To identify the optimal treatment strategy after first-line induction chemotherapy for metastatic colorectal cancer (mCRC).We conducted a meta-analysis of randomized controlled trials comparing bevacizumab-based maintenance therapy, observation, and continuous chemotherapy.We searched the PubMed, Embase, and Cochrane databases for relevant articles published through March 2018. All randomized phase-III trials evaluating bevacizumab-based maintenance treatment after bevacizumab-based induction treatment were eligible for inclusion. The primary and secondary endpoints were progression-free survival (PFS) and overall survival (OS), respectively. Hazard ratios (HRs) with 95% confidence intervals (CIs) or data for calculating HRs with 95% CIs were extracted. The RevMan v5.3  software was used for data analysis.Nine trials (3121 patients) were included in this meta-analysis. Compared with observation alone, bevacizumab-based maintenance therapy significantly improved PFS  and showed a trend toward prolonged OS . The incidence of grade 3/4 toxicity, including hypertension and fatigue, was higher after maintenance therapy than after observation alone. PFS  and OS  did not differ between the maintenance treatment and continuous chemotherapy groups. Grade 3/4 toxicity, including diarrhea and sensory neuropathy, was less common after maintenance therapy than after continuous chemotherapy.Bevacizumab-based maintenance therapy significantly improved PFS, showed a trend toward prolonged OS, and reduced cumulative grade 3/4 toxicity relative to continuous chemotherapy with comparable efficacy. Although maintenance therapy was beneficial, the optimal strategy should be individualized. Moreover, approximately a quarter of all CRC patients present with mCRC at diagnosis. Chemotherapy is the preferred treatment for mCRC patients for whom complete resection cannot be achieved. Over the past few decades, significant advances have been made in mCRC treatment, resulting in improved outcomes and prolonged survival. Several drugs have been developed to treat mCRC, such as oxaliplatin, the fluoropyrimidines 5-fluorouracil (5-FU) and capecitabine, irinotecan, the epidermal growth factor receptor antibodies cetuximab and erlotinib, and the vascular endothelial growth factor (VEGF) antibody bevacizumab. First-line therapy with bevacizumab combined with multi-drug chemotherapeutic regimens  has increased response rates to 50% to 60%, median progression-free survival (PFS) to 9 to 11 months, and median overall survival (OS) to 30 months in patients with unresectable mCRC.Colorectal cancer (CRC) is one of the most commonly diagnosed malignancies. In 2012, there were an estimated 1.36 million new cases of CRC and 694,000 CRC-related deaths worldwide.,13 A comparative assessment of bevacizumab-based maintenance strategies, continuous chemotherapy, and observation alone may help identify the optimal chemotherapeutic regimen for the sequential treatment of mCRC patients who benefit from first-line therapy. We therefore conducted a meta-analysis of randomized controlled trials evaluating the safety and efficacy of the above 3 strategies in terms of PFS and OS in order to identify the optimal follow-up treatment strategy for mCRC patients.However, there is no consensus on the optimal follow-up treatment strategy—maintenance therapy, continuous chemotherapy, or observation alone—for mCRC patients who benefit from first-line therapy. Continuous chemotherapy leads to an increase in drug-related side effects, and long-term exposure to chemotherapeutic drugs reduces cancer cell sensitivity to drugs, resulting in drug resistance. Moreover, treatment interruption significantly reduces the efficacy of chemotherapy and may even affect a patient's PFS and OS. The concept of maintenance treatment envisages a period of high-intensity chemotherapy, after which those agents that are mainly responsible for cumulative toxicity are stopped. The results from 2 large, prospective, observational studies suggest that continued VEGF inhibition with bevacizumab beyond the initial disease progression could play an important role in improving the overall success of therapy in mCRC patients.22.1Potentially relevant studies were independently identified by 2 authors who conducted a structured literature search of the PubMed, Embase, and Cochrane Library databases and the meeting abstracts of American Society of Clinical Oncology and European Society for Medical Oncology published through March 2018. The searches were systematically performed using Medical Subject Headings, and the full-text search terms for the literature search included “colorectal cancer,” “bevacizumab,” and “maintenance.” The online abstracts of the retrieved studies were screened for eligibility. The references of all eligible studies were manually reviewed to find additional relevant studies.2.2The inclusion criteria for the studies were as follows: phase III randomized controlled trials involving patients with histopathologically confirmed CRC; studies comparing bevacizumab-based maintenance therapy with observation alone or those comparing bevacizumab-based maintenance therapy with continuous chemotherapy; studies that reported