A Comprehensive Enhanced Recovery Pathway for Rotator Cuff Surgery Reduces Pain, Opioid Use, and Side Effects.

BACKGROUND: Patients often have moderate to severe pain after rotator cuff surgery, despite receiving analgesics and nerve blocks. There are many suggested ways to improve pain after rotator cuff surgery, but the effects of adopting a pathway that includes formal patient education, a long-acting nerve block, and extensive multimodal analgesia are unclear. QUESTIONS/PURPOSES: (1) Does adoption of a clinical pathway incorporating patient education, a long-acting nerve block, and preemptive multimodal analgesia reduce the worst pain during the first 48 hours after surgery compared with current standard institutional practices? (2) Does adoption of the pathway reduce opioid use? (3) Does adoption of the pathway reduce side effects and improve patient-oriented outcomes? METHODS: From September 2018 to January 2020, 281 patients scheduled for arthroscopic ambulatory rotator cuff surgery were identified for this paired sequential prospective cohort study. Among patients in the control group, 177 were identified, 33% (58) were not eligible, for 11% (20) staff was not available, 56% (99) were approached, 16% (29) declined, 40% (70) enrolled, and 40% (70) were analyzed (2% [4] lost to follow-up for secondary outcomes after postoperative day 2). For patients in the pathway cohort, 104 were identified, 17% (18) were not eligible, for 11% (11) staff was not available, 72% (75) were approached, 5% (5) declined, 67% (70) enrolled, and 67% (70) were analyzed (3% [3] lost to follow-up for secondary outcomes after postoperative day 2). No patients were lost to follow-up for primary outcome; for secondary outcomes, four were lost in the control group and three in the pathway group after postoperative day 2 (p = 0.70). The initial 70 patients enrolled received routine care (control group), and in a subsequent cohort, 70 patients received care guided by a pathway (pathway group). Of the 205 eligible patients, 68% (140) were included in the analysis. This was not a study comparing two tightly defined protocols but rather a study to determine whether adoption of a pathway would alter patient outcomes. For this reason, we used a pragmatic (real-world) study design that did not specify how control patients would be treated, and it did not require that all pathway patients receive all components of the pathway. We developed the pathway in coordination with a group of surgeons and anesthesiologists who agreed to apply the pathway as much as was viewed practical for each individual patient. Patients in both groups received a brachial plexus nerve block with sedation. Major differences between the pathway and control groups were: detailed patient education regarding reasonable pain expectations with a goal of reducing opioid use (no formal educational presentation was given to the control), a long-acting nerve block using bupivacaine with dexamethasone (control patients often received shorter-acting local anesthetic without perineural dexamethasone), and preemptive multimodal analgesia including intraoperative ketamine, postoperative acetaminophen, NSAIDs, and gabapentin at bedtime, with opioids as needed (control patients received postoperative opioids but most did not get postoperative NSAIDS and no controls received gabapentin or separate prescriptions for acetaminophen). The primary outcome was the numerical rating scale (NRS) worst pain with movement 0 to 48 hours after block placement. The NRS pain score ranges from 0 (no pain) to 10 (worst pain possible). The minimum clinically important difference (MCID) [12] for NRS that was used for calculation of the study sample size was 1.3 [18], although some authors suggest 1 [13] or 2 [5] are appropriate; if we had used an MCID of 2, the sample size would have been smaller. Secondary outcomes included NRS pain scores at rest, daily opioid use (postoperative day 1, 2, 7, 14), block duration, patient-oriented pain questions (postoperative day 1, 2, 7, 14), and patient and physician adherence to pathway. RESULTS: On postoperative day 1, pathway patients had lower worst pain with movement (3.3 ± 3.1) compared with control patients (5.6 ± 3.0, mean difference -2.7 [95% CI -3.7 to -1.7]; p < 0.001); lower scores were also seen for pain at rest (1.9 ± 2.3 versus 4.0 ± 2.9, mean difference -2.0 [95% CI -2.8 to -1.3]; p < 0.001). Cumulative postoperative opioid use (0-48 hours) was reduced (pathway oral morphine equivalent use was 23 ± 28 mg versus 44 ± 35 mg, mean difference 21 [95% CI 10 to 32]; p < 0.01). The greatest difference in opioid use was in the first 24 hours after surgery (pathway 7 ± 12 mg versus control 21 ± 21 mg, mean difference -14 [95% CI -19 to -10]; p < 0.01). On postoperative day 1, pathway patients had less interference with staying asleep compared with control patients (0.5 ± 1.6 versus 2.6 ± 3.3, mean difference -2.2 [95% CI -3.3 to -1.1]; p < 0.001); lower scores were also seen for interference with activities (0.9 ± 2.3 versus 1.9 ± 2.9, mean difference -1.1 [95% CI -2 to -0.1]; p = 0.03). Satisfaction with pain treatment on postoperative day 1 was higher among pathway patients compared with control patients (9.2 ± 1.7 versus 8.2 ± 2.5, mean difference 1.0 [95% CI 0.3 to 1.8]; p < 0.001). On postoperative day 2, pathway patients had lower nausea scores compared with control patients (0.3 ± 1.1 versus 1 ± 2.1, mean difference -0.7 [95% CI -1.2 to -0.1]; p = 0.02); lower scores were also seen for drowsiness on postoperative day 1 (1.7 ± 2.7 versus 2.6 ± 2.6, mean difference -0.9 [95% CI - 1.7 to -0.1]; p = 0.03). CONCLUSION: Adoption of the pathway was associated with improvement in the primary outcome (pain with movement) that exceeded the MCID. Patients in the pathway group had improved patient-oriented outcomes and fewer side effects. This pathway uses multiple analgesic drugs, which may pose risks to elderly patients, in particular. Therefore, in evaluating whether to use this pathway, clinicians should weigh the effect sizes against the potential risks that may emerge with large scale use, consider the difficulties involved in adapting a pathway to local practice so that pathway will persist, and recognize that this study only enrolled patients among surgeons and the anesthesiologists that advocated for the pathway; results may have been different with less enthusiastic clinicians. This pathway, based on a long-lasting nerve block, multimodal analgesia, and patient education can be considered for adoption. LEVEL OF EVIDENCE: Level II, therapeutic study.

