The binding process of insulin to its transmembrane receptor entails a sophisticated interplay between two proteins, each possessing two binding sites. Given the difficulties associated with the use of insulin in the treatment of diabetes, despite its remarkable efficacy, there is interest in smaller and more stable compounds than the native hormone that would effectively activate the receptor. Our study adopts a strategy focused on synthesizing extensive combinatorial libraries of bipodal compounds consisting of two distinct peptides linked to a molecular scaffold. These constructs, evaluated in a resin bead-bound format, were designed to assess their binding to the insulin receptor. Despite notable nonspecific binding, our approach successfully generated and tested millions of compounds. Rigorous evaluations via flow cytometry and specific antibodies revealed peptide sequences with specific interactions at either receptor binding Site 1 or 2. Notably, these sequences bear similarity to peptides discovered through phage display by other researchers. This convergence of chemical and biological methods underscores nature's beauty, revealing general principles in peptide binding to the insulin receptor. Overall, our study deepens the understanding of molecular interactions in ligand binding to the insulin receptor, highlighting the challenges of targeting large proteins with small synthetic peptides.
- Klíčová slova
- insulin, library, peptide, receptor, scaffold,
- MeSH
- inzulin metabolismus chemie MeSH
- knihovny malých molekul chemie farmakologie chemická syntéza MeSH
- lidé MeSH
- ligandy MeSH
- molekulární struktura MeSH
- peptidová knihovna MeSH
- peptidy chemie metabolismus chemická syntéza MeSH
- receptor inzulinu * metabolismus chemie MeSH
- techniky kombinatorické chemie * MeSH
- vazba proteinů MeSH
- vazebná místa MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- inzulin MeSH
- knihovny malých molekul MeSH
- ligandy MeSH
- peptidová knihovna MeSH
- peptidy MeSH
- receptor inzulinu * MeSH
BACKGROUND: Amplification of HER2, a receptor tyrosine kinase and a breast cancer-linked oncogene, is associated with aggressive disease. HER2 protein is localised mostly at the cell membrane, but a fraction translocates to mitochondria. Whether and how mitochondrial HER2 contributes to tumorigenicity is currently unknown. METHODS: We enriched the mitochondrial (mt-)HER2 fraction in breast cancer cells using an N-terminal mitochondrial targeting sequence and analysed how this manipulation impacts bioenergetics and tumorigenic properties. The role of the tyrosine kinase activity of mt-HER2 was assessed in wild type, kinase-dead (K753M) and kinase-enhanced (V659E) mtHER2 constructs. RESULTS: We document that mt-HER2 associates with the oxidative phosphorylation system, stimulates bioenergetics and promotes larger respiratory supercomplexes. mt-HER2 enhances proliferation and invasiveness in vitro and tumour growth and metastatic potential in vivo, in a kinase activity-dependent manner. On the other hand, constitutively active mt-HER2 provokes excessive mitochondria ROS generation, sensitises to cell death, and restricts growth of primary tumours, suggesting that regulation of HER2 activity in mitochondria is required for the maximal pro-tumorigenic effect. CONCLUSIONS: mt-HER2 promotes tumorigenicity by supporting bioenergetics and optimal redox balance.
- Klíčová slova
- HER2, cancer, electron transport chain, mitochondria, reactive oxygen species,
- MeSH
- buněčné dýchání fyziologie MeSH
- energetický metabolismus MeSH
- karcinogeneze metabolismus MeSH
- lidé MeSH
- mitochondrie * metabolismus MeSH
- myši MeSH
- nádorové buněčné linie MeSH
- nádory prsu * metabolismus genetika MeSH
- oxidativní fosforylace MeSH
- proliferace buněk MeSH
- reaktivní formy kyslíku metabolismus MeSH
- receptor erbB-2 * metabolismus MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
INTRODUCTION: This study aimed to identify whether microbial invasion of the amniotic cavity and/or intra-amniotic inflammation in women with late preterm prelabor rupture of membranes (PPROM) was associated with changes in concentrations of soluble fms-like tyrosine kinase-1 (sFlt-1), placental growth factor (PlGF) and its ratio in maternal serum, and whether placental features consistent with maternal vascular malperfusion further affect their concentrations. MATERIAL AND METHODS: This historical study included 154 women with singleton pregnancies complicated by PPROM between gestational ages 34+0 and 36+6 weeks. Transabdominal amniocentesis was performed as part of standard clinical management to evaluate the intra-amniotic environment. Women were categorized into two subgroups based on the presence of microorganisms and/or their nucleic acids in amniotic fluid (determined by culturing and molecular biology method) and intra-amniotic inflammation (by amniotic fluid interleukin-6 concentration evaluation): (1) those with the presence of microorganisms and/or inflammation (at least one present) and (2) those with negative amniotic fluid for infection/inflammation (absence of both). Concentrations of sFlt-1 and PlGF were assessed using the Elecsys® sFlt-1 and Elecsys® PlGF immunoassays and converted into multiples of medians. RESULTS: Women with the presence of microorganisms and/or inflammation in amniotic fluid had lower serum concentrations of sFlt-1 and sFlt-1/PlGF ratios and higher concentrations of PlGF compared with those with negative amniotic fluid. (sFlt-1: presence: median 1.0 multiples of the median (MoM), vs negative: median: 1.5 MoM, P = 0.003; PlGF: presence: median 0.7 MoM, vs negative: median 0.4 MoM, P = 0.02; sFlt-1/PlGF: presence: median 8.9 vs negative 25.0, P = 0.001). Higher serum concentrations of sFlt-1 and sFlt-1/PlGF ratios as well as lower concentrations of PlGF were found in the subsets of women with maternal vascular malperfusion than in those without maternal vascular malperfusion. CONCLUSIONS: Among women experiencing late PPROM, angiogenic imbalance in maternal serum is primarily observed in those without both microbial invasion of the amniotic cavity and intra-amniotic inflammation. Additionally, there is an association between angiogenic imbalance and the presence of maternal vascular malperfusion.