one or more of the primary or secondary outcomes; and studies from which we could directly obtain or calculate hazard ratios (HRs) and 95% confidence intervals (95% CIs).The exclusion criteria were as follows: studies that had only a single treatment arm; studies in which data on the primary or secondary outcomes were not available; studies for which we were unable to obtain the full text or those that provided insufficient data; and case reports, meeting abstracts, literature reviews, and animal experiments.2.3Data extraction was performed independently by 2 authors, and the extracted data were entered into a standard data sheet. Data on the following variables were extracted: first author's name, year of publication , journal, affiliated institution, country, study phase, format (full text or abstract), interventional and control treatments, HRs with 95% CIs for PFS and OS, median PFS and OS, randomization method, analysis tool, number of patients randomized, demographic and clinical data , and toxicity (grade 3/4). Any disagreements were resolved by consensus, if necessary, by a third author. When additional information was required, the corresponding authors were contacted via email. All selected trials published as full-text articles were analyzed and classified using the Jadad score when possible. Studies with a Jadad score ≥3 were graded as high quality.2.4All meta-analyses were performed using the software Review Manager, version 5.3 (The Cochrane Collaboration). We performed meta-analyses of PFS, OS, and grade 3/4 toxicities after maintenance treatment versus observation alone and maintenance treatment versus continuous treatment. PFS was defined as the time from maintenance randomization to disease progression or death (not including the induction therapy time). To standardize the data, PFS values from several studies were adjusted to match the above definition. OS was defined as the time from maintenance randomization to death (not including the induction therapy time).α = 0.1. The fixed-effects model was first used to combine the HRs of each group. Significant heterogeneity was deemed present among the studies if the heterogeneity tests yielded a P-value of ≤.10 or an I2 value of >50%. In this case, we applied the random-effects.Heterogeneity between the studies was analyzed using the chi-squared test, with a test boundary value of Statement: Our meta-analysis does not address the subject's life, health, dignity, privacy, and other related issues. All analyses were based on previous published studies, thus no ethical approval or patient consent was required.33.1 CAIRO3, SAKK 41/06, and PRODIGE 9) compared bevacizumab-based maintenance therapy until progression with observation alone, while 5 trials  compared bevacizumab-based maintenance therapy with continuous treatment. The AIO 0207 study had 3 treatment arms, and thus counted as 2 trials. The baseline characteristics of all 9 trials have been summarized in Table A total of 208 articles were retrieved using the initial search query. After a full-text review, 9 trials (in 8 papers), with a combined study population of 3121 mCRC patients, were included in the meta-analysis . Therefore, a random-effects model was used for the analysis  provided PFS. Significant heterogeneity was detected among these trials , hand-and-foot syndrome , and sensory neuropathy  with bevacizumab-based maintenance therapy provided data on PFS. Significant heterogeneity was found among these trials  and showed a trend toward prolonged OS . Although the toxicity of bevacizumab-based maintenance therapy was increased, the patients were well tolerated. However, there were no difference of PFS  and OS  between the bevacizumab alone and combination chemotherapy groups. Furthermore, the toxicity including diarrhea and sensory neuropathy was increased in bevacizumab combination chemotherapy group. Together, these data suggested the bevacizumab alone maintenance therapy maybe the optimal strategy for mCRCs patients.Bevacizumab, a humanized monoclonal antibody against VEGF, selectively blocks VEGF binding to the VEGFR-1 and VEGFR-2 receptors, thereby inhibiting the tumor angiogenesis.Although the OS have not significantly difference between maintenance therapy with bevacizumab alone versus observation alone, there were significant benefits in terms of PFS. However, different follow-up intervals and assessment methods have an impact on PFS data. In the AIO 0207 trial, the median PFS after first-line therapy was slightly but significantly improved by bevacizumab maintenance compared with observation (4.6 vs 3.5 months). Non-inferiority for bevacizumab alone was demonstrated for the primary endpoint in AIO 0207, and during the maintenance phases, CT or MRI scans were done every 6 weeks. In the SAKK 41/06 trial, the median PFS was 4.1 months for the bevacizumab maintenance therapy arm versus 2.9 months for the observation alone arm. The follow-up time and equipment were inconsistent in each trial. Non-inferiority could not be demonstrated for continuing bevacizumab monotherapy, CT scans were also done every 6 weeks until disease progression in SAKK 41/06 trials. CT scans were done every 8 weeks to assess the