One Man's JNK is a Scientist's Treasure

© 2003 Nature Publishing Group CROSS TALKING PATHWAYS: Schematic depicting the TNF-R1-induced pathways modulating apoptosis. Blocking the NF-kB-dependent pathway leads to sustained JNK activation and apoptosis. Conversely, blocking the TNF-a induced JNK pathway promotes cell survival. JNK may induce death by triggering mitochondrial events, either directly or indirectly, but its targets remain unknown. (From G. Franzoso et al., Cell Death Differ, 10:13-15. doi: 10.1038/sj.cdd4401154) Molecular pathways that influence cellular life-and-death decisions increasingly fall under the scientific spotlight. In 1996, researchers discovered that NF-kB is anti-apoptotic, which led to the transcription factor becoming the subject of intense research. More recently, the intracellular signaling enzyme c-jun N-terminal kinase (JNK) emerged as a key promoter of cell suicide. Research into the seemingly opposing roles of these proteins' respective pathways may lead to treatments for several diseases, including organ failure, cancer, and rheumatoid arthritis. In this issue's Hot Papers, Anning Lin and others at Chicago's Ben May Institute for Cancer Research demonstrated that genes controlled by NF-kB inhibit JNK's pro-apoptotic activity.1 Another group at Ben May led by Guido Franzoso proposed that a protein, gadd45b, seems to downregulate JNK's pro-apoptotic activity.2 Two more Hot Papers from groups at San Diego-based Celgene3 and the University of San Diego4 showed that a synthetic JNK inhibitor, SP600125, downregulates expression of inflammatory mediators, including tumor necrosis factor-a (TNF-a), in cell cultures and animal models. These findings underscore the anti-inflammatory role played by apoptosis. These four papers helped establish the pathways' outline; now researchers have to fill in the rest. Data derived from the Science Watch/Hot Papers database and the Web of Science (ISI, Philadelphia) show that Hot Papers are cited 50 to 100 times more often than the average paper of the same type and age. G. Tang et al., "Inhibition of JNK activation through NF-kB target genes," Nature, 414:313-7, 2001. (Cited in 105 papers) E. De Smaele et al., "Induction of gadd45b by NF-kB downregulates pro-apoptotic JNK signaling," Nature, 414:308-13, 2001. (Cited in 95 papers) B.L. Bennett et al., "SP600125, an anthrapyrazolone inhibitor of Jun N-terminal kinase," Proc Natl Acad Sci, 98:13681-6, 2001. (Cited in 162 papers) Z. Han et al., "c-Jun N-terminal kinase is required for metalloproteinase expression and joint destruction in inflammatory arthritis," J Clin Invest, 108:73-81, 2001. (Cited in 92 papers) A "BRAKE BREAKER" Cyto-kines that activate the JNK pathway produce various outcomes orchestrated through an intricate cross-talk between different intracellular messengers. TNF-a upregulates both the pro-apoptotic JNK pathway and the IkB kinase cascade. The IKK pathway ends at NF-kB, whose target genes encode proteins that counter programmed cell death. Lin says that the dual action explains why TNF-a does not typically induce apoptosis unless NF-kB is inhibited. In 2001, Lin's team reported results from experiments using cultures of murine embryonic fibroblasts (MEFs) deficient in various components of the IKK pathway.1 TNF-a induced transient JNK activation in wild-type MEFs. In contrast, TNF-a produced sustained activation in MEFs deficient in either IKKb (a catalytic subunit) or the major transactivating subunit of NF-kB. Moreover, inhibiting protein synthesis converted transient JNK activation to sustained activation. "Taken together, these results suggest that NF-kB activation prevents sustained JNK activation in response to TNF-a, probably through induction of a group of JNK inhibitors," Lin comments. And, NF-kB- mediated inhibition of JNK is highly selective. While signaling pathways for interleukin-1 (IL-1) and TNF-a share many similarities, IL-1-induced JNK activation was unaffected in these experiments. So without NF-kB, sustained JNK activation contributes to TNF-a-induced apoptosis. But, sustained JNK activation alone did not lead to cell death. This may suggest that in the presence of TNF-a, sustained JNK activation removes regulators that suppress cell suicide. "We proposed that JNK activation breaks the brake on apoptosis," Lin says. But, intermediaries are missing. "We haven't identified the in vivo JNK inhibitors induced by NF-kB. And, we don't know why sustained, but not transient, JNK activation contributes to TNF-a-induced apoptosis," Lin says. Despite these uncertainties, Lin's work underscores the need for careful analysis of IKK-JNK cross-talk, says Anthony Manning, global head of arthritis research at Roche Pharmaceuticals, Palo Alto, Calif. For example, the study raises the prospect that JNK inhibition might increase inflammation. INHIBITION BY GADD Using MEFs, Franzoso's group developed a functional screen for isolating NF-kB targets with anti-apoptotic activity. Gadd45b, a factor induced by cytokines and stress, emerged as critical in the prosurvival pathway. NF-kB upregulates transcription of genes encoding gadd45b. The protein appears to disrupt signaling-molecule interactions, thus downregulating the cascade. As a result, gadd45b inhibits the sustained JNK activation linked to apoptosis. But gadd45b does not affect shorter-lived JNK responses, which may underlie cell survival and other critical functions. It is unclear whether MEF findings can be extrapolated to human cells. Manning explains that scaffold proteins seem to organize the JNK pathway into discrete signaling 'wiring' that may differ between cells. "It is hard to know whether the findings from MEFs are relevant to inflammation in [humans]," he says. Nevertheless, Franzoso says that gadd45b might offer a tempting therapeutic target. The group recently found peptides that modulate gadd45b activity, which raises the prospect of new treatments for inflammatory conditions such as rheumatoid arthritis. Moreover, many chemotherapeutic drugs induce gadd45b