A Professor of Orthopaedic Surgery, Ed Greenfield, PhD, joined the Indiana University School of Medicine Department of Orthopaedic Surgery in 2019 after a long career on the faculty at Case Western Reserve University in Cleveland. The Greenfield Lab at Indiana University School of Medicine has long-standing research interests focused on basic and translational aspects of bone biology, and on osteosarcoma metastasis.
How did you decide to focus your career in oncology and specifically what drove your interest in osteosarcoma?
My background is in bone cell biology. When I decided to add cancer to our research program, I chose osteosarcoma rather than the more common cancers that metastasize to bone because we ultimately need to understand the difference between normal cells and the cancer cells, and my background should assist with that understanding regarding osteosarcoma but would be less useful for epithelial cancers that metastasize to bone. Also, osteosarcoma was less studied than the other cancers, but there were good cell and animal models available.
Your lab is focused on basic and translational aspects of bone biology, and on osteosarcoma metastasis. Can you tell us about your active research projects related to osteosarcoma?
Our main osteosarcoma project focuses on repurposed drugs that are FDA-approved for other cancers. It is a close collaboration with Chris Collier, MD, a orthopaedic oncology clinician-scientist whose lab is adjacent to our lab. We initially screened the NCI panel of 114 FDA-approved oncology drugs using the sarcosphere platform that we developed for that purpose. Since then, we have been following up the top hits from that screen (the HDAC inhibitor romidepsin and the proteasome inhibitors bortezomib and carfilzomib) in both the sarcosphere platform. We are evaluating how well sarcosphere responses mimic in vivo responses by comparing sarcosphere responses to romidepsin with responses in murine models of metastatic osteosarcoma and comparing responses to MAP by sarcospheres from pre-chemo biopsies with clinical responses (% necrosis at post-chemo resection).
Infections are a major issue for limb salvage patients. Your lab is studying a novel antibacterial agent called halicin to see if it is effective against bacterial biofilms in vitro and in mice. If effective in pre-clinical models, how might this translate to clinical application?
Infections are a major complication in many areas of orthopaedic surgery in addition to limb salvage patients. If halicin is effective in the pre-clinical models, its initial translational application would likely be the more common infections that occur in ~30% of patients with open fractures. If effective in those patients, we would be very interested in translation to limb salvage patients.
Your current studies utilize 3D sarcosphere cultures, in addition to mouse models. How do you develop the sarcospheres, and what pros/cons have you seen versus mouse models in studying osteosarcoma specifically?
Our protocol to obtain the 3D sarcospheres is based on low-speed centrifugation to form a disc of cells in the bottom of each well of a 96-well plate that were previously coated to prevent attachment. The discs of cells round up into a single tight spheroid per well over the next 24 hours and we begin drug testing at that time. The sarcospheres likely have many advantages:
1. A very simple, relatively high-throughput, method is used to reproducibly obtain a large number of sarcospheres of the size (~400 um diameter) to generate the desired gradients of oxygen, nutrients, metabolic waste products, etc. Sarcosphere size is easily adjusted by selecting the appropriate number of cells in each well, which needs to be determined separately for each cell line and patient sample.
2. The sarcospheres very likely better mimic actual metastases than is achieved by 2D monolayers of cells.
3. The sarcospheres have the potential to direct personalized medicine for osteosarcoma patients as results of drug testing with sarcospheres from an individual patient can be available in 2-3 weeks rather than the many months that would be needed to test PDXs in mice. Moreover, functional testing with sarcospheres is likely to be more successful to direct personalized medicine than genomics for cancers like osteosarcoma where each patient has multiple copy number alterations but few mutations, and the genomics vary greatly between individual patients.
What advice would you give to early career investigators who have an interest in studying osteosarcoma?
My advice would be to identify good mentors, supportive research centers, and projects that excite you and have both translational relevance and interesting biology. Don’t always pick the easiest approach to an experimental question; if it’s worth devoting time and effort, it’s worth doing it the best way that’s feasible rather than the easiest way. Sustained funding is always difficult in our biomedical research system. It’s therefore very important to be persistent and not take rejections personally. It is also very important to carefully evaluate the reviewers’ critiques as they will often have excellent suggestions for your research. If the reviewers appear to have misunderstood portions of your application, it is likely because those portions were not clearly explained and you can do better in future applications.