Experiments show that Hedgehog signaling - a chain of molecular events critical in embryonic development - creates conditions beneficial to the unregulated cell growth associated with cancer in adults. Zinc deficiency is correlated with many cancers; in fact, a hallmark of prostate cancer is zinc deficiency in prostate tissue. But is there a direct link between zinc deficiency and Hedgehog?
A team of researchers led by Rensselaer Polytechnic Institute will examine the link between zinc deficiency, Hedgehog, and prostate cancer in a new study funded by the National Institutes of Health (NIH). Overall, the team will study the structural mechanisms of a special process in Hedgehog signaling, called Hedgehog autoprocessing. One focus is to study how zinc deficiency affects the Hedgehog signaling pathway.
"Although very important in embryonic development, Hedgehog signaling is largely dormant in adults. But in cancer, it is activated and aids in tumor growth," said Chunyu Wang, an associate professor of biological sciences and member of the Center for Biotechnology and Interdisciplinary Studies at Rensselaer. "In test tubes and cell culture studies we can see that zinc deficiency leads to Hedgehog activation. But is this happening in human disease? This is what we want to study."
The team, which includes collaborators from the Icahn School of Medicine at Mount Sinai, Dr. William Oh and Dr. Michael Donovan, and the State University of New York at Binghamton Assistant Professor Brian Callahan, is funded by a $1.8 million NIH Research Project Grant (RO1) titled "Structural Mechanisms of Hedgehog Autoprocessing in Physiology and Disease."
The Hedgehog pathway is a key regulator of cell growth and development that helps to establish the body plan of all animals with bilateral symmetry. In a critical step that launches Hedgehog signaling, the Hedgehog precursor protein divides itself or "self-cleaves" into two parts: the Hedgehog ligand responsible for signaling, and a catalytic domain responsible for the self-cleavage. This step is called Hedgehog autoprocessing. No external catalyst is needed in the autoprocessing reaction, with the Hedgehog catalytic domain acting as the catalyst in the transformation. In research published in 2015, the Wang group, in collaboration with the Callahan group, demonstrated that zinc binds to the active site of the catalytic domain and inhibits the autoprocessing and therefore, the generation of the Hedgehog ligand. Using solution NMR, the Wang lab also identified the specific site where zinc binds to the active site.
The NIH grant will support continuing investigation on three fronts: the relationship between zinc and Hedgehog activation in cancer cells; defining the catalytic mechanism of Hedgehog autoprocessing; and examining the mechanism of the tumor resistance to anti-androgen drugs used in prostate cancer.
To explore the relationship between zinc and Hedgehog in cancer cells, Wang and his collaborators will use tissues from normal and cancerous prostates to search for a correlation between levels of zinc and Hedgehog ligand (a sign of Hedgehog activation). The team will also conduct experiments to determine whether increased zinc can inhibit the Hedgehog pathway in cancer cell cultures.
Using solution NMR, the team will define the catalytic mechanism of Hedgehog autoprocessing, tracking on an atom-by-atom basis the two-step process in which the catalytic domain self-cleaves the precursor protein. In research published in the September 2016 issue of the Journal of the American Chemical Society, Wang produced a detailed mechanism of the D46 residue, a highly conserved residue of the Hedgehog protein throughout the animal kingdom, a sign of its importance. Such detailed understanding of the catalytic mechanism could lead to new strategies to block autoprocessing in cancer.
"Without autoprocessing, there is no Hedgehog signaling, which would inhibit Hedgehog activation in cancer," Wang said. "If we can target Hedgehog autoprocessing, that would be a novel anticancer strategy."
The third area of the grant, led by Callahan, more closely examines the second step of autoprocessing, at which point the ligand is cleaved from the precursor protein, and bonds with a molecule of cholesterol. This research may shed light on why tumors develop resistance to anti-androgen drugs, which closely resemble the cholesterol molecule. The researchers hypothesize Hedgehog autoprocessing may be using the anti-androgen drugs in place of cholesterol, turning a drug intended to inhibit cancer into fuel for tumors.
"These anti-androgen drugs retard cancer for a while, but then chemoresistance emerges," Wang said. "In test tubes and cell cultures, we see the anti-androgen drugs being used in place of cholesterol in Hedgehog autoprocessing, and now we want to see whether it happens in prostate cancer cell cultures."
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