Pyknodysostóza je vzácné genetické onemocnění patřící ke sklerotizujícím kostním dysplaziím. Pyknodysostóza je způsobena ztrátovou mutací v genu pro katepsin K(CTSK), který hraje důležitou roli v odbourávání kostních proteinů, osteoklasty tak nemohou efektivně resorbovat organickou kostní matrix. Důsledkem nedostatku katepsinu K je porucha růstu, dysplazie lebečních a obličejových kostí a článků prstů, osteoskleróza a zvýšená kostní lomivost. Dědičnost je autosomálně recesivní. Prezentovaný pacient byl poslán na antropologii pro poruchu růstu a orofaciální abnormity. Antropometrické vyšetření prokázalo malý vzrůst se zkrácenými končetinami, úzký hrudník a ramena, relativní makrocefalii, opožděný uzávěr velké fontanely a hypoplazii dolní čelisti. Měl krátké distální články prstů a onychodystrofii. Radiologické vyšetření prokázalo difusní osteosklerózu skeletu a akroosteolýzu distálních článků prstů. Na základě těchto nálezů bylo vysloveno podezření na pyknodysostózu. Molekulárně genetické vyšetření pak tuto diagnózu potvrdilo. Terapie tohoto onemocnění je symptomatická a multidisciplinární, včetně neinvazivní dechové podpory pro spánkovou apnoe, ortopedické léčby zlomenin, dentomaxilární rekonstrukce atd. Prognóza je příznivá, onemocnění není progresivní. Znalosti klíčových znaků a časná radiologická diagnóza této vrozené vady umožní prevenci a léčbu obvyklých komplikací.
Pycnodysostosis is arare genetic disorder belonging to sclerosing skeletal dysplasias. Pycnodysostosis is caused by loss of function mutations in the cathepsin K(CTSK) gene which plays an important role in enzymatic digestion of the bone proteins, therefore osteoclasts cannot effectively break down the organic matrix of bone. Deficiency of cathepsin Kresults in dwarfism, dysplasia of the skull and facial bones and phalanges, osteosclerosis, and fragility. It is inherited in an autosomal recessive trait. The presented patient was referred to our department due to growth failure and orofacial abnormality. Anthropometric examination proved short stature with short limbs, narrow chest and shoulder, relative macrocephaly, delayed closure of the anterior fontanelle and mandibular hypoplasia. He has short tips of fingers and onychodystrophy. Radiological examination ascertained diffuse osteosclerosis of the skeleton and acro-osteolysis of the distal phalanges. On the basis of these findings the suspicion on Pycnodysostosis was pronounced. Molecular genetic testing proved this diagnosis. The therapy of this disease is symptomatic and multidisciplinary, including assisted non-invasive ventilation due to sleep apnoea, orthopaedic treatment of fractures, dento-maxillar reconstructions, etc. The prognosis is quite promising, the disease is not progressive. Knowledge of the key signs and early X-ray diagnosis of this inherited disorder will facilitate the prevention and treatment of common complications.
- Publikační typ
- abstrakt z konference MeSH
BACKGROUND AND OBJECTIVE: Hearing loss is the most common sensory deficit in humans. The aim of this study was to clarify the genetic aetiology of nonsyndromic hearing loss in the Moravian-Silesian population of the Czech Republic. PATIENTS AND METHODS: This study included 200 patients (93 males, 107 females, mean age 16.9 years, ranging from 4 months to 62 years) with nonsyndromic sensorineural hearing loss. We screened all patients for mutations in GJB2 and the large deletion del(GJB6-D13S1830). We performed further screening for additional genes (SERPINB6, TMIE, COCH, ESPN, ACTG1, KCNQ4, and GJB3) with Sanger sequencing on a subset of patients that were negative for GJB2 mutations. RESULTS: We detected biallelic GJB2 mutations in 44 patients (22%). Among these patients, 63.6%, 9.1% and 2.3% exhibited homozygous c.35delG, p.Trp24*, and p.Met34Thr mutations, respectively. The remaining 25% of these patients exhibited compound heterozygous c.35delG, c.-23+1G>A, p.Trp24*, p.Val37Ile, p.Met34Thr, p.Leu90Pro, c.235delC, c.313_326del14, p.Ser139Asn, and p.Gly147Leu mutations. We found a monoallelic GJB2 mutation in 12 patients (6.6%). We found no pathogenic mutations in the other tested genes. Conclusions: One fifth of our cohort had deafness related to GJB2 mutations. The del(GJB6-D13S1830), SERPINB6, TMIE, COCH, ESPN, ACTG1, GJB3, and KCNQ4 mutations were infrequently associated with deafness in the Moravian-Silesian population. Therefore, we suggest that del(GJB6-D13S1830) testing should be performed only when patients with deafness carry the monoallelic GJB2 mutation.
