Improved Clinical Outcomes After Partial Repair and In-Situ Biceps Tenodesis for the Treatment of Massive Irreparable Rotator Cuff Tears.

PURPOSE: The aim of this study was to present the short-term outcomes of arthroscopic in-situ biceps tenodesis combined with partial rotator cuff repair in patients with massive irreparable rotator cuff tears (MIRCT) and minimal arthritis. METHODS: A retrospective review was conducted utilizing prospectively maintained institutional databases to identify patients who had undergone a partial rotator cuff repair with in-situ biceps tenodesis between March 2017 to December 2022. Patients were included if they (1) were diagnosed pre- or intra-operatively with MIRCT, and (2) had complete pre-operative and minimum 1-year post-operative patient-reported outcome measures. RESULTS: Thirty-nine patients met the eligibility criteria and were included for analysis. The mean age of the study participants was 65 ± 7 years (range: 46 - 76) with 76.9% (n = 30) being male. The average follow-up was 21 ± 12 months (range: 12 - 58). Patients experienced significant improvement in visual analog scale for pain (VAS), American Shoulder and Elbow Surgeons (ASES) score, and subjective assessment numeric evaluation (SANE) score (p < 0.001 for all comparisons). The pre- to post-operative improvement was 3.1 ± 2.3 for VAS, 27.5 ± 20.6 for ASES, and 31.3 ± 24.8 for SANE. Postoperatively, the average scores for VAS, ASES, and SANE were 1.3 ± 1.5, 79.5 ± 17.0, and 69.6 ± 20.1, respectively. Twenty-six patients (66.7%) achieved the minimal clinically important difference (MCID) for VAS, thirty-three patients (84.6%) achieved the MCID for ASES, and thirty patients (76.9%) achieved the MCID for SANE.. CONCLUSIONS: Partial rotator cuff repair with in-situ biceps tenodesis is an effective treatment for MIRCT, leading to significant improvements in patient-reported outcome and range of motion measures compared to preoperative conditions.

Predictors of poor and excellent outcomes following reverse shoulder arthroplasty for glenohumeral osteoarthritis with an intact rotator cuff.

BACKGROUND: As the indications for reverse total shoulder arthroplasty (RSA) continue to evolve, it has been more commonly utilized for the treatment of glenohumeral osteoarthritis with an intact rotator cuff (GHOA). Given the increased use of RSA for GHOA, it is important to identify factors influential of clinical outcomes. In this study, we sought to identify variables predictive of clinical outcomes following RSA for GHOA. METHODS: Patients undergoing primary RSA for GHOA between 2015 and 2020 were retrospectively identified through a prospectively maintained, single surgeon registry. Eligible patients had complete patient-reported outcome measures and range of motion measurements with a minimum 2-year follow-up. Univariate analysis was utilized to compare characteristics and outcome measures of patients with poor and excellent outcomes, which was defined as postoperative American Shoulder and Elbow Surgeons (ASES) scores in the bottom and top quartiles, respectively. Multivariate linear regression was performed to determine factors independently predictive of postoperative ASES score. RESULTS: A total of 230 patients were included with a mean follow-up of 33.4 months (SD 13.2). The mean age of the study population was 71.9 (SD 6.1). Two hundred twenty-four patients (97.4%) surpassed the minimal clinically important difference and 209 patients (90.1%) achieved substantial clinical benefit for ASES score. Preoperative factors differing between the poor and excellent outcome groups were sex (male: poor 37.9%, excellent 58.6%; P = .041), opioid use (poor 24.1%, excellent 5.2%; P = .009), ASES score (poor 32.9, excellent 41.0; P = .011), and forward elevation (poor 92°, excellent 101°; P = .030). Linear regression demonstrated that Walch B3 glenoids (ß 7.08; P = .010) and higher preoperative ASES scores (ß 0.14; P = .025) were predictors of higher postoperative ASES score, while postoperative complications (ß -18.66; P < .001) and preoperative opioid use (ß -11.88; P < .001) were predictive of lower postoperative ASES scores. CONCLUSION: Over 90% of patients who underwent RSA for GHOA with an intact rotator cuff experienced substantial clinical benefit. An unsurprising handful of factors were associated with postoperative clinical outcomes; higher preoperative ASES scores were slightly associated with higher postoperative ASES, whereas preoperative opioid use and postoperative complications were associated with lower postoperative ASES. Additionally, Walch glenoid type B3 was associated with higher postoperative ASES, indicating that patients with posterior glenoid defects are not predisposed to poor clinical outcomes following RSA. These results serve as a resource to improve preoperative patient counseling and manage postoperative expectations.

