40 research outputs found
Personalized drug testing in human pheochromocytoma/paraganglioma primary cultures
No full text: Aggressive pheochromocytomas and paragangliomas (PPGLs) are difficult to treat, and molecular targeting is being increasingly considered, but with variable results. This study investigates established and novel molecular-targeted drugs and chemotherapeutic agents for the treatment of PPGLs in human primary cultures and murine cell line spheroids. In PPGLs from 33 patients, including 7 metastatic PPGLs, we identified germline or somatic driver-mutations in 79% of cases, allowing us to assess potential differences in drug responsivity between pseudohypoxia-associated cluster 1- (n=10) and kinase signaling-associated cluster 2-related (n=14) PPGL primary cultures. Single anti-cancer drugs were either more effective in cluster 1 (cabozantinib, selpercatinib, 5-FU) or similarly effective in both clusters (everolimus, sunitinib, alpelisib, trametinib, niraparib, entinostat, gemcitabine, AR-A014418, high-dose zoledronic acid). High-dose estrogen and low-dose zoledronic acid were the only single substances more effective in cluster 2. Neither cluster 1- nor cluster 2-related patient primary cultures responded to HIF-2α inhibitors, temozolomide, dabrafenib, or octreotide. We showed particular efficacy of targeted combination treatments (cabozantinib/everolimus, alpelisib/everolimus, alpelisib/trametinib) in both clusters, with higher efficacy of some targeted combinations in cluster 2 and overall synergistic effects (cabozantinib/everolimus, alpelisib/trametinib) or synergistic effects in cluster 2 (alpelisib/everolimus). Cabozantinib/everolimus combination therapy, gemcitabine, and high-dose zoledronic acid appear to be promising treatment options with particularly high efficacy in SDHB-mutant and metastatic tumors. In conclusion, only minor differences regarding drug responsivity were found between cluster 1 and cluster 2: some single anti-cancer drugs were more effective in cluster 1 and some targeted combination treatments were more effective in cluster 2
13C-direct detected NMR experiments for the sequential J-based resonance assignment of RNA oligonucleotides
No full textWe present here a set of 13C-direct detected NMR experiments to facilitate the resonance assignment of RNA oligonucleotides. Three experiments have been developed: (1) the (H)CC-TOCSY-experiment utilizing a virtual decoupling scheme to assign the intraresidual ribose 13C-spins, (2) the (H)CPC-experiment that correlates each phosphorus with the C40 nuclei of adjacent nucleotides via J(C,P) couplings and (3) the (H)CPC-CCH-TOCSY-experiment that correlates the phosphorus nuclei with the respective C10,H10 ribose signals. The experiments were applied to two RNA hairpin structures. The current set of 13C-direct detected experiments allows direct and unambiguous assignment of the majority of the hetero nuclei and the identification of the individual ribose moieties following their sequential assignment. Thus, 13C-direct detected NMR methods constitute useful complements to the conventional 1H-detected approach for the resonance assignment of oligonucleotides that is often hindered by the limited chemical shift dispersion. The developed methods can also be applied to large deuterated RNAs. Keywords: NMR spectroscopy , Direct carbon , detection , RN
Conformational dynamics of bistable RNAs studied by time-resolved NMR spectroscopy
No full textThe structural transition between two alternate conformations of bistable RNAs has been characterized by time-resolved NMR spectroscopy. The mechanism, kinetics, and thermodynamics underlying the global structural transition of bistable RNAs were delineated. Both bistable RNA conformations and a partial unstructured RNA of identical sequence could be trapped using photolabile protecting groups. This trapping allowed for an investigation of the initial folding from an unfolded RNA to one of the preferred conformations of the bistable RNA and of the structural transitions involved. Folding of the secondary structure elements occurs rapidly, while the global structural transition of the bistable RNA occurs on a time scale of minutes and shows marked temperature dependence. Comparison of these results with bistable systems previously investigated leads to the prediction of activation enthalpies (Delta H-double dagger) associated with global structural transitions in RNA.LCA
