Assistant Professor in Dermatology
Brigham and Women's Hospital
Harvard Skin Disease Research Center
77 Ave Louis Pasteur
Harvard Institutes of Medicine, Rm. 650
Boston, MA 02115
The Dimitroff Laboratory, Investigating the Molecular Repertoire in Tissue-Specific Cell Trafficking
Charles J. Dimitroff, Ph.D., Principal Investigator
The Dimitroff Laboratory is led by principal investigator, Charles J. Dimitroff, Ph.D., who is an Assistant Professor of Dermatology at Brigham and Women’s Hospital,
Dr. Dimitroff and his team of postdoctoral fellows, research technicians and students are currently involved in studies that help illuminate how immune cells (T cells) traffic to inflamed skin and how prostate tumor cells metastasize to bone. More specifically, they are investigating the constituency of carbohydrate-bearing receptors on these cells that mediate dynamic binding to the vascular wall and trigger movement of these cells from inside of the bloodstream to the parenchyma of bone or skin. The function of these carbohydrate-bearing receptors or glycoproteins causes a cell adhesion phenomenon called rolling, which is essential for initiating a cascade of successive molecular interactions preceding tissue entry. The Dimitroff Laboratory hypothesizes that these glycoprotein “rolling” receptors are, in part, responsible for development of T cell-dependent dermatitis and for metastasis of prostate cancer to bone (Please see illustration below depicting the role of rolling receptors in cell trafficking to skin and bone). The identity of such receptors ascertained by the Dimitroff Laboratory could provide an opportunity for development of anti-inflammatory/metastatic therapeutics targeting these unique molecules.
A View Into Cell Trafficking Through A Rolling Receptor. A membrane protein displaying key carbohydrate moieties is characteristically expressed by circulating T cells or tumor cells engineered to enter distinct tissues (such as skin and bone as shown). These carbohydrate moieties interact with specific counter-receptors found on the luminal aspect of the microvasculature wall, resulting in target cell rolling and targeted tissue tropisms.
Contrary to what has previously been described on white blood cells, The Dimitroff Laboratory has recently discovered the identity of a glycoprotein, named PSGL-1, on metastatic prostate tumor cells. PSGL-1 is commonly known for its role as a rolling receptor on white blood cells and has been implicated in the trafficking of hematopoietic stem cells to bone and T cells to skin. PSGL-1’s expression on metastatic prostate tumor cells, hence, has led to the hypothesis that prostate tumor cells utilize a similar molecular mechanism as hematopoietic stem cells for trafficking to bone. This emphasis and exciting seminal finding has led to a Project 1 funded by The American Cancer Society.
Project 1: Role Of PSGL-1 In The Bone Metastasis Of Prostate Cancer
Prostate tumor metastasis characteristically occurs in bone and often results in the poor prognosis of patients. Unfortunately, there are no treatments that specifically target the molecular pathogenesis of bone metastasis. Recent studies have indicated that prostate tumor cells exhibit an enhanced adhesion for bone blood vessels compared with adhesion to blood vessel linings in other tissues. The Dimitroff Laboratory has shown that initial adhesive interactions between bone-metastatic prostate tumor cells and bone blood vessels is mediated by PSGL-1. Remarkably, expression analysis of PSGL-1 on tissue micro-arrays (TMA) of normal prostate epithelium and of localized and metastatic prostate tumor tissue has revealed that PSGL-1 is almost exclusively found on metastatic prostate tumor cells with the highest level expressed on tumor cells in bone. The objective of studies outlined in this project is to elucidate the molecular pathogenesis of prostate tumor metastasis to bone and determine whether PSGL-1 promotes bone-metastatic behavior of prostate cancer and whether PSGL-1 possesses structural motif(s) for targeting of novel anti-cancer therapies. The specific aims are: 1.) To analyze the glyco-biochemistry of PSGL-1 on metastatic prostate tumor cells and 2.) To validate the functional role of PSGL-1 in prostate tumor metastasis, in vivo. Experimentation includes innovative glycobiology, biochemistry and cell adhesion technologies to ascertain the critical structures conferring the “rolling” activity of PSGL-1 on metastatic prostate tumor cells. In vivo analysis is being performed using mice transplanted with human bone to explore the efficiency of PSGL-1+ or PSGL-1- human prostate tumor cells to home (traffic) into human bone (Please see illustrations below depicting the transplantation of human bone tissue onto mice and the fidelity of human bone vessel markers). Biochemical perturbation of PSGL-1 structural motifs critical for functional rolling activity with rationally-designed agents is also being performed in this in vivo investigation to reveal potential therapeutic strategies for pharmacologic development. Results from these analyses will offer new insights into the molecular pathogenesis of prostate tumor metastasis and may provide rationale for development of new therapies targeting homing receptors.
