BACKGROUND: Renal cell carcinoma (RCC) is a disease typified by anomalies in cell metabolism. The function of mitochondria, including subunits of mitochondrial respiratory complex II (CII), in particular SDHB, are often affected. Here we investigated the state and function of CII in RCC patients. METHODS: We evaluated tumour tissue as well as the adjacent healthy kidney tissue of 78 patients with RCC of different histotypes, focusing on their mitochondrial function. As clear cell RCC (ccRCC) is by far the most frequent histotype of RCC, we focused on these patients, which were grouped based on the pathological WHO/ISUP grading system to low- and high-grade patients, indicative of prognosis. We also evaluated mitochondrial function in organoids derived from tumour tissue of 7 patients. RESULTS: ccRCC tumours were characterized by mutated von Hippel-Lindau gene and high expression of carbonic anhydrase IX. We found low levels of mitochondrial DNA, protein and function, together with CII function in ccRCC tumour tissue, but not in other RCC types and non-tumour tissues. Mitochondrial content increased in high-grade tumours, while the function of CII remained low. Tumour organoids from ccRCC patients recapitulated molecular characteristics of RCC tissue. CONCLUSIONS: Our findings suggest that the state of CII, epitomized by its assembly and SDHB levels, deteriorates with the progressive severity of ccRCC. These observations hold the potential for stratification of patients with worse prognosis and may guide the exploration of targeted therapeutic interventions.
- MeSH
- antigeny nádorové MeSH
- dospělí MeSH
- karboanhydrasa IX metabolismus genetika MeSH
- karcinom z renálních buněk * patologie metabolismus genetika MeSH
- lidé středního věku MeSH
- lidé MeSH
- mitochondriální DNA genetika metabolismus MeSH
- mitochondrie * metabolismus patologie genetika MeSH
- mutace MeSH
- nádorový supresorový protein VHL genetika metabolismus MeSH
- nádory ledvin * patologie metabolismus genetika MeSH
- respirační komplex II * metabolismus genetika MeSH
- senioři MeSH
- sukcinátdehydrogenasa genetika metabolismus MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
Renální karcinom je histologicky diverzní onemocnění s variabilním a často nepředvídatelným chováním. Profilování genové exprese se jeví jako slibná technika k upřesnění diagnózy a stagingu renálního karcinomu a též k přesnějšímu zacílení léčby (1).
Renal cell carcinoma (RCC) is a histologically diverse disease, with variable and often unpredictable clinical behavior. Gene expression profiling is a promising technique for refining the diagnosis and staging of RCC, as well as for highlighting potential therapeutic targets (1).
- MeSH
- faktor 1 indukovatelný hypoxií analýza MeSH
- genetické testování * metody MeSH
- hepatocytární růstový faktor analýza MeSH
- imunoterapie metody MeSH
- inhibitory kontrolních bodů terapeutické užití MeSH
- karboanhydrasa IX analýza MeSH
- lidé MeSH
- nádory ledvin * farmakoterapie genetika imunologie MeSH
- prediktivní hodnota testů MeSH
- tkáňový inhibitor metaloproteinasy 1 analýza MeSH
- Check Tag
- lidé MeSH
A novel library of human carbonic anhydrase (hCA) inhibitors based on the 2-sulfanilamido[1,2,4]triazolo[1,5-a]pyrimidine skeleton modified at its 7-position was prepared by an efficient convergent procedure. These derivatives were evaluated in vitro for their inhibition properties against a representative panel of hCA isoforms (hCA I, II, IV, IX, and XII). The target tumour-associated isoforms hCA IX and XII were potently inhibited with KIs in the low nanomolar range of 5-96 nM and 4-72 nM, respectively. Compounds 1d, 1j, 1v, and 1x were the most potent hCA IX inhibitors with KIs of 5.1, 8.6, 4.7, and 5.1 nM, respectively. Along with derivatives 1d and 1j, compounds 1r and 1ab potently inhibited hCA XII isoform with KIs in a single-digit nanomolar range of 8.8, 5.4, 4.3, and 9.0 nM, respectively. Compounds 1e, 1m, and 1p exhibited the best selectivity against hCA IX and hCA XII isoforms over off-target hCA II, with selectivity indexes ranging from 5 to 14.
