Non-apoptotic functions of caspases in cellular proliferation and differentiation

The cysteinyl aspartate-specific proteases (caspases) have been identified as key players in the cellular process termed programmed cell death or apoptosis. During apoptosis, activated apoptotic caspases cleave selected target proteins to execute cell death. Additionally to their established function in cell death, a variety of recent publications have provided increasing evidence that apoptotic caspases also participate in several non-apoptotic cellular processes. Activated caspases exhibit functions during T-cell proliferation and cell cycle regulation, but are also involved in the differentiation of a diverse array of cell types. In some cell types, their differentiation can be morphologically viewed as a kind of incomplete apoptosis. Analysis of well-known apoptotic targets of caspases implicates that the cleavage of a limited number of selected substrates plays a major role during non-apoptotic functions of caspases. Selective substrate cleavage might be regulated by activation of anti-apoptotic factors, via a compartmentalized activation of caspases, or through limited activity of caspases during apoptosis-independent functions. The increasing evidence for caspase function in non-apoptotic cellular events suggests that caspases play a much more diverse role than previously assumed.     

A new generation of retinoid drugs for the treatment of dermatological diseases

Dermatological diseases such as acne, psoriasis, and various skin cancers affect approximately 24 million people in the US (17 million acne, 6.4 million psoriasis and 0.7 million skin cancer). In many cases, particularly for acne and psoriasis, these diseases have exhibited favourable responses to single agent or adjuvant retinoid therapy. When administered as both chemotherapeutic and chemopreventive agents, they offer a viable alternative to classical cancer chemotherapy. Although retinoid treatment has shown considerable promise, certain side-effects have limited their utility for chronic administration. Accordingly, there is considerable interest in developing novel retinoids that exhibit improvements in the side-effect profile, particularly for diseases that require chronic administration. Retinoids are potent biological modulators that exert their effects through intracellular receptors (retinoic acid receptors [RARs] and retinoid X receptors [RXRs]) where they regulate cellular proliferation, differentiation and programmed cell death. Currently, using a combination of molecular biology and synthetic chemistry, efforts are underway to fully characterise the biological role of individual retinoid receptor subtypes. Using receptor-selective retinoids, recent discoveries have assisted in clarifying the mechanism and biological function of retinoids and their receptors. It now appears that the RARs are implicated in the regulation of cellular proliferation and differentiation, whereas the RXRs function as modulators of programmed cell death or apoptosis. This suggests that diseases that are associated with abnormal proliferation and/or differentiation may be treatable with RAR active compounds. In fact, the data are supported by successful treatment of psoriasis, epithelial cancer and leukaemia with retinoids that preferentially activate the RARs. In contrast, retinoids that preferentially activate RXRs may be desirable for treatment of diseases for which enhancement of apoptosis is required. For example, they are potentially useful for treatment of cutaneous T-cell lymphoma and as chemopreventive agents [1]. The mechanisms of action of these RXR mediated effects are under investigation. Finally, certain receptor subtypes are implicated in retinoid side-effects. For example, it has become increasingly evident that side-effects such as triglyceridaemia [2] and teratogenesis [3,4] can be correlated to activation of the RARs and not the RXR. These findings offer opportunities for developing a new generation of retinoids which exhibit improved therapeutic indices. Recently, novel treatment strategies have evolved including the application of receptor-selective synthetic retinoids, combination therapies with other hormones or chemotherapeutic agents, and novel formulations. This has resulted in a new generation of retinoids and retinoid treatments which are in clinical development and which exhibit improvements in the therapeutic index.     

Retinoid pathway and cancer therapeutics

The retinoids are a class of compounds that are structurally related to vitamin A. Retinoic acid, which is the active metabolite of retinol, regulates a wide range of biological processes including development, differentiation, proliferation, and apoptosis. Retinoids exert their effects through a variety of binding proteins including cellular retinol-binding protein (CRBP), retinol-binding proteins (RBP), cellular retinoic acid-binding protein (CRABP), and nuclear receptors i.e. retinoic acid receptor (RAR) and retinoid x receptor (RXR). Because of the pleiotropic effects of retinoids, understanding the function of these binding proteins and nuclear receptors assists us in developing compounds that have specific effects. This review summarizes our current understanding of how retinoids are processed and act with an emphasis on the application of retinoids in cancer treatment and prevention.     