Cerebral Oxygenation in the Sitting Position Is Not Compromised During Spontaneous or Positive-Pressure Ventilation.

BACKGROUND: General anesthesia with neuromuscular blockade may facilitate total shoulder arthroplasty but appears to increase risk of cerebral oxygen desaturation. Cerebral desaturation is undesirable and is a proxy for risk of stroke. PURPOSES/QUESTIONS: This study tested the hypothesis that cerebral oxygen desaturation occurs frequently during general anesthesia with neuromuscular blockade and positive-pressure ventilation but does not occur with spontaneous ventilation. Correlations were sought among cerebral oxygen saturation, blood pressure, and cardiac index. METHODS: We designed a prospective, observational, cohort study to measure cerebral oxygenation in 25 patients during general anesthesia, both with and without positive-pressure ventilation. Patients undergoing elective shoulder arthroplasty in the sitting position received an arterial catheter, near-infrared spectroscopic measurement of cerebral oxygenation, and non-invasive cardiac output measurement. Moderate hypotension was allowed. Blood pressure was supported as needed with ephedrine or low-dose epinephrine (but avoiding phenylephrine). Hypercapnia (45 to 55 mmHg) was targeted during positive-pressure ventilation. RESULTS: No cerebral oxygen desaturations occurred, regardless of ventilation mode. Under positive-pressure ventilation, the median (interquartile range: Q1, Q3) cerebral oxygenation was 110% of baseline (104, 113), the mean arterial pressure was 62% of baseline (59, 69), and the cardiac index was 82% of baseline (71, 104). Cerebral oxygenation did not correlate with blood pressure or cardiac index but had moderate correlation with end-tidal carbon dioxide. No strokes occurred. CONCLUSIONS: There were no signs of inadequate brain perfusion during general anesthesia using paralytic agents. Positive-pressure ventilation with moderate hypotension in the sitting position does not endanger patients, in the context of moderate hypercapnia and hemodynamic support using ephedrine or epinephrine.

What Pain Levels Do TSA Patients Experience When Given a Long-acting Nerve Block and Multimodal Analgesia?