Other projects pursued by the Dimitroff Laboratory are based on exciting published findings that a novel fluorosugar analog can downregulate rolling receptor function on T cells and prevent the development of allergic dermatitis. These studies have exposed a contrary view into the putative role of PSGL-1 as “the” rolling receptor on T cell. As described above, PSGL-1 not only has been implicated as a major rolling receptor on white blood cells, but as a major rolling receptor on metastatic prostate tumor cells. However, data from analyses of fluorosugar-inhibitory effects on T cell rolling receptor function suggest that PSGL-1 may not be critical for T cell migration to inflamed skin. The Dimitroff Laboratory has recently discovered that a non-PSGL-1 glycoprotein is critical for the development allergic dermatitis. This exciting finding with implications to the discovery of novel anti-inflammatory target has led to the development of investigations and Project 2.
Project 2: Identification and Characterization of a Novel Skin-Homing Receptor on Inflammatory T cells
Binding interactions between T cell rolling receptors and counter-receptors on blood vessel linings in skin are critical for the development of T cell-mediated dermatitis. The Dimitroff Laboratory hypothesizes that inhibiting the function of T cell receptors can help control T cell entry into inflamed skin. The most widely-studied T cell rolling receptor is PSGL-1. PSGL-1 on in vitro-cultured T cells is specialized to interact with counter-receptors on skin microvessels and therefore is viewed as the “skin-homing” receptor. However, the Dimitroff Laboratory has obtained compelling data demonstrating that PSGL-1 does not function as a major T cell rolling receptor, in vivo. They show that the potential rolling receptor repertoire on T cells freshly-isolated from mice with allergic dermatitis is a previously unrecognized glycoprotein. The objectives of studies outlined in this project are 1.) To investigate the expression and structural biochemistry of candidate rolling receptors on fresh T cells from mice with allergic dermatitis and 2.) To identify and characterize T cell membrane protein(s) that bear critical structural motifs conferring rolling function. Numerous biochemical approaches are being employed to elucidate the expression, identity and structural biochemistry of candidate T cell rolling receptor (s). Innovative cell adhesion assays are also being employed, including the novel “blot rolling” assay, which examines the rolling activity of resolved protein(s) immobilized on Western blots. Results from these analyses could offer a new target for the treatment of T cell-mediated skin diseases, including psoriasis, atopic dermatitis, allergic dermatitis and cutaneous lymphomas.
To appreciate the functional role of T cell rolling receptors in T cell trafficking to skin, the Dimitroff Laboratory has embarked on development of a mutant mouse, which can be induced to produce T cell inflammation in skin. These studies are being pursued in Project 3 as follows.
Project 3: Generation of a Mouse Model of Cutaneous T cell-Mediated Inflammation
Currently, mouse models of allergic contact dermatitis characteristically involve the recruitment of T cells to inflamed skin. In fact, these models are routinely used for evaluating experimental agents that are designed specifically to disrupt the trafficking of T cells to inflamed skin. However, inflammation in these mice require other additional processes, including T cell antigen priming, antigen-presenting cell migration to lymph nodes draining antigen-sensitized skin and naïve T cell entry into skin-draining lymph nodes. To directly address whether experimental agents designed to inhibit T cell rolling receptor function are effective, the Dimitroff Laboratory is interested in developing a mouse model of cutaneous inflammation that is specifically controlled by functional T cell rolling receptor(s). It has been shown that the enzyme, α1,3 fucosyltransferase 7 (FucT-7), is critical for creation of functional T cell rolling receptor(s). The objective of this project is to generate mice that overexpress FucT-7 in all T cells only upon treatment with an inducer drug, tetracycline. They believe that, after induction, mice will develop inflammation in the skin, and, after removal of the inducing agent, mice will revert to normal trafficking behavior. This rapid induction of T cell trafficking to skin in concert with treatments of experimental anti-inflammatory agents will help test the hypothesis that T cell rolling receptors are suitable targets for cutaneous anti-inflammatory/cancer therapeutics.