Effect of vitamin E-enhanced highly cross-linked polyethylene on wear rate and particle debris in anatomic total shoulder arthroplasty: a biomechanical comparison to ultrahigh-molecular-weight polyethylene.

BACKGROUND: Particle-induced osteolysis resulting from polyethylene wear remains a source of implant failure in anatomic total shoulder designs. Modern polyethylene components are irradiated in an oxygen-free environment to induce cross-linking, but reducing the resulting free radicals with melting or heat annealing can compromise the component's mechanical properties. Vitamin E has been introduced as an adjuvant to thermal treatments. Anatomic shoulder arthroplasty models with a ceramic head component have demonstrated that vitamin E-enhanced polyethylene show improved wear compared with highly cross-linked polyethylene (HXLPE). This study aimed to assess the biomechanical wear properties and particle size characteristics of a novel vitamin E-enhanced highly cross-linked polyethylene (VEXPE) glenoid compared to a conventional ultrahigh-molecular-weight polyethylene (UHMWPE) glenoid against a cobalt chromium molybdenum (CoCrMo) head component. METHODS: Biomechanical wear testing was performed to compare the VEXPE glenoid to UHMWPE glenoid with regard to pristine polyethylene wear and abrasive endurance against a polished CoCrMo alloy humeral head in an anatomic shoulder wear-simulation model. Cumulative mass loss (milligrams) was recorded, and wear rate calculated (milligrams per megacycle [Mc]). Under pristine wear conditions, particle analysis was performed, and functional biologic activity (FBA) was calculated to estimate particle debris osteolytic potential. In addition, 95% confidence intervals for all testing conditions were calculated. RESULTS: The average pristine wear rate was statistically significantly lower for the VEXPE glenoid compared with the HXLPE glenoid (0.81 ± 0.64 mg/Mc vs. 7.00 ± 0.45 mg/Mc) (P < .05). Under abrasive wear conditions, the VEXPE glenoid had a statistically significant lower average wear rate compared with the UHMWPE glenoid comparator device (18.93 ± 5.80 mg/Mc vs. 40.47 ± 2.63 mg/Mc) (P < .05). The VEXPE glenoid demonstrated a statistically significant improvement in FBA compared with the HXLPE glenoid (0.21 ± 0.21 vs. 1.54 ± 0.49 (P < .05). CONCLUSIONS: A new anatomic glenoid component with VEXPE demonstrated significantly improved pristine and abrasive wear properties with lower osteolytic particle debris potential compared with a conventional UHMWPE glenoid component. Vitamin E-enhanced polyethylene shows early promise in shoulder arthroplasty components. Long-term clinical and radiographic investigation needs to be performed to verify if these biomechanical wear properties translate to diminished long-term wear, osteolysis, and loosening.

Outcomes after reverse shoulder arthroplasty for the treatment of glenohumeral osteoarthritis in patients under and over 70 years of age: a propensity score-matched analysis.