- Klíčová slova
- amniotic fluid, angiogenic factors, inflammation, microorganism, preterm delivery,
- MeSH
- amniocentéza MeSH
- biologické markery krev MeSH
- chorioamnionitida krev MeSH
- dospělí MeSH
- gestační stáří MeSH
- lidé MeSH
- placentární růstový faktor * krev MeSH
- plodová voda * mikrobiologie metabolismus MeSH
- předčasný odtok plodové vody * krev MeSH
- receptor 1 pro vaskulární endoteliální růstový faktor * krev MeSH
- těhotenství MeSH
- Check Tag
- dospělí MeSH
- lidé MeSH
- těhotenství MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- FLT1 protein, human MeSH Prohlížeč
- PGF protein, human MeSH Prohlížeč
Covalent labeling in combination with mass spectrometry is a powerful approach used in structural biology to study protein structures, interactions, and dynamics. Recently, the toolbox of covalent labeling techniques has been expanded with fast fluoroalkylation of proteins (FFAP). FFAP is a novel radical labeling method that utilizes fluoroalkyl radicals generated from hypervalent Togni reagents for targeting aromatic residues. This report further demonstrates the benefits of FFAP as a new method for structural characterization of therapeutic antibodies and interaction interfaces of antigen-antibody complexes. The results obtained from human trastuzumab and its complex with human epidermal growth factor receptor 2 (HER2) correlate well with previously published structural data and demonstrate the potential of FFAP in structural biology.
- MeSH
- alkylace MeSH
- footprinting proteinů metody MeSH
- halogenace MeSH
- imunokomplex chemie MeSH
- lidé MeSH
- mapování epitopu * metody MeSH
- monoklonální protilátky chemie imunologie MeSH
- receptor erbB-2 * chemie imunologie MeSH
- trastuzumab * chemie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Frequency, distribution and prognostic meaning of ALK-partner genes other than NPM1 in ALK-positive anaplastic large-cell lymphoma (ALCL) are unknown. Forty-nine of 316 ALCL diagnosed in the NHL-BFM study group showed no nuclear ALK expression suggestive of a variant ALK-partner; 41 were analysed by genomic capture high-throughput sequencing or specific RT-PCRs. NPM1::ALK was detected in 13 cases. Among the 28 patients with a non-NPM1::ALK-fusion partner, ATIC (n = 8; 29%) and TPM3 (n = 9; 32%) were the most common. Five of eight patients with ATIC::ALK-positive ALCL relapsed, none of nine with TPM3::ALK. Variant ALK-partners are rare and potentially associated with different prognoses.
