Roles of non-receptor tyrosine kinases in pathogenesis and treatment of depression.

Major depressive disorder is a chronic psychiatric disease with a high prevalence. Brain mechanisms for depression at cellular and molecular levels are far from clear. Increasing evidence from clinical and preclinical studies reveals critical roles of the non-receptor tyrosine kinase (nRTK) superfamily in the pathophysiology, symptomatology, and therapy of depression. To date, several nRTK members from three nRTK subfamilies, i.e., the Src family kinase (SFK), the Janus tyrosine kinase (JAK) and the focal adhesion kinase (FAK) subfamilies, may connect to the intracellular, intranuclear, and synaptic signaling network linking chronic stress to depression- and anxiety-like behavior. These SFK/JAK/FAK nRTKs are abundantly expressed in the prefrontal cortex and hippocampus, two core limbic regions implicated in depression, and are enriched at synaptic sites. In various acute or chronic animal models of depression, the nRTKs were significantly altered (up- or downregulated) in their phosphorylation, expression, subcellular/subsynaptic distribution, and/or function. Stress that precipitates depressive behavior also influenced the interaction of nRTKs with other signaling molecules and downstream substrates, including ionotropic and metabotropic glutamate receptors. The commonly-used antidepressants showed the ability to alter nRTK activity. In sum, the limbic SFK/JAK/FAK nRTKs are sensitive to stress and undergo drastic adaptations in response to chronic depression. These long-lasting adaptations contribute to the remodeling of signaling network or synaptic plasticity critical for the vulnerability to depression and the therapeutic efficacy of antidepressants.

Pyrrole as an Important Scaffold of Anticancer Drugs: Recent Advances.

With the significant increase of patients suffering from different types of cancer, it is evident that prompt measures in the development of novel and effective agents need to be taken. Pyrrole moiety has been found in various active compounds with anti-inflammatory, antiseptic, antibacterial, lipid-lowering and anticancer properties. Recent advances in the exploration of highly active and selective cytotoxic structures containing pyrrole motifs have shown promising data for future investigations. Accordingly, this review presents an overview of recent developments in the pyrrole derivatives as anticancer agents, with a main focus towards the key moieties required for the anti-tumor activities. Pyrrole molecules comprising prominent targeting capacities against microtubule polymerization, tyrosine kinases, cytochrome p450 family 1, histone deacetylase and bcl-2 proteins were reported. In addition, several mechanisms of action, such as apoptosis, cell cycle arrest, inhibiting kinases, angiogenesis, disruption of cell migration, modulation of nuclear receptor responsiveness and others were analyzed. Furthermore, in most of the discussed cases we provided synthesis schemes of the mentioned molecules. Overall, the utilization of pyrrole scaffold for the design and synthesis of novel anticancer drugs could be a promising approach for future investigations.

Analysis of Antidepressant-like Effects and Action Mechanisms of GSB-106, a Small Molecule, Affecting the TrkB Signaling.

Induction of BDNF-TrkB signaling is associated with the action mechanisms of conventional and fast-acting antidepressants. GSB-106, developed as a small dimeric dipeptide mimetic of BDNF, was previously shown to produce antidepressant-like effects in the mouse Porsolt test, tail suspension test, Nomura water wheel test, in the chronic social defeat stress model and in the inflammation-induced model of depression. In the present study, we evaluated the effect of chronic per os administration of GSB-106 to Balb/c mice under unpredictable chronic mild stress (UCMS). It was observed for the first time that long term GSB-106 treatment (1 mg/kg, 26 days) during ongoing UCMS procedure ameliorated the depressive-like behaviors in mice as indicated by the Porsolt test. In addition, chronic per os administration of GSB-106 resulted in an increase in BDNF levels, which were found to be decreased in the prefrontal cortex and hippocampus of mice after UCMS. Furthermore, prolonged GSB-106 treatment was accompanied by an increase in the content of pTrkB706/707 in the prefrontal cortex and by a pronounced increase in the level of pTrkB816 in both studied brain structures of mice subjected to UCMS procedure. In summary, the present data show that chronic GSB-106 treatment produces an antidepressant-like effect in the unpredictable chronic mild stress model, which is likely to be associated with the regulation of the BDNF-TrkB signaling.

Fibroblast growth factor (FGF), FGF receptor (FGFR), and cyclin D1 (CCND1) DNA methylation in head and neck squamous cell carcinomas is associated with transcriptional activity, gene amplification, human papillomavirus (HPV) status, and sensitivity to tyrosine kinase inhibitors.

