Selective, Intrinsically Fluorescent Trk Modulating Probes.

Neurotrophins (NTs) elicit the growth, survival, and differentiation of neurons and other neuroectoderm tissues via activation of Trk receptors. Hot spots for NT·Trk interactions involve three neurotrophin loops. Mimicry of these using "cyclo-organopeptides" comprising loop sequences cyclized onto endocyclic organic fragments accounts for a few of the low molecular mass Trk agonists or modulators reported so far; the majority are nonpeptidic small molecules accessed without molecular design and identified in random screens. It has proven difficult to verify activities induced by low molecular mass substances are due to Trk activation (rather than via other receptors), enhanced Trk expression, enhanced NT expression, or other pathways. Consequently, identification of selective probes for the various Trk receptors (e.g., A, B, and C) has been very challenging. Further, a key feature of probes for early stage assays is that they should be easily detectable, and none of the compounds reported to date are. In this work, we designed novel cyclo-organopeptide derivatives where the organic fragment is a BODIPY fluor and found ones that selectively, though not specifically, activate TrkA, B, or C. One of the assays used to reach this conclusion (binding to live Trk-expressing cells) relied on intrinsic fluorescence in the tested materials. Consequently, this work established low molecular mass Trk-selective probes exhibiting neuroprotective effects.

Withdrawal pain following patients discontinuing Trk inhibitors.

OBJECTIVE: This article aims to expand on the existing literature regarding the incidence of withdrawal pain following discontinuation of Trk inhibitors and to explore strategies that mitigate this withdrawal pain. DATA SOURCE: A retrospective observational study was conducted among patients who were at least 18 years-old or older and had documentation of starting larotrectinib or entrectinib at University of California, San Francisco (UCSF) between November 2018 and November 2022. Data were collected from electronic records and pharmacy databases and a total of 21 patients were identified in this study. DATA SUMMARY: Of the 21 patients included in this study, five patients (24%) experienced pain during temporary or permanent discontinuation of Trk inhibitor with the onset of withdrawal pain ranging from a few hours to three days following discontinuation. Various strategies were implemented to manage this pain including restarting of Trk inhibitor, tapering of Trk inhibitor on discontinuation, minimizing dose interruptions and use of prescription pain medications. CONCLUSION: This article illustrates the presence of withdrawal pain syndrome in patients stopping a Trk inhibitor treatment and highlight the need for patient education to avoid missing any doses and for development of a guideline for Trk inhibitor discontinuation.

Pyrazolo[1,5-a]pyrimidine as a Prominent Framework for Tropomyosin Receptor Kinase (Trk) Inhibitors-Synthetic Strategies and SAR Insights.

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.

The Role of the Neurotrophin Network in Skin Squamous Cell Cancer and the Novel Use of the Zebrafish System.

Cutaneous squamous cell carcinoma (cSCC) is the second most prevalent form of skin cancer. An increasing number of cSCCs are associated with dysregulation of key molecules that control skin homeostasis. These observations have increased interest in the role of neurotrophins and their receptors in the pathogenesis of cSCC. They have been demonstrated to have a considerable impact on the aggressiveness potential of skin cancer by both in vitro and in vivo models. In this context, mouse models are classically used to dissect proliferation versus differentiation balance, but they have some limitations in terms of time, space, and costs. Recently, zebrafish models have been implemented as a new tool to obtain information regarding the invasive capacity and metastasis of neoplastic cells. By xenotransplantation technique, cSCC cells from a patient's biopsy or cell line can be successfully characterized, with or without the presence of genetic manipulation or treatments. In addition, the evaluation of the immune microenvironment contributes to potentially identifying connections and homologies with humans. In this review, we retrace the role of the neurotrophin network in healthy and pathological skin, particularly in cSCC. We review how zebrafish models can be important tools for studying cSCC development, growth, and potential treatments.

Trackins (Trk-Targeting Drugs): A Novel Therapy for Different Diseases.