Independent contribution of three different pathways to ultraviolet-B-induced apoptosis

Ultraviolet-B radiation (UVB) causes a variety of biological effects which include the induction of apoptosis. UVB-induced apoptosis provides a well controlled scavenging mechanism protecting cells from malignant transformation. To induce programmed cell death, UVB uses a variety of cellular signaling pathways. In this context induction of nuclear DNA damage seems to be the predominant pathway, since experimental reduction of DNA damage was associated with a strong suppression of apoptosis. Additionally, UVB has been shown to target cytoplasmatically located or membrane bound components to induce signal transduction. UVB was found to directly activate cell surface death receptors, thereby triggering the apoptotic machinery. Furthermore, UVB-induced intracellular formation of reactive oxygen species (ROS) accompanied by mitochondrial dysfunction and cytochrome c release was demonstrated to be additionally involved in the apoptotic program. The following review will briefly discuss current aspects of the interplay between the different signaling pathways involved in UVB-induced apoptosis.     

Synthetic retinoids as potential antitumour agents

The retinoids, natural or synthetic derivatives of vitamin A, exhibit a variety of biological effects that may have implications in cancer chemoprevention and therapy. The chemopreventative action of classical retinoids has been ascribed to receptor-mediated modulation of a range of biological processes, including cell differentiation and proliferation. The therapeutic interest of several synthetic retinoids (also known as retinoid-related molecules) is related to their proapoptotic activity, which appears to be independent of the receptor-transactivating activity. This review focuses on the relevant features of recently reported synthetic retinoids, with particular reference to their therapeutic potential and current understanding of the mechanism of action. These agents are structurally heterogeneous compounds, including molecules closely related to natural retinoids and molecules with a retinoid-like structure (e.g., atypical retinoids) that include retinoic acid receptor (RAR) selective compounds or RAR antagonists. Promising agents of this class are adamantyl retinoids because of their apoptotic potency, in vivo antitumour activity and low toxicity. Activity of selected adamantyl retinoids (e.g., CD437, MX3350-1, ST1926) has been reported in models of various tumour types, including ovarian, breast, lung, head/neck squamous carcinoma and melanoma. The best in vivo antitumour efficacy was achieved by a protracted treatment. Orally available molecules (e.g., ST-1926) are the most suitable for this treatment schedule. Synthetic retinoids have shown synergistic interaction in combination with a variety of antitumour agents used in clinical therapy. This evidence is expected to expand the therapeutic options for clinical use of retinoids.     

Central effects of clozapine in regulating micturition in anesthetized rats

Background

Retinoids are very potent inducers of cellular differentiation and apoptosis, and are efficient anti-tumoral agents. Synthetic retinoids are designed to restrict their toxicity and side effects, mostly by increasing their selectivity toward each isotype of retinoic acids receptors (RARα,β, γ and RXRα, β, γ). We however previously showed that retinoids displayed very different abilities to activate retinoid-inducible reporter genes, and that these differential properties were correlated to the ability of a given ligand to promote SRC-1 recruitment by DNA-bound RXR:RAR heterodimers. This suggested that gene-selective modulation could be achieved by structurally distinct retinoids.

Results

Using the differential display mRNA technique, we identified several genes on the basis of their differential induction by natural or synthetic retinoids in human cervix adenocarcinoma cells. Furthermore, this differential ability to regulate promoter activities was also observed in murine P19 cells for the RARβ2 and CRABPII gene, showing conclusively that retinoid structure has a dramatic impact on the regulation of endogenous genes.