disease status in the PRODIGE 9 trial. There may be a small difference between trials from the time to start maintenance treatment to the first progression between patients treated with bevacizumab and those in the observation group. and the SAKK41/06 trial. In the CAIRO3 experiment, bevacizumab-based maintenance therapy (CAP-B) cost on average €36,845 more than the observation-alone strategy. In the SAKK41 trial, the average cost (in US dollars) per patient was $37,596  for the bevacizumab maintenance arm and $8180  for the observation group. Compared with observation, maintenance therapy leads to an improvement in the quality of life, but it also leads to an increase in costs. Although there is no consensus on the cost-effectiveness thresholds for cancer treatment, the cost-effectiveness of maintenance treatment cannot be ignored.Cost-effectiveness was mentioned in the CAIRO3 trialThe first-line treatment should also be considered as a potential source of bias. Trials of oxaliplatin-based first-line therapy  that compared maintenance therapy with observation alone showed that maintenance treatment had no significant effect to extend OS. In the PRODIGE 9 trial (100% of patients receiving irinotecan), the irinotecan-based combination with bevacizumab maintenance therapy resulted in prolong the OS. The use of oxaliplatin has cumulative toxicity, especially neurotoxicity. Using irinotecan-based chemotherapy may be more feasible than oxaliplatin-based chemotherapy, which may require more clinical trial on maintenance therapy to further confirm. Based on mature survival data from AIO 0207 trial, patients with left-sided tumors showed a median OS of 24.8 months compared with the right-sided cohort with 18.4 months. In a multivariable model, location of the primary tumor proved to be an prognostic factor. In the AIO 0207 and PRODIGE 9 trials, World Health Organization status ≥2, and more than one metastatic site were associated with a shorter OS. Tumor BRAF mutations were a poor prognostic factor in both trials. In addition, in the CAIRO3 and PRODIGE 9 trials, patients with synchronized metastases who were given bevacizumab-based maintenance therapy had a better OS than the observation-alone group. The PRODIGE 9 trial recommended that patients with poor prognostic molecular markers were unsuitable for maintenance therapy strategies. However, the OPTIMOX series of studies suggested that high-risk patients with poor prognosis can receive maintenance therapy, while observation after first-line therapy may be a more rational strategy in low-risk patients with a good prognosis.CRCs can be characterized by their primary tumor location within the colon. The right and left sides of the colon differ in the clinical features, and chromosomal and molecular characteristics. For the above reasons, in a large number of clinical studies on mCRC patients, the location of the primary colon tumor also has an effect on the therapeutic response.While drawing clear recommendations for optimal maintenance treatment options, we were able to identify key differences in PFS in clinical trials comparing single bevacizumab maintenance therapy with combination chemotherapy. The results of the MACRO trial suggest that single-agent maintenance therapy with bevacizumab may be an appropriate choice for mCRC patients. The “Stop and Go” trial proposes that maintenance therapy with bevacizumab plus capecitabine after first-line chemotherapy with 6 cycles of bevacizumab + XELOX can be considered an appropriate choice. In the Nordic ACT trial and OPTIMOX3 trial, after first-line chemotherapy, maintenance therapy with a combination of erlotinib and bevacizumab demonstrated better PFS than did maintenance therapy with bevacizumab alone. The former is a new non-chemotherapy-based maintenance regimen, whose relatively modest efficacy seems to be outweighed by its significant toxicity, especially rash, diarrhea, and fatigue. Ongoing clinical and translational studies focus on identifying subgroups of patients that may benefit from erlotinib in the maintenance setting. The AIO 0207 trial investigated whether observation strategy or bevacizumab alone are non-inferior to a fluoropyrimidine plus bevacizumab, following first-line treatment with a fluoropyrimidine plus oxaliplatin plus bevacizumab. The results showed that maintenance with a fluoropyrimidine plus bevacizumab provided longer PFS than did de-escalation to bevacizumab monotherapy or to no treatment at all. Furthermore, we found that the incidence of adverse events tended to be higher after the bevacizumab combination first-line treatment regiments than after bevacizumab maintenance treatment or observation alone. Although the 8 trials showed that compared with observation alone, bevacizumab-based maintenance significantly prolonged PFS and improved quality of Life, but without improve OS. Further stratification based on the risk factors, such as primary site of colon cancer, BRAF V600 and RAS mutation status, physical status and number of metastatic sites and so on, the clinical trials of maintenance therapy based on further stratification may be prolong the patient's OS while increasing PFS.The current meta-analysis has several