Clinical and economic outcomes associated with lymph node examination status in early-stage non-small cell lung cancer: a real-world US study using the SEER-Medicare linked database.

Background: Clinical practice guidelines recommend adjuvant therapy for patients with early non-small cell lung cancer (eNSCLC), especially those with lymph node metastasis. This study evaluated the prevalence of lymph node examination and its association with adjuvant treatment rates, overall survival (OS), and healthcare costs among United States (US) Medicare patients with resected eNSCLC. Methods: This retrospective observational cohort study used Surveillance, Epidemiology, and End Results cancer registry data linked with Medicare claims data. Eligible patients were aged ≥65 years with newly diagnosed non-small cell lung cancer (NSCLC) stages IA to IIIB [the American Joint Committee on Cancer (AJCC) Cancer Staging Manual, 7th edition] between January 2010 and December 2017 with surgery ≤1 month prior to or ≤12 months after diagnosis. Patients were grouped by lymph node examination status: no examination (pNX), examination and no metastasis (pN0), or metastasis staging in N1 (pN1) or N2 (pN2). OS and costs were evaluated by examination status and number of lymph node examined. OS was analyzed using extended Cox proportional hazards models for specific time periods and time interaction with examination status, and adjusted for patient characteristics. Adjusted post-surgical healthcare costs per patient per month (PPPM) were analyzed using gamma-log regression models. Results: Among the 14,648 patients included in the study, approximately 11% were pNX, whereas most were pN0 (68%), followed by pN1 (11%) and pN2 (10%). Adjuvant treatment rates were higher for pNX (35%) than pN0 (18%), but lower than pN1 (68%) and pN2 (74%) patients (P<0.001). Unadjusted OS for pNX patients was nearly identical to pN2, and significantly worse compared to pN0 and pN1 (P<0.0001). After adjusting for patient characteristics, pNX patients had higher risk of death relative to pN0 patients (P<0.001). Marginal mean adjusted total costs were comparable across pNX ($15,827 PPPM), pN0 ($12,712 PPPM) and pN1 ($17,089 PPPM), but significantly less for pN0 compared to pN2 ($23,566 PPPM) (P=0.002). Conclusions: Inadequate lymph node examination is associated with underutilization of adjuvant treatment and poor OS in resected NSCLC. In the current era of targeted and immunotherapies, lymph node examination is more important than ever, implicating the need for Quality Improvement practices and multidisciplinary coordination.

Translational Potential of Baicalein in Mitigating RSL3-Induced Ferroptosis in Fibroblasts: Implications for Therapeutic Interventions.

Background: Ferroptosis is an iron-driven cell-death mechanism that plays a central role in various diseases. Recent studies have suggested that baicalein inhibits ferroptosis, making it a promising therapeutic candidate. Materials and Methods: Fibroblast cultures were treated with different agents to determine the effects of baicalein on ferroptosis. Ferroptosis-related gene expression, lipid peroxidation, and post-treatment cellular structural changes were measured using real-time quantitative polymerase chain reaction, C11-BODIPY dye, and transmission electron microscopy, respectively. Results: Baicalein significantly inhibited rat sarcoma virus selective lethal 3-induced ferroptosis in fibroblasts. Moreover, in baicalein-treated groups, reduced ferroptosis-related gene expression, decreased lipid peroxidation, and maintained cell structure was observed when compared with those of the controls. Discussion: The ability of baicalein to counteract RSL3-induced ferroptosis underscores its potential protective effects, especially in diseases characterized by oxidative stress and iron overload in fibroblasts. Conclusion: Baicalein may serve as a potent therapeutic agent against conditions in which ferroptosis is harmful. The compound's efficacy in halting RSL3-triggered ferroptosis in fibroblasts paves the way for further in vivo experiments and clinical trials.

m-nitrobenzyl alcohol supercharging reagent enhances the chromatographic separation and the charging of disulfide bond linked and His-tag peptides.

The linkages of disulfide bond (DSB) play important roles in protein stability and activity. Mass spectrometry-based (MS-based) techniques become accepted tools for DSB analysis in the recent decade. In the bottom-up approach, after enzyme digestion, the neighbouring amino acids of cysteines have great impacts on the physicochemical properties of resulting disulfide bond peptides, determining their retention behaviour on liquid chromatography (LC) and their MS ionization efficiency. In this study, the addition of supercharging reagent in LC mobile phase was used to examine the impact of supercharging reagent on the charge states of disulfide-bond peptides. The results showed that 0.1 % m-nitrobenzyl alcohol (m-NBA) in LC mobile phase increased the sensitivity and charge states of DSB peptides from our model protein, equine Interleukin-5 (eIL5), as well as the resolution of reversed-phase chromatography. Notably, also the sensitivity of C-terminal peptide with His-tag significantly improved. Our findings highlight the effectiveness of employing m-NBA as a supercharging reagent when investigating disulfide-linked peptides and the C-terminal peptide with a His-tag through nano-liquid chromatography mass spectrometry.

