Research Objectives and Rationale: Our research objective is to identify a biomarker that can be used as an early diagnostic test by military doctors to identify patients that are likely to form heterotopic ossification (HO) following orthopaedic trauma related to extremity blast injury in the battlefield setting. We have previously characterized the biology of heterotopic ossification from study of ectopic bone and the adjacent soft tissues from war-injured limbs of soldiers, and have shown that the processes of heterotopic ossification seen in both civilian practice and war trauma share a similar biology several weeks after the initial injury. Moreover, we have developed a unique animal model in a laboratory rat that reproduces the process of HO following blast amputation. Our preliminary data confirm that the biologic events that characterize HO weeks after blast amputation in the rat closely resemble those observed in battle-injured soldiers who develop HO. We now propose to use the same animal model to further our understanding of HO in humans and develop a way to prevent its occurrence. This will be accomplished in three stages:

(1) We will first correlate the presence of molecular and cellular markers from banked human HO tissue specimens with our animal model corresponding to time points several weeks after blast injury.

(2) Next, we will identify early-appearing gene- and protein-level expression at 24 to 72 hours post-blast in the animal model that predicts eventual development of HO.

(3) Finally, after validating similarities of human and animal HO (#1), and identifying early markers of HO in the animal model (#2), we will evaluate the specificity of known early and late candidate biomarkers to predict severe HO formation in human tissues obtained 24 to 72 hours after initial injury.

Background: Heterotopic ossification is the formation of mature bone in the soft tissues and is a frequent complication after orthopaedic trauma; it is commonly seen after total hip replacement and acetabular fracture in civilian practice and occurs in up to two-thirds of patients who sustain severe extremity wartime wounds and multiple trauma by blast injury. In recent military conflicts in Iraq and Afghanistan there has been a marked increase in the number of combat-related blast amputations of one or more extremities. Advanced battlefield evacuation capabilities and widespread use of body armor have substantially reduced mortality for soldiers with these severe limb-compromising events. Having survived the initial trauma and limb amputation, several possible complications often arise in the residual limb related to HO including pain, overlying skin and muscle breakdown, a poorly fitting prosthesis, reoperation for revision of the amputated part, and delayed rehabilitation that can compromise overall function of the residual limb, and jeopardize return to productive civilian life.

Clinical Scenario and Applicability: Soldiers who are currently injured during military operations in Afghanistan are typically transported to the Landstuhl Regional Medical Center in Germany within 24 to 72 hours, where clinicians will have access to the equipment that is necessary to perform testing on the dead and injured tissues removed from the amputated limbs in search of markers that predict likelihood of HO. Our preliminary data indicate that patients who develop HO have already begun to have an abnormal bone-forming response within 2 weeks of their initial injury; we actually expect that the initial cellular events that trigger this bone-making function are turned on within hours of injury. By identifying these early markers of HO within 3 days of injury, we will enable clinicians to begin selective HO prevention measures for these patients within the 5-day window that is known to be necessary for their effectiveness in civilians. Because nearly all therapies are accompanied by some incremental risk of treatment-related complications, and use of NSAIDs and radiation therapy for HO prevention in civilian practice do have some modest associated side-effects, it is important to develop a screening test that will identify those at greatest risk in order to limit HO preventive measures to this group. Additionally, as newer HO therapies emerge, targeting specific at-risk patients early in the disease process will result in the best, most cost-effective outcomes. As seen in civilian practice after total hip replacement, such intervention might be expected to substantially reduce, or even eliminate, the complications attributable to HO in residual limbs after blast amputation. This will expedite prosthetic limb use, rehabilitation, and a return to productive life. After validating this predictive biomarker assay in war-injured patients from LRMC, we plan to initiate an interventional clinical trial for HO prophylaxis using our assay to identify patients at greatest risk. Thus, we anticipate patients will begin benefitting from this study approximately 3 years following project initiation.