Delanzomib Proteasome Inhibiteur

CEP-18770 demonstrates marginal prevention of the tryptic and peptidyl gultamyl activities of the protesome.[1] CEP-18770 inhibits A2780 ovarian cancer cells, PC3 prostate cancer, H460, LoVo colon cancer, RPMI8226 multiple myeloma cancer and HS-Sultan anaplastic non-Hodgkin lymphoma with IC50 values of 13.7, 22.2, 34.2 11.3, 5.6 and 8.2 nM, respectively.[1] CEP-18770 blocks the ubiquitin-proteasome pathway in several MM and in the chronic myelogenous leukemia cell line, K562. CEP-18770 causes an accumulation of ubiquitinated proteins over 4 to 8 hours.[1] IκBα degradation is completely blocked by pretreatment with CEP-18770. CEP-18770 significantly inhibits high levels of NF-κB activity in both RPMI-8226 and U266 cells. The time- and concentration-dependent suppression of NF-kB DNA-binding activity in MM cell lines by CEP-18770 leads to a decrease of expression of several NF-κB-modulated genes mediating the growth and survival of tumor cells including IkBα itself, the X-chromosome-linked inhibitor-of-apoptosis protein (XIAP), the pro-inflammatory cytokines TNF-α and interleukin-1β (IL-1β), the intracellular adhesion molecule (ICAM1), and the pro-angiogeneic factor vascular endothelial growth factor. [1] The expression of these NF-κB–mediated genes are associated with more favorable clinical responsiveness to this agent, highlighting their potential prognostic value in response to CEP-18770 exposure. [1] The proapoptotic activity of CEP-18770 against MM is not limited solely to tumor-derived MM cell lines, but extends to primary MM explants from relapsed or refractory patients. [1] In addition, CEP-18770 in combination produces synergistic inhibition of MM cell viability in vitro.[2]

Sondage de l'activité du Proteasome dans les extraits cellulaires

Les cellules humaines de myélome multiple sont lavées deux fois avec du sérum physiologique tamponné au phosphate froid, centrifugées et lysées avec un volume de billes de verre (<106 microns, lavées à l'acide) et un volume égal de tampon d'homogénéisation (50 mM Tris (pH 7,4), 1 mM dithiothréitol, 5 mM MgCl₂, 2 mM ATP et 250 mM saccharose) par vortexage à haute vitesse pendant 15 à 30 min à 4 °C. Les billes, les fractions membranaires, les noyaux et les débris cellulaires sont ensuite retirés du surnageant par centrifugation à 16 000 g pendant 5 min. La teneur en protéines des extraits est quantifiée à l'aide de l'essai de Bradford. L'activité du Proteasome est mesurée comme décrit ci-dessous. Des quantités égales (typiquement 60 g) de protéines sont dénaturées par ébullition dans un tampon d'échantillon réducteur, séparées par SDS-PAGE à 12,5 % et transférées électriquement sur des membranes de difluorure de polyvinylidène (PVDF). L'immunoblotting est effectué à l'aide d'un anticorps polyclonal dansyl-sulfonamidohexanoyl (1:7 500, lapin) et d'un anticorps secondaire de chèvre ou de porc anti-lapin couplé à la peroxydase de raifort, suivi d'une chimioluminescence améliorée.

CEP-18770 reveals sustained dose-related relative tumor weight inhibition. CEP-18770 leads to dose-related complete tumor regressions, which results in a 50% incidence of CR at its maximally tolerated dose (MTD) of 1.2 mg/kg intravenously. [1] CEP-18770 reveals dose-related increase in the incidence of tumor-free mice by the completion of these studies (120 days after tumor transplantation). Oral administration of CEP-18770 produces a marked decrease in tumor weight and notable dose-related incidence of complete tumor regression with minimal changes in animal body weight over the course of the 120 day studies. [1]_Equiactive doses of CEP-18770 reveal a greater and more sustained dose-related inhibition of tumor proteasome activity, corresponding temporarily with maximum induction of caspase-3 and 7 activity.[1] The maximum apoptotic signal is 2.5 fold greater for CEP-18770. In contrast, proteasome inhibition profiles of CEP-18870 are comparable in the normal peripheral mouse tissues examined (liver, lungs, whole blood, and brain [no activity]) in both their magnitude and their duration.[1] No proteasome inhibition is detected in brain tissue at any time point for either CEP-18770 or. [1] Single agent CEP-18770 PO also shows marked anti-MM effects in these xenograft models[1]