- MeSH
- antigeny nádorové * MeSH
- inhibitory karboanhydras farmakologie MeSH
- karboanhydrasa I metabolismus MeSH
- karboanhydrasa II * metabolismus MeSH
- karboanhydrasa IX metabolismus MeSH
- lidé MeSH
- molekulární struktura MeSH
- protein - isoformy MeSH
- sulfanilamidy MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- MeSH
- bevacizumab terapeutické užití MeSH
- imunohistochemie MeSH
- karboanhydrasa IX metabolismus MeSH
- lidé MeSH
- nádorová hypoxie * fyziologie MeSH
- nádorové biomarkery analýza MeSH
- nádory prsu farmakoterapie patologie MeSH
- prolyl-4-hydroxylasy HIF metabolismus MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
Precancerous lesions of human cervix uteri have a tendency for regression or progression. In cervical intraepithelial neoplasia grade 2 (CINII) case there is an uncertainty if a lesion will progress or regress. The carbonic anhydrase IX (CAIX) enzyme is overexpressed in cervical cancer which is more sensitive to radiotherapy. CAIX is associated with poor prognosis in solid hypoxic tumors. The aim of this study was to determine factors related to elevated soluble CAIX (s-CAIX) in high-grade intraepithelial lesion (HSIL) cases.METHODS: Patients diagnosed with HSIL (N = 77) were included into the research group whereas without HSIL (N = 72)-the control group. Concentration of the soluble CAIX (s-CAIX) in plasma was determined by the DIANA ligand-antibody-based method. C. trachomatis was detected from cervical samples by PCR. Primary outcomes were risk factors elevating s-CAIX level in HSIL group. Non-parametric statistical analysis methods were used to calculate correlations. RESULTS: The s-CAIX level in patients with HSIL was elevated among older participants (rs = 0.27, p = 0.04) and with C. trachomatis infection (p = 0.028). Among heavy smokers with HSIL, the concentration of s-CAIX was higher in older women (rs = 0.52, p = 0.005), but was not related to the age of heavy smokers' controls (τ = 0.18 p = 0.40). CONCLUSION: The concentration of s-CAIX was higher among older, heavy smoking and diagnosed with C. trachomatis patients. All these factors increased the risk for HSIL progression.
A series of 1,3,5-triazinyl aminobenzenesulfonamides substituted by aminoalcohol, aminostilbene, and aminochalcone structural motifs was synthesized as potential human carbonic anhydrase (hCA) inhibitors. The compounds were evaluated on their inhibition of tumor-associated hCA IX and hCA XII, hCA VII isoenzyme present in the brain, and physiologically important hCA I and hCA II. While the test compounds had only a negligible effect on physiologically important isoenzymes, many of the studied compounds significantly affected the hCA IX isoenzyme. Several compounds showed activity against hCA XII; (E)-4-{2-[(4-[(2,3-dihydroxypropyl)amino]-6-[(4-styrylphenyl)amino]-1,3,5-triazin-2-yl)amino]ethyl}benzenesulfonamide (31) and (E)-4-{2-[(4-[(4-hydroxyphenyl)amino]-6-[(4-styrylphenyl)amino]-1,3,5-triazin-2-yl)amino]ethyl}benzenesulfonamide (32) were the most effective inhibitors with KIs = 4.4 and 5.9 nM, respectively. In addition, the compounds were tested against vancomycin-resistant Enterococcus faecalis (VRE) isolates. (E)-4-[2-({4-[(4-cinnamoylphenyl)amino]-6-[(4-hydroxyphenyl)amino]-1,3,5-triazin-2-yl}amino)ethyl]benzenesulfonamide (21) (MIC = 26.33 μM) and derivative 32 (MIC range 13.80-55.20 μM) demonstrated the highest activity against all tested strains. The most active compounds were evaluated for their cytotoxicity against the Human Colorectal Tumor Cell Line (HCT116 p53 +/+). Only 4,4'-[(6-chloro-1,3,5-triazin-2,4-diyl)bis(iminomethylene)]dibenzenesulfonamide (7) and compound 32 demonstrated an IC50 of ca. 6.5 μM; otherwise, the other selected derivatives did not show toxicity at concentrations up to 50 μM. The molecular modeling and docking of active compounds into various hCA isoenzymes, including bacterial carbonic anhydrase, specifically α-CA present in VRE, was performed to try to outline a possible mechanism of selective anti-VRE activity.