BACKGROUND: The pain experience for total shoulder arthroplasty (TSA) patients in the first 2 weeks after surgery has not been well described. Many approaches to pain management have been used, with none emerging as clearly superior; it is important that any approach minimizes postoperative opioid use. QUESTIONS/PURPOSES: (1) With a long-acting nerve block and comprehensive multimodal analgesia, what are the pain levels after TSA from day of surgery until postoperative day (POD) 14? (2) How many opioids do TSA patients take from the day of surgery until POD 14? (3) What are the PainOUT responses at POD 1 and POD 14, focusing on side effects from opioids usage? METHODS: From January 27, 2017 to December 6, 2017, 154 TSA patients were identified as potentially eligible for this prospective, institutional review board-approved observational study. Of those, 46 patients (30%) were excluded (either because they were deemed not appropriate for the study, research staff were not available, patients were not eligible, or they declined to participate), and another six (4%) had incomplete followup data and could not be studied, leaving 102 patients (66%) for analysis here. Median preoperative pain with movement was 7 (interquartile range [IQR], 5-9) and 13 of 102 patients used preoperative opioids. All patients received a single-injection bupivacaine interscalene block with adjuvant clonidine, dexamethasone, and buprenorphine. Multimodal analgesia included acetaminophen, NSAIDs, and opioids. The primary outcome was the Numerical Rating Scale (NRS) pain score with movement on POD 14. The NRS pain score ranges from 0 (no pain) to 10 (worst pain possible). Secondary outcomes included NRS pain scores at rest and with movement (day of surgery, and PODs 1, 3, 7 and 14), daily analgesic use from day of surgery to POD 14 (both oral and intravenous), Opioid-Related Symptom Distress Scale (which assesses 12 symptoms ranging from 0 to 4, with 4 being the most distressing; the composite score is the mean of the 12 symptom-specific scores) on POD 1, and the PainOut questionnaire on POD 1 and POD 14. The PainOut questionnaire includes questions rating nausea, drowsiness, itching from 0 (none) to 10 (severe), as well as rating difficulty staying asleep from 0 (does not interfere) to 10 (completely interferes). RESULTS: The median NRS pain scores with movement were 2 (IQR, 0-5) on POD 1, 5 (IQR, 3-6) on POD 3, and the pain score was 3 (IQR, 1-5) on POD 14. Median total opioid use (converted to oral morphine equivalents) was 16 mg (4-50 mg) for the first 24 hours, 30 mg (8-63 mg) for the third, and 0 mg (0-20 mg) by the eighth 24-hour period, while the most frequent number of activations of the intravenous patient-controlled analgesia device was 0. Median PainOut scores on POD 1 and POD 14 for sleep interference, nausea, drowsiness and itching were 0, and the median composite Opioid-Related Symptom Distress Scale score on day 1 was 0.3 (IQR, 0.1-0.5). CONCLUSIONS: Clinicians using this protocol, which combines a long-acting, single-injection nerve block with multimodal analgesia, can inform TSA patients that their postoperative pain will likely be less than their preoperative pain, and that on average they will stop using opioids after 7 days. Future research could investigate what the individual components of this protocol contribute. Larger cohort studies or registries would document the incidence of rare complications. Randomized controlled trials could directly compare analgesic effectiveness and cost-benefits for this protocol versus alternative strategies, such as perineural catheters or liposomal bupivacaine. Perhaps most importantly, future studies could seek ways to further reduce peak pain and opioid usage on POD 2 and POD 3. LEVEL OF EVIDENCE: Level IV, therapeutic study.

The long non-coding RNA MALAT1 promotes ovarian cancer progression by regulating RBFOX2-mediated alternative splicing.

Ovarian cancer metastasizes via direct seeding, whereby cancer cells shed from the primary site, resist cell death in the peritoneal cavity, then metastasize to peritoneal organs. We sought to identify molecular mechanisms that facilitate ovarian cancer cell anchorage independent survival. Gene expression profiling was performed on ovarian cancer cells grown in attached or forced suspension culture and confirmed by RT-qPCR. Anoikis was measured by Caspase 3/7 assay. Since the long non-coding RNA Metastasis Associated Lung Adenocarcinoma transcript 1 (MALAT1) was among the transcripts most highly increased in forced suspension culture, modified anti-sense oligonucleotides (ASO) were used to inhibit its expression. Knockdown of RBFOX2 and KIF1B was performed using shRNAs. Publically available datasets were analyzed for association of MALAT1 gene expression with clinicopathological variables. In multiple anoikis-resistant ovarian cancer cell lines MALAT1 expression increased after 24 and 48 h in forced suspension culture compared to attached culture. High MALAT1 is associated with increased stage, recurrence, and reduced survival in ovarian cancer, and in a small percentage of ovarian cancers MALAT1 is amplified. MALAT1 knockdown resulted in decreased proliferation, invasion, anchorage-independent growth, and increased anoikis. Suppression of MALAT1 also resulted in decreased expression of RBFOX2, and alternative processing of the pro-apoptotic tumor suppressor gene KIF1B. RBFOX2 suppression resulted in preferential splicing of the pro-apoptotic isoform of KIF1B (KIFB1B-beta) and increased anoikis. The lncRNA MALAT1 facilitates a pro-metastatic phenotype in ovarian cancer by promoting alternative RNA processing and differential expression of anti-apoptosis and epithelial to mesenchymal transition (EMT)-related genes.

Reversal of Triple-Negative Breast Cancer EMT by miR-200c Decreases Tryptophan Catabolism and a Program of Immunosuppression.

Tryptophan-2,3-dioxygenase (TDO2), a rate-limiting enzyme in the tryptophan catabolism pathway, is induced in triple-negative breast cancer (TNBC) by inflammatory signals and anchorage-independent conditions. TNBCs express extremely low levels of the miR-200 family compared with estrogen receptor-positive (ER+) breast cancer. In normal epithelial cells and ER+ breast cancers and cell lines, high levels of the family member miR-200c serve to target and repress genes involved in epithelial-to-mesenchymal transition (EMT). To identify mechanism(s) that permit TNBC to express TDO2 and other proteins not expressed in the more well-differentiated ER+ breast cancers, miRNA-200c was restored in TNBC cell lines. The data demonstrate that miR-200c targeted TDO2 directly resulting in reduced production of the immunosuppressive metabolite kynurenine. Furthermore, in addition to reversing a classic EMT signature, miR-200c repressed many genes encoding immunosuppressive factors including CD274/CD273, HMOX-1, and GDF15. Restoration of miR-200c revealed a mechanism, whereby TNBC hijacks a gene expression program reminiscent of that used by trophoblasts to suppress the maternal immune system to ensure fetal tolerance during pregnancy. IMPLICATIONS: Knowledge of the regulation of tumor-derived immunosuppressive factors will facilitate development of novel therapeutic strategies that complement current immunotherapy to reduce mortality for patients with TNBC.