NMR-spectroscopic characterization of phosphodiester bond cleavage catalyzed by the minimal hammerhead ribozyme
No full textIn order to relate the conformational dynamics of the hammerhead ribozyme to its biological function the cleavage reaction catalyzed by the hammerhead ribozyme was monitored by time-resolved nuclear magnetic resonance (NMR) spectroscopy. For this purpose, the two nucleosides around the scissile phosphodiester bond were selectively C-13 labelled in multi-step organic syntheses starting from uniformly C-13-labelled glucose. The phosphoamidites were incorporated using phosphoamidite chemistry in the hammerhead substrate strand. In addition, the 2'-OH group on the 5'-side of the hammerhead substrate strand was labelled with a photolabile protecting group. This labelling strategy enabled a detailed characterisation of the nucleotides around the scissile phosphodiester bond in the ground state conformation of the hammerhead ribozyme in the absence and presence of Mg2+ ions as well as of the product state. Photochemical induction of the reaction in situ was further characterized by time-resolved NMR spectroscopy. The detailed structural and dynamic investigations revealed that the conformation of the hammerhead ribozyme is significantly affected by addition of Mg2+ leading to an ensemble of conformations where dynamic transitions between energetically similar conformations occur on the ms-timescale in the presence of Mg2+. The dynamic transitions are localized around the catalytic core. Cleavage from this ensemble cannot be described by mono-exponential kinetics but follows bi-exponential kinetics. A model is described to take into account these experimental data.LCA
NMR-spectroscopic characterisation of phosphodiester bond cleavage catalyzed by the minimal hammerhead ribozyme
No full textHealth-Related Quality of Life following Cytoreductive Radical Prostatectomy in Patients with De-Novo Oligometastatic Prostate Cancer
No full text(1) Background: local treatment of the primary tumor has become a valid therapeutic option in de-novo oligo-metastatic prostate cancer (PC). However, evidence regarding radical prostatectomy (RP) in this setting is still subpar, and the effect of cytoreductive RP on postoperative health-related quality of life (HRQOL) is still unclear. (2) Methods: for the current study, patients with de-novo oligo-metastatic PC (cM1-oligo), defined as ≤5 bone lesions in the preoperative staging, were included, and matched cohorts using the variables age, body-mass index (BMI), and pT-stage were generated. Patient-reported outcome measures (PROMS) were assessed pre- and postoperatively using the validated EORTC-QLQ-C30, IIEF-5, and ICIQ-SF questionnaires. The primary endpoint for univariate and multivariable analysis was good general HRQOL defined by previously validated cut-off values. (3) Results: in total, 1268 patients (n = 84 (7%) cM1-oligo) underwent RP between 2012 and 2020 at one tertiary care center. A matched cohort of 411 patients (n = 79 with oligo-metastatic bone disease (cM1-oligo) and n = 332 patients without clinical indication of metastatic disease (cM0)) was created. The median follow-up was 25mo. There was no significant difference in good general HRQOL rates between cM1-oligo-patients and cM0-patients before RP (45.6% vs. 55.2%, p = 0.186), and at time of follow-up (44% vs. 56%, p = 0.811). Global health status (GHS) worsened significantly in cM0-patients compared to baseline (−5, p = 0.001), whereas GHS did not change significantly in cM1-oligo-patients (+3.2, p = 0.381). In multivariate analysis stratified for good erectile function (IIEF5 > 18; OR 5.722, 95% CI 1.89–17.36, p = 0.002) and continence recovery (OR 1.671, 95% CI 1.03–2.70, p = 0.036), cM1-oligo was not an independent predictive feature for general HRQOL (OR 0.821, 95% CI 0.44–1.53, p = 0.536). (4) Conclusions: in this large contemporary retrospective analysis, we observed no significant difference in HRQOL in patients with the oligometastatic bone disease after cytoreductive radical prostatectomy, when compared to patients with localized disease at time of surgery