BACKGROUND: Dysregulation of fibroblast growth factor receptor (FGFR) signaling pathway has been observed in head and neck squamous cell carcinoma (HNSCC) and is a promising therapeutic target for selective tyrosine kinase inhibitors (TKIs). Potential predictive biomarkers for response to FGFR-targeted therapies are urgently needed. Understanding the epigenetic regulation of FGF pathway related genes, i.e. FGFRs, FGFs, and CCND1, could enlighten the way towards biomarker-selected FGFR-targeted therapies. METHODS: We performed DNA methylation analysis of the encoding genes FGFR1, FGFR2, FGFR3, FGFR4, FGF1-14, FGF16-23, and CCND1 at single CpG site resolution (840 CpG sites) employing The Cancer Genome Research Atlas (TCGA) HNSCC cohort comprising N = 530 tumor tissue and N = 50 normal adjacent tissue samples. We correlated DNA methylation to mRNA expression with regard to human papilloma virus (HPV) and gene amplification status. Moreover, we investigated the correlation of methylation with sensitivity to the selective FGFR inhibitors PD 173074 and AZD4547 in N = 40 HPV(-) HNSCC cell lines. RESULTS: We found sequence-contextually nuanced CpG methylation patterns in concordance with epigenetically regulated genes. High methylation levels were predominantly found in the promoter flank and gene body region, while low methylation levels were present in the central promoter region for most of the analyzed CpG sites. FGFRs, FGFs, and CCND1 methylation differed significantly between tumor and normal adjacent tissue and was associated with HPV and gene amplification status. CCND1 promoter methylation correlated with CCND1 amplification. For most of the analyzed CpG sites, methylation levels correlated to mRNA expression in tumor tissue. Furthermore, we found significant correlations of DNA methylation of specific CpG sites with response to the FGFR1/3-selective inhibitors PD 173074 and AZD4547, predominantly within the transcription start site of CCND1. CONCLUSIONS: Our results suggest an epigenetic regulation of CCND1, FGFRs, and FGFs via DNA methylation in HNSCC and warrants further investigation of DNA methylation as a potential predictive biomarker for response to selective FGFR inhibitors in clinical trials.

Abemaciclib is a potent inhibitor of DYRK1A and HIP kinases involved in transcriptional regulation.

Homeodomain-interacting protein kinases (HIPKs) belong to the CMGC kinase family and are closely related to dual-specificity tyrosine phosphorylation-regulated kinases (DYRKs). HIPKs are regulators of various signaling pathways and involved in the pathology of cancer, chronic fibrosis, diabetes, and multiple neurodegenerative diseases. Here, we report the crystal structure of HIPK3 in its apo form at 2.5 Å resolution. Recombinant HIPKs and DYRK1A are auto-activated and phosphorylate the negative elongation factor SPT5, the transcription factor c-Myc, and the C-terminal domain of RNA polymerase II, suggesting a direct function in transcriptional regulation. Based on a database search, we identified abemaciclib, an FDA-approved Cdk4/Cdk6 inhibitor used for the treatment of metastatic breast cancer, as potent inhibitor of HIPK2, HIPK3, and DYRK1A. We determined the crystal structures of HIPK3 and DYRK1A bound to abemaciclib, showing a similar binding mode to the hinge region of the kinase as observed for Cdk6. Remarkably, DYRK1A is inhibited by abemaciclib to the same extent as Cdk4/Cdk6 in vitro, raising the question of whether targeting of DYRK1A contributes to the transcriptional inhibition and therapeutic activity of abemaciclib.

Aberrant B Cell Receptor Signaling in Naïve B Cells from Patients with Idiopathic Pulmonary Fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic and ultimately fatal disease in which an impaired healing response to recurrent micro-injuries is thought to lead to fibrosis. Recent findings hint at a role for B cells and autoimmunity in IPF pathogenesis. We previously reported that circulating B cells from a fraction of patients, compared with healthy controls, express increased levels of the signaling molecule Bruton's tyrosine kinase (BTK). However, it remains unclear whether B cell receptor (BCR) signaling is altered in IPF. Here, we show that the response to BCR stimulation is enhanced in peripheral blood B cells from treatment-naïve IPF patients. We observed increased anti-immunoglobulin-induced phosphorylation of BTK and its substrate phospholipase Cγ2 (PLCγ2) in naïve but not in memory B cells of patients with IPF. In naïve B cells of IPF patients enhanced BCR signaling correlated with surface expression of transmembrane activator and calcium-modulator and cyclophilin ligand interactor (TACI) but not B cell activating factor receptor (BAFFR), both of which provide pro-survival signals. Interestingly, treatment of IPF patients with nintedanib, a tyrosine kinase inhibitor with anti-fibrotic and anti-inflammatory activity, induced substantial changes in BCR signaling. These findings support the involvement of B cells in IPF pathogenesis and suggest that targeting BCR signaling has potential value as a treatment option.