Many routes may lead to the transition from a healthy to a diseased phenotype. However, there are not so many routes to travel in the opposite direction; that is, therapy for different diseases. The following pressing question thus remains: what are the pathogenic routes and how can be they counteracted for therapeutic purposes? Human cells contain >500 protein kinases and nearly 200 protein phosphatases, acting on thousands of proteins, including cell growth factors. We herein discuss neurotrophins with pathogenic or metabotrophic abilities, particularly brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), pro-NGF, neurotrophin-3 (NT-3), and their receptor Trk (tyrosine receptor kinase; pronounced "track"). Indeed, we introduced the word trackins, standing for Trk-targeting drugs, that play an agonistic or antagonistic role in the function of TrkBBDNF, TrkCNT-3, TrkANGF, and TrkApro-NGF receptors. Based on our own published results, supported by those of other authors, we aim to update and enlarge our trackins concept, focusing on (1) agonistic trackins as possible drugs for (1a) neurotrophin-deficiency cardiometabolic disorders (hypertension, atherosclerosis, type 2 diabetes mellitus, metabolic syndrome, obesity, diabetic erectile dysfunction and atrial fibrillation) and (1b) neurodegenerative diseases (Alzheimer's disease, Parkinson's disease, and multiple sclerosis), and (2) antagonistic trackins, particularly TrkANGF inhibitors for prostate and breast cancer, pain, and arrhythmogenic right-ventricular dysplasia. Altogether, the druggability of TrkANGF, TrkApro-NGF, TrkBBDNF, and TrkCNT-3 receptors via trackins requires a further translational pursuit. This could provide rewards for our patients.

Distinct regions of its first intracellular loop contribute to the proper localization, transport activity and substrate-affinity adjustment of the main yeast K+ importer Trk1.

Trk1 is the main K+ importer of Saccharomyces cerevisiae. Its proper functioning enables yeast cells to grow in environments with micromolar amounts of K+. Although the structure of Trk1 has not been experimentally determined, the transporter is predicted to be composed of four MPM (transmembrane segment - pore loop - transmembrane segment) motifs which are connected by intracellular loops. Of those, in particular the first loop (IL1) is unique in its length; it forms more than half of the entire protein. The deletion of the majority of IL1 does not abolish the transport activity of Trk1. However IL1 is thought to be involved in the modulation of the transporter's functioning. In this work, we prepared a series of internally shortened versions of Trk1 that lacked various parts of IL1, and we studied their properties in S. cerevisiae cells without chromosomal copies of TRK genes. Using this approach, we were able to determine that both N- and C-border regions of IL1 are necessary for the proper localization of Trk1. Moreover, the N-border part of IL1 is also important for the functioning of Trk1, as its absence resulted in a decrease in the transporter's substrate affinity. In addition, in the internal part of IL1, we newly identified a stretch of amino-acid residues that are indispensable for retaining the transporter's maximum velocity, and another region whose deletion affected the ability of Trk1 to adjust its affinity in response to external levels of K+.

NTRK-fused central nervous system tumours: clinicopathological and genetic insights and response to TRK inhibitors.

Background Neurotrophic tropomyosin receptor kinase (NTRK) gene fusions are found in 1% of gliomas across children and adults. TRK inhibitors are promising therapeutic agents for NTRK-fused gliomas because they are tissue agnostic and cross the blood-brain barrier (BBB). Methods We investigated twelve NGS-verified NTRK-fused gliomas from a single institute, Seoul National University Hospital. Results The patient cohort included six children (aged 1-15 years) and six adults (aged 27-72 years). NTRK2 fusions were found in ten cerebral diffuse low-grade and high-grade gliomas (DLGGs and DHGGs, respectively), and NTRK1 fusions were found in one cerebral desmoplastic infantile ganglioglioma and one spinal DHGG. In this series, the fusion partners of NTRK2 were HOOK3, KIF5A, GKAP1, LHFPL3, SLMAP, ZBTB43, SPECC1L, FKBP15, KANK1, and BCR, while the NTRK1 fusion partners were TPR and TPM3. DLGGs tended to harbour only an NTRK fusion, while DHGGs exhibited further genetic alterations, such as TERT promoter/TP53/PTEN mutation, CDKN2A/2B homozygous deletion, PDGFRA/KIT/MDM4/AKT3 amplification, or multiple chromosomal copy number aberrations. Four patients received adjuvant TRK inhibitor therapy (larotrectinib, repotrectinib, or entrectinib), among which three also received chemotherapy (n = 2) or proton therapy (n = 1). The treatment outcomes for patients receiving TRK inhibitors varied: one child who received larotrectinib for residual DLGG maintained stable disease. In contrast, another child with DHGG in the spinal cord experienced multiple instances of tumour recurrence. Despite treatment with larotrectinib, ultimately, the child died as a result of tumour progression. An adult patient with glioblastoma (GBM) treated with entrectinib also experienced tumour progression and eventually died. However, there was a successful outcome for a paediatric patient with DHGG who, after a second gross total tumour removal followed by repotrectinib treatment, showed no evidence of disease. This patient had previously experienced relapse after the initial surgery and underwent autologous peripheral blood stem cell therapy with carboplatin/thiotepa and proton therapy. Conclusions Our study clarifies the distinct differences in the pathology and TRK inhibitor response between LGG and HGG with NTRK fusions.

