway-amplified patients than was expected and that an effect of dovitinib regardless of FGR pathway amplification status cannot be excluded, because the population was smaller than expected. TRIAL REGISTRATION: ClinicalTrials.gov identifier: NCT01528345 . Registered 31 January 2012.

Co-clinical trials demonstrate predictive biomarkers for dovitinib, an FGFR inhibitor, in lung squamous cell carcinoma.

BACKGROUND: We conducted co-clinical trials in patient-derived xenograft (PDX) models to identify predictive biomarkers for the multikinase inhibitor dovitinib in lung squamous cell carcinoma (LSCC). METHODS: The PDX01-02 were established from LSCC patients enrolled in the phase II trial of dovitinib (NCT01861197) and PDX03-05 were established from LSCC patients receiving surgery. These five PDX tumors were subjected to in vivo test of dovitinib efficacy, whole exome sequencing and gene expression profiling. RESULTS: The PDX tumors recapitulate histopathological properties and maintain genomic characteristics of originating tumors. Concordant with clinical outcomes of the trial enrolled-LSCC patients, dovitinib produced substantial tumor regression in PDX-01 and PDX-05, whereas it resulted in tumor progression in PDX-02. PDX-03 and -04 also displayed poor antitumor efficacy to dovitinib. Mutational and genome-wide copy number profiles revealed no correlation between genomic alterations of FGFR1-3 and sensitivity to dovitinib. Of note, gene expression profiles revealed differentially expressed genes including FGF3 and FGF19 between PDX-01 and 05 and PDX-02-04. Pathway analysis identified two FGFR signaling-related gene sets, FGFR ligand binding/activation and SHC-mediated cascade pathway were substantially up-regulated in PDX-01 and 05, compared with PDX-02-04. The comparison of gene expression profiles between dovitinib-sensitive versus -resistant lung cancer cell lines in the Cancer Cell Line Encyclopedia database also found that transcriptional activation of 18 key signaling components in FGFR pathways can predict the sensitivity to dovitinib both in cell lines and PDX tumors. These results highlight FGFR pathway activation as a key molecular determinant for sensitivity to dovitinib. CONCLUSIONS: FGFR gene expression signatures are predictors for the response to dovitinib in LSCC.

Efficacy and safety of dovitinib in pretreated patients with advanced squamous non-small cell lung cancer with FGFR1 amplification: A single-arm, phase 2 study.

BACKGROUND: Fibroblast growth factor receptor 1 (FGFR1) amplification is a potential driving oncogene in squamous cell cancer (SCC) of the lung. The current phase 2 study evaluated the efficacy and tolerability of dovitinib, an FGFR inhibitor, in patients with advanced SCC of the lung. METHODS: Patients with pretreated advanced SCC of the lung whose tumors demonstrated FGFR1 amplification of > 5 copies by fluorescence in situ hybridization were enrolled. Dovitinib at a dose of 500 mg was administered orally, once daily, on days 1 to 5 of every week, followed by 2 days off. The primary endpoint was overall response. Secondary endpoints were progression-free survival, overall survival, and toxicity. RESULTS: All 26 patients were men with a median age of 68 years (range, 52-80 years). The majority of patients were ever-smokers. The median duration of dovitinib administration (28 days per cycle) was 2.5 months (range, 0.7-8.6 months). The overall response rate was 11.5% (95% confidence interval [95% CI], 0.8%-23.8%) and the disease control rate was 50% (95% CI, 30.8%-69.2%), with 3 patients achieving partial responses. Response durations for the patients with partial responses were ≥4.5 months, ≥ 5.1 months, and 6.1 months, respectively. After a median follow-up of 15.7 months (range, 1.2-25.6 months), the median overall survival was 5.0 months (95% CI, 3.6-6.4 months) and the median progression-free survival was 2.9 months (95% CI, 1.5-4.3 months). The most common grade 3 or higher adverse events were fatigue (19.2%), anorexia (11.5%), and hyponatremia (11.5%) (event severity was graded based on National Cancer Institute Common Terminology Criteria for Adverse Events [version 4.0]). CONCLUSIONS: Treatment with dovitinib demonstrated modest efficacy in patients with advanced SCC with FGFR1 amplification. Further studies to evaluate other biomarkers correlated with the efficacy of dovitinib in patients with SCC are warranted. Cancer 2016;122:3024-3031. © 2016 American Cancer Society.

