ABSTRACT
Tropomyosin receptor kinases (Trks) are transmembrane receptor tyrosine kinases named TrkA, TrkB, and TrkC and encoded by the NTRK1, NTRK2, and NTRK3 genes, respectively. These kinases have attracted significant attention and represent a promising therapeutic target for solid tumor treatment due to their vital role in cellular signaling pathways. First-generation TRK inhibitors, i.e., Larotrectinib sulfate and Entrectinib, received clinical approval in 2018 and 2019, respectively. However, the use of these inhibitors was significantly limited because of the development of resistance due to mutations. Fortunately, the second-generation Trk inhibitor Repotrectinib (TPX-0005) was approved by the FDA in November 2023, while Selitrectinib (Loxo-195) has provided an effective solution to this issue. Another macrocycle-based analog, along with many other TRK inhibitors, is currently in clinical trials. Two of the three marketed drugs for NTRK fusion cancers feature a pyrazolo[1,5-a] pyrimidine nucleus, prompting medicinal chemists to develop numerous novel pyrazolopyrimidine-based molecules to enhance clinical applications. This article focuses on a comprehensive review of chronological synthetic developments and the structure-activity relationships (SAR) of pyrazolo[1,5-a]pyrimidine derivatives as Trk inhibitors. This article will also provide comprehensive knowledge and future directions to the researchers working in the field of medicinal chemistry by facilitating the structural modification of pyrazolo [1,5-a]pyrimidine derivatives to synthesize more effective novel chemotherapeutics as TRK inhibitors.
Subject(s)
Protein Kinase Inhibitors , Pyrazoles , Pyrimidines , Receptor, trkA , Pyrimidines/chemistry , Pyrimidines/pharmacology , Pyrimidines/chemical synthesis , Humans , Pyrazoles/chemistry , Pyrazoles/pharmacology , Pyrazoles/chemical synthesis , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/therapeutic use , Structure-Activity Relationship , Receptor, trkA/antagonists & inhibitors , Receptor, trkA/metabolism , Receptor, trkA/genetics , Receptor, trkB/antagonists & inhibitors , Receptor, trkB/metabolism , Receptor, trkC/antagonists & inhibitors , Receptor, trkC/genetics , Receptor, trkC/metabolism , Neoplasms/drug therapy , Neoplasms/enzymology , Antineoplastic Agents/pharmacology , Antineoplastic Agents/chemistry , Antineoplastic Agents/chemical synthesisABSTRACT
INTRODUCTION: Tropomyosin related kinase B (TrkB) and C (TrkC) receptor signaling promotes synaptic plasticity and interacts with pathways affected by amyloid beta (Aß) toxicity. Upregulating TrkB/C signaling could reduce Alzheimer's disease (AD)-related degenerative signaling, memory loss, and synaptic dysfunction. METHODS: PTX-BD10-2 (BD10-2), a small molecule TrkB/C receptor partial agonist, was orally administered to aged London/Swedish-APP mutant mice (APPL/S) and wild-type controls. Effects on memory and hippocampal long-term potentiation (LTP) were assessed using electrophysiology, behavioral studies, immunoblotting, immunofluorescence staining, and RNA sequencing. RESULTS: In APPL/S mice, BD10-2 treatment improved memory and LTP deficits. This was accompanied by normalized phosphorylation of protein kinase B (Akt), calcium-calmodulin-dependent kinase II (CaMKII), and AMPA-type glutamate receptors containing the subunit GluA1; enhanced activity-dependent recruitment of synaptic proteins; and increased excitatory synapse number. BD10-2 also had potentially favorable effects on LTP-dependent complement pathway and synaptic gene transcription. DISCUSSION: BD10-2 prevented APPL/S/Aß-associated memory and LTP deficits, reduced abnormalities in synapse-related signaling and activity-dependent transcription of synaptic genes, and bolstered transcriptional changes associated with microglial immune response. HIGHLIGHTS: Small molecule modulation of tropomyosin related kinase B (TrkB) and C (TrkC) restores long-term potentiation (LTP) and behavior in an Alzheimer's disease (AD) model. Modulation of TrkB and TrkC regulates synaptic activity-dependent transcription. TrkB and TrkC receptors are candidate targets for translational therapeutics. Electrophysiology combined with transcriptomics elucidates synaptic restoration. LTP identifies neuron and microglia AD-relevant human-mouse co-expression modules.
