FEATURED INVESTIGATOR
STEPHEN C. BLACKLOW, MD, PHD

Dr. Stephen C. Blacklow’s laboratory is dedicated to research that seeks to answer important, fundamental questions about the structure and function of cell surface receptor molecules at the biochemical level. Among the receptors he studies is the multi-domain protein receptor for low-density lipoprotein—better known as bad cholesterol—and a family of proteins called Notch receptors that communicate signals between neighboring cells. Mutations in these and other cell surface receptors can result in a wide variety of disorders, including heart disease and cancer. Understanding how these receptor molecules function in normal and abnormal states is fundamental to understanding the patho- genesis of these diseases, and thereby provides new opportunities for the development of targeted therapies.

Background

Dr. Blacklow grew up in the greater Boston area. The son and grandson of physicians, he attended undergraduate school at Harvard College, where he majored in the biochemical sciences. After an interim year in the chemistry department at Stanford, he returned to Harvard to complete his M.D. and Ph.D. degrees, carrying out his thesis research in bioorganic chemistry under the guidance of Professor Jeremy Knowles. After a clinical pathology residency at Brigham and Women’s Hospital, Dr. Blacklow completed a postdoctoral fellowship at the Whitehead Institute and then joined the faculty of the department of pathology at Stanford. Recruited back to Boston by the noted former chair of the department of pathology, Dr. Ramzi Cotran, Dr. Blacklow returned to Brigham and Women’s Hospital in January of 1998 to further his research program. Dr. Blacklow is currently an associate professor of pathology at Harvard Medical School and an associate pathologist at Brigham and Women’s Hospital.

Research

“The key to understanding how a protein works is to determine its shape and how it binds other molecules,” explains Dr. Blacklow. “Structure begets function. Moreover, the shape of the protein often yields new insights into how to inhibit or modify its function.” The receptor that removes cholesterol-containing particles from the blood is called the LDL receptor. About one in 500 people have an inherited defect in the LDL receptor, which results in an increased level of cholesterol in the blood. This condition, which is called familial hypercholesterolemia, predisposes patients to heart disease at an early age. Dr. Blacklow’s studies of the LDL receptor have led to new insights into the basic mechanism by which these receptors normally work, and the way the receptors malfunction in familial hypercholesterolemia. In addition, Dr. Blacklow has recently published two landmark papers on the structure and function of human Notch receptors, one in Nature Structure Biology and the other inCell. The Notch signaling pathway normally controls the development and differentiation of many cells and tissues. Normal Notch receptors are activated when they bind to a protein called a ligand that is expressed on a neighboring cell. Binding of ligand somehow makes Notch receptors susceptible to cleavage by two kinds of proteases (a type of scissors for proteins), which liberates the intracellular part of the Notch receptor. Intracellular Notch then travels to the nucleus and turns off the expression of target genes, which influence cellular differentiation and the development of various tissues.

 

Dr. Blacklow’s lab has recently solved the crystalline structure of the extracellular region of Notch where the first cleavage occurs. The cleavage site is normally deeply buried when Notch is in the “off ” state, indicating that there must be a large change in the shape of the protein before cleavage. One likely possibility is that this shape change is caused by a “tug” on the receptor, which is produced by movement of the ligand into the inside of the ligand-expressing cell. If this model is correct, it means that Notch receptors respond to mechanical forces, such as those that are transmitted when cells are stretched or exposed to a flowing fluid (such as the blood). In collaboration with Dr. Jon Aster, Dr. Blacklow’s lab also noted that the mutations in Notch1 that occur in T-cell acute lymphoblastic leukemia (T-ALL) lie within the part of Notch1 that protects the protein from cleavage in the “off ” state. It is likely that these mutations alter the shape of the protein, making it susceptible to cleavage and activation even in the absence of ligand.

 

Beyond explaining how Notch receptors are regulated, the structure of the extracellular domain of Notch suggests that it should be possible to create antibodies that bind this region and selectively activate or inhibit Notch receptors. Drs. Blacklow and Aster have embarked on a collaborative study that seeks to discover such antibodies. Recently, several potent and specific inhibitory antibodies have been discovered, and it is anticipated that such inhibitors could be useful in a wide variety of human cancers, including (T-ALL).