- MeSH
- aktiny genetika MeSH
- dítě MeSH
- dospělí MeSH
- draslíkové kanály KCNQ genetika MeSH
- extracelulární matrix - proteiny genetika MeSH
- hluchota genetika MeSH
- kojenec MeSH
- konexiny genetika MeSH
- lidé středního věku MeSH
- lidé MeSH
- membránové proteiny genetika MeSH
- mikrofilamentové proteiny genetika MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mutace genetika MeSH
- mutační analýza DNA metody MeSH
- percepční nedoslýchavost genetika MeSH
- předškolní dítě MeSH
- serpiny genetika MeSH
- Check Tag
- dítě MeSH
- dospělí MeSH
- kojenec MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- předškolní dítě MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Česká republika MeSH
OBJECTIVE: This study aimed to clarify the molecular epidemiology of hearing loss by identifying the responsible genes in patients without GJB2 mutations. STUDY DESIGN: Prospective genetic study. SETTING: Tertiary referral hospital. PATIENTS: Fifty one patients with bilateral sensorineural hearing loss, 20 men, and 31 women, mean age 24.9 years, range 3 to 64 years, from 49 families. GJB2 and deltaGJB6-D13S1830 mutations were excluded previously. INTERVENTION: Diagnostic. Sixty-nine genes reported to be causative of hearing loss were analyzed. Sequence capture technology, next-generation sequencing, and multiplex ligation-dependent probe amplification (MLPA) were used. Coverage of STRC was screened in Integrative Genomics Viewer software. MAIN OUTCOME MEASURE: Identification of causal pathogenic mutations in genes related to deafness. RESULTS: Five families (10%) had recessive STRC deletions or mutations. Five unrelated patients (10%) had recessive mutations in TMPRSS3, USH2A, PCDH15, LOXHD1, and MYO15A. Three families (6%) had autosomal dominant mutations in MYO6A, KCNQ4, and SIX1. One family (2%) had an X-linked POU3F4 mutation. Thus, we identified the cause of hearing loss in 28% of the families studied. CONCLUSIONS: Following GJB2, STRC was the second most frequently mutated gene in patients from the Czech Republic with hearing loss. To decrease the cost of testing, we recommend STRC deletion screening with MLPA before next-generation sequencing. The existence of a pseudogene and polymorphic STRC regions can lead to false-positive or false-negative results when copy number variation analysis is based on next-generation sequencing data.