Engagement of sialylated glycans with Siglec receptors on suppressive myeloid cells inhibits anticancer immunity via CCL2.

The overexpression of sialic acids on glycans, called hypersialylation, is a common alteration found in cancer cells. Sialylated glycans can enhance immune evasion by interacting with sialic acid-binding immunoglobulin-like lectin (Siglec) receptors on tumor-infiltrating immune cells. Here, we investigated the effect of sialylated glycans and their interaction with Siglec receptors on myeloid-derived suppressor cells (MDSCs). We found that MDSCs derived from the blood of lung cancer patients and tumor-bearing mice strongly express inhibitory Siglec receptors and are highly sialylated. In murine cancer models of emergency myelopoiesis, Siglec-E knockout in myeloid cells resulted in prolonged survival and increased tumor infiltration of activated T cells. Targeting suppressive myeloid cells by blocking Siglec receptors or desialylation strongly reduced their suppressive potential. We further identified CCL2 as a mediator involved in T-cell suppression upon interaction between sialoglycans and Siglec receptors on MDSCs. Our results demonstrated that sialylated glycans inhibit anticancer immunity by modulating CCL2 expression.

Focused ultrasound stimulation of infralimbic cortex attenuates reinstatement of methamphetamine-induced conditioned place preference in rats.

Methamphetamine (MA) use disorder poses significant challenges to both the affected individuals and society. Current non-drug therapies like transcranial direct-current stimulation and transcranial magnetic stimulation have limitations due to their invasive nature and limited reach to deeper brain areas. Transcranial focused ultrasound (FUS) is gaining attention as a noninvasive option with precise spatial targeting, able to affect deeper areas of the brain. This research focused on assessing the effectiveness of FUS in influencing the infralimbic cortex (IL) to prevent the recurrence of MA-seeking behavior, using the conditioned place preference (CPP) method in rats. The study involved twenty male Sprague-Dawley rats. Neuronal activation by FUS was first examined via electromyography (EMG). Rats received alternately with MA or saline, and confined to one of two distinctive compartments in a three compartment apparatus over a 4-day period. After CPP test, extinction, the first reinstatement, and extinction again, FUS was applied to IL prior to the second MA priming-induced reinstatement. Safety assessments were conducted through locomotor and histological function examinations. EMG data confirmed the effectiveness of FUS in activating neurons. Significant attenuation of reinstatement of MA CPP was found, along with successful targeting of the IL region, confirmed through acoustic field scanning, c-Fos immunohistochemistry, and Evans blue dye staining. No damage to brain tissue or impaired locomotor activity was observed. The results of the study indicate that applying FUS to the IL markedly reduced the recurrence of MA seeking behavior, without harming brain tissue or impairing motor skills. This suggests that FUS could be a promising method for treating MA use disorder, with the infralimbic cortex being an effective target for FUS in preventing MA relapse.

Quantifying lower limb muscle stiffness in typically developing children and adolescents using acoustic radiation force impulse shear wave elastography (ARFI/SWE)-a pilot study.

OBJECTIVE: To investigate and quantify age-related changes in lower limb muscle stiffness in typically developing children and adolescents using acoustic radiation force impulse shear wave elastography. MATERIALS AND METHODS: Shear wave velocities of bilateral rectus femoris, tibialis anterior, and medial gastrocnemius muscles at rest were obtained in typically developing children and adolescents aged 3 to 18 years. The participants were classified into three age groups: Group 1 (children), 3 to 7 years old; Group 2, 8 to 12 (pre-adolescent); and Group 3 (adolescent), 13 to 18. The shear wave velocities of muscle were compared across the three age groups, as well as compared between right- and left-side limbs. The correlation between shear wave velocities and body weight or body mass index was assessed. RESULTS: Of the 47 participants, 21 were in Group 1, 17 in Group 2, and 9 in Group 3. There were no significant differences among the three age groups' shear wave velocities of bilateral lower limb muscles, and no significant differences between right and left sides. There was no correlation between muscle stiffness and body weight or body mass index. CONCLUSION: The present pilot study applied acoustic radiation force impulse shear wave elastography to quantify lower limb muscle stiffness in typically developing children and adolescents aged 3 to 18 years, suggesting no marked change in muscle stiffness occurs as they develop.

