A Small Molecule Inhibitor of Human RAD51 Potentiates Breast Cancer Cell Killing by Therapeutic Agents in Mouse Xenografts

<div><p>The homologous recombination pathway is responsible for the repair of DNA double strand breaks. RAD51, a key homologous recombination protein, promotes the search for homology and DNA strand exchange between homologous DNA molecules. RAD51 is overexpressed in a variety of cancer cells. Downregulation of RAD51 by siRNA increases radio- or chemo-sensitivity of cancer cells. We recently developed a specific RAD51 small molecule inhibitor, B02, which inhibits DNA strand exchange activity of RAD51 <i>in</i><i>vitro</i>. In this study, we used human breast cancer cells MDA-MB-231 to investigate the ability of B02 to inhibit RAD51 and to potentiate an anti-cancer effect of chemotherapeutic agents including doxorubicin, etoposide, topotecan, and cisplatin. We found that the combination of B02 with cisplatin has the strongest killing effect on the cancer cells. We then tested the effect of B02 and cisplatin on the MDA-MB-231 cell proliferation in mouse xenografts. Our results showed that B02 significantly enhances the therapeutic effect of cisplatin on tumor cells <i>in</i><i>vivo</i>. Our current data demonstrate that use of RAD51-specific small molecule inhibitor represents a feasible strategy of a combination anti-cancer therapy.</p></div

The effect of cisplatin and B02 on the time course of tumor growth, A.

<p>After 11 days of tumor inoculation, tumors were touchable. Then mice were randomly regrouped (n = 5) and treated on day 11, 13, 15, 17 with either B02 (50 mg/kg), cisplatin (4 mg/kg), or a combination of both. Mice untreated or treated with vehicle (20% DMSO, 20% cremophor, 60% NS) were shown as controls. The tumor volumes were monitored by caliper measurement in each group. The time course of tumor growth presented as a graph. Error bars represent SD. Direct measurement of the weight of tumors tissues dissected from the mice after indicated treatment, B. Mice were sacrificed 43 days after tumor inoculation and the tumors were dissected. The weight of tumors shown in panel B presented as a graph, C. Error bars represent SD.</p

B02 inhibits RAD51 foci formation in MDA-MB-231 cells.

<p>A. MDA-MB-231 cells were treated with cisplatin (32 µM) either alone or in the presence of B02 (in indicated concentrations). RAD51 foci were visualized by immunostaining with RAD51 antibodies. Nuclei were counterstained with DAPI. RAD51 foci were visualized by an Olympus IX70 inverted microscope with a 100× oil objective. Bars indicate 20 µm. B. The mean of RAD51 foci number per nucleus was determined by counting at least 50 cells in each experiment. Experiments were repeated three times. Error bars represent SD.</p

B02 increases sensitivity of MDA-MB-231 cells to DNA damaging agents.

<p>A. The structures of B02, doxorubicin, etoposide, topotecan and cisplatin. B. Survival of MDA-MB-231 cells treated with B02 (○) or with indicated agents in the absence (▴) or presence (▾) of B02 (5 µM). Experiments were repeated at least three times. Error bars represent the standard deviation (SD).</p

B02 in a combination with cisplatin causes a significant inhibition in tumor growth.

<p>A. The tumor images obtained 6(day 0) (top panel) and 32 days after the first treatment (day 32) (bottom panel). Mice were injected with 100 µl of D-luciferin K⁺ (GoldBio) (150 mg/kg) by I.P. and tumor cells were visualized using IVIS Lumina XR (Caliper Life Science). B. The bioluminescent intensities of tumors at day 0 and day 32 after indicated treatments were plotted as a graph. C. The ratios of bioluminescent signals observed at day 0 and day 32. Error bars represent SD.</p

B02 increases sensitivity of 3D-growing MDA-MB-231 cells to cisplatin.

<p>A. MDA-MB-231 cells were exposed to cisplatin (in indicated concentrations) in the absence or presence of B02 (5 µM), the colonies were formed in 6-well plates coated with soft agar and stained with 0.005% crystal violet. B. The effect of B02 on survival of MDA-MB-231 cells plotted as a graph. C. The data from panel A plotted as a graph. Error bars represent SD.</p

Copper-Catalyzed Formal Carbene Migratory Insertion into Internal Olefinic CC Bonds with <i>N</i>‑Tosylhydrazones To Access Iminofuran and 2(3<i>H</i>)‑Furanone Derivatives