Androgen Receptor Supports an Anchorage-Independent, Cancer Stem Cell-like Population in Triple-Negative Breast Cancer.

Preclinical and early clinical trials indicate that up to 50% of triple-negative breast cancers (TNBC) express androgen receptor (AR) and are potentially responsive to antiandrogens. However, the function of AR in TNBC and the mechanisms by which AR-targeted therapy reduces tumor burden are largely unknown. We hypothesized that AR maintains a cancer stem cell (CSC)-like tumor-initiating population and serves as an antiapoptotic factor, facilitating anchorage independence and metastasis. AR levels increased in TNBC cells grown in forced suspension culture compared with those in attached conditions, and cells that expressed AR resisted detachment-induced apoptosis. Culturing TNBC cells in suspension increased the CSC-like population, an effect reversed by AR inhibition. Pretreatment with enzalutamide (Enza) decreased the tumor-initiating capacity of TNBC cells and reduced tumor volume and viability when administered simultaneously or subsequent to the chemotherapeutic paclitaxel; simultaneous treatment more effectively suppressed tumor recurrence. Overall, our findings suggest that AR-targeted therapies may enhance the efficacy of chemotherapy even in TNBCs with low AR expression by targeting a CSC-like cell population with anchorage independence and invasive potential. Cancer Res; 77(13); 3455-66. ©2017 AACR.

Synergy between Androgen Receptor Antagonism and Inhibition of mTOR and HER2 in Breast Cancer.

The androgen receptor (AR) is widely expressed in breast cancer, and evidence suggests dependence on AR signaling for growth and survival. AR antagonists such as enzalutamide and seviteronel have shown success in preclinical models and clinical trials of prostate cancer and are currently being evaluated in breast cancer. Reciprocal regulation between AR and the HER2/PI3K/mTOR pathway may contribute to resistance to HER2- and mTOR-targeted therapies; thus, dual inhibition of these pathways may synergistically inhibit breast cancer growth. HER2+ and triple-negative breast cancer cell lines were treated with AR antagonist plus anti-HER2 mAb trastuzumab or mTOR inhibitor everolimus. Apoptosis, cell proliferation, and drug synergy were measured in vitro Pathway component genes and proteins were measured by qRT-PCR, Western blot, and reverse phase protein array. In vivo, HER2+ breast cancer xenografts were treated with enzalutamide, everolimus, trastuzumab, and combinations of these drugs. AR antagonists inhibited proliferation of both HER2+ and TNBC cell lines. Combining AR antagonist and either everolimus or trastuzumab resulted in synergistic inhibition of proliferation. Dihydrotestosterone caused increased phosphorylation of HER2 and/or HER3 that was attenuated by AR inhibition. Everolimus caused an increase in total AR, phosphorylation of HER2 and/or HER3, and these effects were abrogated by enzalutamide. Growth of trastuzumab-resistant HER2+ xenograft tumors was inhibited by enzalutamide, and combining enzalutamide with everolimus decreased tumor viability more than either single agent. AR antagonists synergize with FDA-approved breast cancer therapies such as everolimus and trastuzumab through distinct mechanisms. Treatment combinations are effective in trastuzumab-resistant HER2+ breast cancer cells in vivoMol Cancer Ther; 16(7); 1389-400. ©2017 AACR.

Perineural dexamethasone with subsartorial saphenous nerve blocks in ACL reconstruction.

PURPOSE: Subsartorial saphenous nerve blockade (SSNB) is an effective analgesic alternative to femoral nerve blockade after anterior cruciate ligament (ACL) reconstruction with bone-tendon-bone (BTB) autograft. It was hypothesized that dexamethasone in a SSNB will prolong analgesia, improve pain and satisfaction, and reduce postoperative opioid requirements and side effects. METHODS: One hundred ninety-five patients undergoing ACL reconstruction with BTB autograft (ages 16-65) were enrolled. Subjects received SSNB with 13 ml of 0.5 % bupivacaine (control group), 1 mg preservative-free dexamethasone +0.5 % bupivacaine (treatment group I), or 4 mg preservative-free dexamethasone +0.5 % bupivacaine (treatment group II). Subjects received identical perioperative management. On postoperative days 1 and 2, subjects reported perceived block duration, pain scores, satisfaction, opioid use, and side effects. Cox-proportional hazards modelling was used to compare block duration, adjusting for body mass index, age, sex, tourniquet time, American Society of Anesthesiologists classification, and intravenous dexamethasone dose. RESULTS: Patient-perceived block duration was significantly increased in treatment group I [hazard ratio (95 % confidence interval [CI]) 0.48 (0.31-0.75); P = 0.001] and treatment group II (hazard ratio (95 % CI): 0.52 (0.33-0.81); P = 0.004) compared to control. The block was extended from a median (95 % CI) of 33.1 (28.4-37.3) to 41.2 (32.4-50.9) and 46.5 (35.8-48.9) hours, respectively. Additionally, patients in treatment group II reported increased time that block provided pain relief, higher patient satisfaction, lower pain scores at rest, and decreased drowsiness and confusion. CONCLUSION: The addition of 1 and 4 mg of dexamethasone to the block injectate significantly increased SSNB duration by 8-13 h compared to control. LEVEL OF EVIDENCE: Therapeutic study, level 1.