Influenza vaccination is associated with lower risk of renal cell carcinoma among chronic kidney disease patients: a population-based cohort study.

Background: Chronic kidney disease (CKD) patients possess a higher risk for renal cell carcinoma (RCC) possibly because of related underlying inflammation and immune dysregulation. In the current population-based cohort study, we evaluate the effects of influenza vaccination on RCC among CKD patients. Methods: We analysed the vaccinated and unvaccinated CKD patients (≥55 years of age) identified from the Taiwan National Health Insurance Database. Propensity score matching was used to reduce the selection bias. Subgroup analyses based on comorbid conditions, dialysis status and vaccinated dosages were also conducted. Results: The incidence of RCC decreased significantly in the vaccinated compared with unvaccinated group {unadjusted hazard ratio [HR] 0.50 [95% confidence interval (CI) 0.31-0.81], P < .01; adjusted HR 0.46 [95% CI 0.28-0.75], P < .01}. Such protective effects of influenza vaccination were noted significantly among those ≥75 years of age [unadjusted HR 0.29 (95% CI 0.12-0.74), P < .01; adjusted HR 0.22 (95% CI 0.08-0.58), P < .01]. A reverse association was noted between the total number of vaccinations and RCC events in both unadjusted and adjusted models. The Kaplan-Meier estimates of the RCC events showed significantly higher free survival rates in the vaccinated as compared with the unvaccinated patients (logrank P = .005). Conclusion: This population-based cohort study found a significant inverse relationship between influenza vaccination and the risk of RCC in CKD patients and the protective effects were more prominent in patients >75 years of age. A possible relation exists between the total number of vaccinations and RCC events. Future randomized clinical and basic studies will be needed to prove these findings and underlying pathophysiological mechanisms.

Acoustic radiation force impulse shear wave elastography quantifies upper limb muscle in patients with Duchenne muscular dystrophy.

We investigated whether the upper limb muscle stiffness quantified by the acoustic radiation force impulse shear wave elastography (ARFI/SWE) is a potential biomarker for age-related muscle alteration and functional decline in patients with Duchenne muscular dystrophy (DMD). 37 patients with DMD and 30 typically developing controls (TDC) were grouped by age (3-8, 9-11, and 12-18 years). ARFI/SWE measured the biceps and deltoid muscle's shear wave velocities (SWVs). Performance of Upper Limb Module (PUL 1.2 module) assessed muscle function in DMD patients. Mann Whitney test compared muscle SWVs between DMD and TDC, stratified by three age groups. We used analysis of variance with Bonferroni correction to compare muscle SWVs between DMD and TDC and correlated muscle SWVs with PUL results in the DMD group. Results showed that the SWVs of biceps differentiated DMD patients from TDC across age groups. Younger DMD patients (3-8 years) exhibited higher SWVs (p = 0.013), but older DMD patients (12-18 years) showed lower SWVS (p = 0.028) than same-aged TDC. DMD patients had decreasing biceps SWVs with age (p < 0.001), with no such age effect in TDC. The SWVs of deltoid and biceps positively correlated with PUL scores (r = 0.527 âˆ¼ 0.897, P < 0.05) and negatively correlated with PUL timed measures (r = -0.425 âˆ¼ -0.542, P < 0.05) in DMD patients. Our findings suggest that ARFI/SWE quantifying the SWVs in upper limb muscle could be a potential biomarker to differentiate DMD from TDC across ages and that DMD patients showed age-related muscle alteration and limb functional decline.

ALK Inhibitor Treatment Patterns and Outcomes in Real-World Patients with ALK-Positive Non-Small-Cell Lung Cancer: A Retrospective Cohort Study.