Antidepressant-like effect of guanosine involves activation of AMPA receptor and BDNF/TrkB signaling.

Guanosine is a purine nucleoside that has been shown to exhibit antidepressant effects, but the mechanisms underlying its effect are not well established. We investigated if the antidepressant-like effect induced by guanosine in the tail suspension test (TST) in mice involves the modulation of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor, voltage-dependent calcium channel (VDCC), and brain-derived neurotrophic factor (BDNF)/tropomyosin receptor kinase B (TrkB) pathway. We also evaluated if the antidepressant-like effect of guanosine is accompanied by an acute increase in hippocampal and prefrontocortical BDNF levels. Additionally, we investigated if the ability of guanosine to elicit a fast behavioral response in the novelty suppressed feeding (NSF) test is associated with morphological changes related to hippocampal synaptogenesis. The antidepressant-like effect of guanosine (0.05 mg/kg, p.o.) in the TST was prevented by DNQX (AMPA receptor antagonist), verapamil (VDCC blocker), K-252a (TrkBantagonist), or BDNF antibody. Increased P70S6K phosphorylation and higher synapsin I immunocontent in the hippocampus, but not in the prefrontal cortex, were observed 1 h after guanosine administration. Guanosine exerted an antidepressant-like effect 1, 6, and 24 h after its administration, an effect accompanied by increased hippocampal BDNF level. In the prefrontal cortex, BDNF level was increased only 1 h after guanosine treatment. Finally, guanosine was effective in the NSF test (after 1 h) but caused no alterations in dendritic spine density and remodeling in the ventral dentate gyrus (DG). Altogether, the results indicate that guanosine modulates targets known to be implicated in fast antidepressant behavioral responses (AMPA receptor, VDCC, and TrkB/BDNF pathway).

Class I HDAC inhibition improves object recognition memory consolidation through BDNF/TrkB pathway in a time-dependent manner.

There is increasing evidence showing that HDACs regulates BDNF (brain-derived neurotrophic factor) expression through its interaction with the Bdnf gene promoter, a key regulator to consolidate memory. Although the nuclear mechanisms regulated by HDACs that control BDNF expression have been partially described recently, the temporal events for memory consolidation remain unknown. Hence, in this work, we studied the temporal pattern for the activation of the BDNF/TrkB pathway through class I HDAC inhibition to enhance object recognition memory (ORM) consolidation. To this end, we inhibited class I HDAC into the insular cortex (IC) and a weak ORM protocol was used to assess temporal expression and function of the BDNF/TrkB pathway in the IC. We found that cortical class I HDAC inhibition enhanced long-term ORM, coincident with a clear peak of BDNF expression at 4 h after acquisition. Furthermore, the tyrosine kinase B (TrkB) receptor blockade at 4 h, but not at 8 h, impaired the consolidation of ORM. These results suggest that histone acetylation regulates the temporal expression of BDNF in cortical circuits potentiating the long-term recognition memory.

Generation of osimertinib-resistant cells from epidermal growth factor receptor L858R/T790M mutant non-small cell lung carcinoma cell line.