Constitutive expression of microRNA-150 in mammary epithelium suppresses secretory activation and impairs de novo lipogenesis.

Profiling of RNA from mouse mammary epithelial cells (MECs) isolated on pregnancy day (P)14 and lactation day (L)2 revealed that the majority of differentially expressed microRNA declined precipitously between late pregnancy and lactation. The decline in miR-150, which exhibited the greatest fold-decrease, was verified quantitatively and qualitatively. To test the hypothesis that the decline in miR-150 is crucial for lactation, MEC-specific constitutive miR-150 was achieved by crossing ROSA26-lox-STOP-lox-miR-150 mice with WAP-driven Cre recombinase mice. Both biological and foster pups nursed by bitransgenic dams exhibited a dramatic decrease in survival compared with offspring nursed by littermate control dams. Protein products of predicted miR-150 targets Fasn, Olah, Acaca, and Stat5B were significantly suppressed in MECs of bitransgenic mice with constitutive miR-150 expression as compared with control mice at L2. Lipid profiling revealed a significant reduction in fatty acids synthesized by the de novo pathway in L2 MECs of bitransgenic versus control mice. Collectively, these data support the hypothesis that a synchronized decrease in miRNAs, such as miR-150, at late pregnancy serves to allow translation of targets crucial for lactation.

Cooperative Dynamics of AR and ER Activity in Breast Cancer.

Androgen receptor (AR) is expressed in 90% of estrogen receptor alpha-positive (ER+) breast tumors, but its role in tumor growth and progression remains controversial. Use of two anti-androgens that inhibit AR nuclear localization, enzalutamide and MJC13, revealed that AR is required for maximum ER genomic binding. Here, a novel global examination of AR chromatin binding found that estradiol induced AR binding at unique sites compared with dihydrotestosterone (DHT). Estradiol-induced AR-binding sites were enriched for estrogen response elements and had significant overlap with ER-binding sites. Furthermore, AR inhibition reduced baseline and estradiol-mediated proliferation in multiple ER+/AR+ breast cancer cell lines, and synergized with tamoxifen and fulvestrant. In vivo, enzalutamide significantly reduced viability of tamoxifen-resistant MCF7 xenograft tumors and an ER+/AR+ patient-derived model. Enzalutamide also reduced metastatic burden following cardiac injection. Finally, in a comparison of ER+/AR+ primary tumors versus patient-matched local recurrences or distant metastases, AR expression was often maintained even when ER was reduced or absent. These data provide preclinical evidence that anti-androgens that inhibit AR nuclear localization affect both AR and ER, and are effective in combination with current breast cancer therapies. In addition, single-agent efficacy may be possible in tumors resistant to traditional endocrine therapy, as clinical specimens of recurrent disease demonstrate AR expression in tumors with absent or refractory ER. IMPLICATIONS: This study suggests that AR plays a previously unrecognized role in supporting E2-mediated ER activity in ER+/AR+ breast cancer cells, and that enzalutamide may be an effective therapeutic in ER+/AR+ breast cancers. Mol Cancer Res; 14(11); 1054-67. ©2016 AACR.

Prospective, Randomized Double-Blind Study: Does Decreasing Interscalene Nerve Block Volume for Surgical Anesthesia in Ambulatory Shoulder Surgery Offer Same-Day Patient Recovery Advantages?