BACKGROUND: Randomized trials have demonstrated that anaplastic lymphoma kinase (ALK) tyrosine kinase inhibitors (TKIs) can be safe and efficacious treatments for patients with ALK-positive advanced non-small-cell lung cancer (aNSCLC). However, their safety, tolerability, effectiveness, and patterns of use in real-world patients remain understudied. OBJECTIVE: We sought to assess the overall treatment pattern characteristics, safety, and effectiveness outcomes of real-world patients with ALK-positive aNSCLC receiving ALK TKIs. PATIENTS AND METHODS: This retrospective cohort study using electronic health record data included adult patients with ALK-positive aNSCLC receiving ALK TKIs between January 2012 and November 2021 at a large tertiary medical center, University of California, San Francisco (UCSF), with alectinib or crizotinib as the initial ALK TKI therapy. Our primary endpoints included the incidence of treatment changes (treatment dose adjustments, interruptions, and discontinuations) during the initial ALK TKI treatment, the count and type of subsequent treatments, rates of serious adverse events (sAEs), and major adverse events (mAEs) leading to any ALK TKI treatment changes. Secondary endpoints included the hazard ratios (HRs) for median mAE-free survival (mAEFS), real-world progression-free survival (rwPFS), and overall survival (OS) when comparing alectinib with crizotinib. RESULTS: The cohort consisted of 117 adult patients (70 alectinib and 47 crizotinib) with ALK-positive aNSCLC, with 24.8%, 17.9%, and 6.0% experiencing treatment dose adjustments, interruptions, and discontinuation, respectively. Of the 73 patients whose ALK TKI treatments were discontinued, 68 received subsequent treatments including newer generations of ALK TKIs, immune checkpoint inhibitors, and chemotherapies. The most common mAEs were rash (9.9%) and bradycardia (7.0%) for alectinib and liver toxicity (19.1%) for crizotinib. The most common sAEs were pericardial effusion (5.6%) and pleural effusion (5.6%) for alectinib and pulmonary embolism (6.4%) for crizotinib. Patients receiving alectinib versus crizotinib as their first ALK TKI treatment experienced significantly prolonged median rwPFS (29.3 versus 10.4 months) with an HR of 0.38 (95% CI 0.21-0.67), while prolonged median mAEFS (not reached versus 91.3 months) and OS (54.1 versus 45.8 months) were observed in patients receiving alectinib versus crizotinib but did not reach statistical significance. Yet, it is worth noting that there was a high degree of cross-over post-progression, which could significantly confound the overall survival measures. CONCLUSIONS: We found that ALK TKIs were highly tolerable, and alectinib was associated with favorable survival outcomes with longer time to adverse events (AE) requiring medical interventions, disease progression, and death, in the context of real-world use. Proactive monitoring for adverse events such as rash, bradycardia, and hepatotoxicity may help further promote the safe and optimal use of ALK TKIs in the treatment of patients with aNSCLC.

The effect of physical activity on sleep disturbance in various populations: a scoping review of randomized clinical trials.

BACKGROUND: Promoting physical activity (PA) in different populations experiencing sleep disturbance may increase population PA levels and improve sleep. This scoping review aimed to examine the effect of various PA intervention strategies on sleep across different populations, identify key sleep outcomes, and analyze knowledge gaps by mapping the relevant literature. METHODS: For this study, we systematically searched articles published till March 2022 from PubMed, Web of Science, Cochrane Library, and Embase databases for randomized clinical trials (RCTs) regarding the effect of physical activity on sleep. Two authors extracted key data and descriptively analyzed the data. Thematic analysis was used to categorize the results into themes by all authors. Arksey and O'Malley's scoping review framework was used to present the findings. RESULTS: Twenty-one randomized controlled trials out of 3052 studies were finally included with 3677 participants (2852 females (78%)). Five trials were conducted in healthy working-age adults with sleep disturbance but without the diagnosis of insomnia, five in healthy older adults, two in perinatal women, four in patients with cancer, three in mental illness related subjects, and another two in other disease-related areas. PA interventions were diverse, including walking, resistance training, aerobic exercise, housework, water exercise, basketball, smartphone/tablet "apps", web, online videos or wearable actigraphy, and self-determined exercise. Three major themes were identified: (1) Sleep environment may be important to address prior to instituting PA interventions, (2) All types of PA were effective for improving sleep in all populations studied, (3) Self-tolerated PA is safe for improving sleep in the elderly and in co-morbid or perinatal populations. CONCLUSIONS: PA is effective and safe for improving sleep in both healthy and co-morbid populations with sleep disturbance by increasing daily activity levels using a variety of strategies, even low intensity, such as housekeeping, sit-to-stand repetitions, along with encouraging PA through web pages, videos, and self-goal setting apps. In addition, this scoping review identifies the need for further therapeutic research and future exploration in populations with sleep initiation or sleep maintenance disturbance.

Non-Contrast-Enhanced MR Arteriography of Potential Living-Related Liver Donor: Using Contrast Enhanced CT Arteriography as Standard Reference.

