Faculty Bibliography

We are developing recombinant atoxic derivatives of botulinum neurotoxin (BoNT) as molecular vehicles that can specifically deliver therapeutics to the cytoplasmic compartment of neurons. For proof-of-concept, we have fused an atoxic derivative of full-length BoNT/C1 (BoNT/C1ad) to a single domain antibody (B8) that specifically inactivates the light chain (LC) protease of BoNT/A1. The antibody fusion protein, termed B8-C1ad, was expressed using a baculovirus Sf9 system, and purified to homogeneity using 2-step tandem affinity chromatography. A murine model of BoNT/A intoxication, was developed to test the effectiveness of B8-C1ad as a function of administration time post-exposure to wt BoNT/A. Mice were intoxicated with 2 LD50 of wt BoNT/A by ip administration. At 3, 6, 8, 12, 16 or 20 h post-intoxication, mice (N = 10 per group) were treated ip with either saline, 10 ug B8-C1ad (0.4 mg/kg), or 1 U of a standard anti-BoNT/A antibody (enough to neutralize 10,000 LD50 of BoNT/A). Treatment with B8-C1ad resulted in 100% survival when administered up to 12 hours post exposure, compared to the standard antibody which was only 30% effective by 12 hours. At 16 and 20 hours post-exposure, B8-C1ad treatment resulted in 80% and 70% survival, respectively, compared to 0% survival in the standard antibody groups. The BoNT/C1ad molecular vehicle and the B8 antibody alone were ineffective under similar conditions. Surviving animals in the B8-C1ad group resumed weight gain approximately 3-4 days post-treatment, and no rebound of symptoms was observed over a 30 day period. These data demonstrate that B8-C1ad can reverse respiratory symptoms of botulism, and enable survival when administered at times post-exposure when available treatments are ineffective. Importantly, the data strongly suggest that B8-C1ad can inactivate the BoNT/A light chain protease exposed inside the presynaptic compartment of intoxicated neurons. To our knowledge this represents the first example of an antibody specifically delivered to cure an intra-neuronal disease, using a molecular vehicle for delivery instead of a virus. B8-C1ad also addresses the practical need for an anti-botulism therapeutic that can rescue patients exposed to BoNT, when mechanical ventilation is impractical

Botulinum neurotoxin (BoNT) binds to and internalizes its light chain into presynaptic compartments with exquisite specificity. While the native toxin is extremely lethal, bioengineering of BoNT has the potential to eliminate toxicity without disrupting neuron-specific targeting, thereby creating a molecular vehicle capable of delivering therapeutic cargo into the neuronal cytosol. Building upon previous work, we have developed an atoxic derivative (ad) of BoNT/C1 through rationally designed amino acid substitutions in the metalloprotease domain of wild type (wt) BoNT/C1. To test if BoNT/C1 ad retains neuron-specific targeting without concomitant toxic host responses, we evaluated the localization, activity, and toxicity of BoNT/C1 ad in vitro and in vivo. In neuronal cultures, BoNT/C1 ad light chain is rapidly internalized into presynaptic compartments, but does not cleave SNARE proteins nor impair spontaneous neurotransmitter release. In mice, systemic administration resulted in the specific co-localization of BoNT/C1 ad with diaphragmatic motor nerve terminals. The mouse LD50 of BoNT/C1 ad is 5 mg/kg, with transient neurological symptoms emerging at sub-lethal doses. Given the low toxicity and highly specific neuron-targeting properties of BoNT/C1 ad, these data suggest that BoNT/C1 ad can be useful as a molecular vehicle for drug delivery to the neuronal cytoplasm.