BACKGROUND AND OBJECTIVES: In this randomized double-blind prospective study in patients undergoing shoulder arthroscopy, we compared the effects of ultrasound-guided interscalene nerve block using 20 mL (intervention group) and 40 mL (control group) of a mepivacaine 1.5% and bupivacaine 0.5% mixture (1:1 volume) on ipsilateral handgrip strength and other postoperative end points. METHODS: One hundred fifty-four patients scheduled for ambulatory shoulder arthroscopy were randomly assigned to receive a single-injection interscalene block under ultrasound guidance with either 40 mL (control) or 20 mL (intervention) and intravenous sedation. The primary outcome was the change in ipsilateral handgrip strength in the postanesthesia care unit (PACU) measured with a dynamometer. Secondary end points were recorded, including negative inspiratory force, incidences of hoarseness and Horner syndrome, time to readiness for discharge from PACU, time to discharge from PACU, patient satisfaction, time to block resolution, and pain scores. RESULTS: Postoperative handgrip strength was greater in the 20-mL group compared with the 40-mL group (difference in means, 2.3 kg [95% confidence interval, 0.6-4.0 kg]; P = 0.009). A smaller proportion of patients in the intervention group experienced hoarseness postoperatively compared with the control group (odds ratio, 0.26 [95% confidence interval, 0.08-0.82]; P = 0.015). Patient satisfaction and duration of analgesia were similar in both groups. CONCLUSIONS: When used for surgical anesthesia for shoulder arthroscopies in the ambulatory setting, a 20-mL volume in an ultrasound-guided interscalene block preserves greater handgrip strength on the ipsilateral side in the PACU compared with 40 mL without significant decrease in block success, duration of analgesia, and patient satisfaction.

Dicer expression in estrogen receptor-positive versus triple-negative breast cancer: an antibody comparison.

Dicer is an RNase III enzyme responsible for cleaving double-stranded RNAs into small interfering RNAs and microRNAs, which either target messenger RNA transcripts for degradation or inhibit translation. Dicer protein levels have been examined in breast cancer with contradictory results. Our goal was to resolve whether Dicer levels differ in breast cancer versus normal breast epithelium and between estrogen receptor-α-positive (ER+) or estrogen receptor-α-negative (ER-) primary breast cancers. We compared 3 different Dicer antibodies: Abcam 4A6, Abcam ab5818, and Sigma HPA000694, using immunohistochemistry and Western blot analyses. All 3 Dicer antibodies detected higher levels of Dicer in ER+ breast cancer cell lines versus ER-, and all 3 recognized exogenous overexpressed Dicer. In clinical specimens, all 3 antibodies detected higher Dicer in ER+ breast cancers versus triple-negative breast cancer (TNBC) but had very different staining patterns by immunohistochemistry on the same tumor samples. Using the optimal antibody, ab5818, selected for its sensitivity and specificity, Dicer protein expression was significantly higher in ER+ versus TNBC clinical specimens of primary tumor (P<.0001, unpaired t test). Dicer was also significantly higher in adjacent normal breast epithelium versus TNBC (P<.0001, paired t test; n=18 pairs). Differences in antibody performance may explain contrasting results observed in the literature regarding Dicer protein in breast cancer. If Dicer becomes more clinically relevant as a prognostic indicator, further antibody optimization and standardization will be critical.

Buprenorphine, Clonidine, Dexamethasone, and Ropivacaine for Interscalene Nerve Blockade: A Prospective, Randomized, Blinded, Ropivacaine Dose-Response Study.

OBJECTIVE: This study investigated interscalene block for shoulder arthroplasty with various ropivacaine concentrations in the presence of clonidine, dexamethasone, and buprenorphine. The goal was prolonged analgesia with minimal motor blockade. DESIGN: Prospective, double-blind, randomized controlled trial. SETTING: University-affiliated orthopedic hospital. METHODS: Patients (20/group) received acetaminophen, ketorolac, pregabalin, opioids, and "Control"; interscalene block, 0.375% ropivacaine, intravenous additives (buprenorphine, clonidine, dexamethasone); "High Dose"; 0.375% ropivacaine, perineural additives; "Medium Dose"; 0.2% ropivacaine, perineural additives; and "Low Dose"; 0.1% ropivacaine, perineural additives. RESULTS: Pain with movement at 24 hours was 4.9 ± 2.5 (mean ± standard deviation [SD]) (Control), 4.5 ± 3.0 (High Dose), 3.4 ± 1.8 (Medium Dose), 4.2 ± 2.4 (Low Dose). The difference between Medium Dose and Control was -1.5 (95% CI: -2.9, -0.1) (P = 0.040). Median time until need for opioids was 16.1 hours (Control) vs 23.7 hours (High Dose); hazard ratio 0.37 [95% CI: 0.17, 0.79]. High Dose had less pain with movement the morning after surgery, vs Control; 2.9 ± 2.5 vs 4.9 ± 2.7; P = 0.027. Pain with movement in the Post-Anesthesia Care Unit was higher in Low Dose, vs Control; 0.9 ± 1.4 vs 0 ± 0, P = 0.009. Low Dose had superior hand strength in the Post-Anesthesia Care Unit (mean ± SD of pre-operative strength: 44.0 ± 20.3%) compared to Control (27.5 ± 24.5%) (P = 0.031). CONCLUSIONS: For maximum pain reduction, combining perineural additives with ropivacaine 0.375% or 0.2% is suggested. To minimize motor blockade, perineural additives can be combined with ropivacaine, 0.1%.

A TDO2-AhR signaling axis facilitates anoikis resistance and metastasis in triple-negative breast cancer.