Traf2- and Nck-interacting kinase (TNIK) is involved in the anti-cancer mechanism of dovitinib in human multiple myeloma IM-9 cells.

Traf2- and Nck-interacting kinase (TNIK) is a member of the germinal center kinase family. TNIK was first identified as a kinase that is involved in regulating cytoskeletal organization in many types of cells, and it was recently proposed as a novel therapeutic target in several types of human cancers. Although previous studies suggest that TNIK plays a pivotal role in cancer cell survival and prognosis, its function in hematological cancer cell survival has not been investigated. Here we investigated the relationship between TNIK function and cell viability in multiple myeloma IM-9 cells using TNIK small interfering RNA (siRNA) transfection and dovitinib treatment. Treatment of IM-9 cells with TNIK siRNA and dovitinib treatment reduced cell proliferation. The ATP competing kinase assay and western blot analysis showed that dovitinib strongly inhibited both the interaction of TNIK with ATP (K i, 13 nM) and the activation of Wnt signaling effectors such as ß-catenin and TCF4. Dovitinib also induced caspase-dependent apoptosis in IM-9 cells without significant cytotoxicity in PBMCs. Our results provide new evidence that TNIK may be involved in the proliferation of multiple myeloma IM-9 cells and in the anti-cancer activity of dovitinib via inhibition of the endogenous Wnt signaling pathway.

Phase 2 study of dovitinib in patients with metastatic or unresectable adenoid cystic carcinoma.

BACKGROUND: The objective of this study was to evaluate the efficacy and safety of dovitinib in patients with adenoid cystic carcinoma (ACC). METHODS: ACC patients with documented disease progression within the past 12 months were eligible. Patients received oral dovitinib (500 mg once daily for 5 consecutive days followed by a 2-day rest every week) until disease progression or unacceptable toxicities. The primary endpoint was the probability of 4-month progression-free survival (PFS). Metabolic response was evaluated with positron emission tomography (PET)/computed tomography (CT) scans performed at the baseline and after 8 weeks of treatment. RESULTS: Between September 2011 and April 2013, 32 patients with metastatic and/or unresectable ACC were enrolled in this prospective, multicenter trial. The 4-month PFS probability was 80.4%, and the median PFS was 6.0 months (95% confidence interval, 4.4-7.6 months). Tumor shrinkage was observed in 22 patients (68.8%), and 1 patient had a confirmed partial response. The disease control rate was 96.9%. Among 26 patients with PET/CT scans both before and after treatment (at 8 weeks), the metabolic activity of ACC was reduced in 13 patients (50.0%), and 5 patients (19.2%) achieved a metabolic partial response, which was defined as a ≥25% reduction in maximum standardized uptake values. Common grade 3 and 4 adverse events were asthenia (50.0%) and neutropenia (25.0%). CONCLUSIONS: Dovitinib shows modest antitumor activity in the treatment of ACC.

Phase 2 study of dovitinib in patients with relapsed or refractory multiple myeloma with or without t(4;14) translocation.

OBJECTIVES: Approximately 15% of patients with multiple myeloma (MM) exhibit a t(4;14) translocation, which often results in constitutive activation of the receptor tyrosine kinase (RTK) fibroblast growth factor receptor 3 (FGFR3). This study evaluated the efficacy and safety of dovitinib, an RTK inhibitor with in vitro inhibitory activity against FGFR, in patients with relapsed or refractory MM with or without t(4;14) translocation. METHODS: Adult patients with relapsed or refractory MM who had received ≥2 prior regimens were enrolled in this multicenter, 2-stage, phase 2 trial. Patients were grouped based on their t(4;14) status. Dovitinib (500 mg/day orally) was administered on a 5-days-on/2-days-off schedule. The primary endpoint was overall response rate by local investigator review (per International Myeloma Working Group criteria). In non-responding patients, treatment could continue with the addition of low-dose dexamethasone. RESULTS: In total, 43 patients (median age, 63 years) were enrolled (13 t(4;14) positive, 26 t(4;14) negative, and 4 t(4;14) status non-interpretable). Patients had received a median of 5 prior regimens. Median duration of treatment was 8.7 weeks in the t(4;14)-positive group and 3.7 weeks in the t(4;14)-negative group. None of the patients on dovitinib had objective responses. The stable disease rate was 61.5% in the t(4;14)-positive group and 34.6% in the t(4;14)-negative group. Overall, 39 patients (90.7%) had adverse events suspected to be related to study drug, most commonly diarrhea (60.5%), nausea (58.1%), vomiting (46.5%), and fatigue (32.6%). CONCLUSION: Dovitinib showed no single-agent activity in relapsed or refractory MM but may stabilize disease in some t(4;14)-positive patients.