INTRODUCTION: As reverse shoulder arthroplasty (RSA) continues to grow in popularity for the treatment of glenohumeral osteoarthritis (GHOA) with an intact rotator cuff, it becomes increasingly important to identify factors that influence postoperative outcome. Although recent studies have demonstrated excellent postoperative range of motion and patient-reported outcome scores following RSA for GHOA, there continues to be surgeon hesitation to adopt RSA as a viable treatment in the younger patient population due to greater functional demands. In this study, we sought to determine the effect of age on clinical outcomes following RSA for GHOA through a comparison of patients over and under the age of 70. METHODS: A retrospective review of prospectively collected data from an institutional registry was performed. Propensity score matching was utilized to match patients under the age of 70 (U-70) to those over 70 (O-70) in a 1:1 ratio based on sex, body mass index (BMI), preoperative ASES score, preoperative active forward elevation (FE), Walch classification, and American Society of Anesthesiologists comorbidity score. Clinical outcomes obtained preoperatively and at a minimum of 2 years postoperatively consisted of Visual Analog Scale (VAS) for pain, Single Assessment Numeric Evaluation (SANE) score, and American Shoulder and Elbow Surgeons (ASES) score, as well as active (FE), internal rotation, and external rotation. Descriptive statistics and univariate analysis were performed to compare cohorts. RESULTS: After matching, each cohort consisted of 66 patients with similar mean follow-up periods (U-70, 28.1 ± 7.5 months vs. O-70, 27.4 ± 7.5 months; P = .887). Mean age of the U-70 cohort was 66.2 ± 3.3 while the O-70 cohort had a mean age of 75.3 ± 3.8. Both groups demonstrated significant improvement in VAS, SANE, and ASES scores, as well as active range of motion in all planes. The only significant difference between cohorts was greater postoperative FE in younger patients (143 ± 16° vs. 136 ± 15°; P = .017), though the baseline-to-postoperative improvement in FE was similar between cohorts (50 ± 29° vs. 43 ± 29°, P = .174). CONCLUSION: RSA is a successful surgical treatment for GHOA regardless of age. Aside from greater postoperative FE in younger patients, there were no other differences in clinical outcomes between younger and older patients in this retrospective analysis, which compared patients who were matched by sex, BMI, and Walch classification, among other factors. Based on our results, 70 years of age should not be used as a threshold in preoperative counseling when determining whether a patient with GHOA with an intact rotator cuff is indicated for reverse shoulder arthroplasty.

Clinically significant outcome thresholds and rates of achievement by shoulder arthroplasty type and preoperative diagnosis.

BACKGROUND: Clinically significant outcome (CSO) benchmarks have been previously established for shoulder arthroplasty by assimilating preoperative diagnoses and arthroplasty types. The purpose of this study was to establish unique CSO thresholds and compare the time-to-achievement of these for reverse shoulder arthroplasty (RSA) for osteoarthritis (GHOA), RSA for rotator cuff arthropathy (RCA), and total shoulder arthroplasty (TSA) for GHOA. MATERIALS AND METHODS: Consecutive patients who underwent elective RSA for GHOA, TSA for GHOA, or RSA for RCA between February 2015 and May 2020, with 2-year minimum follow-up, were retrospectively identified from a prospectively maintained single surgeon registry. The American Shoulder and Elbow Surgeons (ASES) score was administered preoperatively and postoperatively at 2-week, 6-week, 3-month, 6-month, 1-year, and 2-year timepoints. Satisfaction and subjective overall improvement anchor questionnaires were administered at the time of final follow-up. Distribution-based methods were used to calculate the Minimal Clinically Important Difference (MCID), and anchor-based methods were used to calculate the Substantial Clinical Benefit (SCB) and the Patient Acceptable Symptom State (PASS) for each patient group. Median time to achievement, individual incidence of achievement at each time point, and cumulative incidence of achievement calculated using Kaplan-Meier survival curve analysis with interval censoring were compared between groups for each CSO. Cox-regression analyses were also performed to determine which patient factors were significantly associated with early or delayed achievement of CSOs. RESULTS: There were 471 patients eligible for study analysis: 276 RSA for GHOA, 107 TSA for GHOA, and 88 RSA for RCA. The calculated MCID, SCB, and PASS scores differed for each group. There were no significant differences in median time to achievement of any CSO between groups. Log-rank testing revealed that cumulative achievements significantly differed between groups for MCID (P = .014) but not for SCB (P = .053) or PASS (P = .620). On cox regression analysis, TSA patients had earlier achievement of SCB, whereas TSA and RSA for GHOA patients had earlier achievement of MCID. At 2-years, a significantly higher percentage of RSA for GHOA patients achieved MCID and SCB compared to RSA for RCA (MCID:100%, 95.5%, P = .003, SCB:94.6%, 86.4%, P = .036). CONCLUSION: Calculated CSO thresholds differ according to preoperative diagnosis and shoulder arthroplasty type. Patients undergoing TSA and RSA for GHOA achieve CSOs earlier than RSA for RCA patients, and a significantly higher percentage of RSA for GHOA patients achieve CSOs by 2 years compared to RSA for RCA patients.

Predictors of dislocations after reverse shoulder arthroplasty: a study by the ASES complications of RSA multicenter research group.