Subject(s)
Alzheimer Disease , Microglia , Receptor, trkB , Synapses , Animals , Male , Mice , Alzheimer Disease/drug therapy , Disease Models, Animal , Hippocampus/drug effects , Hippocampus/metabolism , Long-Term Potentiation/drug effects , Mice, Transgenic , Microglia/drug effects , Microglia/metabolism , Neuronal Plasticity/drug effects , Receptor, trkB/metabolism , Receptor, trkC/metabolism , Receptor, trkC/genetics , Synapses/drug effects , Transcriptome/drug effectsABSTRACT
Cancer patients undergoing treatment with antineoplastic drugs often experience chemotherapy-induced neuropathic pain (CINP), and the therapeutic options for managing CINP are limited. Here, we show that systemic paclitaxel administration upregulates the expression of neurotrophin-3 (Nt3) mRNA and NT3 protein in the neurons of dorsal root ganglia (DRG), but not in the spinal cord. Blocking NT3 upregulation attenuates paclitaxel-induced mechanical, heat, and cold nociceptive hypersensitivities and spontaneous pain without altering acute pain and locomotor activity in male and female mice. Conversely, mimicking this increase produces enhanced responses to mechanical, heat, and cold stimuli and spontaneous pain in naive male and female mice. Mechanistically, NT3 triggers tropomyosin receptor kinase C (TrkC) activation and participates in the paclitaxel-induced increases of C-C chemokine ligand 2 (Ccl2) mRNA and CCL2 protein in the DRG. Given that CCL2 is an endogenous initiator of CINP and that Nt3 mRNA co-expresses with TrkC and Ccl2 mRNAs in DRG neurons, NT3 likely contributes to CINP through TrkC-mediated activation of the Ccl2 gene in DRG neurons. NT3 may be thus a potential target for CINP treatment.
Subject(s)
Chemokine CCL2 , Ganglia, Spinal , Neuralgia , Neurons , Neurotrophin 3 , Paclitaxel , Receptor, trkC , Animals , Female , Male , Mice , Antineoplastic Agents/adverse effects , Chemokine CCL2/metabolism , Chemokine CCL2/genetics , Ganglia, Spinal/metabolism , Ganglia, Spinal/drug effects , Neuralgia/chemically induced , Neuralgia/metabolism , Neuralgia/genetics , Neurons/metabolism , Neurons/drug effects , Neurotrophin 3/metabolism , Neurotrophin 3/genetics , Paclitaxel/adverse effects , Paclitaxel/pharmacology , Receptor, trkC/metabolism , Receptor, trkC/genetics , RNA, Messenger/metabolism , RNA, Messenger/genetics , Nerve Growth Factors/genetics , Nerve Growth Factors/metabolismABSTRACT
Neurotrophic receptor tyrosine kinase 3 (NTRK3) has pleiotropic functions: it acts not only as an oncogene in breast and gastric cancers but also as a dependence receptor in tumor suppressor genes in colon cancer and neuroblastomas. However, the role of NTRK3 in upper tract urothelial carcinoma (UTUC) is not well documented. This study investigated the association between NTRK3 expression and outcomes in UTUC patients and validated the results in tests on UTUC cell lines. A total of 118 UTUC cancer tissue samples were examined to evaluate the expression of NTRK3. Survival curves were generated using Kaplan-Meier estimates, and Cox regression models were used for investigating survival outcomes. Higher NTRK3 expression was correlated with worse progression-free survival, cancer-specific survival, and overall survival. Moreover, the results of an Ingenuity Pathway Analysis suggested that NTRK3 may interact with the PI3K-AKT-mTOR signaling pathway to promote cancer. NTRK3 downregulation in BFTC909 cells through shRNA reduced cellular migration, invasion, and activity in the AKT-mTOR pathway. Furthermore, the overexpression of NTRK3 in UM-UC-14 cells promoted AKT-mTOR pathway activity, cellular migration, and cell invasion. From these observations, we concluded that NTRK3 may contribute to aggressive behaviors in UTUC by facilitating cell migration and invasion through its interaction with the AKT-mTOR pathway and the expression of NTRK3 is a potential predictor of clinical outcomes in cases of UTUC.