limitations. First of all, this is a meta-analysis at study level. We could not obtain individual patient data from the publication, thus we could not incorporate patients variables into the analysis. Second, there were heterogeneities in the trial design . Furthermore, the variability in the baseline patient characteristics  could not be controlled for. This necessitated adjusting the data according to the study design, which should be considered as a potential source of bias.5Compared with observation alone, bevacizumab-based maintenance therapy significantly prolonged the PFS of mCRC patients. Bevacizumab-based maintenance therapy seems have comparable effectiveness (in terms of PFS and OS) to single drug maintenance chemotherapy with lower cumulative grade 3/4 toxicity. Thus, maintenance therapy with bevacizumab may be a valid option for mCRC patients. Although maintenance therapy has demonstrated significant benefits in clinical studies, the treatment should still be individualized. Irinotecan-based first-line chemotherapy may be more likely to prolong OS than oxaliplatin-based induction chemotherapy, more clinical studies are needed to confirm. Clinical studies are conducted on the basis of further stratification, which may prolong the OS of patients with mCRC.Funding acquisition: Hongbo Ma.Investigation: Hongbo Ma, Nan Tang, Yanyan Li.Methodology: Miaomiao Tao, Xianquan Zhang.Project administration: Hongbo Ma, Qi Zhou.Resources: Hongbo Ma, Yanyan Li.Software: Hongbo Ma, Xiaoli Wu, Miaomiao Tao.Writing – original draft: Hongbo Ma.Writing – review & editing: Hongbo Ma, Xianquan Zhang, Qi Zhou.Qi Zhou orcid: 0000-0002-6391-0453Hongbo Ma orcid: 0000-0003-0712-9926.


Smartphone apps are a potential mechanism for development of self-management skills in people with persistent pain. However, the inclusion of best-practice content items in available pain management apps fostering core self-management skills for self-management support is not known.The aim of the study was to evaluate the contents of smartphone apps providing information on pain management strategies for people with persistent pain facilitating self-management support and to appraise the app quality.A systematic search was performed in the New Zealand App Store and Google Play Store. Apps were included if they were designed for people with persistent pain, provided information on pain self-management strategies, and were available in English. App contents were evaluated using an a priori 14-item self-management support (SMS-14) checklist. App quality was assessed using the 23-item Mobile Apps Rating Scale.Of the 939 apps screened, 19 apps met the inclusion criteria. Meditation and guided relaxation were the most frequently included self-management strategies. Overall, the included apps met a median of 4 (range 1-8) of the SMS-14 checklist. A total of 3 apps  met the largest number of items (8 out of 14) to foster self-management of pain. Self-monitoring of symptoms (n=11) and self-tailoring of strategies (n=9) were frequently featured functions, whereas a few apps had features facilitating social support and enabling communicating with clinicians. No apps provided information tailored to the cultural needs of the user. The app quality mean scores using Mobile Apps Rating Scale ranged from 2.7 to 4.5 (out of 5.0). Although use of 2 apps (Headspace and SuperBetter) has been shown to improve health outcomes, none of the included apps have been evaluated in people with persistent pain.Of the 3 apps  that met the largest number of items to support skills in self-management of pain, 2 apps  were free, suggesting the potential for using apps as a scalable, wide-reaching intervention to complement face-to-face care. However, none provided culturally tailored information. Although 2 apps (Headspace and SuperBetter) were validated to show improved health outcomes, none were tested in people with persistent pain. Both users and clinicians should be aware of such limitations and make informed choices in using or recommending apps as a self-management tool. For better integration of apps in clinical practice, concerted efforts are required among app developers, clinicians, and people with persistent pain in developing apps and evaluating for clinical efficacy. Persistent, noncancer pain is the leading cause of disability worldwide, affecting 1 in 5 people . Persistshared decision making), individuals learn to identify meaningful goals  to improve pain and functioning  [self-monitoring) and tailor strategies that work best for them (self-tailoring) [problem solving) and set meaningful goals [self-efficacy building) to engage in meaningful activities despite pain (acceptance), thereby fostering sustained behavioral change [The current best practice for management of persistent pain involves group-based, multidisciplinary, cognitive behavioral interventions focusing on fostering self-management -7. Self-setting) . Practiciloring) . With thul goals . Positivul goals  ultimatel change .Specialized pain services including multidisciplinary, cognitive behavioral interventions are resource intensive and are often only provided in secondary or tertiary care settings . TherefoSmartphone apps are