expression, which inhibits apoptosis and may contribute to some treatment-resistant cancers. Therefore, inhibiting gadd45b might offer a new tool to tackle the perennial problem of resistant cancers. "It's a long way from being proven yet, but we are excited about the prospects," Franzoso concludes. TESTING, TESTING A third Hot Paper describes an anthrapyrazolone, SP600125, that specifically inhibits JNK.3 "SP600125 was a pioneering compound," says Brydon Bennett, team leader for Celgene's JNK drug discovery program. "[It] was the first small-molecule inhibitor of JNK that did not inhibit related MAP kinases. So, SP600125 provided a way to directly test a JNK hypothesis using a pharmacological agent." According to Celgene's research, SP600125 is greater than 20-fold more selective for JNK than for several other kinases and enzymes. In cell culture, its actions include downregulating expression of genes encoding inflammatory mediators, including COX-2, IL-2, interferon-k, and TNF-a. The JNK cascade ends at the transcription factor AP-1, which controls expression of these mediators. In animal studies, SP600125 blocked expression of TNF-a induced by bacterial lipopolysaccharide, and it inhibited apoptosis. "For us, the exciting result was that this small molecule inhibitor supported the earlier genetic work describing the role of JNK in cell response to stress," says Bennett. "Therefore, small molecules that inhibit JNK could be highly effective therapies for multiple diseases, including inflammatory conditions, organ failure, and cancer. While JNK is widely expressed, its activity is most frequently induced following stress or immune stimuli. In theory, correct dosing of an inhibitor allows us to dampen this activity to background levels." As JNK contributes to fundamental survival responses, researchers inevitably worry about toxicity. "To our ongoing encouragement we are seeing remarkably little target-based toxicity with candidate compounds after several weeks of daily dosing in preclinical toxicology studies," Bennett reports. The lead chemical that emerged from the work on SP600125--CC-401--is currently completing Phase 1 clinical trials. This issue's final Hot Paper built on previous studies by Gary Firestein's group at the University of San Diego, showing that JNK activation was higher in the synoviocytes of rheumatoid arthritis than in those from osteoarthritic joints. And, the JNK activation correlated with matrix metalloproteinase (MMP) expression. "Our hope is that selectively targeting certain isoforms will limit toxicity," he says. "Of course, this is still theoretical and we are very concerned about the role of signaling molecules like MAPK in normal cell processes." But, Firestein notes, similar issues were raised about cytokine inhibitors. "Despite their importance in host defense, TNF inhibition has been a key advance in the treatment of inflammatory disease," he remarks. In synoviocyte culture, SP600125 suppressed molecular hallmarks of inflammation, such as AP-1 binding and collagenase mRNA expression.4 (Collagenase is an MMP that damages rheumatoid joints. AP-1 regulates collagenase gene expression.) SP600125 decreased paw swelling in a rat model of arthritis, as well as the radiographic damage associated with decreased AP-1 activity and collagenase-3 expression. Firestein says that JNK inhibition blocks cytokine-mediated MMP induction but not basal expression. This maintains the homeostatic functions of MMPs, which isn't the case with direct MMP inhibitors. "By carefully selecting the kinase that is important in a particular cell type or disease, one can hopefully alter the natural history of inflammatory diseases," he says. Although JNK inhibitors have potential for treatment of arthritis and autoimmune diseases, Manning notes that SP600125's anti-inflammatory effects in the model were modest and that the pharmacokinetic properties are not optimal. The study suggests that NF-kB modulates inflammation, while AP-1 controls joint destruction. Manning says that in this model, however, more advanced JNK inhibitors seem to suppress inflammation to a greater extent than SP600125. These and other papers highlight JNK's importance in cellular differentiation, proliferation, and survival. But seeing the pathways at work in various tissues of healthy and diseased models is a different story, says Carsten Weiss, University of Mainz, Germany. "Further experiments in knockout mice will possibly provide new insights into the underlying mechanisms of how JNK signaling determines cellular fate. In the end, these might lead us towards an understanding of the role of this pathway in human physiology and disease." Mark Greener (markgreener1@aol.com) is a freelance writer in Cambridge, UK. References 1. G. Tang et al., "Inhibition of JNK activation through NF-kB target genes," Nature, 414:313-7, 2001. (Cited in 105 papers) 2. E. De Smaele et al., "Induction of gadd45b by NF-kB downregulates pro-apoptotic JNK signaling," Nature, 414:308-13, 2001. (Cited in 95 papers) 3. B.L. Bennett et al., "SP600125, an anthrapyrazolone inhibitor of Jun N-terminal kinase," Proc Natl Acad Sci, 98:13681-6, 2001. (Cited in 162 papers) 4. Z. Han et al., "c-Jun N-terminal kinase is required for metalloproteinase expression and joint destruction in inflammatory arthritis," J Clin Invest, 108:73-81, 2001. (Cited in 92 papers) function sendData() { document.frm.pathName.value = location.pathname; result = false if (document.frm.score[0].checked) result = true; if (document.frm.score[1].checked) result = true; if (document.frm.score[2].checked) result = true; if (document.frm.score[3].checked) result = true; if (document.frm.score[4].checked) result = true; if (!result) alert("Please select") return result; } .myradio{background-color : #94bee5} Please indicate on a 1 - 5 scale how strongly you would recommend this article to your colleagues? Not recommended 1 2 3 4 5 Highly recommended Please register your vote

One Man's JNK is a Scientist's Treasure

Interested in reading more?

Become a Member of