BACKGROUND: Contrast-enhanced computed tomography angiography (CTA) and magnetic resonance angiography (MRA) are the primary modalities to assess donors' vessels before transplant surgery. Radiation and contrast medium are potentially harmful to donors. PURPOSE: To compare the image quality and visualization scores of hepatic arteries on CTA and balanced steady-state free-precession (bSSFP) non-contrast-enhanced MRA (NC-MRA), and to evaluate if bSSFP NC-MRA can potentially be a substitute for CTA. STUDY TYPE: Prospective. POPULATION: Fifty-six consecutive potential living-related liver donors (30.9 ± 8.4 years; 31 men). FIELD STRENGTH/SEQUENCE: 1.5T; four bSSFP NC-MRA sequences: respiratory-triggered (Inhance inflow inversion recovery [IFIR]) and three breath-hold (BH); and CTA. ASSESSMENT: The artery-to-liver contrast (Ca-l) was quantified. Three radiologists independently assigned visualization scores using a four-point scale to potential origins, segments, and branches of the hepatic arteries, determined the anatomical variants based on Hiatt's classification, and assessed the image quality of NC-MRA sequences. STATISTICAL TESTS: Fleiss' kappa to evaluate the readers' agreement. Repeat measured ANOVA or Friedman test to compare Ca-l of each NC-MRA. Friedman test to compare overall image quality and visualization scores; post hoc analysis using Wilcoxon signed-rank test. P-value <0.05 was considered statistically significant. RESULTS: Inhance IFIR Ca-l was significantly higher than all BH bSSFP Ca-l (0.56 [0.45-0.64] vs. 0.37 [0.29-0.47] to 0.41 [0.23-0.51]). Overall image quality score of BH bSSFP TI1200 was significantly higher than other NC-MRA (4 [4-4] vs. 4 [3 to 4-4]). The median visualization scores of almost all arteries on CTA were significantly higher than on NC-MRA (4 [3 to 4-4] vs. 1 [1-2] to 4 [4-4]). The median visualization scores were all 4 [4-4 ] on Inhance IFIR with >92.3% observed scores ≥3, except the segment 4 branch (3 [1-4], 53.6%). The identification rates of arterial variants were 92.9%-97% on Inhance IFIR. DATA CONCLUSIONS: Although CTA is superior to the NC-MRA, all NC-MRA depict the donor arterial anatomy well. Inhance IFIR can potentially be an alternative image modality for CTA to evaluate the arterial variants of living donors. LEVEL OF EVIDENCE: 3 TECHNICAL EFFICACY: Stage 2.

A new technique for separating platelet-rich plasma by a copolymer device － without a centrifugation process.

BACKGROUND: Platelet-rich plasma (PRP) is beneficial for clinical applications in various medical fields. Although commercial PRP preparation kits are already available in the market, most of these kits employ centrifugation. METHODS: We used a new cationic copolymer coating on a polyurethane (PU) sponge to promote platelet separation from the blood. This copolymer showed no cytotoxicity against cell viability or hemolysis. We further evaluated the efficiency of the new PRP preparation device by comparing it with that of a commercially available kit (RegenKit-THT). RESULTS: We demonstrated that PRP obtained using copolymer device contains high concentrations of platelets and angiogenic growth factors (epidermal growth factor, vascular endothelial growth factor-A, growth differentiation factor 2, and interleukin-8). The separated PRP also displayed beneficial effects on cell migration, angiogenesis, and matrix metalloproteinase gene expression. CONCLUSION: Based on these results, we developed a cationic copolymer-coated PU sponge as a PRP preparation device without the need for any centrifugation.

State-of-the-art of ultrasound-triggered drug delivery from ultrasound-responsive drug carriers.

INTRODUCTION: Delivering sufficient therapeutics at the target site without off-target effects is a major goal of drug delivery technology innovation. Among the established methods, ultrasound (US) with US-responsible carriers holds great promise and demonstrates on-demand delivery of a variety of functional substances with spatial precision of several millimeters in deep-seated tissues in animal models and humans. These properties have motivated several explorations of US with US responsible-responsible carriers as a modality for neuromodulation and the treatment of various diseases, such as stroke and cancer. AREAS COVERED: We briefly discuss three specific mechanisms that enhance in vivo drug delivery via US with US-responsible carriers: 1) permeabilizing cellular membrane, 2) increasing the permeability of vessels, and 3) promoting cellular endocytotic uptake. We then reviewed the state-of-the-art materials for US-triggered drug delivery, including conventional US contrast agents, and nanocarrier formulations, such as inorganic nanoparticles and gas vesicles. EXPERT OPINION: In this article, we summarized recent progress for each of US-responsible drug carrier, focusing on the routes of enhancing delivery and applications. The mechanisms of interaction between US-responsible carriers and US waves, such as cavitation, streaming, hyperthermia, and ROS, as well as how those interactions can improve drug release and cell/tissue uptake.