BACKGROUND: Instability after reverse shoulder arthroplasty (RSA) is one of the most frequent complications and remains a clinical challenge. Current evidence is limited by small sample size, single-center, or single-implant methodologies that limit generalizability. We sought to determine the incidence and patient-related risk factors for dislocation after RSA, using a large, multicenter cohort with varying implants. METHODS: A retrospective, multicenter study was performed involving 15 institutions and 24 American Shoulder and Elbow Surgeons members across the United States. Inclusion criteria consisted of patients undergoing primary or revision RSA between January 2013 and June 2019 with minimum 3-month follow-up. All definitions, inclusion criteria, and collected variables were determined using the Delphi method, an iterative survey process involving all primary investigators requiring at least 75% consensus to be considered a final component of the methodology for each study element. Dislocations were defined as complete loss of articulation between the humeral component and the glenosphere and required radiographic confirmation. Binary logistic regression was performed to determine patient predictors of postoperative dislocation after RSA. RESULTS: We identified 6621 patients who met inclusion criteria with a mean follow-up of 19.4 months (range: 3-84 months). The study population was 40% male with an average age of 71.0 years (range: 23-101 years). The rate of dislocation was 2.1% (n = 138) for the whole cohort, 1.6% (n = 99) for primary RSAs, and 6.5% (n = 39) for revision RSAs (P < .001). Dislocations occurred at a median of 7.0 weeks (interquartile range: 3.0-36.0 weeks) after surgery with 23.0% (n = 32) after a trauma. Patients with a primary diagnosis of glenohumeral osteoarthritis with an intact rotator cuff had an overall lower rate of dislocation than patients with other diagnoses (0.8% vs. 2.5%; P < .001). Patient-related factors independently predictive of dislocation, in order of the magnitude of effect, were a history of postoperative subluxations before radiographically confirmed dislocation (odds ratio [OR]: 19.52, P < .001), primary diagnosis of fracture nonunion (OR: 6.53, P < .001), revision arthroplasty (OR: 5.61, P < .001), primary diagnosis of rotator cuff disease (OR: 2.64, P < .001), male sex (OR: 2.21, P < .001), and no subscapularis repair at surgery (OR: 1.95, P = .001). CONCLUSION: The strongest patient-related factors associated with dislocation were a history of postoperative subluxations and having a primary diagnosis of fracture nonunion. Notably, RSAs for osteoarthritis showed lower rates of dislocations than RSAs for rotator cuff disease. These data can be used to optimize patient counseling before RSA, particularly in male patients undergoing revision RSA.

Indication matters: effect of indication on clinical outcome following reverse total shoulder arthroplasty-a multicenter study.

BACKGROUND: As the utilization and success of reverse total shoulder arthroplasty (RTSA) have continued to grow, so have its surgical indications. Despite the adoption of RTSA for the treatment of glenohumeral osteoarthritis (GHOA) with an intact rotator cuff and irreparable massive rotator cuff tears (MCTs) without arthritis, the literature remains sparse regarding the differential outcomes after RTSA among these varying indications. Thus, the purpose of this study was to examine the postoperative clinical outcomes of RTSA based on indication. METHODS: A retrospective review of 2 large institutional databases was performed to identify all patients who underwent RTSA between 2015 and 2019 with minimum 2-year follow-up. Patients were stratified by indication into 3 cohorts: GHOA, rotator cuff tear arthropathy (CTA), and MCT. Baseline demographic characteristics were collected to determine differences between the 3 cohorts. Clinical outcomes were measured preoperatively and postoperatively, including active range of motion, American Shoulder and Elbow Surgeons (ASES) score, Single Assessment Numeric Evaluation score, and visual analog scale pain score. Multivariate linear regression was performed to determine the factors independently predictive of the postoperative ASES score. RESULTS: A total of 625 patients (383 with GHOA, 164 with CTA, and 78 with MCTs) with a mean follow-up period of 33.4 months were included in the analysis. Patients with GHOA had superior ASES scores (85.6 ± 15.7 vs. 76.6 ± 20.8 in CTA cohort [P < .001] and 75.9 ± 19.9 in MCT cohort [P < .001]), Single Assessment Numeric Evaluation scores (86 ± 20.9 vs. 76.7 ± 24.1 in CTA cohort [P < .001] and 74.2 ± 25.3 in MCT cohort [P < .001]), and visual analog scale pain scores (median [interquartile range], 0.0 [0.0-1.0] vs. 0.0 [0.0-2.0] in CTA cohort [P < .001] and 0.0 [0.0-2.0] in MCT cohort [P < .001]) postoperatively. Postoperative active forward elevation (P < .001) and improvement in active external rotation (P < .001) were greatest in the GHOA cohort compared with other indications. Multivariate linear regression demonstrated that the factors independently associated with the postoperative ASES score included a diagnosis of GHOA (ß coefficient, 7.557 [P < .001]), preoperative ASES score (ß coefficient, 0.114 [P = .009]), female sex (ß coefficient, -4.476 [P = .002]), history of surgery (ß coefficient, -3.957 [P = .018]), and postoperative complication (ß coefficient, -13.550 [P < .001]). CONCLUSION: RTSA for the treatment of GHOA generally has superior patient-reported and functional outcomes when compared with CTA and MCTs without arthritis. Long-term follow-up is needed to identify the lasting implications of such outcome differences.

