Inhibitors of Protein Methyltransferases as Chemical Tools

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Neurodegenerative disorders, including Alzheimers disease, belong to the group of the most difficult and challenging conditions with very limited treatment options

Neurodegenerative disorders, including Alzheimers disease, belong to the group of the most difficult and challenging conditions with very limited treatment options. natural products and their derivatives, we identified two compounds, 8 and 9, with dual activity and balanced IC50 values of 10 and 5 M at AChE, and 34 and 14 M at 7 nAChR, respectively. This is the first report presenting successful use of virtual screening in finding compounds with dual mode of action inhibiting both the AChE enzyme and the 7 nAChR and shows that computational methods can be a valuable tool in the early lead discovery process. oocytes and against AChE using the Ellmans colorimetric assay. 2.1. Virtual Screening We focused the search on natural products and natural product derivatives, which are a valuable source of active compounds against both targets. We screened a database consisting of 87,250 natural products and natural product derivatives from the ZINC database merged with an in-house database containing 250 lycopodium alkaloids. The substances had been screened practically, at two homology types of the 7 nAChR 1st, and then the very best hits had been docked towards the energetic site of the co-crystal structure from the AChE enzyme destined with galantamine. Galantamine was determined amongst the best scoring hits in every displays. After post-docking filtering, 78 substances had been left out, which a sub-set of 13 substances (Desk 1), i.e., galantamine analogues (6, 7) and structurally unrelated substances (8C18), had been purchased and selected for biological evaluation. Desk 1 Constructions and related docking results and in vitro activities of chosen research and strikes substances. All substances except sources galantamine and physostigmine were initially evaluated at 100 Balovaptan M. For compounds exceeding 70% inhibition at the AChE and 90% at the 7 nAChR, IC50 values were determined. oocytes using two-electrode voltage-clamp electrophysiology. The oocytes were pre-incubated with Balovaptan the test compound and subsequently the test compound was co-applied with an EC20 concentration of ACh, KRT17 an experimental design adopted to facilitate identification of PAMs. We first confirmed the ability to identify PAMs by testing NS1738, a well-established 7 PAM [16]. As evident from Figure 2, robust potentiation (440% at 31.6 M) of (30 M) ACh-evoked currents was observed. Unexpectedly, galantamine did not show any PAM activity at concentrations ranging from 10 nM to 100 M, instead, inhibition of the (30 M) ACh-evoked response was observed. At the highest concentration, galantamine inhibited ACh by 67.3%. These results triggered an Balovaptan in depth evaluation of galantamine effects at the 7 and the 42 nAChRs. The outcome of this study was published recently [23], with the conclusion that galantamine is not a PAM of the investigated nAChRs. Out of the 13 tested compounds in the present study, all except compound 18 inhibited the 7 nAChR and compounds 9C13 showed more than 90% inhibition at 100 M. Open in a separate window Figure 2 Representative current traces for NS1738, galantamine, and selected compounds from 7 nAChRs expressed in oocytes. Cells were subjected to two-electrode voltage-clamp electrophysiology experiments; the oocyte membrane potential was clamped at ?60 mV. All experiments involved a pre-incubation protocol that consisted of 25 s application of the test solution (or a saline solution for the reference trace) followed by 20 s co-application with 30 M ACh. The representative traces were baseline subtracted, and the bars above each trace represent the application periods, and concentrations of the test solutions appear above the bars. The majority of the washing periods (3 min) between each trace are omitted. For compounds exceeding 70% and 90% inhibition at 100 M at AChE and the 7 nAChR, respectively, IC50 values were determined based on full concentration response relationships (Table 1, Figure 2 and Figure 3A). To verify the AChE assay, Balovaptan we first tested physostigmine (5) and galantamine inhibition and found IC50 Balovaptan values of 0.78 and 0.68 M, respectively, that are in agreement using the released results [24,25,26,27]. Two galantamine analogues, 6.


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