"Joel's lab and my lab worked very closely together on this," said Balch. "That's what's great about Scripps Research-its collaborative nature. People just walk in next door and two hours later you are doing an experiment together!"

Working with Ray Frizzell and Joe Pilewski, cystic fibrosis investigators at the University of Pittsburg School of Medicine, the Balch laboratory treated human lung epithelial cells isolated from patients with the devastating ?”F508 mutation with known HDAC inhibitors. Intriguingly, the FDA-approved HDAC inhibitor SAHA was shown to be most effective in restoring surface channel activity-one of the main markers of cystic fibrosis that is responsible for rehydration of the cells' surface. Control cells demonstrated negligible levels of channel activity, while SAHA-treated cells were restored to 28 percent of the normal level found in healthy individuals.

"It's a pretty solid rescue with some intriguing properties," commented Balch.

Using a bioinformatics approach led by the Gerard Manning laboratory at the Salk Institute of Biological Sciences, the team showed that the compound increased the functioning of mutated CFTR proteins at multiple levels in the proteostasis network. Not only were mutant CFTR proteins more protected from destruction in the endoplasmic reticulum, they were also more efficiently transported to the lung cell surface where they were found at comparable levels to that of wild-type (normal) CFTR. In addition, once at the surface of HDAC inhibitor-treated cells, mutant CFTR proteins were better able to resist destruction by additional degradation pathways than DF508 CFTR proteins in untreated cells.

"By rebalancing the proteostasis program to provide a more supportive cellular environment," said Balch, "the cells appear to treat the mutation more like a polymorphism [genetic differences that are responsible for individual diversity] rather than something dangerous needing to be completely eliminated."

Balch also likened this process to evolutionary adaptations to changes in protein structure, which support mutations providing a selective functional advantage.

The study showed that one specific HDAC-HDAC7, one of 18 known human HDACs-appeared to be largely responsible for the effects on the treated cells. Little is known about the function of HDAC7 in human physiology and efforts are currently under way by the laboratories of Balch, Gottesfeld, Manning, and Scripps Research Professor John Yates to understand its mechanism of action.

Mindful that dosing would be a key issue in any attempts at drug development, the researchers also began to address dosing issues in the current study.

"We know that a compound won't make it into the clinic if patients have to take the equivalent of a cereal bowl of it several times a day," said Balch. "That's especially true because, given the nature of cystic fibrosis, this would need to be a sustained, life-long treatment to protect the patient from disease."

Remarkably, the team found that low doses of SAHA not only worked in cultures of cystic fibrosis lung cells, but also offered some significant advantages over acute doses. While acute doses of SAHA produced an increase in the surface channel activity the next day, the effects stopped soon after the drug was withdrawn. In contrast, much smaller doses began working efficiently after six to eight days and strong channel activity was observed after the drug was withdrawn, gradually declining over the following week. This feature is reminiscent of its potential mechanism of action, Balch said, perhaps involving chromatin remodeling leading to an altered, protective proteostasis environment in the lung cell that could be sustainable.

While thrilled with the results, Balch cautions that "there is much work to do"-including further drug development, preclinical work, and clinical trials-before any new therapy for cystic fibrosis becomes a reality using this approach. The FDA-approved drug SAHA, while initially approved as an acute dose regimen for cancer therapeutics, remains to be carefully examined for use in a low-dose, chronic treatment regimen that would be required for protecting cystic fibrosis patients from disease over a lifetime.

Source: Scripps Research Institute