Preparation of the C-ribose-labeled 2′--TOM protected ribonucleoside phosphoramidites –
No full text<p><b>Copyright information:</b></p><p>Taken from "Short, synthetic and selectively C-labeled RNA sequences for the NMR structure determination of protein–RNA complexes"</p><p>Nucleic Acids Research 2006;34(11):e79-e79.</p><p>Published online 28 Jun 2006</p><p>PMCID:PMC1904103.</p><p>© 2006 The Author(s)</p> Abbreviations: Ac = acetyl, Bz = benzoyl, Ibu = isobutyryl, CE = cyanoethyl, DMT = (4,4′-dimethoxy)trityl, TOM = (triisopropylsilyl)oxymethyl. Reagents and conditions: () Adapted from Saito (), detailed procedure in Supplementary Data: 1. FeCl, MgSO, acetone, 20°; 2. pyridinium dichromate, AcO, CHCl, reflux; 3. HIO, THF, 20°; 4. NaBH, THF/EtOH 1:1, 20°; 5. BzCl, pyridine, 20°; 6. AcO, AcOH, HSO, 20°. () 6-Chloro--isobutyrylpurine-2-amine, ,-bis(trimethylsilyl)acetamide (BSA), MeSiOTf, 1,2-dichloroethane, 65°. () 1. Allyl alcohol, DABCO, DBU, 20°; 2. NaOH, THF/MeOH/HO, 0°; 3. DMT-Cl, pyridine, 20°. () Pd(PhP), HNEt, PPh, CHCl, 20°. () Synthesis of : 1. -benzoyladenine, BSA, SnCl, 1,2-dichloroethane, 65°; 2. NaOH, THF/MeOH/HO, 0°; 3. DMT-Cl, pyridine, 20°; synthesis of : 1. uracil, BSA, MeSiOTf, MeCN, 60°; 2. MeNH, EtOH, 20°; 3. DMT-Cl, pyridine, 20°. () BuSnCl, PrNEt, TOM-Cl, 1,2-dichloroethane, 80° according to (). () 1. AcO, DMAP, pyridine 25°; 2. 4-chlorophenyl phosphorodichloridate, 1-1,2,4-triazole, PrNEt, MeCN, 4° → 20°; 3. aqueous NH, dioxane/MeCN, 20°; 4. NaOH, THF/MeOH/HO, 4° 5. AcO, DMF, 20°, according to (). () 2-Cyanoethyldiisopropylphosphoramidochloridite, PrNEt, CHCl, 20°, according to ()
Radium-223 for primary bone metastases in patients with hormone-sensitive prostate cancer after radical prostatectomy
No full textRadium-223 dichloride (Ra-223) is the first bone-targeting agent showing improvement in overall survival in patients with castration-resistant prostate cancer (CRPC) and bone metastases. We aimed to assess feasibility of Ra-223 treatment in patients with metastatic hormone-sensitive prostate cancer (mHSPC). Ten patients with primary bone metastases received Ra-223 following radical prostatectomy (RP). Changes in alkaline phosphatase (ALP) and prostate-specific antigen (PSA) were recorded, while pain intensity was evaluated using the self-reporting Brief Pain Inventory (BPI) questionnaire. Bone scintigraphy (BS) was performed to assess treatment response. Seven patients completed six cycles of Ra-223. Discontinuation was due to leuko-and lymphopenia, progressive lymph node metastasis or newly diagnosed liver metastasis. Treatment-related adverse events occurred in three patients and included leuko-and lymphopenia, fatigue, abdominal discomfort and nausea. Overall, a median decrease of 28% in ALP and a median decrease of 83% in PSA were noted at follow-up. However, PSA progressed in five patients at follow-up. Improvement of pain was observed in all patients (median decrease of 36% after 3 cycles and of 40% at the end of therapy). On BS, three patients showed remission, four had stable disease, and one showed progressive disease at follow-up. Our results suggest that Ra-223 for primary bone metastases in patients with mHSPC after RP is feasible and alleviates pain. ALP, rather than PSA, may be a good marker for assessing treatment response. Ra-223 could therefore be taken into consideration as part of a multimodal approach for carefully selected patients with advanced prostate cancer