The ability of a cancer cell to develop resistance to anoikis, a programmed cell death process triggered by substratum detachment, is a critical step in the metastatic cascade. Triple-negative breast cancers (TNBC) exhibit higher rates of metastasis after diagnosis, relative to estrogen-positive breast cancers, but while TNBC cells are relatively more resistant to anoikis, the mechanisms involved are unclear. Through gene expression and metabolomic profiling of TNBC cells in forced suspension culture, we identified a molecular pathway critical for anchorage-independent cell survival. TNBC cells in suspension upregulated multiple genes in the kynurenine pathway of tryptophan catabolism, including the enzyme tryptophan 2,3-dioxygenase (TDO2), in an NF-κB-dependent manner. Kynurenine production mediated by TDO2 in TNBC cells was sufficient to activate aryl hydrocarbon receptor (AhR), an endogenous kynurenine receptor. Notably, pharmacologic inhibition or genetic attenuation of TDO2 or AhR increased cellular sensitivity to anoikis, and also reduced proliferation, migration, and invasion of TNBC cells. In vivo, TDO2 inhibitor-treated TNBC cells inhibited colonization of the lung, suggesting that TDO2 enhanced metastatic capacity. In clinical specimens of TNBC, elevated expression of TDO2 was associated with increased disease grade, estrogen receptor-negative status, and shorter overall survival. Our results define an NF-κB-regulated signaling axis that promotes anoikis resistance, suggest functional connections with inflammatory modulation by the kynurenine pathway, and highlight TDO2 as an attractive target for treatment of this aggressive breast cancer subtype.

Androgen Receptor Biology in Triple Negative Breast Cancer: a Case for Classification as AR+ or Quadruple Negative Disease.

Triple negative breast cancer (TNBC) is an aggressive breast cancer subtype that lacks estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2 (HER2) amplification. Due to the absence of these receptors, TNBC does not respond to traditional endocrine or HER2-targeted therapies that improve patient prognosis in other breast cancer subtypes. TNBC has a poor prognosis, and currently, there are no effective targeted therapies. Some TNBC tumors express androgen receptor (AR) and may benefit from AR-targeted therapies. Here, we review the literature on AR in TNBC and propose that TNBC be further sub-classified as either AR+ TNBC or quadruple negative breast cancer since targeting AR may represent a viable therapeutic option for a subset of TNBC.

Multiple molecular subtypes of triple-negative breast cancer critically rely on androgen receptor and respond to enzalutamide in vivo.

Triple-negative breast cancer (TNBC) has the lowest 5-year survival rate of invasive breast carcinomas, and currently there are no approved targeted therapies for this aggressive form of the disease. The androgen receptor (AR) is expressed in up to one third of TNBC and we find that all AR(+) TNBC primary tumors tested display nuclear localization of AR, indicative of transcriptionally active receptors. While AR is most abundant in the "luminal AR (LAR)" molecular subtype of TNBC, here, for the first time, we use both the new-generation anti-androgen enzalutamide and AR knockdown to demonstrate that the other non-LAR molecular subtypes of TNBC are critically dependent on AR protein. Indeed, AR inhibition significantly reduces baseline proliferation, anchorage-independent growth, migration, and invasion and increases apoptosis in four TNBC lines (SUM159PT, HCC1806, BT549, and MDA-MB-231), representing three non-LAR TNBC molecular subtypes (mesenchymal-like, mesenchymal stem-like, and basal-like 2). In vivo, enzalutamide significantly decreases viability of SUM159PT and HCC1806 xenografts. Furthermore, mechanistic analysis reveals that AR activation upregulates secretion of the EGFR ligand amphiregulin (AREG), an effect abrogated by enzalutamide in vitro and in vivo. Exogenous AREG partially rescues the effects of AR knockdown on proliferation, migration, and invasion, demonstrating that upregulation of AREG is one mechanism by which AR influences tumorigenicity. Together, our findings indicate that non-LAR subtypes of TNBC are AR dependent and, moreover, that enzalutamide is a promising targeted therapy for multiple molecular subtypes of AR(+) TNBC.

Postoperative Analgesia with Saphenous Block Appears Equivalent to Femoral Nerve Block in ACL Reconstruction.

BACKGROUND: Adequate pain control following anterior cruciate ligament reconstruction (ACL) often requires regional nerve block. The femoral nerve block (FNB) has been traditionally employed. Ultrasound application to regional nerve blocks allows for the use of alternatives such as the saphenous nerve block following ACL reconstruction. QUESTIONS/PURPOSES: This study evaluated postoperative analgesia provided by the subsartorial saphenous nerve block (SSNB) compared to that provided by the traditional FNB for patients undergoing ACL reconstruction with patellar tendon (bone-tendon-bone (BTB)) autografts. METHODS: A randomized, blinded, controlled clinical trial was conducted using 80 ASA I-III patients, ages 16-65, undergoing ACL reconstruction with BTB. The individuals assessing all outcome measures were blinded to the treatment group. Postoperatively, all patients received cryotherapy and parenteral hydromorphone to achieve numeric rating scale pain scores less than 4. At discharge, patients were given prescriptions for oral opioid analgesics and a scheduled NSAID. Patients were instructed to complete pain diaries and record oral opioid utilization. Patients were contacted on postoperative days (POD) 1 and 2 to ascertain the level of patient satisfaction with the analgesic regimen. RESULTS: No differences between the two groups were found. Patient demographics and postoperative pain scores at rest were not different. In addition, there was no difference in opioid use, as measured in daily oral morphine equivalents between groups. A small but statistically significant report of higher patient satisfaction with the FNB was found on POD 1 but not on POD 2. CONCLUSION: These data support our hypothesis that the SSNB provides similar and adequate postoperative analgesia when compared to the FNB, following arthroscopic ACL reconstruction with patellar tendon autograft.

