Amputation of a limb can often result in problems beyond the obvious life-changing loss of the use of the limb. These problems can occur because of uncontrolled growth of nerves adjacent to the amputation site (neuroma), as well as bone growth outside of the skeleton (heterotopic ossification), presumably as an attempt by the body to replace the missing limb. Both processes present major therapeutic challenges to clinicians in orthopedic and trauma surgery. Recent reports from the military suggest that limb pain after amputation is linked to the induction of bone formation (heterotopic ossification) in limb stump, which occurs in approximately 63% of wounded warriors receiving amputation. Our own studies have also demonstrated a central role for pain mediators in inducing this type of aberrant bone formation. In addition to the potential of unwanted bone leading to pain, almost all amputations result in the formation of neuromas within the stump. A neuroma is a benign tumor within the nerve, that can, 10%-25% of the time, lead to significant pain. The formation of neuroma and HO cannot only be painful, but can lead to difficulties fitting and maintaining prosthetics. This ultimately reduces mobility, resulting in significantly lower quality of life for these wounded warriors, who are already dealing with the loss of a limb. Finally, in approximately 5%-10% of these wounded warriors, the pain and sensitivity is so significant that even standard analgesics do not seem to help. Unfortunately, our understanding of the peripheral nervous system and the regulation of pain, particularly with respect to regeneration of tissues, is still in its infancy.

Recent work in our laboratory found that the first step in inducing bone formation involves changes in peripheral nerves involved in sensory perception. These sensory nerves respond to growth factors known to be released from bone matrix during injury, to undergo release of pain mediators, leading to significant pain. However, these pain mediators, during normal wound repair, actually aid in the regenerative process, because they appear to expand progenitors within the nerve that are essential to heal the bone. We hypothesize that in patients receiving amputation, these bone growth factors are released and sensory nerves attempt to respond and repair. However, these sensory nerves have also been injured during the amputation and thus an aberrant repair process is likely responsible in part for neuroma and/or heterotopic bone formation or both.

Our laboratory has recently characterized this pathway and identified a novel adult stem cell within the nerve that appears to coordinate regeneration. In this application we propose that this cell coordinates repair in traumatic injury or amputation under ordinary circumstances, but during amputation becomes deregulated, which leads to adverse painful events. If we can successfully isolate these cells, then we propose that they can be used to both potentially repair peripheral nerves as well as other tissues such as bone and blood vessels, allowing for increased limb salvage. Additionally, we should be able to regulate this pathway so that, either, we can suppress this process after amputation or we can activate it during regeneration.

We have assembled a team including physicians in orthopedic surgery and neurology along with translational scientists to develop these systems for rapid clinical use. The work proposed will provide key data to complement other data already obtained, and allow us to put together the strong evidence in two animal models needed for preclinical data and assembly of an investigational new drug (IND) application with the FDA. This would be the first step in translating these novel cells and function into the clinic. Our Center for Cell and Gene Therapy (CAGT) has the manufacturing facilities to produce cell products that can be used in clinical trials. Currently, the CAGT has approval from the FDA to test several therapies for different indications that would be in the identical category as the system we currently propose. Therefore, we believe that we have the infrastructure to be able to bring a cellular therapy to the clinic in a safe and effective manner. In addition, the concepts demonstrated by completion of this work would be paradigm-shifting and could potentially cross to a wide variety of additional therapeutic areas such as diabetes and other diseases of peripheral nerves as well as tissue regeneration.