Targeting Signaling Transduction Pathways in Bladder Cancer.

Systemic therapy for urothelial carcinoma (UC) of the bladder has largely revolved around cytotoxic chemotherapy regimens. However, several recent clinical trials have explored the roles of targeted therapies which specifically inhibit signal transduction pathways. Simultaneously, a rationale for such therapies has come to the forefront of management of this disease because an overabundance of signaling pathways are genetically deranged as a result of point mutation or copy number alteration (CNA) as identified by several recent next generation sequencing (NGS) studies. Importantly, these derangements are found in all stages of disease, and therefore targeted therapies hold promise as a next step in the evolution of the medical management of both localized and metastatic UCC. We review the rationale for and progress in studying inhibition of signal transduction as a means of treatment of UCC.

Dovitinib (CHIR258, TKI258): structure, development and preclinical and clinical activity.

Dovitinib is an oral multikinase inhibitor targeting FGF receptors, PDGF receptors and VEGF receptors. Its activity against FGF receptors suggests its usefulness in treating cancers after the failure of VEGF/VEGF receptor-targeting agents. The identified dose and schedule to be used in further studies was 500 mg orally for 5 days on and 2 days off. Biological considerations and the results achieved in a Phase I/II trial suggested its activity in advanced renal cell carcinoma patients pretreated with a tyrosine kinase inhibitor and an mTOR inhibitor. Surprisingly, in a randomized controlled Phase III trial versus sorafenib in the same setting, dovitinib failed to demonstrate any superiority. At present, dovitinib is being tested in different tumor types. However, molecular-based patient selection seems to be key to fully exploit the activity of this drug.

Analysis of serum protein biomarkers, circulating tumor DNA, and dovitinib activity in patients with tyrosine kinase inhibitor-refractory gastrointestinal stromal tumors.

BACKGROUND: An exploratory translational analysis was conducted as part of a phase II study of dovitinib to assess the relevance of soluble serum proteins and circulating tumor (ct) DNA (ctDNA) as biomarkers in patients with tyrosine kinase inhibitor (TKI)-refractory gastrointestinal stromal tumors (GISTs). PATIENTS AND METHODS: Predose serum samples were collected from 30 patients on day 1 of cycle 1 and cycle 2. Serum levels of angiogenesis-related proteins were assessed by enzyme-linked immunosorbent assay, and Beads, emulsions, amplification, and magnetics (BEAMing) assays were carried out to detect mutations in serum ctDNA. RESULTS: Dovitinib increased vascular endothelial growth factor (VEGF)165 (1.26-fold, P = 0.006), VEGF-A (1.27-fold, P = 0.004), placental growth factor (6.0-fold, P = 0.002), fibroblast growth factor 23 (1.45-fold, P = 0.02), and interleukin 8 (1.75-fold, P = 0.04) levels, and decreased soluble vascular endothelial growth factor receptor (sVEGFR)-2 levels (0.8-fold, P = 0.001). The changes in sVEGFR-2 were significantly associated with metabolic response determined by positron emission tomography (P = 0.02) and progression-free survival (PFS; P = 0.02). Secondary kinase mutations were identified in the ctDNA of 11 patients (41%), and these patients all had mutations involving KIT exon 17. Patients with secondary KIT mutations had significantly worse overall survival {median, 5.5 months [95% confidence interval (CI) 3.8-7.2 months]} than those with no detectable secondary mutations [9.8 months (95% CI 9.6-10.0 months); hazard ratio = 2.7 (95% CI 1.0-7.3); P = 0.047]. CONCLUSIONS: Changes in sVEGFR-2 levels were associated with dovitinib-mediated antitumor activity. Genotyping of serum ctDNA with BEAMing is useful for the identification of resistant mutations potentially associated with poor prognosis in patients with GISTs.

Preformulation studies of dovitinib free base: Solubility, lipophilicity and stability.

