Methods and compositions for modulating apoptosis
OwnerGuido Franzoso, Enrico DeSmaele +2inventors
InventorsGuido Franzoso, Enrico DeSmaele, Francesca Zazzeroni, Salvatore Papa
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Abstract
Methods and compositions for modulating apoptosis by acting on the c-Jun-N-terminal kinase (JNK) pathway. Assays for the isolation of agents capable of modulating apoptosis, including modulators of the JNK pathway. A member of the Gadd45 protein family that inhibits JNK signaling is a target. Methods and compositions are presented for the preparation and use of novel therapeutic compositions for modulating diseases and conditions associated with elevated or decreased apoptosis.
Claims34
A method for modulating pathways leading to programmed cell death, said method comprising:
- (a) selecting a target within the JNK pathway;
- and
- (b) interfering with said target by an agent that either upregulates or downregulate the JNK pathway.
A method to identify agents that modulate JNK signaling, said method comprising:
- (a) determining whether the agent binds to Gadd45β;
- and
- (b) assaying for activity of the bound Gadd45β to determine the effect on JNK signaling.
A method for obtaining a mimetic that is sufficient to suppress JNK activation by interacting with JNKK2, said method comprising:
- (a) designing the mimetic to mimic the function of a Gadd45 protein;
- (b) contacting the mimetic to a system that comprises the JNK pathway;
- and
- (c) determining whether there is suppression of JNK signaling.
A method for screening and identifying an agent that modulates JNK pathway in vitro, said method comprising:
- (a) obtaining a target component of the JNK pathway;
- (b) exposing a cell to the agent;
- and
- (c) determining the ability of the agent to modulate the JNK pathway.
A method for screening and identifying an agent that modulates JNK activity in vivo, said method comprising:
- (a) obtaining a candidate agent;
- (b) administering the agent to a non-human animal;
- and
- (c) determining the level of JNK activity in the animal compared to JNK activity in animals not receiving the agent.
A method for identifying an agent that prevents Gadd45β from blocking apoptosis, said method comprising:
- (a) contacting cells that express high levels of Gadd45β which are protected against TNFα-induced apoptosis with the agent and TNFα;
- (b) comparing apoptosis in the cells in (a) with control cells exposed to the agent but not to TNFα;
- and
- (c) inferring from differences in apoptosis in treated versus control cells, whether the agent prevents Gadd45β from blocking apoptosis.
A method for screening for a modulator of the JNK pathway, said method comprising:
- (a) obtaining a candidate modulator of the JNK pathway, wherein the candidate is potentially any agent capable of modulating a component of the JNK pathway, including peptides, mutant proteins, cDNAs, anti-sense oligonucleotides or constructs, synthetic or natural chemical compounds;
- (b) administering the candidate agent to a cancer cell;
- (c) determining the ability of the candidate substance to modulate the JNK pathway, including either upregulation or downregulation of the JNK pathway and assaying the levels of up or down regulation.
A method of treating degenerative disorders and other conditions caused by effects of apoptosis in affected cells, said method comprising:
- (a) obtaining a molecule that interferes with the activation of JNK pathways;
- and
- (b) contacting the affected cells with the molecule.
A method of aiding the immune system to kill cancer cells by augmenting JNK signaling, said method comprising:
- (a) obtaining an inhibitor to block JNK signaling;
- and
- (b) contacting the cancer cells with the inhibitor.
A method for transactivating a gadd45β promoter, said method comprising:
- (a) binding NF-κB complexes to promoter elements of gadd45β;
- and
- (b) assaying for gadd45β gene expression.
A method for treating cancer, said method comprising:
- (a) increasing JNK activity by inhibiting Gadd45β function;
- and
- (b) administering inhibitors that interfere with Gadd45β function.
A method to determine agents that interfere with binding between Gadd45 protein and JNKK2, said method comprising:
- (a) obtaining an agent that binds to Gadd45 protein;
- (b) contacting a cell with the agent under conditions that would induce transient JNK activation;
- and
- (c) comparing cells contacted with the agent to cells not contacted with the agent to determine if the JNK pathway is activated.
A molecule with a nucleotide sequence having Gene Bank Acc. # AF441860 that functions as a gadd45β promoter.
A molecule with a nucleotide sequence that is an element of the promoter at amino acid positions selected from the group consisting of positions −447/−438 (κβ-1), −426/−417 (κβ-2), −377/−368 (κβ-3) according to FIG. 8.
A molecule comprising a region of Gadd45β, characterized by the amino acid sequence from positions 60-114 of the full length of Gadd45β protein.
A molecule comprising a binding region of JNKK2 characterized by the amino acid sequence from positions 132-156˜231-244 of full length JNKK2.
Dependencies are inferred from the claim text.