Efficient copper-catalyzed formal carbene migratory insertion into the olefinic CC bonds of internal olefins, that is, α-oxo ketene <i>N</i>,<i>S</i>-acetals, has been achieved by means of <i>N</i>-tosylhydrazones of ketones as the carbene precursors. Iminofuran derivatives were obtained and further transformed to the corresponding 2­(3<i>H</i>)-furanones and 4-oxobutanoates (γ-ketoesters), respectively, under mild conditions. In a similar fashion, α-thioxo ketene <i>N</i>,<i>S</i>-acetals reacted with <i>N</i>-tosylhydrazones of ketones to afford iminothiophenes. It is suggested that formal carbene migratory insertion into the olefinic CC bond is involved in the overall catalytic cycle, demonstrating a new type of carbene insertion reaction for five-membered heterocycle construction

Copper-Catalyzed Formal Carbene Migratory Insertion into Internal Olefinic CC Bonds with <i>N</i>‑Tosylhydrazones To Access Iminofuran and 2(3<i>H</i>)‑Furanone Derivatives

Efficient copper-catalyzed formal carbene migratory insertion into the olefinic CC bonds of internal olefins, that is, α-oxo ketene <i>N</i>,<i>S</i>-acetals, has been achieved by means of <i>N</i>-tosylhydrazones of ketones as the carbene precursors. Iminofuran derivatives were obtained and further transformed to the corresponding 2­(3<i>H</i>)-furanones and 4-oxobutanoates (γ-ketoesters), respectively, under mild conditions. In a similar fashion, α-thioxo ketene <i>N</i>,<i>S</i>-acetals reacted with <i>N</i>-tosylhydrazones of ketones to afford iminothiophenes. It is suggested that formal carbene migratory insertion into the olefinic CC bond is involved in the overall catalytic cycle, demonstrating a new type of carbene insertion reaction for five-membered heterocycle construction

Molecular Insight into 6FD Polyimide-Branched Poly(phenylene) Copolymers: Synthesis, Block Compatibility, and Gas Transport Study

A series of 6FDA-DABA (6FD) polyimide and branched poly(phenylene) (PP) block copolymers and homopolymers were successfully synthesized using Diels–Alder and polycondensation reactions. PP and 6FD homopolymer blends in tetrahydrofuran were immiscible. The result coincides with their large chemical dissimilarity and theoretical solubility parameter differences of 25.47 and 33.17 (MJ/m3)1/2. However, 6FD-PP block copolymer solutions were clear, and thin films were robust and creasable. Densities and fractional free volumes (FFV) (0.162–0.346) largely obeyed the rule of mixing, suggesting a “blend-like” morphology. At moderate PP block lengths, two distinct glass transition temperatures (340 and 420 °C) were evident, while large PP block lengths suppressed first-order polyimide transitions entirely. A small-angle X-ray scattering and atomic force microscopy morphological analysis revealed two distinct domains, with separation lengths increasing with the PP block length. Their gas permeation, diffusion, sorption, and separation properties were thoroughly investigated and exhibited a strong correlation with polymer chemistry, block length, and FFV. A block copolymer had an O2 permeability roughly between 6FD and PP, resulting in a 30% increase in O2/N2 selectivity. The N2/CH4 selectivities ranged from 4.2 to 0.58, suggesting that this 6FD-PP system could be efficiently tuned from highly N2-selective to CH4-selective performance. Five structural models, rule-of-mixture, Maxwell, equivalent box model, laminate, and blend, were used to predict gas transport properties. Compared with experimental data, the miscible blend model provided the best results for the 6FD-PP block copolymer system. Block copolymerization by combining highly selective polyimide and highly permeable branched poly(phenylene) provides an opportunity for gas separation tunability and improvement in selected gas pairs

Amide Bond Formation Assisted by Vicinal Alkylthio Migration in Enaminones: Metal- and CO-Free Synthesis of α,β-Unsaturated Amides

Amide bond formation is one of the most important transformations in organic synthesis, drug development, and materials science. Efficient construction of amides has been among the most challenging tasks for organic chemists. Herein, we report a concise methodology for amide bond (−CONH−) formation assisted by vicinal group migration in alkylthio-functionalized enaminones (α-oxo ketene <i>N</i>,<i>S</i>-acetals) under mild conditions. Simple treatment of such enaminones with PhI­(OAc)₂ at ambient temperature in air afforded diverse multiply functionalized α,β-unsaturated amides including β-cyclopropylated acrylamides, in which a wide array of functional groups such as aryl, (hetero)­aryl, alkenyl, and alkyl can be conveniently introduced to a ketene moiety. The reaction mechanism was investigated by exploring the origins of the amide oxygen and carbon atoms as well as isolation and structural characterization of the reaction intermediates. The amide bond formation reactions could also be efficiently performed under solventless mechanical milling conditions