- MeSH
- antibakteriální látky farmakologie MeSH
- enterokoky rezistentní vůči vankomycinu účinky léků MeSH
- HCT116 buňky MeSH
- inhibitory karboanhydras farmakologie MeSH
- karboanhydrasa I antagonisté a inhibitory MeSH
- karboanhydrasa II antagonisté a inhibitory MeSH
- karboanhydrasa IX antagonisté a inhibitory MeSH
- karboanhydrasy účinky léků MeSH
- lidé MeSH
- nádory farmakoterapie MeSH
- protinádorové látky farmakologie terapeutické užití MeSH
- simulace molekulární dynamiky MeSH
- simulace molekulového dockingu MeSH
- sulfonamidy farmakologie MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Simple molecular descriptors of extensive series of 1,3,5-triazinyl sulfonamide derivatives, based on the structure of sulfonamides and their physicochemical properties, were designed and calculated. These descriptors were successfully applied as inputs for artificial neural network (ANN) modelling of the relationship between the structure and biological activity. The optimized ANN architecture was applied to the prediction of the inhibition activity of 1,3,5-triazinyl sulfonamides against human carbonic anhydrase (hCA) II, tumour-associated hCA IX, and their selectivity (hCA II/hCA IX).
- MeSH
- antigeny nádorové metabolismus MeSH
- inhibitory karboanhydras chemie metabolismus MeSH
- karboanhydrasa II antagonisté a inhibitory metabolismus MeSH
- karboanhydrasa IX antagonisté a inhibitory metabolismus MeSH
- lidé MeSH
- neuronové sítě * MeSH
- racionální návrh léčiv MeSH
- sulfonamidy chemie metabolismus MeSH
- triaziny chemie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Human carbonic anhydrase IX (CA IX), a protein specifically expressed on the surface of solid tumour cells, represents a validated target both for anticancer therapy and diagnostics. We recently identified sulfonamide dicarbaboranes as promising inhibitors of CA IX with favourable activities both in vitro and in vivo. To explain their selectivity and potency, we performed detailed X-ray structural analysis of their interactions within the active sites of CA IX and CA II. Series of compounds bearing various aliphatic linkers between the dicarbaborane cluster and sulfonamide group were examined. Preferential binding towards the hydrophobic part of the active site cavity was observed. Selectivity towards CA IX lies in the shape complementarity of the dicarbaborane cluster with a specific CA IX hydrophobic patch containing V131 residue. The bulky side chain of F131 residue in CA II alters the shape of the catalytic cavity, disrupting favourable interactions of the spherical dicarbaborane cluster.