Subject(s)
Cell Movement , Receptor, trkC , Urologic Neoplasms , Female , Humans , Male , Cell Line, Tumor , Gene Expression Regulation, Neoplastic , Kaplan-Meier Estimate , Phosphatidylinositol 3-Kinases/metabolism , Phosphatidylinositol 3-Kinases/genetics , Proto-Oncogene Proteins c-akt/metabolism , Proto-Oncogene Proteins c-akt/genetics , Receptor, trkC/metabolism , Receptor, trkC/genetics , Signal Transduction , TOR Serine-Threonine Kinases/metabolism , TOR Serine-Threonine Kinases/genetics , Urologic Neoplasms/genetics , Urologic Neoplasms/metabolism , Urologic Neoplasms/pathologyABSTRACT
Context: Two-thirds of metastatic differentiated thyroid cancer (DTC) patients have radioiodine (RAI)-resistant disease, resulting in poor prognosis and high mortality. For rare NTRK and RET fusion-positive metastatic, RAI-resistant thyroid cancers, variable success of re-induction of RAI avidity during treatment with NTRK or RET inhibitors has been reported. Case presentation and results: We report two cases with RAI-resistant lung metastases treated with larotrectinib: an 83-year-old male presenting with an ETV6::NTRK3 fusion-positive tumor with the TERT promoter mutation c.-124C>T, and a 31-year-old female presenting with a TPR::NTRK1 fusion-positive tumor (and negative for TERT promoter mutation). Post larotrectinib treatment, diagnostic I-123 whole body scan revealed unsuccessful RAI-uptake re-induction in the TERT-positive tumor, with a thyroid differentiation score (TDS) of -0.287. In contrast, the TERT-negative tumor exhibited successful I-131 reuptake with a TDS of -0.060. Conclusion: As observed for RAI-resistance associated with concurrent TERT and BRAF mutations, the co-occurrence of TERT mutations and NTRK fusions may also contribute to re-sensitization failure.
Subject(s)
Iodine Radioisotopes , Thyroid Neoplasms , Humans , Iodine Radioisotopes/therapeutic use , Thyroid Neoplasms/genetics , Thyroid Neoplasms/pathology , Thyroid Neoplasms/radiotherapy , Male , Female , Adult , Aged, 80 and over , Pyrimidines/therapeutic use , Oncogene Proteins, Fusion/genetics , Lung Neoplasms/genetics , Lung Neoplasms/pathology , Lung Neoplasms/radiotherapy , Lung Neoplasms/secondary , Pyrazoles/therapeutic use , Receptor, trkA/genetics , Telomerase/genetics , Receptor, trkC/genetics , Receptor, trkC/metabolism , Repressor Proteins/genetics , Proto-Oncogene Proteins c-ets/genetics , Mutation , ETS Translocation Variant 6 ProteinSubject(s)
Receptor, trkA , Sarcoma , Uterine Neoplasms , Humans , Female , Adult , Uterine Neoplasms/genetics , Uterine Neoplasms/pathology , Uterine Neoplasms/metabolism , Middle Aged , Receptor, trkA/genetics , Receptor, trkA/metabolism , Sarcoma/genetics , Sarcoma/pathology , Sarcoma/metabolism , Gene Rearrangement , Receptor, trkC/genetics , Receptor, trkC/metabolism , Retrospective Studies , Cyclin D1/metabolism , Cyclin D1/genetics , S100 Proteins/metabolism , S100 Proteins/geneticsABSTRACT
Fear-related pathologies are among the most prevalent psychiatric conditions, having inappropriate learned fear and resistance to extinction as cardinal features. Exposure therapy represents a promising therapeutic approach, the efficiency of which depends on inter-individual variation in fear extinction learning, which neurobiological basis is unknown. We characterized a model of extinction learning, whereby fear-conditioned mice were categorized as extinction (EXT)-success or EXT-failure, according to their inherent ability to extinguish fear. In the lateral amygdala, GluN2A-containing NMDAR are required for LTP and stabilization of fear memories, while GluN2B-containing NMDAR are required for LTD and fear extinction. EXT-success mice showed attenuated LTP, strong LTD and higher levels of synaptic GluN2B, while EXT-failure mice showed strong LTP, no LTD and higher levels of synaptic GluN2A. Neurotrophin 3 (NT3) infusion in the lateral amygdala was sufficient to rescue extinction deficits in EXT-failure mice. Mechanistically, activation of tropomyosin receptor kinase C (TrkC) with NT3 in EXT-failure slices attenuated lateral amygdala LTP, in a GluN2B-dependent manner. Conversely, blocking endogenous NT3-TrkC signaling with TrkC-Fc chimera in EXT-success slices strengthened lateral amygdala LTP. Our data support a key role for the NT3-TrkC system in inter-individual differences in fear extinction in rodents, through modulation of amygdalar NMDAR composition and synaptic plasticity.