- MeSH
- dítě MeSH
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- membránové proteiny genetika MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mutace MeSH
- percepční nedoslýchavost vrozené genetika MeSH
- předškolní dítě MeSH
- prospektivní studie MeSH
- Check Tag
- dítě MeSH
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- předškolní dítě MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Česká republika MeSH
Bohemine and roscovitine are the most important representatives of the group of compounds structurally derived from olomoucine. Biologically they function as inhibitors of cyclin-dependent kinases (CDKs), the key regulators of cell cycle, which is often disrupted in cancer cells resulting in uncontrollable proliferation. Bohemine and roscovitine have demonstrated their cytostatic and cytotoxic in vitro and also in vivo effects. Currently the phase II clinical trials for roscovitine are underway. The aim of the study was to evaluate the potential in vitro radiosensitising effect of bohemine (BOH) and roscovitine (ROS). Clonogenic survival assay and human lung adenocarcinoma cell line A549 were used. Tested schedules were: A-pretreatment, B-concomitant application and C-posttreatment. Concentrations corresponded to IC10, IC25 and IC50 for BOH/ROS (0.1-30 microM). The radiation doses were 1, 2 and 3 Gy. Flow cytometry and western blot analysis were used to characterize cell cycle distribution, BrdU incorporation and DNA repair processes. The highest in vitro radiosensitising effect of BOH/ROS was observed for Schedule A in all tested concentrations (SER(37%) 1.46-3.20). Cell cycle analysis showed an inclination towards G0/G1 delay 48 hours posttreatment and unaltered level of apoptosis. Changes in the DNA repair processes were observed - inhibition of DNA-PK kinase, inhibition of BrdU incorporation, strong and enduring induction of p21 protein and long-lasting phosphorylation of gammaH2AX(Ser139). Certain low concentration activities of BOH/ROS in monotherapy were detected, mainly the activation of DNA-PK kinase. The results demonstrated strong in vitro radiosensitising effect of BOH/ROS that is concentration and especially schedule dependent. The strong cytostatic effect of the pretreatment schedule is mediated through the inhibition/rearrangements of DNA repair processes.
- MeSH
- adenokarcinom farmakoterapie patologie radioterapie MeSH
- apoptóza účinky léků účinky záření MeSH
- buněčný cyklus účinky léků účinky záření MeSH
- cyklin-dependentní kinasy antagonisté a inhibitory metabolismus MeSH
- inhibitory proteinkinas farmakologie MeSH
- kinetin farmakologie MeSH
- lidé MeSH
- nádorové buňky kultivované účinky léků MeSH
- nádory plic farmakoterapie radioterapie MeSH
- průtoková cytometrie MeSH
- puriny farmakologie MeSH
- radiosenzibilizující látky farmakologie MeSH
- rentgenové záření MeSH
- western blotting MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Inhibitors of cyclin-dependent kinases (CDKs) undergoing clinical trials as anticancer agents usually target several CDKs in cells. Some of them are also able to increase cellular levels of p53 protein and to activate p53-regulated transcription. To define the role of p53 in the anticancer effect of selective CDK inhibitors, two related compounds roscovitine and olomoucine II were studied. Roscovitine differs functionally from its congener olomoucine II only in the selectivity towards transcriptional CDK9. Action of both compounds on proliferation, cell-cycle progression, and apoptosis was examined in RPMI-8226 cells expressing the temperature-sensitive mutant of p53 and in MCF-7 cells with wild-type p53. Both compounds blocked proliferation, decreased phosphorylation of RNA polymerase II, downregulated antiapoptotic protein Mcl-1 in both cell lines in a dose-dependent manner, and also activated p53 in MCF-7 cells. Moreover, we showed that the anticancer efficiency of CDK inhibitors was enhanced by active p53 in RPMI-8226 cells kept at permissive temperature, where downregulation of Mcl-1, fragmentation of PARP-1, and increased caspase-3 activity was detected with lower doses of the compounds. The results confirm that functional p53 protein may enhance the anticancer activity of roscovitine that could be beneficial for anticancer therapy.
- MeSH
- antitumorózní látky farmakologie MeSH
- buněčná smrt MeSH
- buněčný cyklus MeSH
- cyklin-dependentní kinasy antagonisté a inhibitory metabolismus MeSH
- fosforylace MeSH
- inhibitory proteinkinas farmakologie MeSH
- kaspasy metabolismus MeSH
- lidé MeSH
- nádorové buněčné linie MeSH
- nádorový supresorový protein p53 metabolismus MeSH
- proliferace buněk MeSH
- puriny farmakologie MeSH
- RNA-polymerasa II metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
Cytokinins are a class of plant hormones that regulate the cell cycle and diverse developmental and physiological processes. Several compounds have been identified that antagonize the effects of cytokinins. Based on structural similarities and competitive inhibition, it has been assumed that these anticytokinins act through a common cellular target, namely the cytokinin receptor. Here, we examined directly the possibility that various representative classical anticytokinins inhibit the Arabidopsis cytokinin receptors CRE1/AHK4 (cytokinin response 1/Arabidopsis histidine kinase 4) and AHK3 (Arabidopsis histidine kinase 3). We show that pyrrolo[2,3-d]pyrimidine and pyrazolo[4,3-d]pyrimidine anticytokinins do not act as competitors of cytokinins at the receptor level. Flow cytometry and microscopic analyses revealed that anticytokinins inhibit the cell cycle and cause disorganization of the microtubular cytoskeleton and apoptosis. This is consistent with the hypothesis that they inhibit regulatory cyclin-dependent kinase (CDK) enzymes. Biochemical studies demonstrated inhibition by selected anti-cytokinins of both Arabidopsis and human CDKs. X-ray determination of the crystal structure of a human CDK2-anticytokinin complex demonstrated that the antagonist occupies the ATP-binding site of CDK2. Finally, treatment of human cancer cell lines with anticytokinins demonstrated their ability to kill human cells with similar effectiveness as known CDK inhibitors.