Alignment of health plan coverage policies for somatic multigene panel testing with clinical guidelines in select solid tumors.

Aim: Commercial plan coverage policies for multigene panel tests may vary and could result in geographic variation in coverage due to the fragmented nature of the commercial insurance market. This study aimed to characterize the alignment of multigene panel tests coverage policies to that of clinical guidelines, overall and by state. Materials & methods: We reviewed NCCN Guidelines® for four tumors. Public coverage policies were identified via web search. Payer policies included those with the largest or second largest number of commercial lives in each state. Policies were classified as 'more restrictive' or 'consistent' with the guidelines. Results: Of 38 plans/policies reviewed, 71% were classified as 'more restrictive' than the guidelines, with variation in the number of commercial lives by state. Among these, 52% restricted on panel size and 63% restricted in all or select tumors. Conclusion: Most coverage policies were more restrictive. Clinical guideline clarity and state policies may improve alignment to guidelines and geographic variations.

The level of coverage provided by commercial plan policies for tests that simultaneously assess multiple genes for cancer (multigene panel tests) may vary, and the availability of these tests differs depending on geographic location. This may be due to the variable nature of the commercial insurance market. This study aimed to understand how well aligned the coverage for multigene panel tests provided by insurance policies is with recommendations from clinical guidelines, both overall and by US state. NCCN Guidelines^® for four cancer types were reviewed and public insurance coverage policies were identified via internet searches. The insurance policies included those with the largest or second largest number of commercial lives in each state. Policies were judged to be either 'more restrictive' or 'consistent' with the clinical guidelines. Of the 38 plans/policies we reviewed, almost three-quarters (71%) were judged to be 'more restrictive' than the guidelines, with differences in the number of commercial lives by state. About half (52%) were restricted on the number of genes to be tested and almost two-thirds (63%) restricted in all or select types of cancer. We found that most insurance policies were more restrictive than the guidelines ­ therefore covering fewer genes in each test, a different set of genes in each test or restricted to certain types of cancer. Clearer clinical guideline and state policies may help to improve alignment to guidelines and geographic differences.

Recovery of Dysregulated Genes in Cancer-Related Lower Limb Lymphedema After Supermicrosurgical Lymphaticovenous Anastomosis - A Prospective Longitudinal Cohort Study.

PURPOSE: This study aims at profiling the expression of dysregulated genes in circulating monocytes of patients with cancer-related lower limb lymphedema before and after treatment with supermicrosurgical lymphaticovenous anastomosis (LVA). MATERIALS AND METHODS: This prospective longitudinal cohort study enrolled 51 women with post-treatment gynecological cancer, including those with unilateral lymphedema (study group, n = 25) and those without (control group, n = 26). Venous blood samples obtained from the study group before and after LVA and those from the controls were sent for next-generation sequencing, which was validated by real-time PCR. Dysregulated gene expression in the study group, relative to expression in the controls, was recorded before LVA. After one month, postoperative changes in the expression of the identified genes were evaluated. Protein-protein interaction (PPI) was used to investigate dysregulated genes whose expression returned to baseline levels after LVA. RESULTS: Of the 148 preoperative dysregulated genes, which comprised 108 up- and 40 down-regulated genes, 78 genes, consisting of 69 up- and 9 down-regulated genes, showed post-LVA recovery to baseline levels. Through PPI analysis, five functional modules involving immunity, lipid metabolism, oxidative stress, transcriptional regulators, and tumor suppression, as well as six hub genes (CCL2, LPL, PDK4, FOXO3, EGR1, and DUSP5), were identified. Cross-linking and co-regulated genes between modules were also identified. CONCLUSION: Localized lymphedema leads to dysregulated gene expression in circulating monocytes. The current study is the first to identify the hub genes related to lymphedema and demonstrate the recovery of some dysregulated genes after LVA.

Real-world adherence and persistence with anaplastic lymphoma kinase inhibitors in non-small cell lung cancer.