Subject(s)
Amygdala , Extinction, Psychological , Fear , Individuality , Mice, Inbred C57BL , Neuronal Plasticity , Neurotrophin 3 , Receptor, trkC , Receptors, N-Methyl-D-Aspartate , Animals , Fear/physiology , Extinction, Psychological/physiology , Amygdala/metabolism , Amygdala/physiology , Mice , Neuronal Plasticity/physiology , Male , Receptors, N-Methyl-D-Aspartate/metabolism , Receptor, trkC/metabolism , Neurotrophin 3/metabolism , Long-Term Potentiation/physiology , Signal Transduction/physiology , Conditioning, Classical/physiologyABSTRACT
The seat of higher-order cognitive abilities in mammals, the neocortex, is a complex structure, organized in several layers. The different subtypes of principal neurons are distributed in precise ratios and at specific positions in these layers and are generated by the same neural progenitor cells (NPCs), steered by a spatially and temporally specified combination of molecular cues that are incompletely understood. Recently, we discovered that an alternatively spliced isoform of the TrkC receptor lacking the kinase domain, TrkC-T1, is a determinant of the corticofugal projection neuron (CFuPN) fate. Here, we show that the finely tuned balance between TrkC-T1 and the better known, kinase domain-containing isoform, TrkC-TK+, is cell type-specific in the developing cortex and established through the antagonistic actions of two RNA-binding proteins, Srsf1 and Elavl1. Moreover, our data show that Srsf1 promotes the CFuPN fate and Elavl1 promotes the callosal projection neuron (CPN) fate in vivo via regulating the distinct ratios of TrkC-T1 to TrkC-TK+. Taken together, we connect spatio-temporal expression of Srsf1 and Elavl1 in the developing neocortex with the regulation of TrkC alternative splicing and transcript stability and neuronal fate choice, thus adding to the mechanistic and functional understanding of alternative splicing in vivo.
Subject(s)
Neocortex , Receptor, trkC , Animals , Alternative Splicing , Mammals/metabolism , Neocortex/metabolism , Neurons/metabolism , Protein Isoforms/genetics , Protein Isoforms/metabolism , Receptor, trkC/chemistry , Receptor, trkC/genetics , Receptor, trkC/metabolism , Mice , Cell Line, TumorABSTRACT
Learned fear is orchestrated by a brain fear network that comprises the amygdala, hippocampus and the medial prefrontal cortex. Synaptic plasticity within this network is critical for the formation of proper fear memories. Known for their role in the promotion of synaptic plasticity, neurotrophins position as obvious candidates in the regulation of fear processes. Indeed, recent evidence from our laboratory and others associates dysregulated signalling through neurotrophin-3 and its receptor TrkC with the pathophysiology of anxiety and fear-related disorders. Here, we put wild-type C57Bl/6J mice through a contextual fear conditioning paradigm in order to characterize TrkC activation and expression in the main brain regions involved in (learned) fear - amygdala, hippocampus, and prefrontal cortex - during the formation of a fear memory. We report an overall decreased activation of TrkC in the fear network during fear consolidation and reconsolidation. During reconsolidation, hippocampal TrkC downregulation was accompanied by a decrease in the expression and activation of Erk, a critical signalling pathway in fear conditioning. Moreover, we did not find evidence that the observed decrease of TrkC activation was caused by altered expression of dominant negative form of TrkC, neurotrophin-3, or the PTP1B phosphatase. Our results indicate hippocampal TrkC inactivation through Erk signalling as a potential mechanism in the regulation of contextual fear memory formation.
Subject(s)
Brain , Fear , Animals , Mice , Brain/metabolism , Fear/physiology , Hippocampus/metabolism , Nerve Growth Factors/metabolism , Prefrontal Cortex/metabolism , Receptor, trkC/metabolismABSTRACT
Neurotrophins are a family of closely related secreted proteins that promote differentiation, development, and survival of neurons, which include nerve growth factor (NGF), brain-derived neurotrophic factor, neurotrophin-3, and neurotrophin-4. All neurotrophins signal through tropomyosin receptor kinases (TrkA, TrkB, and TrkC) which are more selective to NGF, brain-derived neurotrophic factor, and neurotrophin-3, respectively. NGF is the most studied neurotrophin in the ocular surface and a human recombinant NGF has reached clinics, having been approved to treat neurotrophic keratitis. Brain-derived neurotrophic factor, neurotrophin-3, and neurotrophin-4 are less studied neurotrophins in the ocular surface, even though brain-derived neurotrophic factor is well characterized in glaucoma, retina, and neuroscience. Recently, neurotrophin analogs with panTrk activity and TrkC selectivity have shown promise as novel drugs for treating dry eye disease. In this review, we discuss the biology of the neurotrophin family, its role in corneal homeostasis, and its use in treating ocular surface diseases. There is an unmet need to investigate parenteral neurotrophins and its analogs that activate TrkB and TrkC selectively.