 

The second structure that Dr. Blacklow’s lab has solved is a complex consisting of part of the intracellular portion of Notch1 and two nuclear proteins that bind Notch, called CSL and Mastermind-like 1. The shape of this complex and the way the proteins fit together have provided key insights into how Notch turns on the expression of genes, which is the key to understanding how Notch acts in normal cells and in tumors. Drs. Blacklow and Aster have identified a small piece of the Mastermind-like 1 protein that acts as a very specific inhibitor of Notch action in the nucleus. This mastermind-like 1 peptide is under development as another new type of Notch inhibitor.

Laboratory

The size of Dr. Blacklow’s laboratory is currently 7 researchers and one technician. Among the researchers are post-doctoral fellows and graduate students. Most of his post-doctoral fellows have a strong background in chemistry, molecular biology, and structural biology, including X-ray crystallography.

Collaborations

Dr. Blacklow works closely with several members of the Biomedical Research Institute—Cancer Research Center and area institutions. In addition to the collaboration with Dr. Aster, he has close working relationships with Dr. Piotr Sliz, who oversees the management of the computing infrastructure for Harvard Medical School faculty; Dr. Michael Eck’s group at the Dana Farber Cancer Institute; and Dr. Tim Springer at the Center for Blood Research. The infrastructure for research in structural biology and protein biochemistry is costly, and Dr. Blacklow is grateful to colleagues elsewhere in the Harvard Medical area for granting access to a number of specialized instruments (most notably, Drs. Michael Eck at the Dana Farber Cancer Institute and Dr. Stephen Harrison at the Harvard Medical School). Additionally, he has numerous other regional, national, and international collaborations, notably, with Dr. Warren Pear of the University of Pennsylvania.

FUNDING

Dr. Blacklow’s laboratory is principally funded by the National Institutes of Health.

Importance of Being at the Brigham

“The environment at the Brigham is very strong,” Dr. Blacklow relates. “My colleagues make a huge difference in my work. People are our biggest strength at the Brigham. We have incredibly talented people here who, given the appropriate resources, can make almost anything happen. You can have all the technology and machines you want, but in the end, it’s the personnel that matters.”

Future

Dr. Blacklow believes that it will be possible to design small molecules or develop antibodies that target Notch on the cell surface and in the nucleus. As is the case with human immunodeficiency virus (HIV), targeting the Notch pathway at more than one point may prevent the development of resistance in tumors. Dr. Blacklow adds, “Clearly, one can make a case for regulating Notch activity in women’s cancers, as unbridled Notch signaling has also been implicated in ovarian cancer and breast cancer.” He envisions that ongoing research efforts at Brigham and Women’s Hospital will result in translational drugs and therapies directed at the Notch pathway that are relevant to many forms of cancer.

Selected References

Beglova N, Jeon H, Fisher C, Blacklow SC. Cooperation between fixed and low pHinducible interfaces controls lipoprotein release by the LDL receptor. Molecular Cell. 2004;16:281-292.

 

Nam Y, Sliz P, Song L, Aster JC, Blacklow SC. Structural basis for cooperativity in recruitment of MAML co-activators to Notch transcription complexes. Cell. 2006;124:973-983.

 

Fisher C, Beglova N, Blacklow SC. A general mode for recognition of ligands by lipoprotein receptors from the structure of an LDLR-RAP complex. Molecular Cell.  2006;22:277-283.

 

Malecki MJ, Sanchez-Irizarry C, Mitchell JL, Xu M, Histen G, Aster, Blacklow SC.Leukemia-associated mutations within the NOTCH1 heterodimerization domain fall into at least two distinct mechanistic classes. Mol Cell Biol. 2006;26:4642-4651.

 

Nam Y, Sliz P, Pear WS, Aster JC, Blacklow SC. Cooperative assembly of higher order Notch complexes functions as a switch to induce transcription. Proc Natl Acad Sci.2007;104:2103-2108.

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