a potential mechanism for the development of self-management skills. Interventions delivered via smartphones are more likely to be easily accessible to a wider population and are scalable -16, withPrevious reviews have examined the quality and content of self-management apps for people with persistent pain ,24. HoweIn light of such limitations, this review extends previous work by adopting a comprehensive International Classification of Diseases (ICD-11) definition for persistent pain  and evalThis review was reported according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines for systematic reviews . The revPersistent pain was defined as recurrent pain for more than 3 months .pain, pain management, and chronic pain. An updated search was conducted on December 07, 2018, using the term pain management in both Google Play and App stores  and the App Store (iOS)—via the website fnd.io  —on Novea fnd.io ) to idenApps were included if they were (1) smartphone apps targeted to adults (>16 years) with persistent pain, (2) capable of being used for any persistent pain condition, (3) able to provide information on at least one or more of the following recommended strategies for pain management: pain education, activity pacing, thought and behavioral management, exercises, relaxation or breathing, meditation or mindfulness, distraction techniques, and (4) available in English.Apps were excluded if (1) a specific pain condition was targeted , (2) only pain monitoring function was provided, and (3) the app was an advertisement for a specific clinic or product.lite in the title) and fully featured paid version of a same app were available, we assessed only the fully featured paid version so that the full functionality of the app was evaluated. A list of the final included apps was collated into an Excel spreadsheet (Microsoft Corp), with metadata about each app extracted from the relevant app stores. This included information on developer, price, app size (in megabytes), app version, and a brief summary of app contents.App names and descriptions from the app search in Google Play Store and the App Store were screened against a priori selection criteria. Next, eligible apps were downloaded for further screening using either OnePlus 3 smartphone running Android 8.0.0 for Google Play Store apps or Apple iPhone 6S running iOS 11.1.2 for the iOS platform. Apps found on both platforms were only downloaded on the iOS platform . DF condCurrently, there are no widely accepted frameworks or clinical guidelines recommending the key components of a pain self-management program. Therefore, a customized 14-item self-management support (SMS-14) checklist ,8,24,36 The 23-item Mobile Apps Rating Scale (MARS)  was usedThe initial search identified 600 unique Android apps from the Google Play Store and 339 unique iOS apps from the App Store—A summary of key characteristics of included apps is presented in Overall, the 19 apps met a median of 4 (range 1-8) of the SMS-14 checklist . Free apThe app quality mean scores from MARS evaluation ranged from 2.72 to 4.54 . CurableAlthough this app review identified 19 apps that purported to support self-management of persistent pain, the comprehensiveness of app contents fostering ongoing self-management skills and functions was limited. Of interest, 2 of the 3 apps  including the largest number of items to support self-management of pain were both free, suggesting the potential for using apps as a scalable, wide-reaching intervention to complement face-to-face care. None of the included apps provided information tailored to cultural needs of the user, and none had been evaluated in people with pain. Furthermore, features facilitating individual goal setting, peer support, and communicating health information with clinicians were infrequent.A majority of the included apps n=11) focused on delivering a single self-management strategy  for symptom management but lacked the features to facilitate core self-management skills . Apps with more features facilitating core self-management skills have the potential for facilitating behavioral change , as iden focused Self-management strategies delivered for symptom management were predominantly mindfulness-based cognitive interventions. Mindfulness-based interventions such as stress reduction and cognitive therapy have been shown to improve long-term health outcomes in people with persistent pain . MindfulPain education and activity pacing, 2 widely used self-management strategies were infrequently present in the included apps. For the purpose of the review, pain education was defined relative to concepts of pain neuroscience education, information on pain-stress-depression, medication, and sleep management. Pain neuroscience education has been shown to improve long-term improvements in pain-related disability and functioning . Except quests, power ups, and bad guys to foster goal setting. Despite the differences, further research is required to identify best-practice delivery of the goal-setting features in pain apps that would address the needs of users [Goal setting was found in only 3 of the included apps, a finding similar to previous app-reviews on persistent pain ,31. It