Subject(s)
Brain-Derived Neurotrophic Factor , Eye Injuries , Nerve Growth Factor , Receptor Protein-Tyrosine Kinases , Humans , Brain-Derived Neurotrophic Factor/metabolism , Brain-Derived Neurotrophic Factor/pharmacology , Eye/metabolism , Eye/pathology , Ligands , Nerve Growth Factor/metabolism , Nerve Growth Factor/pharmacology , Receptor Protein-Tyrosine Kinases/metabolism , Receptors, Nerve Growth Factor/metabolism , Receptor, trkA/metabolism , Receptor, trkB/metabolism , Receptor, trkC/metabolism , Eye Injuries/drug therapy , Eye Injuries/genetics , Eye Injuries/metabolismABSTRACT
None of the previous studies has systematically explored how upregulation of TrkC plays a central role in the pathogenesis of hepatocellular carcinoma (HCC) by regulating the underlying mechanisms that promote invasion and metastasis. In this report, we demonstrated the possible association between upregulation of TrkC and acquisition of cancer stem cells traits or chemoresistance in HCC. We show that upregulation of TrkC is closely associated with the survival and progression of HCC in vivo and in vitro. Most strikingly, activation of STAT3 by TrkC-mediated inhibition of DJ-1 degradation significantly enhances the efficacy of invasion and metastasis during the progression of HCC cells. Acquiring the traits of cancer stem cells (CSCs) by TrkC/DJ-1/STAT3 signaling pathway leads to the induction of chemoresistance via upregulation of ABC transporters and anti-apoptotic genes. Also, activating the epithelial-mesenchymal transition (EMT) program by inducing EMT-transcription factor (TF)s by TrkC/DJ-1/STAT3 signaling pathway is the direct cause of multiple tumor malignancies of HCC. Thus, understanding the mechanisms by which acquisition of anticancer drug resistance by TrkC-mediated inhibition of DJ-1 degradation can help enhance the efficacy of anticancer therapies.
Subject(s)
Antineoplastic Agents , Carcinoma, Hepatocellular , Liver Neoplasms , Protein Deglycase DJ-1/metabolism , Receptor, trkC/metabolism , ATP-Binding Cassette Transporters/genetics , Antineoplastic Agents/pharmacology , Carcinoma, Hepatocellular/pathology , Cell Line, Tumor , Epithelial-Mesenchymal Transition , Gene Expression Regulation, Neoplastic , Humans , Liver Neoplasms/pathologyABSTRACT
Fusion genes are abnormal genes resulting from chromosomal translocation, insertion, deletion, inversion, etc. ETV6, a rather promiscuous partner forms fusions with several other genes, most commonly, the NTRK3 gene. This fusion leads to the formation of a constitutively activated tyrosine kinase which activates the Ras-Raf-MEK and PI3K/AKT/MAPK pathways, leading the cells through cycles of uncontrolled division and ultimately resulting in cancer. Targeted therapies against this ETV6-NTRK3 fusion protein are much needed. Therefore, to find a targeted approach, a transcription factor RBPJ regulating the ETV6 gene was established and since the ETV6-NTRK3 fusion gene is downstream of the ETV6 promoter/enhancer, this fusion protein is also regulated. The regulation of the ETV6 gene via RBPJ was validated by ChIP analysis in human glioblastoma (GBM) cell lines and patient tissue samples. This study was further followed by the identification of an inhibitor, Furamidine, against transcription factor RBPJ. It was found to be binding with the DNA binding domain of RBPJ with antitumorigenic properties and minimal organ toxicity. Hence, a new target RBPJ, regulating the production of ETV6 and ETV6-NTRK3 fusion protein was found along with a potent RBPJ inhibitor Furamidine.
Subject(s)
DNA-Binding Proteins , Glioblastoma , DNA-Binding Proteins/genetics , Glioblastoma/drug therapy , Glioblastoma/genetics , Humans , Immunoglobulin J Recombination Signal Sequence-Binding Protein , Oncogene Proteins, Fusion/genetics , Oncogene Proteins, Fusion/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-ets/genetics , Receptor, trkC/genetics , Receptor, trkC/metabolism , Repressor Proteins/chemistry , Repressor Proteins/genetics , Transcription Factors/geneticsABSTRACT
Solid tumors resulting from oncogenic stimulation of neurotrophin receptors (TRK) by chimeric proteins are a group of rare tumors of various localization that respond to therapy with targeted drugs entrectinib and larotrectinib. The standard method for detecting chimeric TRK genes in tumor samples today is considered to be next generation sequencing with the determination of the prime structure of the chimeric transcripts. We hypothesized that expression of the chimeric tyrosine kinase proteins in tumors can determine the specific transcriptomic profile of tumor cells. We detected differentially expressed genes allowing distinguishing between TRK-dependent tumors papillary thyroid cancer (TC) from other molecular variants of tumors of this type. Using PCR with reverse transcription (RT-PCR), we identified 7 samples of papillary TC carrying a EVT6-NTRK3 rearrangement (7/215, 3.26%). Using machine learning and the data extracted from TCGA, we developed of a recognition function for predicting the presence of rearrangement in NTRK genes based on the expression of 10 key genes: AUTS2, DTNA, ERBB4, HDAC1, IGF1, KDR, NTRK1, PASK, PPP2R5B, and PRSS1. The recognition function was used to analyze the expression data of the above genes in 7 TRK-dependent and 10 TRK-independent thyroid tumors obtained by RT-PCR. On the test samples from TCGA, the sensitivity was 72.7%, the specificity - 99.6%. On our independent validation samples tested by RT-PCR, sensitivity was 100%, specificity - 70%. We proposed an mRNA profile of ten genes that can classify TC in relation to the presence of driver NTRK-chimeric TRK genes with acceptable sensitivity and specificity.