Conclusions

Our findings therefore show that some degree of selective induction or repression of gene expression may be achieved when using appropriately designed ligands for retinoic acid receptors, extending the concept of selective modulators from estrogen and peroxisome proliferator activated receptors to the class of retinoid receptors.     

Differentiation of embryonic stem cells for pharmacological studies on adipose cells.

The ongoing global explosion in the incidence of obesity has focused attention on the development of adipose cells. Severe obesity is the result of an increase in fat cell size in combination with increased fat cell number. New fat cells arise from a pre-existing pool of adipose stem cells that are present irrespective of age. The development of established preadipocyte cell lines has facilitated the study of different steps leading to terminal differentiation. However, these systems are limited for studying early events of differentiation as they represent cells which are already determined for the adipogenic lineage. In vitro differentiation of mouse embryonic stem (ES) cells towards the adipogenic lineage provides an alternative source of adipocytes for study in tissue culture and offers the possibility to investigate regulation of the first steps of adipose cell development. In this review, we describe the sequential requirement of retinoic acid and PPARgamma during adipogenesis in ES cells. Stimulation of ES cells with synthetic retinoids which are selective ligands of the retinoic acid receptor isotypes allowed the investigation of the contribution of the different retinoic receptors on the RA-dependent differentiation. The effects of thiazolidinediones, a new class of pharmacological agents used for the treatment of type 2 diabetes, and of statins, drugs used in therapy for lowering cholesterol, on the differentiation of ES cells into adipocytes or osteoblasts are described. Finally, we propose a model in which PPARgamma plays a key role in the decision of stem cells to undergo differentiation into adipocytes or osteoblasts, two closely related lineages.     

Intracellular signal transduction pathways activated by ceramide and its metabolites.

The sphingolipid ceramide has proven to be a powerful second-signal effector molecule that regulates diverse cellular processes including apoptosis, cell senescence, the cell cycle, and cellular differentiation. Ceramide has been shown to activate a number of enzymes involved in stress signaling cascades including both protein kinases and protein phosphatases. Ceramide kinase targets include stress-activated protein kinases (SAPKs) such as the jun kinases (JNKs), kinase suppressor of Ras (KSR), and the atypical protein kinase C (PKC) isoform, PKC zeta. Ceramide also is capable of activating protein phosphatases such as protein phosphatase 1 (PP1) and protein phosphatase 2A (PP2A). It is through these protein phosphatases that ceramide can indirectly inhibit kinases that are key components of pro-growth signaling processes such as the classical and novel PKC isoforms and protein kinase B (PKB; also known as Akt). However, the mechanisms how ceramide directly activates enzymes such as JNK and PP2A are still not clear. Elucidation of these mechanisms will reveal how ceramide functions in stress signaling cascades and will provide important information on cellular processes such as apoptosis. It is becoming clear that the ceramide generation is a near universal feature of programmed cell death. It is possible that during at least some apoptotic events, ceramide may be required to activate stress-signal cascades that lead to cell death, while concurrently, suppressing growth and survival pathways in the dying cell. Such a versatile role for ceramide is not unreasonable since ceramide has been implicated as having a role in both intrinsic (i.e. mitochondrial) and extrinsic (i.e. death receptor-mediated) apoptotic pathways. The recent data suggesting that aberrant glycosylation of ceramide (i.e. inactivation of the molecule) may be an important cause of drug resistance in certain cancers suggests that ceramide-mediated signaling cascades are critical components of chemotherapy-induced cell killing. Taken together, these properties of ceramide suggest that this important second-signal molecule may be an important target in anti-neoplastic strategies.     

维甲类化合物构效关系研究──维甲X受体特异性配体的二维和三维定量构效关系研究

定向地设计作用于特定受体的维甲配体分子对于阐明维甲作用的分子机制、合成新作用类型的维甲类化合物有重要意义。本文在研究系列维甲X受体特异性配体的二维与三维构效关系的基础上,考察了维甲X核受体对其配体的结构及电性需求,以进一步指导设计和合成特异性的维甲X受体激动剂。     

Heterocycle-containing retinoids. Discovery of a novel isoxazole arotinoid possessing potent apoptotic activity in multidrug and drug-induced apoptosis-resistant cells.