Quinoline Compounds Targeting the c-Ring of ATP Synthase Inhibit Drug-Resistant Pseudomonas aeruginosa.

Pseudomonas aeruginosa (PA) is a Gram-negative, biofilm-forming bacterium and an opportunistic pathogen. The growing drug resistance of PA is a serious threat that necessitates the discovery of novel antibiotics, ideally with previously underexplored mechanisms of action. Due to their central role in cell metabolism, bacterial bioenergetic processes are of increasing interest as drug targets, especially with the success of the ATP synthase inhibitor bedaquiline to treat drug-resistant tuberculosis. Like Mycobacterium tuberculosis, PA requires F1Fo ATP synthase for growth, even under anaerobic conditions, making the PA ATP synthase an ideal drug target for the treatment of drug-resistant infection. In previous work, we conducted an initial screen for quinoline compounds that inhibit ATP synthesis activity in PA. In the present study, we report additional quinoline derivatives, including one with increased potency against PA ATP synthase in vitro and antibacterial activity against drug-resistant PA. Moreover, by expressing the PA ATP synthase in Escherichia coli, we show that mutations in the H+ binding site on the membrane-embedded rotor ring alter inhibition by the reported quinoline compounds. Identification of a potent inhibitor and its probable binding site on ATP synthase enables further development of promising quinoline derivatives into a viable treatment for drug-resistant PA infection.

Risk factors of acromial and scapular spine stress fractures differ by indication: a study by the ASES Complications of Reverse Shoulder Arthroplasty Multicenter Research Group.

BACKGROUND: Both patient and implant related variables have been implicated in the incidence of acromial (ASF) and scapular spine fractures (SSF) following reverse shoulder arthroplasty (RSA); however, previous studies have not characterized nor differentiated risk profiles for varying indications including primary glenohumeral arthritis with intact rotator cuff (GHOA), rotator cuff arthropathy (CTA), and massive irreparable rotator cuff tear (MCT). The purpose of this study was to determine patient factors predictive of cumulative ASF/SSF risk for varying preoperative diagnosis and rotator cuff status. METHODS: Patients consecutively receiving RSA between January 2013 and June 2019 from 15 institutions comprising 24 members of the American Shoulder and Elbow Surgeons (ASES) with primary, preoperative diagnoses of GHOA, CTA and MCT were included for study. Inclusion criteria, definitions, and inclusion of patient factors in a multivariate model to predict cumulative risk of ASF/SSF were determined through an iterative Delphi process. The CTA and MCT groups were combined for analysis. Consensus was defined as greater than 75% agreement amongst contributors. Only ASF/SSF confirmed by clinical and radiographic correlation were included for analysis. RESULTS: Our study cohort included 4764 patients with preoperative diagnoses of GHOA, CTA, or MCT with minimum follow-up of 3 months (range: 3-84). The incidence of cumulative stress fracture was 4.1% (n = 196). The incidence of stress fracture in the GHOA cohort was 2.1% (n = 34/1637) compared to 5.2% (n = 162/3127) (P < .001) in the CTA/MCT cohort. Presence of inflammatory arthritis (odds ratio [OR] 2.90, 95% confidence interval [CI] 1.08-7.78; P = .035) was the sole predictive factor of stress fractures in GHOA, compared with inflammatory arthritis (OR 1.86, 95% CI 1.19-2.89; P = .016), female sex (OR 1.81, 95% CI 1.20-2.72; P = .007), and osteoporosis (OR 1.56, 95% CI 1.02-2.37; P = .003) in the CTA/MCT cohort. CONCLUSION: Preoperative diagnosis of GHOA has a different risk profile for developing stress fractures after RSA than patients with CTA/MCT. Though rotator cuff integrity is likely protective against ASF/SSF, approximately 1/46 patients receiving RSA with primary GHOA will have this complication, primarily influenced by a history of inflammatory arthritis. Understanding risk profiles of patients undergoing RSA by varying diagnosis is important in counseling, expectation management, and treatment by surgeons.