- MeSH
- apoptóza MeSH
- Arabidopsis metabolismus MeSH
- buněčný cyklus MeSH
- chronická myeloidní leukemie metabolismus patologie MeSH
- cyklin-dependentní kinasa 2 antagonisté a inhibitory metabolismus MeSH
- cytokininy antagonisté a inhibitory metabolismus MeSH
- cytoskelet MeSH
- financování organizované MeSH
- krystalografie rentgenová MeSH
- lidé MeSH
- nádorové buňky kultivované MeSH
- nádory kostí metabolismus patologie MeSH
- nádory prsu metabolismus patologie MeSH
- osteosarkom metabolismus patologie MeSH
- proliferace buněk účinky léků MeSH
- proteinkinasy metabolismus MeSH
- proteiny huseníčku metabolismus MeSH
- průtoková cytometrie MeSH
- pyrimidiny farmakologie MeSH
- receptory buněčného povrchu metabolismus MeSH
- regulace genové exprese u rostlin MeSH
- transportní proteiny MeSH
- Check Tag
- lidé MeSH
- MeSH
- antitumorózní látky MeSH
- cyklin-dependentní kinasy * antagonisté a inhibitory chemie MeSH
- lidé MeSH
- puriny * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
In a routine screening of our small-molecule compound collection we recently identified 4-arylazo-3,5-diamino-1H-pyrazoles as a novel group of ATP antagonists with moderate potency against CDK2-cyclin E. A preliminary SAR study based on 35 analogues suggests ways in which the pharmacophore could be further optimized, for example, via substitutions in the 4-aryl ring. Enzyme kinetics studies with the lead compound and X-ray crystallography of an inhibitor-CDK2 complex demonstrated that its mode of inhibition is competitive. Functional kinase assays confirmed the selectivity toward CDKs, with a preference for CDK9-cyclin T1. The most potent inhibitor, 4-[(3,5-diamino-1H-pyrazol-4-yl)diazenyl]phenol 31b (CAN508), reduced the frequency of S-phase cells of the cancer cell line HT-29 in antiproliferation assays. Further observed cellular effects included decreased phosphorylation of the retinoblastoma protein and the C-terminal domain of RNA polymerase II, inhibition of mRNA synthesis, and induction of the tumor suppressor protein p53, all of which are consistent with inhibition of CDK9.
- MeSH
- antimetabolity diagnostické užití MeSH
- azosloučeniny farmakologie chemická syntéza MeSH
- bromodeoxyuridin diagnostické užití MeSH
- buněčný cyklus účinky léků MeSH
- cyklin-dependentní kinasa 2 antagonisté a inhibitory MeSH
- cyklin-dependentní kinasy antagonisté a inhibitory MeSH
- financování organizované MeSH
- imunoblotting MeSH
- inhibitory enzymů farmakologie chemická syntéza MeSH
- krystalografie rentgenová MeSH
- lidé MeSH
- molekulární modely MeSH
- nádorové buněčné linie MeSH
- polymerázová řetězová reakce s reverzní transkripcí MeSH
- proliferace buněk účinky léků MeSH
- pyrazoly farmakologie chemická syntéza MeSH
- reverzní transkripce účinky léků MeSH
- RNA biosyntéza izolace a purifikace MeSH
- substrátová specifita MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH