Histones are a bit like paper clips for chromosomes.
They are part of the machinery that compacts DNA strands to fit into the cell nucleus. When a stretch of DNA needs to be read, histones are modified to allow the DNA to open up and become accessible to a cell's transcription machinery. Because of their role in regulating gene expression, the enzymes that modify histones are an obvious potential target for therapeutic intervention.
That brings us to histone deacetylase, which is capable of up- or down-regulating gene expression, depending on both the cell type and the gene that is manipulated. Histone deacetylase inhibitors are the subject of both preclinical and clinical investigations for their promise in treating a number of diseases, including several cancers as well as spinal muscular atrophy. (See BioWorld Today, July 14, 2004.)
Suberoylanilide Hydroxamic Acid, or SAHA, is a histone deacetylase inhibitor in Phase II and III trials for the treatment of advanced cutaneous T-cell lymphoma. Now scientists from a number of institutions have co-published a study that shows SAHA might be useful in a different disease: lupus-associated kidney damage. While its symptoms and severity vary widely, lupus leads to kidney problems in about 50 percent of patients.
How's this for collaborative research? The work, which was reported in the Sept. 15, 2004, issue of the Journal of Immunology and bears the title "Modulation of renal disease in MRL/lpr mice by Suberoylanilide Hydroxamic Acid," was carried out by researchers at the Wake Forest University School of Medicine; the Virginia-Maryland Regional College of Veterinary Medicine; the Virginia Polytechnic Institute and State University and Edmund Via College of Osteopathic Medicine; the Charleston, S.C.-based Veteran's Administration Medical Center; the Medical University of South Carolina; the University of Miami Medical Center; Aton Pharmaceuticals, which licensed SAHA patents from Columbia University and the Memorial-Sloan Kettering Center (the company was bought by Merck & Co. Inc. in March); and the Memorial-Sloan Kettering Center itself.
"We were testing SAHA and another HDAC inhibitor, trichostatin A [or TSA], in the same animal model," said Nilamadhab Mishra, instructor in internal medicine at Wake Forest's School of Medicine and a co-author of the paper. The results from those studies, published in 2003 in the Journal of Clinical Investigation, were positive, and the researchers decided to focus their efforts on SAHA because it is "less costly, and it has already been in two trials for cancer and shown to be safe."
In the work reported here, the researchers set out specifically to test what SAHA's effects would be on kidney disease.
The researchers first isolated mesangial cells from the kidneys of MRL/lpr mice, an animal model for lupus. The cells are specific to the kidneys and play a dual role there, regulating blood flow, as well as presenting antigens. In lupus, interferon (IFN)-gamma stimulates the mesangial cells, which in turn produce and release a variety of inflammatory compounds, including both cytokines and free radicals.
Groups of cells were cultured, pretreated with various doses of SAHA and exposed to LPS and IFN-gamma, both of which act as stimulators of mesangial cells. Overall, SAHA "down-regulates inflammatory cytokines in vitro," Mishra told BioWorld Today. The scientists found that SAHA dose-dependently reversed the production of interleukin-6, TNF-alpha and nitric oxide that was caused by stimulation with IFN-gamma.
Smaller Spleen, Healthier Kidneys In Vivo
Next, Mishra and his colleagues treated 10- to 20-week-old MRL/lpr mice with daily injections of either SAHA or cyclophosphamide, which is used to treat lupus patients with kidney disease. (The mouse model normally develops kidney and skin disease, as well as arthritis, by the age of 12 to 14 weeks.)
SAHA treatment in vivo had several effects. Compared to control animals, the researchers found decreased renal pathology, as well as a decreased spleen weight in the treated animals.
Mishra believes that those gross anatomy findings can be explained by discoveries at the cellular level, namely a decrease in so-called double-negative (DN) T cells - regulatory T cells expressing neither CD4 nor CD8 proteins on their surface. That type of cell is "known to be overexpressed in both mouse models and humans with lupus," Mishra said.
DN T cells produce - wouldn't you know it - IFN-gamma.
"IFN-inducible gene expression is very high in lupus patients," Mishra said, and treatment with SAHA "may affect these genes."
The researchers did not observe a decrease in autoantibodies, or antibodies directed against an individual's tissue. Asked whether he found that surprising, Mishra said the researchers were "not really surprised, because there are reports that some autoantibodies are protective."
"We were more surprised that we did not see a decrease in autoantibodies with cyclophosphamide," which a number of previous studies had shown. Mishra thought that difference might be due to the relatively young age of the animals used in the study.
The researchers plan to investigate more thoroughly the details of genes regulated by SAHA, in particular whether the drug acts by affecting histone modifications. They also are hoping to initiate a Phase I study on SAHA for lupus in the next year. Mishra said that "another mouse model shows the same benefits" as the MRL/lpr mice, a finding he called "very preliminary, but very encouraging."