Subject(s)
Proto-Oncogene Proteins c-ets , Receptor, trkC , Receptors, Nerve Growth Factor , Repressor Proteins , Thyroid Neoplasms , High-Throughput Nucleotide Sequencing , Humans , Proto-Oncogene Proteins c-ets/genetics , Proto-Oncogene Proteins c-ets/metabolism , Receptor Protein-Tyrosine Kinases/genetics , Receptor Protein-Tyrosine Kinases/metabolism , Receptor, trkC/genetics , Receptor, trkC/metabolism , Receptors, Nerve Growth Factor/genetics , Receptors, Nerve Growth Factor/metabolism , Repressor Proteins/genetics , Repressor Proteins/metabolism , Thyroid Cancer, Papillary/genetics , Thyroid Cancer, Papillary/metabolism , Thyroid Neoplasms/genetics , Thyroid Neoplasms/metabolism , ETS Translocation Variant 6 ProteinABSTRACT
In the central nervous system, most neurons co-express TrkB and TrkC, the tyrosine kinase receptors for brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT3). As NT3 can also activate TrkB, it has been difficult to understand how NT3 and TrkC can exert unique roles in the assembly of neuronal circuits. Using neurons differentiated from human embryonic stem cells expressing both TrkB and TrkC, we compared Trk activation by BDNF and NT3. To avoid the complications resulting from TrkB activation by NT3, we also generated neurons from stem cells engineered to lack TrkB. We found that NT3 activates TrkC at concentrations lower than those of BDNF needed to activate TrkB. Downstream of Trk activation, the changes in gene expression caused by TrkC activation were found to be similar to those resulting from TrkB activation by BDNF, including a number of genes involved in synaptic plasticity. At high NT3 concentrations, receptor selectivity was lost as a result of TrkB activation. In addition, TrkC was down-regulated, as was also the case with TrkB at high BDNF concentrations. By contrast, receptor selectivity as well as reactivation were preserved when neurons were exposed to low neurotrophin concentrations. These results indicate that the selectivity of NT3/TrkC signalling can be explained by the ability of NT3 to activate TrkC at concentrations lower than those needed to activate TrkB. They also suggest that in a therapeutic perspective, the dosage of Trk receptor agonists will need to be taken into account if prolonged receptor activation is to be achieved.
Subject(s)
Brain-Derived Neurotrophic Factor , Membrane Glycoproteins/metabolism , Receptor, trkB/metabolism , Receptor, trkC/metabolism , Brain-Derived Neurotrophic Factor/metabolism , Down-Regulation , Humans , Neurons/metabolism , Neurotrophin 3/metabolism , Receptor Protein-Tyrosine Kinases/metabolism , Receptor, trkB/genetics , Receptor, trkC/genetics , Receptors, Nerve Growth Factor/genetics , Receptors, Nerve Growth Factor/metabolismABSTRACT
In a recent issue of Cell Reports, Morelli et al. (2021) identify a subpopulation of mechanosensitive peripheral sensory neurons that coexpress tyrosine hydroxylase (TH) and tropomyosin receptor kinase C (TrkC) and innervate cutaneous arterioles. They show that activation of TrkC sensory neurons causes cutaneous vasoconstriction and, most remarkably, that their lesion is associated with sudden death of an undetermined cause, preceded by a progressive drop in blood pressure, and conclude that TrkC+ TH+ neurons represent a baroreceptor class of homeostatic enteroceptor. This represents a radical departure from current consensus models for the central control of blood pressure. Here, we offer an alternative perspective on their findings and suggest priorities for further investigation. This Matters Arising paper is in response to Morelli et al. (2021), published in Cell Reports. See also the response by Heppenstall et al. (2022), published in this issue.