- MeSH
- antigeny nádorové genetika MeSH
- HEK293 buňky MeSH
- hydrofobní a hydrofilní interakce MeSH
- inhibitory karboanhydras chemie farmakologie MeSH
- karboanhydrasa IX antagonisté a inhibitory genetika MeSH
- katalytická doména MeSH
- krystalografie rentgenová MeSH
- lidé MeSH
- protinádorové látky chemie farmakologie MeSH
- screeningové testy protinádorových léčiv MeSH
- sekvence aminokyselin MeSH
- sloučeniny boru chemie MeSH
- sulfonamidy chemie farmakologie MeSH
- vazba proteinů MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Solid tumors, including breast cancer, are characterized by the hypoxic microenvironment, extracellular acidosis, and chemoresistance. Hypoxia marker, carbonic anhydrase IX (CAIX), is a pH regulator providing a selective survival advantage to cancer cells through intracellular neutralization while facilitating tumor invasion by extracellular acidification. The expression of CAIX in breast cancer patients is associated with poor prognosis and metastases. Importantly, CAIX-positive hypoxic tumor regions are enriched in cancer stem cells (CSCs). Here we investigated the biological effects of CA9-silencing in breast cancer cell lines. We found that CAIX-downregulation in hypoxia led to increased levels of let-7 (lethal-7) family members. Simultaneously with the increase of let-7 miRNAs in CAIX-suppressed cells, LIN28 protein levels decreased, along with downstream metabolic pathways: pyruvate dehydrogenase kinase 1 (PDK1) and phosphorylation of its substrate, pyruvate dehydrogenase (PDH) at Ser-232, causing attenuation of glycolysis. In addition to perturbed glycolysis, CAIX-knockouts, in correlation with decreased LIN28 (as CSC reprogramming factor), also exhibit reduction of the further CSC-associated markers NANOG (Homeobox protein NANOG) and ALDH1 (Aldehyde dehydrogenase isoform 1). Oppositely, overexpression of CAIX leads to the enhancement of LIN28, ALDH1, and NANOG. In conclusion, CAIX-driven regulation of the LIN28/let-7 axis augments glycolytic metabolism and enhances stem cell markers expression during CAIX-mediated adaptation to hypoxia and acidosis in carcinogenesis.
- MeSH
- antigeny nádorové genetika MeSH
- glykolýza MeSH
- hypoxie buňky MeSH
- karboanhydrasa IX genetika MeSH
- koncentrace vodíkových iontů MeSH
- lidé MeSH
- MFC-7 buňky MeSH
- mikro RNA genetika MeSH
- nádorové buněčné linie MeSH
- nádorové kmenové buňky metabolismus MeSH
- nádory prsu genetika metabolismus MeSH
- přeprogramování buněk MeSH
- proteiny vázající RNA genetika MeSH
- stanovení celkové genové exprese MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
The DNA-linked inhibitor antibody assay (DIANA) has been recently validated for ultrasensitive enzyme detection and for quantitative evaluation of enzyme inhibitor potency. Here we present its adaptation for high-throughput screening of human carbonic anhydrase IX (CAIX), a promising drug and diagnostic target. We tested DIANA's performance by screening a unique compound collection of 2816 compounds consisting of lead-like small molecules synthesized at the Institute of Organic Chemistry and Biochemistry (IOCB) Prague ("IOCB library"). Additionally, to test the robustness of the assay and its potential for upscaling, we screened a pooled version of the IOCB library. The results from the pooled screening were in agreement with the initial nonpooled screen with no lost hits and no false positives, which shows DIANA's potential to screen more than 100,000 compounds per day.All DIANA screens showed a high signal-to-noise ratio with a Z' factor of >0.89. The DIANA screen identified 13 compounds with Ki values equal to or better than 10 µM. All retested hits were active also in an orthogonal enzymatic assay showing zero false positives. However, further biophysical validation of identified hits revealed that the inhibition activity of several hits was caused by a single highly potent CAIX inhibitor, being present as a minor impurity. This finding eventually led us to the identification of three novel CAIX inhibitors from the screen. We confirmed the validity of these compounds by elucidating their mode of binding into the CAIX active site by x-ray crystallography.
- MeSH
- antigeny nádorové genetika MeSH
- biotest * MeSH
- DNA účinky léků genetika MeSH
- inhibitory karboanhydras izolace a purifikace terapeutické užití MeSH
- karboanhydrasa IX antagonisté a inhibitory genetika MeSH
- katalytická doména účinky léků MeSH
- léčivé přípravky MeSH
- lidé MeSH
- rychlé screeningové testy * MeSH
- simulace molekulového dockingu MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