BACKGROUND: Lung cancer contributes to high cancer mortality worldwide with 80% of total cases diagnosed as non-small cell lung cancer (NSCLC). Epidermal growth factor receptor (EGFR) tyrosine kinase (TK) domain serves as a druggable target in NSCLC patients with exon 19 deletion and L858R mutation. However, patients eventually succumbed to resistance to first- and second-generation EGFR-TK inhibitors through activation of T790M mutation. Third-generation EGFR-TKI, Osimertinib exhibits high efficacy in patients with exon 19 deletion/L858R/T790M mutation but they experienced acquired resistance thereafter. Available treatment options in NSCLC patients remains a challenge due to unknown molecular heterogeneity responsible for acquired resistance to EGFR-TKI. In this study, we aim to generate Osimertinib-resistant (OR) cells from H1975 carrying L858R/T790M double mutation which can be used as a model to elucidate mechanism of resistance. METHODS: OR cells were established via stepwise-dose escalation and limiting single-cell dilution method. We then evaluated Osimertinib resistance potential via cell viability assay. Proteins expression related to EGFR-signalling, epithelial to mesenchymal transition (EMT), and autophagy were analyzed via western blot. RESULTS: OR cell lines exhibited increased drug resistance potential compared to H1975. Distinguishable mesenchymal-like features were observed in OR cells. Protein expression analysis revealed EGFR-independent signaling involved in the derived OR cells as well as EMT and autophagy activity. CONCLUSION: We generated OR cell lines in-vitro as evidenced by increased drug resistance potential, increased mesenchymal features, and enhanced autophagy activity. Development of Osimertinib resistance cells may serve as in-vitro model facilitating discovery of molecular aberration present during acquired mechanism of resistance.

Epoxyquinophomopsins A and B from endophytic fungus Phomopsis sp. and their activity against tyrosine kinase.

Two new quinone derivatives, epoxyquinophomopsins A (1) and B (2), were purified from the EtOAc extract of endophytic fungus Phomopsis sp isolated from Morus cathayana. The structures of both compounds were determined based on 1D and 2D NMR and mass spectral data, as well as by x-ray diffraction analysis for 1. Compounds 1 and 2 were screened against eight receptor- (RTKs) and eight non-receptor tyrosine kinases (nRTKs). Both compounds showed strong inhibitory properties against Bruton's Tyrosine Kinase (nRTK) with their kinase activity were 19% and 20%, respectively. Only compound 1 that showed strong inhibitory properties against RTKs EGFR and HER-4 with its kinase activity were 16 and 15%, respectively. Thus, both compounds have potential as tyrosine kinase inhibitors.

Rapid-acting and long-lasting antidepressant-like action of (R)-ketamine in Nrf2 knock-out mice: a role of TrkB signaling.

The transcription nuclear factor-erythroid factor 2-related factor 2 (Nrf2) plays a key role in inflammation that is involved in depression. We previously reported that Nrf2 knock-out (KO) mice exhibit depression-like phenotypes through systemic inflammation. (R)-ketamine, an enantiomer of ketamine, has rapid-acting and long-lasting antidepressant-like effects in rodents. We investigated whether (R)-ketamine can produce antidepressant-like effects in Nrf2 KO mice. Effects of (R)-ketamine on the depression-like phenotypes in Nrf2 KO mice were examined. Furthermore, the role of TrkB in the antidepressant-like actions of (R)-ketamine was also examined. In the tail-suspension test (TST) and forced swimming test (FST), (R)-ketamine (10 mg/kg) significantly attenuated the increased immobility times of TST and FST in the Nrf2 KO mice. In the sucrose preference test (SPT), (R)-ketamine significantly ameliorated the reduced preference of SPT in Nrf2 KO mice. Decreased expression of synaptic proteins (i.e., GluA1 and PSD-95) in the medial prefrontal cortex (mPFC) of Nrf2 KO mice was significantly ameliorated after a single injection of (R)-ketamine. Furthermore, the pre-treatment with the TrkB antagonist ANA-12 (0.5 mg/kg) significantly blocked the rapid and long-lasting antidepressant-like effects of (R)-ketamine in Nrf2 KO mice. Furthermore, ANA-12 significantly antagonized the beneficial effects of (R)-ketamine on decreased expression of synaptic proteins in the mPFC of Nrf2 KO mice. These findings suggest that (R)-ketamine can produce rapid and long-lasting antidepressant-like actions in Nrf2 KO mice via TrkB signaling.

Novel antiplatelet strategies targeting GPVI, CLEC-2 and tyrosine kinases.

Antiplatelet medications comprise the cornerstone of treatment for diseases that involve arterial thrombosis, including acute coronary syndromes (ACS), stroke and peripheral arterial disease. However, antiplatelet medications may cause bleeding and, furthermore, thrombotic events may still recur despite treatment. The interaction of collagen with GPVI receptors on the surface of platelets has been identified as one of the major players in the pathophysiology of arterial thrombosis that occurs following atherosclerotic plaque rupture. Promisingly, GPVI deficiency in humans appears to have a minimal impact on bleeding. These findings together suggest that targeting platelet GPVI may provide a novel treatment strategy that provides additional antithrombotic efficacy with minimal disruption of normal hemostasis compared to conventional antiplatelet medications. CLEC-2 is gaining interest as a therapeutic target for a variety of thrombo-inflammatory disorders including deep vein thrombosis (DVT) with treatment also predicted to cause minimal disruption to hemostasis. GPVI and CLEC-2 signal through Src, Syk and Tec family tyrosine kinases, providing additional strategies for inhibiting both receptors. In this review, we summarize the evidence regarding GPVI and CLEC-2 and strategies for inhibiting these receptors to inhibit platelet recruitment and activation in thrombotic diseases.

Transcriptional response of human articular chondrocytes treated with fibronectin fragments: an in vitro model of the osteoarthritis phenotype.

OBJECTIVE: Fibronectin is a matrix protein that is fragmented during cartilage degradation in osteoarthritis (OA). Treatment of chondrocytes with fibronectin fragments (FN-f) has been used to model OA in vitro, but the system has not been fully characterized. This study sought to define the transcriptional response of chondrocytes to FN-f, and directly compare it to responses traditionally observed in OA. DESIGN: Normal human femoral chondrocytes isolated from tissue donors were treated with either FN-f or PBS (control) for 3, 6, or 18 h. RNA-seq libraries were compared between time-matched FN-f and control samples in order to identify changes in gene expression over time. Differentially expressed genes were compared to a published OA gene set and used for pathway, transcription factor motif, and kinome analysis. RESULTS: FN-f treatment resulted in 3,914 differentially expressed genes over the time course. Genes that are up- or downregulated in OA were significantly up- (P < 0.00001) or downregulated (P < 0.0004) in response to FN-f. Early response genes were involved in proinflammatory pathways, whereas many late response genes were involved in ferroptosis. The promoters of upregulated genes were enriched for NF-κB, AP-1, and IRF motifs. Highly upregulated kinases included CAMK1G, IRAK2, and the uncharacterized kinase DYRK3, while growth factor receptors TGFBR2 and FGFR2 were downregulated. CONCLUSIONS: FN-f treatment of normal human articular chondrocytes recapitulated many key aspects of the OA chondrocyte phenotype. This in vitro model is promising for future OA studies, especially considering its compatibility with genomics and genome-editing techniques.

FOXM1 drives HPV+ HNSCC sensitivity to WEE1 inhibition.

Head and neck squamous cell carcinoma (HNSCC) associated with high-risk human papilloma virus (HPV) infection is a growing clinical problem. The WEE1 kinase inhibitor AZD1775 (WEE1i) overrides cell cycle checkpoints and is being studied in HNSCC regimens. We show that the HPV16 E6/E7 oncoproteins sensitize HNSCC cells to single-agent WEE1i treatment through activation of a FOXM1-CDK1 circuit that drives mitotic gene expression and DNA damage. An isogenic cell system indicated that E6 largely accounts for these phenotypes in ways that extend beyond p53 inactivation. A targeted genomic analysis implicated FOXM1 signaling downstream of E6/E7 expression and analyses of primary tumors and The Cancer Genome Atlas (TCGA) data revealed an activated FOXM1-directed promitotic transcriptional signature in HPV+ versus HPV- HNSCCs. Finally, we demonstrate the causality of FOXM1 in driving WEE1i sensitivity. These data suggest that elevated basal FOXM1 activity predisposes HPV+ HNSCC to WEE1i-induced toxicity and provide mechanistic insights into WEE1i and HPV+ HNSCC therapies.

Valproic Acid Promotes Early Neural Differentiation in Adult Mesenchymal Stem Cells Through Protein Signalling Pathways.

Regenerative medicine is a rapidly expanding area in research and clinical applications. Therapies involving the use of small molecule chemicals aim to simplify the creation of specific drugs for clinical applications. Adult mesenchymal stem cells have recently shown the capacity to differentiate into several cell types applicable for regenerative medicine (specifically neural cells, using chemicals). Valproic acid was an ideal candidate due to its clinical stability. It has been implicated in the induction of neural differentiation; however, the mechanism and the downstream events were not known. In this study, we showed that using valproic acid on adult mesenchymal stem cells induced neural differentiation within 24 h by upregulating the expression of suppressor of cytokine signaling 5 (SOCS5) and Fibroblast growth factor 21 (FGF21), without increasing the potential death rate of the cells. Through this, the Janus Kinase/Signal Transducer and Activator of Transcription (JAK/STAT) pathway is downregulated, and the mitogen-activated protein kinase (MAPK) cascade is activated. The bioinformatics analyses revealed the expression of several neuro-specific proteins as well as a range of functional and structural proteins involved in the formation and development of the neural cells.

Observational Study of Sequential Afatinib and Osimertinib in EGFR Mutation-Positive NSCLC: Patients Treated with a 40-mg Starting Dose of Afatinib.

INTRODUCTION: Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) are the standard of care for patients with EGFR mutation-positive non-small cell lung cancer (NSCLC). However, questions remain about the optimal treatment sequence of EGFR TKIs. The global, observational GioTag study demonstrated prolonged time on treatment with sequential afatinib and osimertinib therapy in patients who acquired the T790M mutation. Here, we assessed outcomes in patients who received the approved 40-mg starting dose of afatinib, as used in the clinical trial setting. METHODS: In the non-interventional, global, multicenter GioTag study, patients had T790M-positive disease following first-line afatinib and started osimertinib treatment ≥ 10 months prior to data entry. Primary outcome was time on treatment. This subanalysis assessed outcomes in patients who received afatinib 40 mg. RESULTS: In 169 patients who received an afatinib starting dose of 40 mg, median time on treatment was 27.6 months (90% confidence interval [CI] 26.3-31.3). Benefit was seen across patient subgroups, particularly those with Del19-positive disease and Asian patients; median time on treatment was 29.9 months (90% CI 27.6-46.7) in patients with Del19-positive disease and 46.7 months (90% CI 28.4-not reached) in Asian patients. The 2-year overall survival rate was 80%. CONCLUSIONS: These real-world results support the overall study results and demonstrate prolonged time on treatment with sequential afatinib and osimertinib. The results suggest that sequential afatinib and osimertinib is a feasible therapeutic strategy for patients who acquire the T790M mutation, particularly those with Del19-positive disease or Asian patients. TRIAL REGISTRATION NUMBER: NCT03370770.

Pharmacologic and genetic approaches define human pancreatic ß cell mitogenic targets of DYRK1A inhibitors.

Small molecule inhibitors of dual specificity, tyrosine phosphorylation-regulated kinase 1A (DYRK1A), including harmine and others, are able to drive human ß cell regeneration. While DYRK1A is certainly a target of this class, whether it is the only or the most important target is uncertain. Here, we employ a combined pharmacologic and genetic approach to refine the potential mitogenic targets of the DYRK1A inhibitor family in human islets. A combination of human ß cell RNA sequencing, DYRK1A inhibitor kinome screens, pharmacologic inhibitors, and targeted silencing of candidate genes confirms that DYRK1A is a central target. Surprisingly, however, DYRK1B also proves to be an important target: silencing DYRK1A results in an increase in DYRK1B. Simultaneous silencing of both DYRK1A and DYRK1B yields greater ß cell proliferation than silencing either individually. Importantly, other potential kinases, such as the CLK and the GSK3 families, are excluded as important harmine targets. Finally, we describe adenoviruses that are able to silence up to 7 targets simultaneously. Collectively, we report that inhibition of both DYRK1A and DYRK1B is required for induction of maximal rates of human ß cell proliferation, and we provide clarity for future efforts in structure-based drug design for human ß cell regenerative drugs.

Targeting Kaposi's Sarcoma-Associated Herpesvirus ORF21 Tyrosine Kinase and Viral Lytic Reactivation by Tyrosine Kinase Inhibitors Approved for Clinical Use.