Anatomic and reverse shoulder arthroplasty for management of type B2 and B3 glenoids: a matched-cohort analysis.

BACKGROUND: Severe glenohumeral osteoarthritis (GHOA) with posterior glenoid erosion remains challenging to address for shoulder surgeons. Whereas anatomic total shoulder arthroplasty (TSA) has historically been the treatment of choice, reverse shoulder arthroplasty (RSA) offers an alternative option. Limited evidence exists directly comparing these 2 treatments in a similar patient population. The purpose of this study was to compare the clinical outcomes of patients with GHOA and Walch type B2 and B3 glenoid morphologies treated with TSA vs. RSA. METHODS: We performed a multicenter retrospective cohort study of patients with GHOA who were treated with primary shoulder arthroplasty and had a minimum follow-up period of 2 years. Preoperative computed tomography was used to determine type B2 and B3 glenoid morphology as described by the modified Walch classification. Three-dimensional perioperative planning software was used to characterize glenoid retroversion and humeral subluxation. Patients were categorized based on type of arthroplasty (TSA or RSA) and were matched 1:1 by sex, Walch classification, and age. Patient-reported outcome measures, active range of motion, presence and severity of glenoid loosening, and complications were compared. The percentage of patients who reached previously established clinically significant thresholds of the minimal clinically important difference and substantial clinical benefit for the American Shoulder and Elbow Surgeons score was also comparatively assessed. RESULTS: In total, 202 patients (101 per group) with GHOA and type B2 or B3 glenoids were included in the 1:1 matched analysis. The mean length of follow-up (± standard deviation) was 39 ± 18.7 months. The cohorts were well matched, with no differences in sex, age, American Society of Anesthesiologists score, body mass index, preoperative glenoid morphology (Walch classification), glenoid retroversion, or posterior subluxation (P > .05). RSA was associated with a lower postoperative visual analog scale pain score (0.5 in RSA group vs. 1.2 in TSA group, P = .036); however, no other no other significant differences in patient-reported significant differences in patient-reported outcome measures were found. Most patients in both groups (95.0% in TSA group vs. 98.0% in RSA group, P = .436) reached the minimal clinically important difference, and 82% of TSA patients and 90% of RSA patients reached the substantial clinical benefit value (P = .292). No significant differences in the overall complication rate (P = .781) and active range of motion were found, with the exception of internal rotation (scored on a numeric scale) being worse in the RSA group (2.7 preoperatively and 5.2 postoperatively in RSA group vs. 3.9 and 6.5, respectively, in TSA group; P < .001). Baseplate loosening occurred in 2 RSA cases, and 29 TSA cases had glenoid radiolucencies (P < .001), with 3 grossly loose glenoid components. CONCLUSION: Primary RSA results in short-term outcomes largely comparable to those of TSA in patients with Walch type B2 or B3 glenoid morphology. Both TSA and RSA provide substantial clinical benefit to patients with significant posterior glenoid wear.

Stimulating TAM-mediated anti-tumor immunity with mannose-decorated nanoparticles in ovarian cancer.