Subject(s)
Ganglia, Spinal , Receptor, trkC , Carrier Proteins , Ganglia, Spinal/metabolism , Receptor, trkC/metabolism , Sensory Receptor Cells/metabolism , Tyrosine 3-MonooxygenaseABSTRACT
Tropomyosin receptor kinases (TrkA, TrkB, and TrkC) are attractive therapeutic targets for multiple cancers. Two first-generation small-molecule Trks inhibitors, larotrectinib and entrectinib, have just been approved to use clinically. However, the drug-resistance mutations of Trks have already emerged, which calls for new-generation Trks inhibitors. Herein, we report the structural optimization and structure-activity relationship studies of 6,6-dimethyl-4-(phenylamino)-6H-pyrimido[5,4-b][1,4]oxazin-7(8H)-one derivatives as a new class of pan-Trk inhibitors. The prioritized compound 11g exhibited low nanomolar IC50 values against TrkA, TrkB, and TrkC and various drug-resistant mutants. It also showed good kinase selectivity. 11g displayed excellent in vitro antitumor activity and strongly suppressed Trk-mediated signaling pathways in intact cells. In in vivo studies, compound 11g exhibited good antitumor activity in BaF3-TEL-TrkA and BaF3-TEL-TrkCG623R allograft mouse models without exhibiting apparent toxicity. Collectively, 11g could be a promising lead compound for drug discovery targeting Trks and deserves further investigation.
Subject(s)
Oxazines/chemistry , Protein Kinase Inhibitors/chemistry , Receptor, trkA/antagonists & inhibitors , Receptor, trkB/antagonists & inhibitors , Receptor, trkC/antagonists & inhibitors , Animals , Antineoplastic Agents/chemistry , Antineoplastic Agents/metabolism , Antineoplastic Agents/pharmacology , Antineoplastic Agents/therapeutic use , Binding Sites , Cell Line, Tumor , Cell Proliferation/drug effects , Drug Design , Drug Resistance, Neoplasm/drug effects , Half-Life , Humans , Mice , Molecular Docking Simulation , Neoplasms/drug therapy , Neoplasms/pathology , Oxazines/metabolism , Oxazines/pharmacology , Oxazines/therapeutic use , Protein Kinase Inhibitors/metabolism , Protein Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/therapeutic use , Rats , Receptor, trkA/genetics , Receptor, trkA/metabolism , Receptor, trkB/genetics , Receptor, trkB/metabolism , Receptor, trkC/genetics , Receptor, trkC/metabolism , Signal Transduction/drug effects , Structure-Activity RelationshipABSTRACT
OBJECTIVE: The aim of the study was to investigate the mechanism by which p75 neurotrophin receptor (p75NTR) affects mitochondrial damage and neuronal apoptosis in spinal cord injury (SCI). METHODS: After the establishment of SCI rat models, short hairpin (sh) RNA of p75NTR and control sh-RNA were injected into SCI rats, respectively. On days 1, 7 and 21 after SCI, the severity of SCI and cell apoptosis in SCI rats were determined as well as the recovery of hind limb performance and p75NTR expression. After spinal cord neurons were transfected with p75NTR overexpression plasmid or empty plasmid vector or cotransfected with overexpression plasmids of p75NTR and neurotrophic tyrosine receptor kinase3 (NTRK3), the expression levels of p75NTR and NTRK3 were quantified. Moreover, we detected the apoptosis and proliferation rates of the neurons in addition to the levels of reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) in the neurons. The binding between p75NTR and NTRK3 was confirmed via Co-immunoprecipitation (Co-IP). RESULTS: The rat spinal cords in the Model group were notably damaged after SCI accompanied by increased apoptosis and decreased locomotor function. The expression of p75NTR was significantly upregulated after SCI. The aforementioned injuries were remarkably ameliorated in response to injection of sh-p75NTR. p75NTR overexpression induced mitochondrial damage and neuronal apoptosis in spinal cord neurons, while the promotive effects were perturbed by NTRK3 overexpression. Furthermore, p75NTR directly bound to and downregulated NTRK3. CONCLUSION: Both in vivo and in vitro experiments showed that p75NTR aggravates mitochondrial damage and neuronal apoptosis in SCI through downregulating NTRK3.