Discovery of novel indazole derivatives as second-generation TRK inhibitors.

NTRK gene fusion leads to the activation of downstream signaling pathways, which is a oncogenic driver in various cancers. NTRK fusion-positive cancers can be treated with the first-generation TRK inhibitors, larotrectinib and entrectinib. Unfortunately, the patients eventually face the dilemma of no drugs available as the emergence of certain resistance mutations. The development of efficient and broad-spectrum second-generation TRK inhibitors is still of great significance. Here, we analyzed the binding modes of compounds 6, 10 with TRKA protein, respectively, a series of novel indazole TRK inhibitors were designed and synthesized using molecular hybridization strategy. Among them, the optimal compound B31 showed strong antiproliferative activities against Km-12, Ba/F3-TRKAG595R, and Ba/F3-TRKAG667C cell lines with IC50 values of 0.3, 4.7, and 9.9 nM, respectively. And the inhibitory effect against TRKAG667C (IC50 = 9.9 nM) was better than that of selitrectinib (IC50 = 113.1 nM). Further, compound B31 exhibited moderate kinase selectivity and excellent plasma stability (t1/2 > 480 min). In vivo pharmacokinetic studies in Sprague-Dawley rats showed that B31 had acceptable pharmacokinetic properties.

Plain language summary of a clinical study: anamorelin for treatment of Japanese people with gastrointestinal cancer and cachexia.

What is this summary about? This is a plain language summary of a clinical research study. The study investigated the effects of a drug called anamorelin for people in Japan with advanced gastrointestinal cancer (colon, rectum, stomach or pancreas) who also had a condition called cachexia.People with cachexia have a loss of appetite, severe weight loss, loss of body fat, and loss of muscle. Anamorelin has been shown in previous research studies to improve appetite and increase lean body mass (the total weight of the body, not counting fat) and overall body weight. In this study, participants were given anamorelin (100 mg daily) for 12 weeks.What did the researchers find out? 31 out of 49 participants (63%) had an increase in their lean body mass, with an average increase of almost 2 kg, which was seen from week 3 of the study. Participants also experienced an improvement in their appetite and overall body weight. The researchers found that anamorelin did not cause a large number of serious side effects.What do the results mean? This study found that anamorelin can help people with advanced gastrointestinal cancer and cachexia to increase their lean body mass and overall body weight, regain their appetite and improve their nutritional status. Since this study was done, the use of anamorelin for cachexia in people with gastrointestinal cancer has been approved for use in Japan.

Effects of oral administration of nonselective Trk inhibitor on bladder overactivity in rodent models of prostatic inflammation.

BACKGROUND: Our research focused on the assessment of the impact of systemic inhibition of Trk receptors, which bind to nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF), on bladder hypersensitivity in two distinct rodent models of prostatic inflammation (PI). METHODS: Male Sprague-Dawley rats were divided into three groups (n = 6 each): the control group (no PI, vehicle administration), the untreated group (PI, vehicle administration), and the treated group (PI, nonselective Trk inhibitor, GNF 5837, administration). PI in rats was induced by a intraprostatic injection of 5% formalin. Posttreatment, we carried out conscious cystometry and a range of histological and molecular analyses. Moreover, the study additionally evaluated the effects of a nonselective Trk inhibitor on bladder overactivity in a mouse model of PI, which was induced by prostate epithelium-specific conditional deletion of E-cadherin. RESULTS: The rat model of PI showed upregulations of NGF and BDNF in both bladder and prostate tissues in association with bladder overactivity and inflammation in the ventral lobes of the prostate. GNF 5837 treatment effectively mitigated these PI-induced changes, along with reductions in TrkA, TrkB, TrkC, and TRPV1 mRNA expressions in L6-S1 dorsal root ganglia. Also, in the mouse PI model, GNF 5837 treatment similarly improved bladder overactivity. CONCLUSIONS: The findings of our study suggest that Trk receptor inhibition, which reduced bladder hypersensitivity and inflammatory responses in the prostate, along with a decrease in overexpression of Trk and TRPV1 receptors in sensory pathways, could be an effective treatment strategy for male lower urinary tract symptoms associated with PI and bladder overactivity.