In a search for retinoic acid (RA) receptor ligands endowed with potent apoptotic activity, a series of novel arotinoids were prepared. Because the stereochemistry of the C9-alkenyl portion of natural 9-cis-RA and the olefinic moiety of the previously synthesized isoxazole retinoid 4 seems to have particular importance for their apoptotic activity, novel retinoid analogues with a restricted or, vice versa, a larger flexibility in this region were designed and prepared. The new compounds were evaluated in vitro for their ability to activate natural retinoid receptors and for their differentiation-inducing activity. Cytotoxic and apoptotic activities were, in addition, evaluated. In general, these analogues showed low cytotoxicity, with the restricted structures being slightly more active than the more flexible ones. As an exception, however, the isoxazole retinoid 15b proved to be particularly able to induce apoptosis at concentrations <5 microM, showing a higher activity than the classical retinoids such as all-trans-RA, 13-cis-RA, and 9-cis-RA and the previously described synthetic retinoid 4. 15b also exhibited a good affinity for the retinoid receptors. Interestingly, another important property of 15b was its ability to induce apoptosis in the HL60R multidrug-resistant (MDR) cell line, at the same concentration as is effective in HL60. Therefore, 15b represents a new retinoid possessing high apoptotic activity in an MDR cell line. The ability of 15b to act on K562 and HL60R cells suggests that this compound may have important implications in the treatment of different leukemias, and its structure could offer an interesting model for the design of new compounds endowed with apoptotic activity on MDR- and retinoid-resistant malignancies.     

Therapeutic applications for ligands of retinoid receptors

Synthetic retinoids, ligands for the RAR and RXR members of the steroid/thyroid superfamily of nuclear hormone receptors, are used for the treatment of psoriasis, acne, photoaging and cancer. Retinoid mechanisms of action for these conditions largely involve effects on epithelial differentiation and modulation of inflammation with some impact on the immune system. Retinoid medicinal chemistry in recent years has identified ligands highly specific for one of the three RAR subtypes (RAR-alpha) and for the RXR family of receptors, as well as antagonists for the RARs, RARalpha and the RXRs. Structure-activity relationships among the novel retinoid classes are reviewed along with potential therapeutic activities and side effects. RAR-alpha specific retinoids inhibit cancer cell growth but lack other retinoid toxicities, including skin irritation now ascribed to RAR-gama. RXR-specific retinoids lower blood glucose in animal models of type 2 diabetes albeit with a potential for mild hypothyroidism. Function-selective retinoids, especially a class of RAR antagonists called inverse agonists, have unexpected gene regulatory activity. Given the diverse properties and tissue distributions of the retinoid receptors, synthesis of additional classes of receptor-specific and function-selective ligands has the potential to produce novel therapeutic applications.     

Recent developments of retinoids as therapeutic agents

Retinoids, a group of natural and synthetic retinol (vitamin A) analogues, play an important role in regulating pleiotropic biological events, including growth, differentiation and death of normal, premalignant and malignant cell types, which appears to account for their therapeutic or preventive effects in acne, psoriasis, photoageing, cancer and other diseases. Nuclear retinoic acid receptors and retinoid X receptors are thought to mediate the majority of retinoid biological effects. One effective strategy is to design and synthesise retinoids with receptor selectivity restricted to specific retinoic acid receptors or retinoid X receptor subtypes (α, β and γ) in order to develop novel retinoids with a more favourable therapeutic index and with reduced adverse effects and teratogenic risk. Indeed, retinoid medicinal chemistry has identified ligands that include highly specific antagonists for one of the three RAR subtypes and for retinoid X receptors. Since the retinoid X receptors also serve as heterodimer partners for several other nuclear receptors, including thyroid hormone receptors, vitamin D receptors, peroxisomal proliferator-activator receptors, Farnesoid X receptors and liver X receptors, retinoid X receptor-selective retinoids may have clinical applications for the prevention and treatment of diseases other than dermatological diseases and cancer, such as diabetes, obesity and atherosclerosis.     

