Imidazo[1,2-a]quinoxalines Derivatives Grafted with Amino Acids: Synthesis and Evaluation on A375 Melanoma Cells.

Imiqualines (imidazoquinoxaline derivatives) are anticancer compounds with high cytotoxic activities on melanoma cell lines. The first generation of imiqualines, with two lead compounds (EAPB0203 and EAPB0503), shows remarkable in vitro (IC50 = 1 570 nM and IC50 = 200 nM, respectively, on the A375 melanoma cell line) and in vivo activity on melanoma xenografts. The second generation derivatives, EAPB02302 and EAPB02303, are more active, with IC50 = 60 nM and IC50 = 10 nM, respectively, on A375 melanoma cell line. The aim of this study was to optimize the bioavailability of imiqualine derivatives, without losing their intrinsic activity. For that, we achieved chemical modulation on the second generation of imiqualines by conjugating amino acids on position 4. A new series of twenty-five compounds was efficiently synthesized by using microwave assistance and tested for its activity on the A375 cell line. In the new series, compounds 11a, 9d and 11b show cytotoxic activities less than second generation compounds, but similar to that of the first generation ones (IC50 = 403 nM, IC50 = 128 nM and IC50 = 584 nM, respectively). The presence of an amino acid leads to significant enhancement of the water solubility for improved drugability.

Liquid chromatography-electrospray ionization-tandem mass spectrometry method for quantitative estimation of new imiqualine leads with potent anticancer activities in rat and mouse plasma. Application to a pharmacokinetic study in mice.

Imidazoquinoxaline derivatives (imiqualines) are a new series of anticancer compounds. Two lead compounds (EAPB0203 and EAPB0503) with remarkable in vitro and in vivo activity on melanoma and T-cell lymphomas have been previously identified. The modulation of the chemical structure of the most active compound, EAPB0503, has led to the synthesis of two compounds, EAPB02302 and EAPB02303, 7 and 40 times more active than EAPB0503 against A375 human melanoma cancer cell line, respectively. The aim of this study was to develop and validate a sensitive and accurate liquid chromatography-electrospray ionization-tandem mass spectrometry method to simultaneously quantify EAPB02303 and its potential active metabolite, EAPB02302, in rat and mouse plasma. Analytes were detected in multiple reaction monitoring acquisition mode using an electrospray ionization detector in positive ion mode. Following a liquid-liquid extraction with ethyl acetate, analytes and internal standard were separated by HPLC reversed-phase on a C18 RP18 Nucleoshell column (2.7µm, 4.6×100mm). The method was validated according to FDA and EMA Bioanalytical Method Validation guidelines. The robustness of the method was assessed by introducing small variations in nine nominal analytical parameters. Statistical interpretation was performed by mean of the Student's t-test. Standard curves were generated via unweighted quadratic regression of calibrators (EAPB02303: 1.95-1000ng/mL, EAPB02302: 7.81-1000ng/mL in rat plasma; EAPB02303: 0.98-1000ng/mL, EAPB02302: 1.95-1000ng/mL in mouse plasma). From the analysis of QC samples, intra- and inter-assay precision and accuracy studies demonstrated %R.S.Ds. <12.5% and percent deviation from nominal concentration <7%. Matrix effects (mean matrix factors from 91.8-108.5% in rat plasma; and from 90.4-102.4% in mouse plasma) and stability assays (recoveries >87%) were acceptable and in accordance with the guidelines. No quantifiable carryover effect was observed. The LLOQs were 1.95ng/mL for EAPB02303 and 7.81ng/mL for EAPB02302 in rat plasma, and 0.98ng/mL and 1.95ng/mL for the two compounds in mouse plasma, respectively. This method was successfully implemented to support a mouse pharmacokinetic study following a single intraperitoneal administration of EAPB02303 in male C57Bl/6 mice. The obtained pharmacokinetic parameters of EAPB02303 would be useful to optimize the dosing and the rhythm of administration for subsequent preclinical in vivo activity studies.

Lipid nanocapsules formulation and cellular activities evaluation of a promising anticancer agent: EAPB0503.

OBJECTIVE: EAPB0503, lead compound of imiqualines, presented high antitumor activities but also a very low water solubility which was critical for further preclinical studies. To apply to EAPB0503, a robust and safe lipid formulation already used for poor soluble anticancer agents for injectable administration at a concentration higher than 1 mg/mL. MATERIALS AND METHODS: Physicochemical properties of EAPB0503 were determined to consider an adapted formulation. In a second time, lipid nanocapsules (LNC) formulations based on the phase-inversion process were developed for EAPB0503 encapsulation. Then, EAPB0503 loaded-LNC were tested in vitro on different cell lines and compared to standard EAPB0503 solutions. RESULTS: Optimized EAPB0503 LNC displayed an average size of 111.7 ± 0.9 nm and a low polydispersity index of 0.059 ± 0.002. The obtained loading efficiency was higher than 96% with a drug loading of 1.7 mg/mL. A stability study showed stability during 4 weeks stored at 25°C. In vitro results highlighted similar efficiencies between LNC and standard EAPB0503 solutions prepared in dimethyl sulfoxide. CONCLUSION: In view of results obtained for loading efficiency and drug loading, the use of a LNC formulation is very interesting to permit the solubilization of a lipophilic drug and to improve its bioavailability. Preliminary tested pharmaceutical formulation applied to EAPB0503 significantly improved its water solubility and will be soon considered for future preclinical in vivo studies.

New IKK inhibitors: Synthesis of new imidazo[1,2-a]quinoxaline derivatives using microwave assistance and biological evaluation as IKK inhibitors.

The inhibition of the NF-κB-dependent pathways by IKK inhibitors plays an important role in immunity, inflammation, and cancer. New imidazoquinoxalines tricyclic derivatives are prepared using microwave assistance and their biological activities as IKK inhibitors are described. Compounds 6a present a potent inhibition activity and selectivity for IKK2. Docking studies in the IKK2 binding site allowed identification of residues most likely to interact with theses inhibitors and explain their potent IKK2 inhibition activity and selectivity.