CD34/S100 protein-positive, NTRK1-rearranged infantile fibrosarcoma-like tumors in genitourinary system: two cases expanding the clinicopathologic spectrum and illustrating the diagnostic dilemma.

Infantile fibrosarcoma (IFS) is malignant fibroblastic tumor of infants characterized genetically by ETV6::NTRK3 fusion. Tumors that show morphology indistinguishable from IFS but harbor alternative genetic alterations are uncommon, which have been designated as IFS-like tumors. We report two cases of IFS-like tumors harboring an NTRK1 rearrangement and arsing from genitourinary system. The patients aged 3 and 14 years. One arose in the kidney and one in the paratesticular region. The tumors measured 13 and 3.5 cm in greatest dimension. Both tumors were composed of cellular, mildly atypical, spindle to ovoid cells arranged haphazardly or in intersecting fascicles within a collagenized to myxoid stroma. Mitoses numbered 3 and 5/10 high-power fields. Tumor cells in both neoplasms demonstrated variable co-expression of CD34 and S100 protein, and diffuse and strong cytoplasmic staining for pan-TRK and TrkA. Fluorescence in-situ hybridization demonstrated NTRK1 rearrangement in both tumors. Targeted RNA-sequencing identified CPSF6::NTRK1 fusion and TMP3::NTRK1 fusion. Limited follow-up showed no tumor recurrences or metastases. We expand the clinicopathologic spectrum of IFS-like tumors harboring alternative NTRK1 fusions.

Type II & III inhibitors of tropomyosin receptor kinase (Trk): a 2020-2022 patent update.

INTRODUCTION: The Trk family proteins are membrane-bound kinases predominantly expressed in neuronal tissues. Activated by neurotrophins, they regulate critical cellular processes through downstream signaling pathways. Dysregulation of Trk signaling can drive a range of diseases, making the design and study of Trk inhibitors a vital area of research. This review explores recent advances in the development of type II and III Trk inhibitors, with implications for various therapeutic applications. AREAS COVERED: Patents covering type II and III inhibitors targeting the Trk family are discussed as a complement of the previous review, Type I inhibitors of tropomyosin receptor kinase (Trk): a 2020-2022 patent update. Relevant patents were identified using the Web of Science database, Google, and Google Patents. EXPERT OPINION: While type II and III Trk inhibitor development has advanced more gradually compared to their type I counterparts, they hold significant promise in overcoming resistance mutations and achieving enhanced subtype selectivity - a critical factor in reducing adverse effects associated with pan-Trk inhibition. Recent interdisciplinary endeavors have marked substantial progress in the design of subtype selective Trk inhibitors, with impressive success heralded by the type III inhibitors. Notably, the emergence of mutant-selective Trk inhibitors introduces an intriguing dimension to the field, offering precise treatment possibilities.

NTRK gene fusion testing and management in lung cancer.

Neurotrophic tyrosine receptor kinase (NTRK) gene fusions are recurrent oncogenic drivers found in a variety of solid tumours, including lung cancer. Several tropomyosin receptor kinase (TRK) inhibitors have been developed to treat tumours with NTRK gene fusions. Larotrectinib and entrectinib are first-generation TRK inhibitors that have demonstrated efficacy in patients with TRK fusion lung cancers. Genomic testing is recommended for all patients with metastatic non-small cell lung cancer for optimal drug therapy selection. Multiple testing methods can be employed to identify NTRK gene fusions in the clinic and each has its own advantages and limitations. Among these assays, RNA-based next-generation sequencing (NGS) can be considered a gold standard for detecting NTRK gene fusions; however, several alternatives with minimally acceptable sensitivity and specificity are also available in areas where widespread access to NGS is unfeasible. This review highlights the importance of testing for NTRK gene fusions in lung cancer, ideally using the gold-standard method of RNA-based NGS, the various assays that are available, and treatment algorithms for patients.