Kaposi's sarcoma-associated herpesvirus (KSHV) is the cause of three human malignancies: Kaposi's sarcoma, primary effusion lymphoma, and the plasma cell variant of multicentric Castleman disease. Previous research has shown that several cellular tyrosine kinases play crucial roles during several steps in the virus replication cycle. Two KSHV proteins also have protein kinase function: open reading frame (ORF) 36 encodes a serine-threonine kinase, while ORF21 encodes a thymidine kinase (TK), which has recently been found to be an efficient tyrosine kinase. In this study, we explore the role of the ORF21 tyrosine kinase function in KSHV lytic replication. By generating a recombinant KSHV mutant with an enzymatically inactive ORF21 protein, we show that the tyrosine kinase function of ORF21/TK is not required for the progression of the lytic replication in tissue culture but that it is essential for the phosphorylation and activation to toxic moieties of the antiviral drugs zidovudine and brivudine. In addition, we identify several tyrosine kinase inhibitors, already in clinical use against human malignancies, which potently inhibit not only ORF21 TK kinase function but also viral lytic reactivation and the development of KSHV-infected endothelial tumors in mice. Since they target both cellular tyrosine kinases and a viral kinase, some of these compounds might find a use in the treatment of KSHV-associated malignancies.IMPORTANCE Our findings address the role of KSHV ORF21 as a tyrosine kinase during lytic replication and the activation of prodrugs in KSHV-infected cells. We also show the potential of selected clinically approved tyrosine kinase inhibitors to inhibit KSHV TK, KSHV lytic replication, infectious virion release, and the development of an endothelial tumor. Since they target both cellular tyrosine kinases supporting productive viral replication and a viral kinase, these drugs, which are already approved for clinical use, may be suitable for repurposing for the treatment of KSHV-related tumors in AIDS patients or transplant recipients.

Protein tyrosine kinase 2: a novel therapeutic target to overcome acquired EGFR-TKI resistance in non-small cell lung cancer.

BACKGROUND: Protein tyrosine kinase 2 (PTK2) expression has been reported in various types of human epithelial cancers including lung cancer; however, the role of PTK2 in epidermal growth factor receptor (EGFR)-mutant non-small cell lung cancer (NSCLC) has not been elucidated. We previously reported that pemetrexed-resistant NSCLC cell line PC-9/PEM also acquired EGFR-TKI resistance with constitutive Akt activation, but we could not find a therapeutic target. METHODS: Cell viability in EGFR-mutant NSCLC cell lines was measured by the WST-8 assay. Phosphorylation antibody array assay for receptor tyrosine kinases was performed in PC-9 and PC-9/PEM cell lines. We evaluated the efficacy of EGFR and PTK2 co-inhibition in EGFR-TKI-resistant NSCLC in vitro. Oral defactinib and osimertinib were administered in mice bearing subcutaneous xenografts to evaluate the efficacy of the treatment combination in vivo. Both the PTK2 phosphorylation and the treatment combination efficacy were evaluated in erlotinib-resistant EGFR-mutant NSCLC cell lines. RESULTS: PTK2 was hyperphosphorylated in PC-9/PEM. Defactinib (PTK2 inhibitor) and PD173074 (FGFR inhibitor) inhibited PTK2 phosphorylation. Combination of PTK2 inhibitor and EGFR-TKI inhibited Akt and induced apoptosis in PC-9/PEM. The combination treatment showed improved in vivo therapeutic efficacy compared to the single-agent treatments. Furthermore, erlotinib-resistant NSCLC cell lines showed PTK2 hyperphosphorylation. PTK2 inhibition in the PTK2 hyperphosphorylated erlotinib-resistant cell lines also recovered EGFR-TKI sensitivity. CONCLUSION: PTK2 hyperphosphorylation occurs in various EGFR-TKI-resistant NSCLCs. Combination of PTK2 inhibitor and EGFR-TKI (defactinib and osimertinib) recovered EGFR-TKI sensitivity in the EGFR-TKI-resistant NSCLC. Our study result suggests that this combination therapy may be a viable option to overcome EGFR-TKI resistance in NSCLC.

Fostamatinib disodium hexahydrate: a novel treatment for adult immune thrombocytopenia.

Immune thrombocytopenia (ITP) is an autoimmune disease associated with substantial heterogeneity and varying outcomes. Significant bleeding, including intracranial hemorrhage, is a persistent risk for patients with ITP, along with cardiovascular disease. ITP has also been associated with decreased patient functionality and quality of life. The primary goal of ITP therapy is to lower the risk of bleeding and associated complications by raising platelet counts to levels that provide adequate hemostasis with minimal treatment-related toxicity. Current first-line treatments include corticosteroids, as well as intravenous and anti-D immunoglobulin. Despite the availability of several second-line options, the need for additional treatment options that can provide a stable, long-term response with few adverse effects is critical and ongoing. Fostamatinib disodium hexahydrate is an oral spleen tyrosine kinase inhibitor that produces a rapid, durable response in patients who have failed one or other treatments. Additionally, fostamatinib is well tolerated, and adverse effects can be actively mitigated through dose reduction, dose interruption, or standard therapeutic approaches.