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Drug Targeting Chromatin-Interacting Bromodomain Proteins Makes Sleeping Sickness Parasite “Think” It’s in TseTse Fly, Not Human Host; Parasite Stops Changing Glycoprotein Coat & Becomes Vulnerable to Attack

Some infectious diseases are particularly difficult to treat because of their ability to evade the immune system. One such illness, African sleeping sickness, is caused by the parasite Trypanosoma brucei, transmitted by the tsetse fly, and is fatal if left untreated. The trypanosome parasite is transmitted to mammals through fly bites and eventually invades major organs such as the brain, where it disrupts the sleep cycle, and causes other problems. Trypanosomes exist in different forms. When inhabiting a fly, they are covered with proteins called procyclins. But upon entering the bloodstream of a mammal, these parasites acquire a dense layer of glycoproteins that continually change, allowing the parasite to dodge attacks from the host's immune system that target specific glycoproteins in the parasite coat. Now, new research from postdoctoral scientists Dr. Danae Schulz and Dr. Erik Debler, working in the labs of Nina Papavasiliou, Ph.D., and in Nobelist Günter Blobel's (M.D., Ph.D.) lab at Rockefeller University in New York City, reveals a method by which to manipulate trypanosomes in the mammalian bloodstream so as to cause them to acquire fly-life-stage characteristics, a state that makes it easier for the human immune system to eliminate the invader. The findings suggest that inhibiting specific proteins that interact with chromatin—i.e, the mass of DNA and proteins that packages a cell's genetic information--can "trick" the parasite into differentiating to a different stage of its lifecycle, from the bloodstream stage to the fly stage of that cycle. The study was published on December 8, 2015 in the open-access journal PLOS Biology. The article is titled “Bromodomain Proteins Contribute to Maintenance of Bloodstream Form Stage Identity in the African Trypanosome.” "By blocking these chromatin-interacting proteins, we have found a way to make the parasite visible to the immune system," says Dr. Nina Papavasiliou, Ph.D., Head of the Laboratory of Lymphocyte Biology at Rockefeller. "The bloodstream form of the parasite is constantly switching protein coats, so the immune system can't recognize and eliminate it. This new method makes the parasite think it's in the fly, where it doesn't need to worry about the immune system attacking it."

[BioQuick Editor’s Note: Dr. Blobel was the sole recipient of the 1999 Nobel Prize in Physiology or Medicine for his discovery that proteins have intrinsic signals that govern their transport and localization in the cell. He also received the King Faisal International Prize in 1996, the Albert Lasker Award for Basic Medical Research in 1993, the Louisa Gross Horwitz Prize in 1989, and the Gairdner Foundation International Award in 1982. A native of Germany, Dr. Blobel received his M.D. from the University of Tübingen in 1960 and his Ph.D. in 1967 from the University of Wisconsin, Madison, where he worked with Van R. Potter in the McArdle Laboratory for Cancer Research. He did postdoctoral work at The Rockefeller University in the laboratory of 1974 Nobelist George E. Palade and has been at the Rockefeller since then. He was named the John D. Rockefeller Jr. Professor in 1992 and became an investigator at the Howard Hughes Medical Institute (HHMI) in 1986. As a further note, Dr. Palade shared the 1974 Nobel Prize for Physiology or Medicine equally with Albert Claude and Christian de Duve (de Duve was also at the Rockefeller at the time of this award) for discoveries concerning the structural and functional organization of the cell.]

EPIGENETIC REGULATION

Regulatory proteins interact with chromatin to either unwind it or package it more tightly, affecting which genes are expressed. Some of these regulatory proteins contain a region called the bromodomain, which recognizes a specific signal on chromatin and induces changes in gene expression.

Recent findings in mice have indicated that bromodomains are involved in cell differentiation, which led Dr. Papavasiliou and colleagues to hypothesize that such epigenetic mechanisms may drive the trypanosome to change from one life-stage form to another.

"The changes in gene expression that accompany the transition between the different parasite [life-stage] forms had been well established," said Dr. Schulz, the lead author of the study.

"But we didn't understand if there was some type of regulation happening at DNA, at the level of chromatin.”

“Whether chromatin-altering mechanisms might be important for differentiation hadn't really been studied before."

To investigate this, the researchers inhibited bromodomain proteins in cells by introducing genetic mutations in their DNA or by exposing the cells to a small-molecule drug called I-BET151, which is known to block bromodomains in mammals.

When these perturbations were made, the investigators observed changes in gene expression levels that resembled those seen in cells differentiating from the trypanosome bloodstream form to the trypanosome fly form. They also saw that the parasites developed a procyclin coat such as normally found on the fly form.

Based on these findings, Dr. Papavasiliou and colleagues suggest that proteins with bromodomains maintain the bloodstream form of trypanosomes, and inhibiting these bromodomain proteins causes the parasite to progress in its development toward the fly form. They believe bromodomains could serve as a potential therapeutic target to treat African sleeping sickness.

HARNESSING THE NATURAL IMMUNE SYSTEM

To explore whether I-BET151 could be used to combat the disease, the researchers used drug-treated trypanosomes to infect mice. The mice infected with drug-treated trypanosomes survived significantly longer than those infected with untreated trypanosomes, indicating that the virulence of the parasite--its ability to invade the host--was diminished in the presence of I-BET151.

"When bromodomains are inhibited, the variant protein coat is replaced with an unvarying coat on the surface of the trypanosome cell," says Dr. Schulz.

"This means that the parasite surface is no longer a moving target, giving the immune system enough time to eliminate it."

I-BET151 is not effective enough to be used in the clinic, but a crystal structure determined by Dr. Debler and published as part of this study provides direct clues for how an optimized drug could be designed to bind parasite bromodomains in a highly specific manner, limiting side-effects.

"Current treatments for this disease are limited and they have substantial side-effects, including very high mortality rates," says Dr. Papavasiliou. "This study, and recent work by others, demonstrate that targeting chromatin-interacting proteins offers a promising new avenue to develop therapeutics."

This could apply not only to African sleeping sickness, she adds, but to a number of related parasitic diseases like Chagas or malaria, with disease burdens that are far more substantial than those caused by Trypanosoma brucei.

[Rockefeller press release] [PLOS Biology article] [Dr. Papavasiliou's lab] [Dr. Blobel's lab]