Two series of potent retinoid X receptor (RXR)-selective compounds were designed and synthesized based upon recent observation that (E)-4-[2-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-2-naphthalenyl)-1- propenyl]benzoic acid (TTNBP) binds and transactivates only the retinoic acid receptor (RAR) subtypes whereas (E)-4-[2-(3,5,5,8,8-pentamethyl-5,6,7,8- tetrahydro-2-naphthalenyl)-1-propenyl]benzoic acid (3-methyl TTNPB) binds and transactivates both the RAR and RXR subfamilies. Addition of functional groups such as methyl, chloro, bromo, or ethyl to the 3 position of the tetrahydronaphthalene moiety of 4-[(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-2-naphthyl)carbonyl]benzoic acid (5a) and 4-[1-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-2- naphthyl)ethenyl]benzoic acid (6a) results in compounds which elicit potent and selective activation of the RXR class. Such RXR-selective compounds offer pharmacological tools for elucidating the biological role of the individual retinoid receptors with which they interact. Activation profiles in cotransfection and competitive binding assays as well as molecular modeling calculations demonstrate critical structural determinants that confer selectivity for members of the RXR subfamily. The most potent compound of these series, 4-[1-(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)ethenyl]ben zoi c acid (6b), is the first RXR-selective retinoid (designated as LGD1069) to enter clinical trials for cancer indications.

Structural modifications of the retinoid X receptor (RXR) selective compound 4-[1-(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2- naphthyl)ethenyl]benzoic acid (LGD1069), which is currently in phase I/IIA clinical trials for cancer and dermatological indications, have resulted in the identification of increasingly potent retinoids with > 1000-fold selectivity for the RXRs. This paper describes the design and preparation of a series of RXR selective retinoids as well as the biological data obtained from cotransfection and competitive binding assays which were used to evaluate their potency and selectivity. The most potent and selective of the analogs is 6-[1-(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydronaphthalen-2- yl)cyclopropyl]nicotinic acid (12d; LG100268). This compound has proven useful for investigating RXR dependent biological pathways including the induction of programmed cell death (PCD) and transglutaminase (TGase) activity. Our studies indicate that the induction of PCD and TGase in human leukemic myeloid cells is dependent upon activation of RXR-mediated pathways.

In vivo interactions between neutrophils and endothelial cells (EC) follow a multistep process involving two distinct neutrophil adhesion receptors. L-selectin, constitutively functional on resting neutrophils, mediates an activation-independent primary interaction resulting in rolling along the venular wall. Subsequent activation of rolling neutrophils induces upregulation and functional activation of beta 2-integrins (CD11/CD18) leading to firm attachment. Based on previous findings we hypothesized that, under shear force, rolling may be essential for successful neutrophil-EC recognition. Here we report results of our studies of human neutrophil behavior in interleukin (IL)-1-activated rabbit mesentery venules, an interaction that requires both L-selectin and beta 2-integrins. Rolling of human neutrophils is L-selection mediated; it was strongly reduced by monoclonal antibody inhibition or enzymatic removal of L-selectin. Furthermore, activation induced L-selectin shedding and, in a dose- and time-dependent fashion, rendered neutrophils unable to recognize inflamed EC despite expression of active beta 2-integrins, which promoted adhesion in vitro. Neutrophils activated for 5 min or longer lost most of their ability to roll. However, 1-3 min after activation, rolling was reduced (not abolished), and cells that were still able to roll displayed a significant tendency for a CD18-dependent transition from rolling to sticking. The whole sequence of events, rolling, sticking, and transendothelial migration, could be observed if an extravascular chemotactic stimulus was applied by superfusing mesenteries with leukotriene B4. Under such conditions, sticking and emigration was blocked when rolling was inhibited by enzymatic removal of L-selectin. Our results indicate that primary neutrophil interaction with inflamed EC through the L-selectin is a prerequisite for neutrophil function at physiological shear rates in vivo.

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Eighty percent of the single-strand DNA breaks induced by y-irradiation were prevented by the hydroxyl radical (-OH) scavenger dimethyl sulfoxide (Me2SO); CH4 was generated in the process as a product of the interaction of -OH and Me2SO. In contrast, Me SO completely blocked DNA nicking by an iron/ H202 system which produces -OH but smaller amounts of CH4 from Me2SO. Because Me2SO prevented DNA breaks from the more efficient iron/H202 system but only blocked 80% of irradiation-mediated nicking, the results sugest that -OH is responsible for 80% ofthe DNA single-strand breaks and the remaining 20% is due to interactions not involving OH.vents the formation of 80% of the single-strand breaks in DNA introduced by ionizing radiation in the presence of oxygen. During both of these processes, CH4, a product of the interaction of OH and Me2SO (8,10), is generated. The results indicate that at least 80% ofsingle-strand breaks introduced in DNA by ionizing radiation are due to an indirect effect and that 80% ofbreaks are probably generated by -OH; 20% ofbreaks are due to some process other than -OH, but our results do not indicate whether this process is direct or indirect.Lethal damage to cells exposed to ionizing radiation has been attributed mostly to effects on the structure of cellular DNA. For this reason the radiochemistry of DNA and its component parts has been studied extensively in the past 3 decades (for reviews, see refs. 1 and 2). Damage to DNA from ionizing radiation might occur directly ifenergy were transferred, without intermediates, to ionize or to excite components of the DNA. Damage might also develop indirectly if irradiation of water generated toxic products which then reacted with the DNA. One recent analysis ofthese processes based on theoretical considerations concluded that both direct and indirect damage to DNA occurs and that about 45% ofthe damaged nucleotides are derived from the direct interaction (3). This is important because radioprotective agents, which are believed to act by scavenging the toxic products such as -OH and other free radicals, are likely to interfere only with the indirect processes. Experiments with many such agents have found that about 70% of the single-strand breaks introduced into DNA by ionizing radiation can be eliminated, suggesting that only 30% of the breaks result from direct action of radiation of the DNA (4, 5). In these studies the frequency ofradiation-induced strand breaks was estimated from the sedimentation rate of damaged DNA on alkaline sucrose gradients. It is known that alkali-labile lesions in irradiated DNA can be converted into interruptions in the DNA phosphodiester backbone (6, 7). Measurements in alkali therefore overestimate the number ofbreaks. We have reinvestigated this question using procedures that assess the direct formation ofsingle-strand breaks more accurately. We also have studied in detail the radioprotective effect ofthe -OH scavenger dimethyl sulfoxide (Me2SO) (8-11). Animals (12) or cells (13) irradiated in the presence ...