hSocial support can foster self-management , but it None of the apps provided culturally tailored information, a finding similar to our previous review on pain self-management websites . Althougevidence base for improved health outcomes in people with anxiety and depression [The included apps had high rating on the MARS. Apps in general scored highly in esthetics and often earned higher scores in engagement, resulting in apps that are easy to use, interactive, and promoted repeat use. Although 2 of our included apps (Headspace and SuperBetter) scored on the MARS item 19 for pression ,39, no apression . Furtherpression . DiffereThe majority of included apps required payment  ranging from a single payment of NZD $2.99 to NZD $25.99 per month, raising the question if pain apps can be an equitable intervention. This is particularly important when considering existing inequities among people with persistent pain. As those in lower socioeconomic areas are overrepresented in this clinical group , they arAlthough this is the first review to comprehensively assess the app contents using best-practice items to support self-management skills, including cultural tailoring in people with persistent pain, the following limitations need to be acknowledged in interpreting the results from this review. First, during our app screening process, we identified a number of popular apps for symptom monitoring , which failed to meet our inclusion criteria because the present review had an inclusion criterion that required apps to provide information on pain management strategies. Although pain intensity assessment is an important aspect of overall self-management, pain intensity alone is a poor indicator of suffering and disability associated with persistent pain . FurtherSecondly, our customized SMS-14 checklist, although based on evidence-based recommendations, has not been tested for validity. However, evidence suggests that technological interventions with more features facilitating core self-management skills  and support functions  have the potential to foster long-term adaptive behavioral change -42. TherThis app review evaluated the contents of apps fostering self-management support for people with persistent pain. Of the 3 apps  that met the largest number of items to support skills in self-management of pain, 2 apps  were free, suggesting the potential for using apps as a scalable, wide-reaching intervention to complement face-to-face care. However, none provided culturally tailored information. Although 2 apps (Headspace and SuperBetter) were validated to show improved health outcomes, none were tested in people with persistent pain. Both users and clinicians have to be aware of such limitations and make informed choices in using and recommending apps as a self-management tool. Concerted efforts are required among app developers, clinicians, and people with persistent pain in developing and testing apps for better integration of such technologies in clinical practice.


What is the prevalence of periodontal disease and citrullinating periodontopathic bacteria in anti–cyclic citrullinated protein–positive at-risk individuals (CCP+ at-risk) compared with a healthy control group and patients with early rheumatoid arthritis (RA)?Porphyromonas gingivalis at healthy periodontal sites compared with the control group and patients with early RA.This cross-sectional study identified an increased prevalence of periodontal disease sites, clinical periodontitis, and periodontal inflamed surface area in CCP+ at-risk individuals and those with early RA compared with a control group. Results showed that CCP+ at-risk individuals had increased abundance of P gingivalis were increased before joint disease and may be a target for prevention.In individuals at risk of RA, periodontitis and  This cross-sectional study estimates the prevalence of periodontal disease and citrullinating periodontopathic bacteria in individuals who test positive for anti–cyclic citrullinated protein (anti-CCP) antibody without rheumatoid arthritis compared with groups of healthy control participants and patients with early rheumatoid arthritis. The prevalence of periodontitis is increased in patients with rheumatoid arthritis (RA) and periodontopathic bacteria can citrullinate proteins. Periodontitis may, therefore, be an initiator of RA and a target for prevention. Periodontal disease and periodontal bacteria have not been investigated in at-risk individuals with RA autoimmunity but no arthritis.To examine periodontal disease and periodontopathic bacteria in anti–cyclic citrullinated protein (anti-CCP) antibody–positive at-risk individuals without arthritis.This cross-sectional study took place at a teaching hospital from April 27, 2015, to May 8, 2017. Forty-eight anti-CCP–positive individuals without arthritis (CCP+ at-risk) were recruited nationally. Twenty-six patients with early RA (ERA) and 32 healthy control individuals were recruited locally. Data were analyzed between June 1, 2017, and December 1, 2017.Periodontal assessment and examination of joints using ultrasonography.Prevalence of diseased periodontal sites, clinical periodontitis, and periodontal inflamed surface area in CCP+ at-risk