Subject(s)
Nerve Tissue Proteins/metabolism , Receptor, trkC/metabolism , Receptors, Growth Factor/metabolism , Spinal Cord Injuries , Animals , Apoptosis , Rats , Receptor, Nerve Growth Factor/genetics , Receptors, Amino Acid , Spinal Cord/metabolism , Spinal Cord Injuries/geneticsABSTRACT
Atypical Spitz tumors (AST) deviate from stereotypical Spitz nevi for one or more atypical features and are now regarded as an intermediate category of melanocytic tumors with uncertain malignant potential. Activating NTRK1/NTRK3 fusions elicit oncogenic events in Spitz lesions and are targetable with kinase inhibitors. However, their prevalence among ASTs and the optimal approach for their detection is yet to be determined. A series of 180 ASTs were screened with pan-TRK immunohistochemistry and the presence of NTRK fusions was confirmed using FISH, two different RNA-based NGS panels for solid tumors, and a specific real time RT-PCR panel. Overall, 26 ASTs showed pan-TRK immunostaining. NTRK1 fusions were detected in 15 of these cases showing cytoplasmic immunoreaction, whereas NTRK3 was detected in one case showing nuclear immunoreaction. Molecular tests resulted all positive in only two ASTs (included the NTRK3 translocated), RNA-based NGS and real time RT-PCR were both positive in three cases, and FISH and real time RT-PCR in another two cases. In seven ASTs NTRK1 fusions were detected only by FISH and in two cases only by real time RT-PCR. The frequency of NTRK fusions in ASTs is 9%, with a clear prevalence of NTRK1 compared to NTRK3 alterations. Pan-TRK immunohistochemistry is an excellent screening test. Confirmation of NTRK fusions may require the use of different molecular techniques.
Subject(s)
Nevus, Epithelioid and Spindle Cell/genetics , Nevus, Epithelioid and Spindle Cell/metabolism , Oncogene Fusion , Receptor, trkA/genetics , Receptor, trkA/metabolism , Receptor, trkC/genetics , Receptor, trkC/metabolism , Skin Neoplasms/genetics , Skin Neoplasms/metabolism , Adolescent , Adult , Child , Child, Preschool , Data Accuracy , Female , High-Throughput Nucleotide Sequencing/methods , Humans , Immunohistochemistry/methods , In Situ Hybridization, Fluorescence/methods , Male , Middle Aged , Oncogene Proteins, Fusion/genetics , Oncogene Proteins, Fusion/metabolism , Real-Time Polymerase Chain Reaction/methods , Sequence Analysis, RNA/methods , Young AdultABSTRACT
Tropomyosin receptor kinases (TRKA, TRKB, TRKC) are transmembrane receptor tyrosine kinases, which are respectively encoded by NTRK1, NTRK2, and NTRK3 genes. Herein, we reported the design, synthesis and Structure-Activity Relationship (SAR) investigation of a series of macrocyclic derivatives as new TRK inhibitors. Among these compounds, compound 9e exhibited strong kinase inhibitory activity (TRKG595R IC50 = 13.1 nM) and significant antiproliferative activity in the Ba/F3-LMNA-NTRK1 cell line (IC50 = 0.080 µM) and compound 9e has shown a better inhibitory effect (IC50 = 0.646 µM) than control drug LOXO-101 in Ba/F3-LMNA-NTRK1-G595R cell line. These results indicate that compound 9e is a potential TRK inhibitor for further investigation.
Subject(s)
Antineoplastic Agents/pharmacology , Drug Design , Macrocyclic Compounds/pharmacology , Membrane Glycoproteins/antagonists & inhibitors , Protein Kinase Inhibitors/pharmacology , Receptor, trkA/antagonists & inhibitors , Receptor, trkB/antagonists & inhibitors , Receptor, trkC/antagonists & inhibitors , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Cell Line, Tumor , Cell Proliferation/drug effects , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Humans , Macrocyclic Compounds/chemical synthesis , Macrocyclic Compounds/chemistry , Membrane Glycoproteins/metabolism , Molecular Structure , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/chemistry , Receptor, trkA/metabolism , Receptor, trkB/metabolism , Receptor, trkC/metabolism , Structure-Activity RelationshipABSTRACT
Neurotrophins (NTs) and their signal-transducing Trk receptors play a crucial role in the development and maintenance of specific neuronal subpopulations in nervous and sensory systems. NTs are supposed to regulate two sensory systems in fish, the inner ear and the lateral line system (LLS). The latter is one of the major mechanosensory systems in fish. Considering that annual fishes of the genus Nothobranchius, with their short life expectancy, have become a suitable model for aging studies and that the occurrence and distribution of neurotrophin Trk receptors have never been investigated in the inner ear and LLS of killifish (Nothobranchius guentheri), our study aimed to investigate the localization of neurotrophin-specific Trk receptors in mechanosensory systems of N. guentheri. For histological and immunohistochemical analysis, adult specimens of N. guentheri were processed using antibodies against Trk receptors and S100 protein. An intense immunoreaction for TrkA and TrkC was found in the sensory cells of the inner ear as well as in the hair cells of LLS. Moreover, also the neurons localized in the acoustic ganglia displayed a specific immunoreaction for all Trk receptors (TrkA, B, and C) analyzed. Taken together, our results demonstrate, for the first time, that neurotrophins and their specific receptors could play a pivotal role in the biology of the sensory cells of the inner ear and LLS of N. guentheri and might also be involved in the hair cells regeneration process in normal and aged conditions.