Our group focuses on discovering and understanding fundamental relationships between molecular structure and photophysical behavior. Because of their remarkable versatility, we are particularly interested in establishing these correlations for archetype donor-acceptor dyes and their analogs. We use these findings to develop improved chromophores for use in biophysical, biological and medical applications such as single molecule detection, fluorescent reporting and photodynamic therapy. Our approach encompasses nearly every aspect that is essential to such an undertaking including computer-aided design, chemical synthesis, photophysical characterization and in-vivo application of the resulting dyes.
Over the years post doctoral fellows and other scientists have contributed to the our labs endeavors. Please see our group pages for a listing of these fellows. We have participated in collaborative research with many outstanding intramural and extramural colleagues. We particularly enjoy sharing our expertise and knowledge of dye chemistry with researchers inexperienced in that field.
Primary brain tumors account for 2-3% of all diagnosed cancers each year in the United States and are the sixth leading cause of cancer induced mortality in the adult population. Gliomas, the most common type of malignant brain tumor, are aggressive infiltrative tumors that damage the brain by disruptive local effects. There are no known treatments that are curative for this disease. Diagnosis of grade 4 glioblastoma invariably leads to death with a survival time of less than one year; methods that are palliative or which extend life by several months are considered successful. Many in the medical community believe that patients could realize significant benefit if complete tumor resection could be achieved. This objective remains a daunting challenge because the tumor infiltrates the surrounding normal brain tissue with small tendrilous colonies of neoplastic cells which are extremely difficult to detect; this problem is exacerbated by the constraint that healthy brain tissue cannot be indiscriminately resected at these tumor margins for fear of causing harm or death to the patient.
Because of the obvious importance of the problem, a variety of approaches for improving tumor demarcation are currently being investigated. In light of the great sensitivity possible with fluorescence techniques, there have been surprisingly few reports of the use of endogenous chromophores and exogenous fluorescent agents as tumor margin contrast markers; furthermore, the exogenous dyes that have been investigated for this purpose, with few exceptions, have been developed as photodynamic agents and, as a consequence, have the potential to damage normal brain tissue. With this in mind we participated in a collaborative effort with Dr. Demetrios Nikas and Dr. Peter Black, colleagues at Harvard Medical School (Children's and Brigham and Women's Hospitals), with the goal of developing fluorescent, non-photosensitizing dyes as selective tumor stains to aid the surgeon in the real time demarcation of tumor margins during resection. The successful results of that study using an intracranial glioma in a murine model were recently reported in "Lasers in Surgery and Medicine" 29:11-17 (2001).
Poorna Mahalingam is a postdoctoral fellow working on new fluorescent dyes. A list of previous lab members includes: Steven Song, Joel Blush, Louis Cincotta, Barbara Fournier, Angela Frimberger, Daniel Gloster, and Sara Tuttle.
Our primary objective in this area has been the rational development of drugs for the treatment of solid tumors and for the eradication of pathogens in whole blood. When we entered the field of photodynamic therapy (PDT), there were few systematic efforts to develop drugs from classes of compounds other than those belonging to the porphyrin family. This remains true today. Our research is based on the premise that any effort to extend and/or improve the applicability of PDT would require the use of photosensitizers that have inherently different physicochemical and pharmacological properties from the porphyrins. We chose to study the benzophenoxazine family of dyes because they possess many of the properties that characterize a good PDT agent. For example, they preferentially localize in neoplastic cells, are relatively non-toxic in the dark, absorb red light efficiently and are rapidly eliminated from the body. A key finding is that some of the novel photosensitizers we have developed inactivate tumors via a different mechanism than is operative with the porphyrins: benzophenothiazine PDT drugs, such as EtNBS, mediate the destruction of neoplasia by directly damaging intracellular organelles whereas porphyrin derivatives act by shutting down the vasculature that support tumor tissue. Thus the two approaches are complimentary and, when used simultaneously, synergistic.
In collaboration with physicians from Tufts School of Veterinary Medicine we recently demonstrated the effectiveness of the benzophenothiazine dye EtNBS against spontaneous tumors in pet cats and dogs (Clinical Cancer Research 4: 2207-2218 (1998). Below we present pre- and post-treatment photographs of Blaze, one of the successes of this study.
2mg/Kg EtNBS (i.v.)
Two Yrs. Post Initial PDT
Many seminal advances in modern molecular biology and medicine are possible because of the availability of sophisticated fluorescent reporter compounds that, with exquisite selectivity and sensitivity, provide the means to observe structure-function relationships in biological systems. Although this use of organic fluorophore reporters is not new, it remains a dynamic and growing technology that continues to be used to solve important new problems. Currently there is considerable interest in employing this methodology in single molecule detection to study individual events in biochemicals. As is the case with any technology, the use of fluorescent probes has limitations that may frustrate the experimentalist from taking full advantage of the promise of the method. One of the most severe is the constraint related to fluorophore photodecomposition (bleaching) which sets strict limits both on the time that chromophores can be monitored and on the illumination intensity that can be used to interrogate the sample. As a consequence, researchers are often precluded from exploiting one of the most powerful tenets of fluorescence spectroscopy which teaches that the signal strength observed for a fluorophore (within limits) increases linearly with the intensity of the excitation.
Renewed demands for dyes better suited for fluorescence reporting has stimulated us to initiate a research program in collaboration with our Rowland colleague Dr. Amit Meller, a biophysicist with expertise in single molecule detection, with the goal of developing a series of improved fluorophores having much better light- and dark-stabilities, more ideal photophysical characteristics (i.e. high quantum yields that are invariant with temperature or environment) and which collectively have absorption and emission bands that span the visible and near-infrared spectral regions.