Discovery of 9H-pyrimido[4,5-b]indole derivatives as dual RET/TRKA inhibitors.

Aberrant RET kinase signaling is activated in numerous cancers including lung, thyroid, breast, pancreatic, and prostate. Recent approvals of selective RET inhibitors, pralsetinib and selpercatinib, has shifted the focus of RET kinase drug discovery programs towards the development of selective inhibitors. However, selective inhibitors invariably lose efficacy as the selective nature of the inhibitor places Darwinian-like pressure on the tumor to bypass treatment through the selection of novel oncogenic drivers. Further, selective inhibitors are restricted for use in tumors with specific genetic backgrounds that do not encompass diverse patient classes. Here we report the identification of a pyrimido indole RET inhibitor found to also have activity against TRK. This selective dual RET/TRK inhibitor can be utilized in tumors with both RET and TRK genetic backgrounds and can also provide blockade of NTRK-fusions that are selected for from RET inhibitor treatments. Efforts towards developing dual RET/TRK inhibitors can be beneficial in terms of encompassing more diverse patient classes while also achieving blockade against emerging resistance mechanisms.

Acquired NF2 mutation confers resistance to TRK inhibition in an ex vivo LMNA::NTRK1-rearranged soft-tissue sarcoma cell model.

Genomic rearrangements of the neurotrophic receptor tyrosine kinase genes (NTRK1, NTRK2, and NTRK3) are the most common mechanism of oncogenic activation for this family of receptors, resulting in sustained cancer cell proliferation. Several targeted therapies have been approved for tumours harbouring NTRK fusions and a new generation of TRK inhibitors has already been developed due to acquired resistance. We established a patient-derived LMNA::NTRK1-rearranged soft-tissue sarcoma cell model ex vivo with an acquired resistance to targeted TRK inhibition. Molecular profiling of the resistant clones revealed an acquired NF2 loss of function mutation that was absent in the parental cell model. Parental cells showed continuous sensitivity to TRK-targeted treatment, whereas the resistant clones were insensitive. Furthermore, resistant clones showed upregulation of the MAPK and mTOR/AKT pathways in the gene expression based on RNA sequencing data and increased sensitivity to MEK and mTOR inhibitor therapy. Drug synergy was seen using trametinib and rapamycin in combination with entrectinib. Medium-throughput drug screening further identified small compounds as potential drug candidates to overcome resistance as monotherapy or in combination with entrectinib. In summary, we developed a comprehensive model of drug resistance in an LMNA::NTRK1-rearranged soft-tissue sarcoma and have broadened the understanding of acquired drug resistance to targeted TRK therapy. Furthermore, we identified drug combinations and small compounds to overcome acquired drug resistance and potentially guide patient care in a functional precision oncology setting. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

NTRK3 exhibits a pro-oncogenic function in upper tract urothelial carcinomas.

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.

Therapeutic benefit of larotrectinib over the historical standard of care in patients with locally advanced or metastatic infantile fibrosarcoma (EPI VITRAKVI study).

BACKGROUND: Patients with infantile fibrosarcoma (IFS) have shown strong and long-lasting responses to larotrectinib, a tropomyosin receptor kinase inhibitor (TRKi), in single-arm clinical trials. Conventional chemotherapy has also shown important efficacy. But, until now, no comparative data exist. This study aims to assess the therapeutic benefit of larotrectinib over the current standard of care (SOC) of chemotherapy in paediatric patients with locally advanced or metastatic IFS. PATIENTS AND METHODS: EPI VITRAKVI is a retrospective, observational, externally controlled study (NCT05236257). Data of patients aged ≤21 years with locally advanced or metastatic IFS treated with larotrectinib in the phase I/II SCOUT trial (NCT02637687) were compared with those of an external historical control group (data of Institut Curie and Cooperative Weichteilsarkom Studiengruppe) treated with a chemotherapy-based regimen. Between-group differences were assessed after balancing groups using inverse probability of treatment weighting (IPTW). RESULTS: In total, 93 patients were compared, 51 in the larotrectinib arm and 42 in the external control arm. After therapy, 4 patients (7.8%) in the larotrectinib group had a medical treatment failure event [start of new systemic treatment (2 cases), mutilating surgery (2 cases)] versus 15 (35.7%) in the external control group [start of new systemic treatment (6 cases), mutilating surgery (5 cases), radiation therapy (2 cases), and death (2 cases)]. Larotrectinib was associated with an 80% reduced likelihood of encountering a medical treatment failure event, when compared to the external control group (weighted and stratified hazard ratio 0.20, 95% confidence interval 0.06-0.63, P = 0.0060). These results were confirmed by sensitivity analyses, including exact matching, and subgroup analyses for number of lines of treatment. CONCLUSIONS: Treatment with larotrectinib reduced the need of subsequent therapies compared to SOC with chemotherapy in children with locally advanced or metastatic IFS, regardless of the line of treatment.