BACKGROUND: Lung cancer is the leading cause of cancer-related deaths in the United States. Several anaplastic lymphoma kinase (ALK) rearrangement inhibitors have been approved for the treatment of metastatic ALK-positive non-small cell lung cancer (NSCLC). Effective disease management requires an understanding of how these treatments are used in clinical practice, since low treatment adherence and/or early discontinuation have been associated with poor patient outcomes. Owing to the recency of approvals, real-world data on the use of ALK inhibitors in patients with ALKpositive NSCLC are currently limited; this represents a notable gap in our understanding of ALK treatment use. OBJECTIVE: To assess real-world adherence and persistence with ALK inhibitors in patients with ALK-positive NSCLC. METHODS: This retrospective observational study used US commercial claims for patients aged at least 18 years with lung cancer receiving ALK inhibitors (alectinib, brigatinib, ceritinib, crizotinib) between July 1, 2015, and December 31, 2018. Patients' first and any subsequent ALK inhibitor uses were categorized into ALK inhibitor-naive and ALK inhibitor-pretreated cohorts, respectively. Adherence was measured by medication possession ratio and persistence by time from treatment initiation to discontinuation (earliest of a treatment switch or greater than a 60-day gap). Descriptive statistics were used to summarize patient characteristics. Cohort comparisons were made using chi-square tests and t-tests. Persistence and time to next ALK inhibitor were analyzed using Kaplan-Meier methods and the log-rank test. Poisson and Cox regression models of adherence and persistence, respectively, were applied to compare ALK inhibitors. RESULTS: We identified 1,482 patients treated with alectinib (n = 445) or crizotinib (n = 1,037) in the ALK inhibitor-naive cohort; 604, 142, and 134 patients received alectinib, brigatinib, or ceritinib in the ALK inhibitor-pretreated cohort. Adherence during the treatment period (95%-97%) and the proportion of patients with a medication possession ratio of at least 0.8 (92%-95%) were similar for all ALK inhibitors. In the ALK inhibitor-naive cohort, median time to treatment discontinuation with alectinib and crizotinib was 27.1 and 8.8 months, respectively; patients receiving alectinib were 46% less likely to discontinue than patients receiving crizotinib (adjusted hazard ratio [aHR] [95% CI]: 0.54 [0.44-0.65]; P < 0.0001). In the ALK inhibitor-pretreated cohort, the discontinuation risk for alectinib was 64% lower than for ceritinib (aHR [95% CI]: 0.36 [0.27-0.49]; P < 0.0001) and 34% lower than for brigatinib (aHR [95% CI]: 0.66 [0.42-1.02]; P = 0.062). CONCLUSIONS: To our knowledge, this study is the first to address a current research gap by assessing real-world adherence and persistence with ALK inhibitors among patients with ALK-positive NSCLC in real-world clinical practice. Alectinib was associated with longer real-world persistence than other ALK inhibitors, despite similar adherence. Further research with more patients and longer follow-up is needed to link persistence to real-world clinical outcomes. DISCLOSURES: This study was funded by Genentech Inc. Ganti has received research support from Takeda and has provided consulting services to Genentech Inc., AstraZeneca, Flagship Biosciences, Cardinal Health, BioGene, Mirati Therapeutics, Blueprint Medicines, and G1 Therapeutics. Lin, Wong, and Ogale are employees of Genentech Inc. and may own stock in F. Hoffmann-La Roche. Yang was employed by Genentech Inc. at the time of this study. Part of the study findings were presented as a poster at the NCCN 2020 Virtual Annual Conference, April 9, 2020.

Exosomes Secreted by Adipose-Derived Stem Cells Following FK506 Stimulation Reduce Autophagy of Macrophages in Spine after Nerve Crush Injury.

Macrophages emerge in the milieu around innervated neurons after nerve injuries. Following nerve injury, autophagy is induced in macrophages and affects the regulation of inflammatory responses. It is closely linked to neuroinflammation, while the immunosuppressive drug tacrolimus (FK506) enhances nerve regeneration following nerve crush injury and nerve allotransplantation with additional neuroprotective and neurotrophic functions. The combined use of FK506 and adipose-derived stem cells (ADSCs) was employed in cell therapy for organ transplantation and vascularized composite allotransplantation. This study aimed to investigate the topical application of exosomes secreted by ADSCs following FK506 treatment (ADSC-F-exo) to the injured nerve in a mouse model of sciatic nerve crush injury. Furthermore, isobaric tags for relative and absolute quantitation (iTRAQ) were used to profile the potential exosomal proteins involved in autophagy. Immunohistochemical analysis revealed that nerve crush injuries significantly induced autophagy in the dorsal root ganglia and dorsal horn of the spinal segments. Locally applied ADSC-F-exo significantly reduced autophagy of macrophages in the spinal segments after nerve crush injury. Proteomic analysis showed that of the 22 abundant exosomal proteins detected in ADSC-F-exo, heat shock protein family A member 8 (HSPA8) and eukaryotic translation elongation factor 1 alpha 1 (EEF1A1) are involved in exosome-mediated autophagy reduction.