Cyto-012 is a recombinant derivative of Botulinum neurotoxin Type A (BoNT/A). It primarily differs from wild type (wt) BoNT/A1 in that it incorporates two amino acid substitutions in the catalytic domain of the light chain (LC) metalloprotease (E224 > A and Y366 > A), designed to provide a safer clinical profile. Cyto-012 is specifically internalized into rat cortical and hippocampal neurons, and cleaves Synaptosomal-Associated Protein 25 (SNAP-25), the substrate of wt BoNT/A, but exhibits slower cleavage kinetics and therefore requires a higher absolute dose to exhibit pharmacologic activity. The pharmacodynamics of Cyto-012 and wt BoNT/A have similar onset and duration of action using the Digital Abduction Assay (DAS). Intramuscular LD50 values for Cyto-012 and wt BoNT/A respectively, were 0.63 ug (95% CI = 0.61, 0.66) and 6.22 pg (95% CI = 5.42, 7.02). ED50 values for Cyto-012 and wt BoNT/A were respectively, 0.030 ug (95% CI = 0.026, 0.034) and 0.592 pg (95% CI = 0.488, 0.696). The safety margin (intramuscular LD50/ED50 ratio) for Cyto-012 was found to be improved 2-fold relative to wt BoNT/A (p < 0.001). The DAS response to Cyto-012 was diminished when a second injection was administered 32 days after the first. These data suggest that the safety margin of BoNT/A can be improved by modulating their activity towards SNAP-25.

BACKGROUND: Preoperative risk stratification and optimization of preoperative care may be helpful in reducing readmission rates after primary total joint arthroplasty. Assessment of the predictive value of individual modifiable risk factors without a tool to assess cumulative risk may not provide proper risk stratification of patients with regard to potential readmissions. As part of a Perioperative Orthopaedic Surgical Home model, we developed a scoring system, the Readmission Risk Assessment Tool (RRAT), which allows for risk stratification in patients undergoing elective primary total joint arthroplasty at our institution. The purpose of this study was to analyze the relationship between the RRAT score and readmission after primary hip or knee arthroplasty. METHODS: The RRAT, which is scored incrementally on the basis of the number and severity of modifiable comorbidities, was used to generate readmission scores for a cohort of 207 readmitted patients and two cohorts (one random and one age-matched) of 234 non-readmitted patients each. Regression analysis was performed to assess the strength of association of individual risk factors and the RRAT score with readmissions. We also calculated the odds and odds ratio (OR) at each RRAT score level to identify patients with relatively higher risk of readmission. RESULTS: There were 207 (2.08%) readmissions among 9930 patients over a six-year period (2008 through 2013). Surgical site infection was the most common cause of readmission (ninety-three cases, 45%). The median RRAT scores were 3 (IQR [interquartile range], 1 to 4) and 1 (IQR, 0 to 2) for readmitted and non-readmitted groups, respectively. An RRAT score of >/=3 was significantly associated with higher odds of readmission. CONCLUSIONS: Population health management, cost-effective care, and optimization of outcomes to maximize value are the new maxims for health-care delivery in the United States. We found that the RRAT score had a significant association with readmission after joint arthroplasty and could potentially be a clinically useful tool for risk mitigation.