Sepsis: links between pathogen sensing and organ damage

The host's inflammatory response to sepsis can be divided into two phases, the initial detection and response to the pathogen initiated by the innate immune response, and the persistent inflammatory state characterized by multiple organ dysfunction syndrome (MODS). New therapies aimed at pathogen recognition receptors (PRRs) particularly the TLRs and the NOD-like receptors offer hope to suppress the initial inflammatory response in early sepsis and to bolster this response in late sepsis. The persistence of MODS after the initial inflammatory surge can also be a determining factor to host survival. MODS is due to the cellular damage and death induced by sepsis. The mechanism of this cell death depends in part upon mitochondrial dysfunction. Damaged mitochondria have increased membrane permeability prompting their autophagic removal if few mitochondria are involved but apoptotic cell death may occur if the mitochondrial losses are more extensive. In addition. severe loss of mitochondria results in low cell energy stores, necrotic cell death, and increased inflammation driven by the release of cell components such as HMGB1. Therapies, which aim at improving cellular energy reserves such as the promotion of mitochondrial biogenesis by insulin, may have a role in future sepsis therapies. Finally, both the inflammatory responses and the susceptibility to organ failure may be modulated by nutritional status and micronutrients, such as zinc, Therapies aimed at micronutrient repletion may further augment approaches targeting PRR function and mitochondrial viability.     

Inhibition of the Low Density Lipoprotein-Induced Vascular Smooth Muscle Cell Growth by PPARgamma Ligands

Low density lipoprotein has been shown to induce vascular smooth muscle cell proliferation and, therefore, to directly contribute to the development of atherosclerosis. Peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptors involved in various cellular processes, such as cell cycle regulation and carcinogenesis. PPARgamma, the best characterized of the PPARs, plays a crucial role in the regulation of cell proliferation, adipocyte differentiation, insulin sensitivity, energy expenditure, development of atherosclerosis and carcinogenesis. In the present review we discuss recent findings showing that a new class of antidiabetic drugs, the PPARgamma ligands thiazolidinediones, might have potential antiatherosclerotic effects. (c) 2002 Prous Science. All rights reserved.     

Enniatin B-induced cell death and inflammatory responses in RAW 267.4 murine macrophages

The mycotoxin enniatin B (EnnB) is predominantly produced by species of the Fusarium genera, and often found in grain. The cytotoxic effect of EnnB has been suggested to be related to its ability to form ionophores in cell membranes. The present study examines the effects of EnnB on cell death, differentiation, proliferation and pro-inflammatory responses in the murine monocyte-macrophage cell line RAW 264.7. Exposure to EnnB for 24 h caused an accumulation of cells in the G0/G1-phase with a corresponding decrease in cyclin D1. This cell cycle-arrest was possibly also linked to the reduced cellular ability to capture and internalize receptors as illustrated by the lipid marker ganglioside GM1. EnnB also increased the number of apoptotic, early apoptotic and necrotic cells, as well as cells with elongated spindle-like morphology. The Neutral Red assay indicated that EnnB induced lysosomal damage; supported by transmission electron microscopy (TEM) showing accumulation of lipids inside the lysosomes forming lamellar structures/myelin bodies. Enhanced levels of activated caspase-1 were observed after EnnB exposure and the caspase-1 specific inhibitor ZYVAD-FMK reduced EnnB-induced apoptosis. Moreover, EnnB increased the release of interleukin-1beta (IL-1β) in cells primed with lipopolysaccharide (LPS), and this response was reduced by both ZYVAD-FMK and the cathepsin B inhibitor CA-074Me.In conclusion, EnnB was found to induce cell cycle arrest, cell death and inflammation. Caspase-1 appeared to be involved in the apoptosis and release of IL-1β and possibly activation of the inflammasome through lysosomal damage and leakage of cathepsin B.