BACKGROUND: Current cancer immunotherapies have made tremendous impacts but generally lack high response rates, especially in ovarian cancer. New therapies are needed to provide increased benefits. One understudied approach is to target the large population of immunosuppressive tumor-associated macrophages (TAMs). Using inducible transgenic mice, we recently reported that upregulating nuclear factor-kappaB (NF-κB) signaling in TAMs promotes the M1, anti-tumor phenotype and limits ovarian cancer progression. We also developed a mannose-decorated polymeric nanoparticle system (MnNPs) to preferentially deliver siRNA payloads to M2, pro-tumor macrophages in vitro. In this study, we tested a translational strategy to repolarize ovarian TAMs via MnNPs loaded with siRNA targeting the inhibitor of NF-κB alpha (IκBα) using mouse models of ovarian cancer. METHODS: We evaluated treatment with MnNPs loaded with IκBα siRNA (IκBα-MnNPs) or scrambled siRNA in syngeneic ovarian cancer models. ID8 tumors in C57Bl/6 mice were used to evaluate consecutive-day treatment of late-stage disease while TBR5 tumors in FVB mice were used to evaluate repetitive treatments in a faster-developing disease model. MnNPs were evaluated for biodistribution and therapeutic efficacy in both models. RESULTS: Stimulation of NF-κB activity and repolarization to an M1 phenotype via IκBα-MnNP treatment was confirmed using cultured luciferase-reporter macrophages. Delivery of MnNPs with fluorescent payloads (Cy5-MnNPs) to macrophages in the solid tumors and ascites was confirmed in both tumor models. A three consecutive-day treatment of IκBα-MnNPs in the ID8 model validated a shift towards M1 macrophage polarization in vivo. A clear therapeutic effect was observed with biweekly treatments over 2-3 weeks in the TBR5 model where significantly reduced tumor burden was accompanied by changes in immune cell composition, indicative of reduced immunosuppressive tumor microenvironment. No evidence of toxicity associated with MnNP treatment was observed in either model. CONCLUSIONS: In mouse models of ovarian cancer, MnNPs were preferentially associated with macrophages in ascites fluid and solid tumors. Evidence of macrophage repolarization, increased inflammatory cues, and reduced tumor burden in IκBα-MnNP-treated mice indicate beneficial outcomes in models of established disease. We have provided evidence of a targeted, TAM-directed approach to increase anti-tumor immunity in ovarian cancer with strong translational potential for future clinical studies.

Kupffer cell release of platelet activating factor drives dose limiting toxicities of nucleic acid nanocarriers.

This work establishes that Kupffer cell release of platelet activating factor (PAF), a lipidic molecule with pro-inflammatory and vasoactive signaling properties, dictates dose-limiting siRNA nanocarrier-associated toxicities. High-dose intravenous injection of siRNA-polymer nano-polyplexes (si-NPs) elicited acute, shock-like symptoms in mice, associated with increased plasma PAF and consequently reduced PAF acetylhydrolase (PAF-AH) activity. These symptoms were completely prevented by prophylactic PAF receptor inhibition or Kupffer cell depletion. Assessment of varied si-NP chemistries confirmed that toxicity level correlated to relative uptake of the carrier by liver Kupffer cells and that this toxicity mechanism is dependent on carrier endosome disruptive function. 4T1 tumor-bearing mice, which exhibit increased circulating leukocytes, displayed greater sensitivity to these toxicities. PAF-mediated toxicities were generalizable to commercial delivery reagent in vivo-jetPEI® and an MC3 lipid formulation matched to an FDA-approved nanomedicine. These collective results establish Kupffer cell release of PAF as a key mediator of siRNA nanocarrier toxicity and identify PAFR inhibition as an effective strategy to increase siRNA nanocarrier tolerated dose.

Increased canonical NF-kappaB signaling specifically in macrophages is sufficient to limit tumor progression in syngeneic murine models of ovarian cancer.