- Klíčová slova
- ALK positive ALCL, non‐Hodgkin's lymphoma, variant ALK,
- MeSH
- anaplastická lymfomová kináza * genetika analýza MeSH
- anaplastický velkobuněčný lymfom * genetika patologie MeSH
- dítě MeSH
- fúzní onkogenní proteiny genetika MeSH
- jaderné proteiny genetika metabolismus MeSH
- kojenec MeSH
- lidé MeSH
- mladiství MeSH
- nukleofosmin * MeSH
- předškolní dítě MeSH
- prognóza MeSH
- tropomyosin MeSH
- Check Tag
- dítě MeSH
- kojenec MeSH
- lidé MeSH
- mladiství MeSH
- mužské pohlaví MeSH
- předškolní dítě MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- ALK protein, human MeSH Prohlížeč
- anaplastická lymfomová kináza * MeSH
- fúzní onkogenní proteiny MeSH
- jaderné proteiny MeSH
- NPM1 protein, human MeSH Prohlížeč
- nukleofosmin * MeSH
- TPM3 protein, human MeSH Prohlížeč
- tropomyosin MeSH
Alterations in kinase genes such as NTRK1/2/3, RET, and BRAF underlie infantile fibrosarcoma (IFS), the emerging entity 'NTRK-rearranged spindle cell neoplasms' included in the latest WHO classification, and a growing set of tumors with overlapping clinical and pathological features. In this study, we conducted a comprehensive clinicopathological and molecular analysis of 22 cases of IFS and other kinase gene-altered spindle cell neoplasms affecting both pediatric and adult patients. Follow-up periods for 16 patients ranged in length from 10 to 130 months (mean 38 months). Six patients were treated with targeted therapy, achieving a partial or complete response in five cases. Overall, three cases recurred and one metastasized. Eight patients were free of disease, five were alive with disease, and two patients died. All cases showed previously reported morphological patterns. Based on the cellularity and level of atypia, cases were divided into three morphological grade groups. S100 protein and CD34 were at least focally positive in 12/22 and 14/22 cases, respectively. Novel PWWP2A::RET, NUMA1::RET, ITSN1::RAF1, and CAPZA2::MET fusions, which we report herein in mesenchymal tumors for the first time, were detected by RNA sequencing. Additionally, the first uterine case with BRAF and EGFR mutations and CD34 and S100 co-expression is described. DNA sequencing performed in 13 cases uncovered very rare additional genetic aberrations. The CNV profiles showed that high-grade tumors demonstrate a significantly higher percentage of copy number gains and losses across the genome compared with low- and intermediate-grade tumors. Unsupervised clustering of the tumors' methylation profiles revealed that in 8/9 cases, the methylation profiles clustered with the IFS methylation class, irrespective of their clinicopathological or molecular features. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.
- Klíčová slova
- BRAF mutation, CDKN2A, NTRK‐rearranged spindle cell neoplasms, entrectinib, infantile fibrosarcoma, kinase gene fusion, larotrectinib, methylation profiling, selpercatinib,
- MeSH
- dítě MeSH
- dospělí MeSH
- fibrosarkom * genetika patologie MeSH
- fúzní onkogenní proteiny genetika MeSH
- lidé MeSH
- lokální recidiva nádoru genetika MeSH
- nádorové biomarkery genetika analýza MeSH
- nádory měkkých tkání * genetika MeSH
- nádory z pojivové a měkké tkáně * MeSH
- protoonkogenní proteiny B-Raf genetika MeSH
- receptor trkA genetika MeSH
- Check Tag
- dítě MeSH
- dospělí MeSH
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- fúzní onkogenní proteiny MeSH
- nádorové biomarkery MeSH
- protoonkogenní proteiny B-Raf MeSH
- receptor trkA MeSH
Targeting Anaplastic lymphoma kinase (ALK) is a promising therapeutic strategy for aberrant ALK-expressing malignancies including neuroblastoma, but resistance to ALK tyrosine kinase inhibitors (ALK TKI) is a distinct possibility necessitating drug combination therapeutic approaches. Using high-throughput, genome-wide CRISPR-Cas9 knockout screens, we identify miR-1304-5p loss as a desensitizer to ALK TKIs in aberrant ALK-expressing neuroblastoma; inhibition of miR-1304-5p decreases, while mimics of this miRNA increase the sensitivity of neuroblastoma cells to ALK TKIs. We show that miR-1304-5p targets NRAS, decreasing cell viability via induction of apoptosis. It follows that the farnesyltransferase inhibitor (FTI) lonafarnib in addition to ALK TKIs act synergistically in neuroblastoma, inducing apoptosis in vitro. In particular, on combined treatment of neuroblastoma patient derived xenografts with an FTI and an ALK TKI complete regression of tumour growth is observed although tumours rapidly regrow on cessation of therapy. Overall, our data suggests that combined use of ALK TKIs and FTIs, constitutes a therapeutic approach to treat high risk neuroblastoma although prolonged therapy is likely required to prevent relapse.