Dovitinib has been investigated as an anti-tumor drug due to its ability to inhibit multiple receptor tyrosine kinases. Dovitinib free base has a poor water solubility leading to poor absorption. Salts and lipid-based formulations have been used to improve drug availability. Here, we investigated the physiochemical properties of the dovitinib free base in the presence of some pharmaceutical excipients. We sought to study the effect of acidic counterions on the aqueous solubility and lipophilicity of dovitinib and how pH, buffer species, and cyclodextrin (CD) influenced dovitinib stability. pH-solubility studies were performed by titration against five different acids. Aqueous solubility of dovitinib salt depended on the counterion. Lactic acid greatly increased the aqueous solubility of dovitinib. The counterion effect on the solubility was also investigated in the aqueous complexing media. Unexpected synergistic solubilization was found with γ-CD/phosphoric acid and γ-CD/maleic acid. The counterion did not affect the lipophilicity of dovitinib at physiological pH. Accelerated degradation of dovitinib was carried out at high temperature. Stability was studied across a range of pH values, buffer species and in the presence of two CDs. Dovitinib was most stable at pH 4 in the phosphate buffer species. γ-CD stabilized the drug at relatively low pH.

In Vivo Evaluation of Fibroblast Growth Factor Receptor Inhibition in Mouse Xenograft Models of Gastrointestinal Stromal Tumor.

Advanced gastrointestinal stromal tumors (GIST) are typically treated with tyrosine kinase inhibitors, and imatinib is the most commonly used standard of care in first line treatments. The use of this and other tyrosine kinase inhibitors is associated with objective tumor responses and prolongation of progression-free and overall survival, but the treatment of metastatic disease is non-curative due to the selection or acquisition of secondary mutations and the activation of alternative kinase signaling pathways, leading to resistance and disease progression after an initial response. The present preclinical study evaluated the potential use of the fibroblast growth factor receptor inhibitors infigratinib and dovitinib alone or in combination with the mitogen-activated protein kinase inhibitor binimetinib in mouse models of GIST with different sensitivity or resistance to imatinib. Patient- and cell-line-derived GIST xenografts were established by bilateral, subcutaneous transplantation of human GIST tissue in female adult nu/nu NMRI mice. The mice were treated with dovitinib, infigratinib, or binimetinib, either alone or in combination with imatinib. The safety of treated animals was assessed by well-being inspection and body weight measurement. Antitumor effects were assessed by caliper-based tumor measurement. H&E staining and immunohistochemistry were used for assessing anti-mitotic and pro-apoptotic activity of the experimental treatments. Western blotting was used for assessing effects of the agents on kinase signaling pathways. Anti-angiogenic activity was assessed by measuring tumor vessel density. Dovitinib was found to have antitumor efficacy in GIST xenografts characterized by different imatinib resistance patterns. Dovitinib had better efficacy than imatinib (both at standard and increased dose) and was found to be well tolerated. Dovitinib had better efficacy in a KIT exon 9 mutant model, highlighting a role of patient selection in clinical GIST trials with the agent. In a model with KIT exon 11 and 17 mutations, dovitinib induced tumor necrosis, most likely due to anti-angiogenic effects. Additive effects combining dovitinib with binimetinib were limited.

Targeted FGFR/VEGFR/PDGFR inhibition with dovitinib enhances the effects of nab-paclitaxel in preclinical gastric cancer models.

Standard chemotherapy regimens for gastric adenocarcinoma (GAC) have limited efficacy and considerable toxicity profiles. Nab-paclitaxel has shown promising antitumor benefits in previous GAC preclinical studies. Dovitinib inhibits members of the receptor tyrosine kinase family including FGFR, VEGFR and PDGFR, and has exhibited antitumor effects in many solid tumors including GAC. Based on the antimitotic, antistromal and EPR effects of nab-paclitaxel, we investigated augmentation of nab-paclitaxel response by dovitinib in multiple GAC preclinical models. In MKN-45 subcutaneous xenografts, inhibition in tumor growth by nab-paclitaxel and dovitinib was 75% and 76%, respectively. Dovitinib plus nab-paclitaxel had an additive effect on tumor growth inhibition and resulted in tumor regression (85% of its original value). Dovitinib monotherapy resulted in minimal improvement in animal survival (25 days) compared to control (23 days), while nab-paclitaxel monotherapy or dovitinib plus nab-paclitaxel combination therapy led to a clinically significant lifespan extension of 83% (42 days) and 187% (66 days), respectively. IHC analysis of subcutaneous tumors exhibited reduced tumor cell proliferation and tumor vasculature by dovitinib. In vitro studies demonstrated that dovitinib and nab-paclitaxel individually reduced tumor cell proliferation, with an additive effect from combination therapy. Immunoblot analyses of MKN-45 and KATO-III cells revealed that dovitinib decreased phospho-FGFR, phospho-AKT, phospho-ERK, phospho-p70S6K, phospho-4EBP1, Bcl-2 and increased cleaved PARP-1, cleaved-caspase-3, p27, Bax, Bim, with an additive effect from combination therapy. These results demonstrate that the FGFR/VEGFR/PDGFR inhibitor, dovitinib, has the potential to augment the antitumor effects of nab-paclitaxel, with implications for use in the advancement of clinical GAC therapy.