Boosting BDNF in muscle rescues impaired axonal transport in a mouse model of DI-CMTC peripheral neuropathy.

Charcot-Marie-Tooth disease (CMT) is a genetic peripheral neuropathy caused by mutations in many functionally diverse genes. The aminoacyl-tRNA synthetase (ARS) enzymes, which transfer amino acids to partner tRNAs for protein synthesis, represent the largest protein family genetically linked to CMT aetiology, suggesting pathomechanistic commonalities. Dominant intermediate CMT type C (DI-CMTC) is caused by YARS1 mutations driving a toxic gain-of-function in the encoded tyrosyl-tRNA synthetase (TyrRS), which is mediated by exposure of consensus neomorphic surfaces through conformational changes of the mutant protein. In this study, we first showed that human DI-CMTC-causing TyrRSE196K mis-interacts with the extracellular domain of the BDNF receptor TrkB, an aberrant association we have previously characterised for several mutant glycyl-tRNA synthetases linked to CMT type 2D (CMT2D). We then performed temporal neuromuscular assessments of YarsE196K mice modelling DI-CMT. We determined that YarsE196K homozygotes display a selective, age-dependent impairment in in vivo axonal transport of neurotrophin-containing signalling endosomes, phenocopying CMT2D mice. This impairment is replicated by injection of recombinant TyrRSE196K, but not TyrRSWT, into muscles of wild-type mice. Augmenting BDNF in DI-CMTC muscles, through injection of recombinant protein or muscle-specific gene therapy, resulted in complete axonal transport correction. Therefore, this work identifies a non-cell autonomous pathomechanism common to ARS-related neuropathies, and highlights the potential of boosting BDNF levels in muscles as a therapeutic strategy.

Trk-fused gene plays a critical role in diet-induced adipose tissue expansion and is also involved in thyroid hormone action.

Mutations in the Trk-fused gene (TFG) cause hereditary motor and sensory neuropathy with proximal dominant involvement, which reportedly has high co-incidences with diabetes and dyslipidemia, suggesting critical roles of the TFG in metabolism as well. We found that TFG expression levels in white adipose tissues (WATs) were elevated in both genetically and diet-induced obese mice and that TFG deletion in preadipocytes from the stromal vascular fraction (SVF) markedly inhibited adipogenesis. To investigate its role in vivo, we generated tamoxifen-inducible adipocyte-specific TFG knockout (AiTFG KO) mice. While a marked down-regulation of the peroxisome proliferator-activated receptor gamma target, de novo lipogenesis (DNL), and mitochondria-related gene expressions were observed in subcutaneous WAT (scWAT) from AiTFG KO mice, these effects were blunted in SVF-derived adipocytes when the TFG was deleted after differentiation into adipocytes, implying cell nonautonomous effects. Intriguingly, expressions of thyroid hormone receptors, as well as carbohydrate responsive element-binding protein ß, which mediates the metabolic actions of thyroid hormone, were drastically down-regulated in scWAT from AiTFG KO mice. Reduced DNL and thermogenic gene expressions in AiTFG KO mice might be attributable to impaired thyroid hormone action in vivo. Finally, when adipocyte TFG was deleted in either the early or the late phase of high-fat diet feeding, the former brought about an impaired expansion of epididymal WAT, whereas the latter caused prominent adipocyte cell death. TFG deletion in adipocytes markedly exacerbated hepatic steatosis in both experimental settings. Collectively, these observations indicate that the TFG plays essential roles in maintaining normal adipocyte functions, including an enlargement of adipose tissue, thyroid hormone function, and thermogenic gene expressions, and in preserving hypertrophic adipocytes.