individuals compared with patients with ERA and healthy individuals matched for age and smoking. Paired-end sequencing of DNA from subgingival plaque from diseased and healthy periodontal sites was performed and DNA was profiled and analyzed.2 [81-504 mm2] vs 40 mm2 [12-205 mm2]). Patients with CCP+ at-risk had increased relative abundance of Porphyromonas gingivalis (but not Aggregatibacter actinomycetemcomitans) at healthy periodontal sites compared with healthy individuals  and patients with ERA .A total of 48 CCP+ at-risk individuals , 26 patients with ERA , and 32 healthy individuals  were recruited. Of 48 CCP+ at-risk individuals, 46 had no joint inflammation on ultrasonography. Thirty-five CCP+ at-risk individuals (73%), 12 healthy individuals (38%), and 14 patients with ERA (54%) had clinical periodontitis. The median (interquartile range) percentage of periodontal sites with disease was greater in CCP+ at-risk individuals compared with healthy individuals (3.3% [0%-11.3%] vs 0% [0%-0.7%]) and similar to patients with ERA (1.1% [0%-13.1%]). Median (interquartile range) periodontal inflamed surface area was higher in CCP+ at-risk individuals compared with healthy individuals (221 mmP gingivalis in CCP+ at-risk individuals. This suggests periodontitis and P gingivalis are associated with disease initiation and could be targets for preventive interventions in RA.This study found increased prevalence of periodontitis and  Such observations suggest a preclinical phase of RA and, importantly, raise the possibility of disease prevention. The enrichment of serum IgA anticitrullinated protein antibodies (ACPA) in individuals at risk of RA suggests mucosal sites  may be important in the earliest phase of RA.5 There is good evidence that periodontitis and RA are clinically associated.8 Furthermore, periodontitis is associated with a specific bacterial signature characterized by the increased abundance of the pathogenic organism Porphyromonas gingivalis alongside a community of other, predominantly anaerobic, organisms.9Porphyromonas gingivalis is capable of citrullinating local antigens by virtue of its peptidylarginine deiminase enzyme.10 In a putative etiological model, virulent strains of P gingivalis at inflamed periodontal sites generate novel citrullinated antigens that trigger a mucosal immune response in certain individuals, possibly those with genetic predispositions.11 Recent data suggest the periodontopathic bacterium Aggregatibacter actinomycetemcomitans may also directly induce neutrophil citrullination at the periodontium12 and therefore potentially initiate ACPA.Autoantibodies associated with rheumatoid arthritis (RA) can be detected in the serum years before patients develop joint inflammation,Despite these observations, to our knowledge, periodontitis and citrullinating bacteria have not been described in individuals at risk of RA. We sought to comprehensively measure periodontitis and the abundance of key citrullinating bacteria in individuals who were ACPA positive , patients with anti-CCP–positive early RA (ERA), and healthy control individuals. We hypothesized that (1) periodontitis would be similarly increased in CCP+ at-risk individuals and those with ERA compared with healthy individuals and (2) there would be an increased abundance of citrullinating periodontopathic bacteria in CCP+ at-risk individuals and patients with ERA compared with healthy individuals.A cross-sectional study of periodontal and clinical parameters was performed in CCP+ at-risk individuals, patients with ERA, and healthy individuals between April 27, 2015, and May 8, 2017. In this exploratory study, we aimed for 30 participants per group, in line with recommendations for pilot studies. Groups were approximately frequency matched during recruitment for age, sex, and smoking status. After 20 CCP+ at-risk individuals, 20 healthy individuals, and 10 patients with ERA were recruited, demographic and smoking data were reviewed. Approximate frequency matching was performed to recruit remaining healthy individuals and patients with ERA within the age range of 31 to 70 years and to recruit balanced numbers within the tertiles of age calculated in the first 20 CCP+ at-risk individual . We approximately matched the proportion of participants who had ever smoked, which was 60% in the first 20 CCP+ at-risk individuals.A shotgun metagenomic analysis was performed on subgingival plaque samples collected during the periodontal assessments.STROBE) reporting guideline.13Ethical approval for this study was provided by the National Research Ethics Service Committee Yorkshire and the Humber, Leeds West. Written informed consent was received from all participants. This study followed the Strengthening the Reporting of Observational Studies in Epidemiology . Those with a positive test result were invited to a dedicated research clinic in Leeds where recruitment for this study was undertaken. Patients from the Leeds early arthritis clinic who were anti-CCP–positive but did not have clinical synovitis were also recruited. Patients with ERA were all anti-CCP–positive and within the first 3 months of disease-modifying antirheumatic drug (DMARD) therapy. Control participants