BACKGROUND: Greater levels of self-reported pain, pain catastrophizing, and depression have been shown to be associated with persistent pain and functional limitation after surgeries such as TKA. It would be useful for clinicians to be able to measure these factors efficiently. QUESTIONS/PURPOSES: We asked: (1) What is the association of whole-body pain with osteoarthritis (OA)-related knee pain, function, pain catastrophizing, and mental health? (2) What is the sensitivity and specificity for different cutoffs for body pain diagram region categories in relation to pain catastrophizing? METHODS: Patients (n = 267) with knee OA undergoing elective TKA at one academic center and two community orthopaedic centers were enrolled before surgery in a prospective cohort study. Questionnaires included the WOMAC Pain and Function Scales, Pain Catastrophizing Scale (PCS), Mental Health Inventory-5 (MHI-5), and a pain body diagram. The diagram documents pain in 19 anatomic areas. Based on the distribution of the anatomic areas, we established six different body regions. Our analyses excluded the index (surgically treated) knee. Linear regression was used to evaluate the association between the total number of nonindex painful sites on the whole-body pain diagram and measures of OA-related pain and function, mental health, and pain catastrophizing. Generalized linear regression was used to evaluate the association between the number of painful nonindex body regions (categorized as 0; 1-2; or 3-6) with our measures of interest. All models were adjusted for age, sex, and number of comorbid conditions. The cohort included 63% females and the mean age was 66 years (SD, 9 years). With removal of the index knee, the median pain diagram score was 2 (25th, 75th percentiles, 1, 4) with a range of 0 to 15. The median number of painful body regions was 2 (25th, 75th percentiles, 1, 3). RESULTS: After adjusting for age, sex, and number of comorbid conditions, we found modest associations between painful body region categories and mean scores for WOMAC physical function (r = 0.22, p < 0.001), WOMAC pain (r = 0.20, p = 0.001), MHI-5 (r = -0.31, p < 0.001), and PCS (r = 0.27, p < 0.001). A nonindex body pain region score greater than 0 had 100% (95% CI, 75%-100%) sensitivity for a pain catastrophizing score greater than 30 but a specificity of just 23% (95% CI, 18%-29%) . A score of 3 or greater had greater specificity (73%; 95% CI, 66%-79%) but lower sensitivity (53%; 95% CI, 27%-78%). CONCLUSIONS: We found modest associations between the number of painful sites on a whole-body pain diagram and the number of painful body regions and measures of OA-related pain, function, pain catastrophizing, and mental health. Patients with higher self-reported body pain region scores might benefit from further evaluation for depression and pain catastrophizing. LEVEL OF EVIDENCE: Level III, therapeutic study.

The Centers for Medicare & Medicaid Services considers readmissions within 30 days of discharge to be a quality indicator. Hospitals' and eventually physicians' readmission rates will be used to determine payment for services. It is imperative that health care providers understand which patients are at risk for readmission so that they can apply the appropriate preventive interventions. The research team analyzed all orthopedic admissions and readmissions at their institution from September 2008 to April 2011 in this study. Preparing for the next stage in health care reform, identifying any preoperative factors that may place certain patients into a "high-risk" category for readmission following an orthopedic procedure is of paramount importance. This data analysis of more than 13 000 patients noted that race-based and income-based risk factors did not translate into significant risk factors or predictors of 30-day readmission following orthopedic admission.

OBJECTIVES: Osteoarthritis of the knee is a chronic disease associated with pain and reduced quality of life. The ability to reliably measure patient-reported symptoms is important for clinical decision making and evaluation of outcomes. Electronic and web-based tools can eliminate much of the labor-intensive aspects of questionnaire administration and enables both real-time evaluation of responses by physicians and integration of data from multiple sites. This article describes the results of implementing a single integrated electronic questionnaire system into routine orthopedic practice at 2 diverse institutions. STUDY DESIGN: Case study. METHODS: A web-based version of a general quality-of-life questionnaire (EuroQol 5-dimension [EQ-5D]) and the pain domain of a disease-specific questionnaire (Knee Osteoarthritis Outcome Score [KOOS]) were administered in the office waiting room to (n = 666) patients at 2 centers over a 9-month period using touchscreen devices. Data were analyzed and descriptive statistics were calculated to assess feasibility of integration into the distinct work flows and to assess the agreement of the results. RESULTS: The electronic questionnaire had a completion rate of 93% to 95%. Average questionnaire completion times were 3 to 5 minutes at each institution. Mean EQ-5D and KOOS scores for patients pre- and postsurgery were also consistent with prior literature studies. CONCLUSIONS: Lessons learned for future adoption of questionnaire systems elsewhere include the need for baseline assessment of clinic work flows to identify the optimal point of administration and the need for IT support. This study demonstrates the feasibility of routinely collecting patient-reported data as part of standard care, which will become increasingly important as the nationwide emphasis on tracking quality and cost-effectiveness of treatments in orthopedics grows.