Antitumoral effects of dovitinib in triple-negative breast cancer are synergized by calcitriol in vivo and in vitro.

Chemotherapy is a standard therapeutic option for triple-negative breast cancer (TNBC); however, its effectiveness is often compromised by drug-related toxicity and resistance development. Herein, we aimed to evaluate whether an improved antineoplastic effect could be achieved in vitro and in vivo in TNBC by combining dovitinib, a multi-kinase inhibitor, with calcitriol, a natural anticancer hormone. In vitro, cell proliferation and cell-cycle distribution were studied by sulforhodamine B-assays and flow cytometry. In vivo, dovitinib/calcitriol effects on tumor growth, angiogenesis, and endothelium activation were evaluated in xenografted mice by caliper measures, Itgb3/VEGFR2-immunohistochemistry and 99mTc-Ethylenediamine-N,N-diacetic acid/hydrazinonicotinamyl-Glu[cyclo(Arg-Gly-Asp-D-Phe-Lys)]2 (99mTc-RGD2)-tumor uptake. The drug combination elicited a synergistically improved antiproliferative effect in TNBC-derived cells, which allowed a 7-fold and a 3.3-fold dovitinib dose-reduction in MBCDF-Tum and HCC-1806 cells, respectively. Mechanistically, the co-treatment induced a cell cycle profile suggestive of cell death and DNA damage (accumulation of cells in SubG1, S, and G2/M phases), increased the number of multinucleated cells and inhibited tumor growth to a greater extent than each compound alone. Tumor uptake of 99mTc-RGD2 was reduced by dovitinib, suggesting angiogenesis inhibition, which was corroborated by decreased endothelial cell growth, tumor-vessel density and VEGFR2 expression. In summary, calcitriol synergized dovitinib anticancer effects in vitro and in vivo, allowing for a significant dose-reduction of dovitinib while maintaining its antiproliferative potency. Our results suggest the beneficial convergence of independent antitumor mechanisms of dovitinib and calcitriol to inhibit TNBC-tumor growth.

Effects of FGFR Tyrosine Kinase Inhibition in OLN-93 Oligodendrocytes.

Fibroblast growth factor (FGF) signaling is involved in the pathogenesis of multiple sclerosis (MS). Data from neuropathology studies suggest that FGF signaling contributes to the failure of remyelination in MS. In MOG35-55-induced EAE, oligodendrocyte-specific deletion of FGFR1 and FGFR2 resulted in a less severe disease course, reduced inflammation, myelin and axon degeneration and changed FGF/FGFR and BDNF/TrkB signaling. Since signaling cascades in oligodendrocytes could not be investigated in the EAE studies, we here aimed to characterize FGFR-dependent oligodendrocyte-specific signaling in vitro. FGFR inhibition was achieved by application of the multi-kinase-inhibitor dovitinib and the FGFR1/2/3-inhibitor AZD4547. Both substances are potent inhibitors of FGF signaling; they are effective in experimental tumor models and patients with malignancies. Effects of FGFR inhibition in oligodendrocytes were studied by immunofluorescence microscopy, protein and gene analyses. Application of the tyrosine kinase inhibitors reduced FGFR1, phosphorylated ERK and Akt expression, and it enhanced BDNF and TrkB expression. Furthermore, the myelin proteins CNPase and PLP were upregulated by FGFR inhibition. In summary, inhibition of FGFR signaling in oligodendrocytes can be achieved by application of tyrosine kinase inhibitors. Decreased phosphorylation of ERK and Akt is associated with an upregulation of BDNF/TrkB signaling, which may be responsible for the increased production of myelin proteins. Furthermore, these data suggest that application of FGFR inhibitors may have the potential to promote remyelination in the CNS.