Increased Angiogenesis by Exosomes Secreted by Adipose-Derived Stem Cells upon Lipopolysaccharide Stimulation.

Exosomes secreted by adipose-derived stem cells (ADSCs) enhance angiogenesis and wound healing. However, in clinical settings, wounds may be infected by various bacteria or pathogens. We investigated whether human ADSCs stimulated with lipopolysaccharide (LPS) secrete exosomes (ADSC-LPS-exo) that augment the angiogenesis of human umbilical vein endothelial cells (HUVECs). ExoQuick-TC exosome precipitation solution was used to purify exosomes from human ADSC culture media in the presence or absence of 1 µg/mL LPS treatment for 24 h. The uptake of ADSC-LPS-exo significantly induced the activation of cAMP response element binding protein (CREB), activating protein 1 (AP-1), and nuclear factor-κB (NF-κB) signaling pathways and increased the migration of and tube formation in HUVECs. RNA interference with CREB, AP-1, or NF-κB1 significantly reduced the migration of and tube formation in HUVECs treated with ADSC-LPS-exo. An experiment with an antibody array for 25 angiogenesis-related proteins revealed that only interleukin-8 expression was significantly upregulated in HUVECs treated with ADSC-LPS-exo. In addition, proteomic analysis revealed that eukaryotic translation initiation factor 4E, amyloid beta A4 protein, integrin beta-1, and ras-related C3 botulinum toxin substrate 1 may be potential candidates involved in ADSC-LPS-exo-mediated enhanced angiogenesis.

Identification and characterization of hADSC-derived exosome proteins from different isolation methods.

Exosomes are secreted into the extracellular space by most cell types and contain various molecular constituents, which play roles in many biological processes. Adipose-derived mesenchymal stem cells (ADSCs) can differentiate into a variety of cell types and secrete a series of paracrine factors through exosomes. ADSC-derived exosomes have shown diagnostic and therapeutic potential in many clinical diseases. The molecular components are critical for their mechanisms. Several methods have been developed for exosome purification, including ultracentrifugation, ultrafiltration, density gradient purification, size-based isolation, polymer precipitation and immuno-affinity purification. Thus, we employed four methods to isolate exosomes from the hADSC culture medium, including ultracentrifugation, size exclusion chromatography, ExoQuick-TC precipitation and ExoQuick-TC ULTRA isolation. Following exosome isolation, we performed quantitative proteomic analysis of the exosome proteins using isobaric tags for relative and absolute quantification (iTRAQ) labelling, combined with 2D-LC-MS/MS. There were 599 universal and 138 stably expressed proteins in hADSC-derived exosomes. We proved that these proteins were potential hADSC-derived exosomes markers, including CD109, CD166, HSPA4, TRAP1, RAB2A, RAB11B and RAB14. From the quantitative proteomic analysis, we demonstrated that hADSC-derived exosome protein expression varied, with lipopolysaccharide (LPS) treatment, in the different isolation methods. Pathway analysis and proliferation, migration and endothelial tube formation assays showed varying effects in cells stimulated with hADSC-derived exosomes from different isolation methods. Our study revealed that different isolation methods might introduce variations in the protein composition in exosomes, which reflects their effects on biological function. The pros and cons of these methods are important points to consider for downstream research applications.

Quantifying Lower Limb Muscle Stiffness as Ambulation Function Declines in Duchenne Muscular Dystrophy with Acoustic Radiation Force Impulse Shear Wave Elastography.

Duchenne muscular dystrophy (DMD) is a progressive muscular disease, but validated imaging tools to quantify muscle microstructure alteration as mobility declines are lacking. We aimed to determine the feasibility of using acoustic radiation force impulse shear-wave elastography (ARFI/SWE) in the quantitative assessment of lower limb muscle stiffness in DMD patients. Shear wave velocities (SWVs) of lower limbs were measured in 39 DMD patients and 36 healthy controls aged 3-20 y. Mean SWV values of the controls and of the DMD patients at different ambulatory stages were compared using analysis of variance with Bonferroni correction. The DMD group had increased lower limb muscle stiffness compared with controls. Stiffness of the tibialis anterior and medial gastrocnemius muscle decreased from ambulatory to early non-ambulatory stages, whereas stiffness of the rectus femoris muscle increased from ambulatory to late non-ambulatory stages. We describe how SWV changes in lower limb muscles have the potential to predict ambulatory decline in DMD.