A clinical pathway for total shoulder arthroplasty-a pilot study.

BACKGROUND: Appropriate pain management after total shoulder arthroplasty (TSA) facilitates rehabilitation and may improve clinical outcomes. QUESTIONS/PURPOSES: This prospective, observational study evaluated a multimodal analgesia clinical pathway for TSA. METHODS: Ten TSA patients received an interscalene nerve block (25 cm(3) 0.375% ropivacaine) with intraoperative general anesthesia. Postoperative analgesia included regularly scheduled non-opioid analgesics (meloxicam, acetaminophen, and pregabalin) and opioids on demand (oral oxycodone and intravenous patient-controlled hydromorphone). Patients were evaluated twice daily to assess pain, anterior deltoid strength, handgrip strength, and sensory function. RESULTS: The nerve block lasted an average of 18 h. Patients had minimal pain after surgery; 0 (median score on a 0-10 scale) in the Post-Anesthesia Care Unit (PACU) but increased on postoperative day (POD) 1 to 2.3 (0.0, 3.8; median (25%, 75%)) at rest and 3.8 (2.1, 6.1) with movement. Half of the patients activated the patient-controlled analgesia four or fewer times in the first 24 h after surgery. Operative anterior deltoid strength was 0 in the PACU but returned to 68% by POD 1. Operative hand strength was 0 (median) in the PACU, but the third quartile (75%) had normalized strength 49% of preoperative value. CONCLUSIONS: Patients did well with this multimodal analgesic protocol. Pain scores were low, half of the patients used little or no intravenous opiate, and some patients had good handgrip strength. Future research can focus on increasing duration of analgesia from the nerve block, minimizing motor block, lowering pain scores, and avoiding intravenous opioids.

Association of obesity with inflammation and pain after total hip arthroplasty.

BACKGROUND: The prevalence of obesity is increasing, and obesity often leads to degenerative joint disease requiring total hip arthroplasty (THA). Obesity is a proinflammatory state associated with an increase in chronic, low-grade inflammatory response. As such, it may augment the postoperative inflammatory response, which has been associated with postoperative pain and complications. QUESTIONS/PURPOSES: We determined whether severity of obesity was associated with (1) severity of inflammatory response, as measured by the in vivo circulating levels of cytokines and ex vivo functional reactivity of mononuclear blood cells, and (2) severity of pain, as measured by verbal pain scores and analgesic consumption, in the first 24 hours after THA. METHODS: We studied 60 patients (20 normal weight, 20 overweight, 20 obese) undergoing elective primary unilateral THA in this prospective cross-sectional study. Blood samples were collected for C-reactive protein and cytokine levels, including IL-1ß, IL-2, IL-6, IL-8, and tumor necrosis factor α (TNF-α), from patients before and 24 hours after surgery. Cytokine response of whole blood was evaluated ex vivo with or without two standard activators, phorbol-12-myristate-13-acetate and lipopolysaccharide, using standardized blood sample from patients at 24 hours. These standard immune activators are implicated in the inflammatory response to gram-negative infection, translocation of microbial products, pathophysiology of septic shock syndrome in human, and tumor promotion. Pain response was gauged using verbal pain scores (on a 0- to 10-point scale, where 0 = no pain and 10 = worst pain) at rest and with activity at 24 hours after surgery and analgesic consumption of volume of epidural analgesic solution for the first 24 hours after surgery. RESULTS: No correlation was found between BMI and postoperative spontaneous circulating cytokine levels. However, after activation of blood leukocytes with lipopolysaccharide, there was a significant positive correlation between the BMI and IL-1ß, IL-6, and TNF-α levels (r = 0.26-0.32; p = 0.03, p = 0.03, and p = 0.01, respectively), suggesting priming of the innate immune system in obesity and potential for excessive postoperative inflammatory response. Obesity was not associated with increased pain or analgesic consumption in the first 24 hours after surgery. CONCLUSIONS: Obesity is associated with a proinflammatory state after THA as demonstrated by enhanced cytokine reactivity. Larger studies exploring the specific impact of obesity and inflammation on surgical outcomes, including pain, are warranted. LEVEL OF EVIDENCE: Level II, therapeutic study. See the Instructions for Authors for a complete description of levels of evidence.