BACKGROUND: New treatment options for ovarian cancer are urgently required. Tumor-associated macrophages (TAMs) are an attractive target for therapy; repolarizing TAMs from M2 (pro-tumor) to M1 (anti-tumor) phenotypes represents an important therapeutic goal. We have previously shown that upregulated NF-kappaB (NF-κB) signaling in macrophages promotes M1 polarization, but effects in the context of ovarian cancer are unknown. Therefore, we aimed to investigate the therapeutic potential of increasing macrophage NF-κB activity in immunocompetent mouse models of ovarian cancer. METHODS: We have generated a transgenic mouse model, termed IKFM, which allows doxycycline-inducible overexpression of a constitutively active form of IKK2 (cIKK2) specifically within macrophages. The IKFM model was used to evaluate effects of increasing macrophage NF-κB activity in syngeneic murine TBR5 and ID8-Luc models of ovarian cancer in two temporal windows: 1) in established tumors, and 2) during tumor implantation and early tumor growth. Tumor weight, ascites volume, ascites supernatant and cells, and solid tumor were collected at sacrifice. Populations of macrophages and T cells within solid tumor and/or ascites were analyzed by immunofluorescent staining and qPCR, and soluble factors in ascitic fluid were analyzed by ELISA. Comparisons of control versus IKFM groups were performed by 2-tailed Mann-Whitney test, and a P-value < 0.05 was considered statistically significant. RESULTS: Increased expression of the cIKK2 transgene in TAMs from IKFM mice was confirmed at the mRNA and protein levels. Tumors from IKFM mice, regardless of the timing of doxycycline (dox) administration, demonstrated greater necrosis and immune infiltration than control tumors. Analysis of IKFM ascites and tumors showed sustained shifts in macrophage populations away from the M2 and towards the anti-tumor M1 phenotype. There were also increased tumor-infiltrating CD3+/CD8+ T cells in IKFM mice, accompanied by higher levels of CXCL9, a T cell activating factor secreted by macrophages, in IKFM ascitic fluid. CONCLUSIONS: In syngeneic ovarian cancer models, increased canonical NF-κB signaling in macrophages promoted anti-tumor TAM phenotypes and increased cytotoxic T cell infiltration, which was sufficient to limit tumor progression. This may present a novel translational approach for ovarian cancer treatment, with the potential to increase responses to T cell-directed therapy in future studies.

PEGylated PLGA Nanoparticle Delivery of Eggmanone for T Cell Modulation: Applications in Rheumatic Autoimmunity.

BACKGROUND: Helper T cell activity is dysregulated in a number of diseases including those associated with rheumatic autoimmunity. Treatment options are limited and usually consist of systemic immune suppression, resulting in undesirable consequences from compromised immunity. Hedgehog (Hh) signaling has been implicated in the activation of T cells and the formation of the immune synapse, but remains understudied in the context of autoimmunity. Modulation of Hh signaling has the potential to enable controlled immunosuppression but a potential therapy has not yet been developed to leverage this opportunity. METHODS: In this work, we developed biodegradable nanoparticles to enable targeted delivery of eggmanone (Egm), a specific Hh inhibitor, to CD4+ T cell subsets. We utilized two FDA-approved polymers, poly(lactic-co-glycolic acid) and polyethylene glycol, to generate hydrolytically degradable nanoparticles. Furthermore, we employed maleimide-thiol mediated conjugation chemistry to decorate nanoparticles with anti-CD4 F(ab') antibody fragments to enable targeted delivery of Egm. RESULTS: Our novel delivery system achieved a highly specific association with the majority of CD4+ T cells present among a complex cell population. Additionally, we have demonstrated antigen-specific inhibition of CD4+ T cell responses mediated by nanoparticle-formulated Egm. CONCLUSION: This work is the first characterization of Egm's immunomodulatory potential. Importantly, this study also suggests the potential benefit of a biodegradable delivery vehicle that is rationally designed for preferential interaction with a specific immune cell subtype for targeted modulation of Hh signaling.

Optimizing Mannose "Click" Conjugation to Polymeric Nanoparticles for Targeted siRNA Delivery to Human and Murine Macrophages.

"Smart", dual pH-responsive, and endosomolytic polymeric nanoparticles have demonstrated great potential for localized drug delivery, especially for siRNA delivery to the cytoplasm of cells. However, targeted delivery to a specific cell phenotype requires an additional level of functionality. Copper-catalyzed azide-alkyne cycloaddition (CuAAC) is a highly selective bioconjugation reaction that can be performed in conjunction with other polymerization techniques without adversely affecting reaction kinetics, but there exists some concern for residual copper causing cytotoxicity. To alleviate these concerns, we evaluated conjugation efficiency, residual copper content, and cell viability in relation to copper catalyst concentration. Our results demonstrated an optimal range for minimizing cytotoxicity while maintaining high levels of conjugation efficiency, and these conditions produced polymers with increased targeting to M2-polarized macrophages, as well as successful delivery of therapeutic siRNA that reprogrammed the macrophages to a proinflammatory phenotype.