- MeSH
- anaplastická lymfomová kináza * genetika metabolismus antagonisté a inhibitory MeSH
- apoptóza účinky léků genetika MeSH
- chemorezistence genetika účinky léků MeSH
- dibenzocyklohepteny * MeSH
- farnesyltranstransferasa * antagonisté a inhibitory metabolismus MeSH
- GTP-fosfohydrolasy * genetika metabolismus MeSH
- inhibitory proteinkinas * farmakologie terapeutické užití MeSH
- lidé MeSH
- membránové proteiny metabolismus genetika MeSH
- mikro RNA * genetika metabolismus MeSH
- mutace MeSH
- myši MeSH
- nádorové buněčné linie MeSH
- neuroblastom * farmakoterapie genetika patologie metabolismus MeSH
- piperidiny * farmakologie terapeutické užití MeSH
- pyridiny * farmakologie terapeutické užití MeSH
- regulace genové exprese u nádorů účinky léků MeSH
- synergismus léků MeSH
- xenogenní modely - testy antitumorózní aktivity MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- ALK protein, human MeSH Prohlížeč
- anaplastická lymfomová kináza * MeSH
- dibenzocyklohepteny * MeSH
- farnesyltranstransferasa * MeSH
- GTP-fosfohydrolasy * MeSH
- inhibitory proteinkinas * MeSH
- lonafarnib MeSH Prohlížeč
- membránové proteiny MeSH
- mikro RNA * MeSH
- NRAS protein, human MeSH Prohlížeč
- piperidiny * MeSH
- pyridiny * MeSH
Acute myeloid leukaemia (AML) is a complex haematological malignancy characterised by diverse genetic alterations leading to abnormal proliferation of myeloid precursor cells. One of the most significant genetic alterations in AML involves mutations in the FLT3 gene, which plays a critical role in haematopoiesis and haematopoietic homeostasis. This review explores the current understanding of FLT3 gene mutations and isoforms and the importance of the FLT3 protein in AML. FLT3 mutations, including internal tandem duplications (FLT3-ITD) and point mutations in the tyrosine kinase domain (FLT3-TKD), occur in 25-30% in AML and are associated with poor prognosis. FLT3-ITD mutations lead to constitutive activation of the FLT3 signalling pathway, promoting cell survival and proliferation. FLT3-TKD mutations affect the tyrosine kinase domain and affect AML prognosis in various ways. Furthermore, FLT3 isoforms, including shorter variants, contribute to the complexity of FLT3 biology. Additionally, nonpathological polymorphisms in FLT3 are being explored for their potential impact on AML prognosis and treatment response. This review also discusses the development of molecular treatments targeting FLT3, including first-generation and next-generation tyrosine kinase inhibitors, highlighting the challenges of resistance that often arise during therapy. The final chapter describes FLT3 protein domain rearrangements and their relevance to AML pathogenesis.
- Klíčová slova
- Acute myeloid leukaemia, FLT3, Gene isoforms, Mutations,
- MeSH
- akutní myeloidní leukemie * genetika MeSH
- lidé MeSH
- mutace genetika MeSH
- protein - isoformy genetika MeSH
- tyrosinkinasa 3 podobná fms genetika MeSH
- tyrosinkinasy MeSH
- viabilita buněk MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
- Názvy látek
- FLT3 protein, human MeSH Prohlížeč
- protein - isoformy MeSH
- tyrosinkinasa 3 podobná fms MeSH
- tyrosinkinasy MeSH
The differential signaling of multiple FGF ligands through a single fibroblast growth factor (FGF) receptor (FGFR) plays an important role in embryonic development. Here, we use quantitative biophysical tools to uncover the mechanism behind differences in FGFR1c signaling in response to FGF4, FGF8, and FGF9, a process which is relevant for limb bud outgrowth. We find that FGF8 preferentially induces FRS2 phosphorylation and extracellular matrix loss, while FGF4 and FGF9 preferentially induce FGFR1c phosphorylation and cell growth arrest. Thus, we demonstrate that FGF8 is a biased FGFR1c ligand, as compared to FGF4 and FGF9. Förster resonance energy transfer experiments reveal a correlation between biased signaling and the conformation of the FGFR1c transmembrane domain dimer. Our findings expand the mechanistic understanding of FGF signaling during development and bring the poorly understood concept of receptor tyrosine kinase ligand bias into the spotlight.
- Klíčová slova
- FGFR, biased signaling, molecular biophysics, none, signal transduction, structural biology,
- MeSH
- fibroblastové růstové faktory * MeSH
- fosforylace MeSH
- lidé MeSH
- ligandy MeSH
- receptor fibroblastových růstových faktorů, typ 1 genetika MeSH
- signální transdukce * MeSH
- těhotenství MeSH
- zkreslení výsledků (epidemiologie) MeSH
- Check Tag
- lidé MeSH
- těhotenství MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- FGFR1 protein, human MeSH Prohlížeč
- fibroblastové růstové faktory * MeSH
- ligandy MeSH
- receptor fibroblastových růstových faktorů, typ 1 MeSH
- MeSH
- biologické markery MeSH
- lidé MeSH
- novorozenec MeSH
- placentární růstový faktor MeSH
- předčasná porodní činnost * MeSH
- prediktivní hodnota testů MeSH
- preeklampsie * MeSH
- receptor 1 pro vaskulární endoteliální růstový faktor MeSH
- Check Tag
- lidé MeSH
- novorozenec MeSH
- ženské pohlaví MeSH
- Publikační typ
- dopisy MeSH
- Názvy látek
- biologické markery MeSH
- placentární růstový faktor MeSH
- receptor 1 pro vaskulární endoteliální růstový faktor MeSH