had no joint disease or history of inflammatory arthritis (and no affected first-degree relatives). Control individuals included coworkers at the University of Leeds  and people from the local community . Control participants were typical of the general population with a range of socioeconomic groups represented.We recruited CCP+ at-risk participants from the Leeds at-risk cohort.Demographic details and a serum IgG anti-CCP2 test  were taken at the time of the periodontal assessment.Periodontal assessments were performed at Chapel Allerton Hospital, Leeds. Periodontal status was assessed by 3 experienced dentists . Dentists were blinded to the RA status of the participants. A full-mouth examination of 6 sites on each natural tooth (recorded as a 6-point pocket chart) was performed on each participant. The 6 sites measured were the 4 corners of the tooth and the midpoint between the buccal and lingual aspects of the tooth. The following parameters were recorded at each available dental site: probing pocket depth (PPD) (millimeters), clinical attachment level (CAL) (millimeters), and presence of bleeding on probing (BOP) (indicated as present or absent). The PCP10 periodontal probe (Hu-Friedy) was used for assessment of BOP and PPD. Measurements of PPD were taken along the vertical axis of the tooth at each site using approximately 0.25 N of force.16 To ensure high sensitivity, thresholds for CAL and PPD were deliberately set so that sites with mild, moderate, and severe periodontitis would all be captured. Periodontal sites with 2 mm or greater CAL and 4 mm or greater PPD were defined as periodontitis sites (PDD) and considered to represent sites of current or past (including treated) periodontitis. Periodontal sites with 2 mm or greater CAL, 4 mm or greater PPD, and BOP were defined as active PDD and considered to represent current active periodontitis.Periodontal disease sites were determined according to the recent update to the 1999 American Academy of Periodontology Classification of Periodontal Diseases and Conditions.16 taking into account the distribution, extent, and severity of periodontitis and also the need for treatment.In addition to these parameters, dentists also classified all participants according to overall clinical periodontal status. The periodontal medical record for each participant was reviewed by the 3 dentists who were blinded to all patient details. In each case, clinical periodontal status was agreed by consensus and was classified as follows: (1) healthy (no periodontitis or gingivitis), (2) gingivitis only (no periodontitis), or (3) periodontitis with or without gingivitis. Classification was based on clinical judgment and the update to the 1999 American Academy of Periodontology Classification of Periodontal Diseases and Conditions,17 This index has been proposed as a way of more accurately quantifying inflamed periodontal tissues.17To quantify the total burden of periodontal inflammation, the total periodontal inflamed surface area (PISA) was calculated for each participant from PPD and CAL measurements at each dental site using the method described by Nesse et al.19Ultrasonography (US) evaluation was performed on all CCP+ at-risk individuals by 2 experienced musculoskeletal sonographers (J.L.N. and L.H.). A standardized 38-joint US protocol was used  as per manufacturer’s instructions and quantified by using PicoGreen dsDNA Reagent and Kits (Thermo Fisher Scientific) (eAppendix 2 in the The DNA was sheared to 200 base pairs (bp) in a small glass vial (microTUBE AFA Fiber Pre-Slit Snap-Cap 6 × 16 mm) by using a S220 Focused-ultrasonicator (Covaris). The size distribution of 4-fold diluted samples was evaluated on the Agilent 2200 TapeStation controlled by Agilent 2200 TapeStation Software A.01.05, using the Agilent High Sensitivity D1000 ScreenTape and Reagents.Depending on the concentration of sheared DNA in the samples, either NEBNext Ultra DNA Library Prep Kit for Illumina or NEBNext Ultra II DNA Library Prep Kit for Illumina (New England Biolabs) was used for library construction, including end preparation, adaptor ligation, and polymerase chain reaction enrichment. AxyPrep Mag PCR Clean-up beads (Corning) were used for the clean-up steps during and after the library preparation to remove unincorporated adaptors, primers, adaptor dimers, primer dimers, and other contaminants. The size distribution and the quantity of the DNA libraries were checked using the method described. We pooled DNA libraries tagged with different index primers together and paired-end sequenced on the Hiseq 3000 machine (Illumina).20 Low-quality regions  and reads shorter than 15 bp were discarded. Artificial replicate sequences and host-specific species sequences  were also removed. Taxonomic abundance profiles at species level were generated by annotation against the RefSeq database housed within MG-RAST with the threshold of 95% identity21 using representative hit strategy. Abundance profiles for P gingivalis, A actinomycetemcomitans, and a control species, Filifactor alocis, were selected and extracted for analysis. We selected F alocis because it is a periodontal pathogen with no known ability to induce citrullination.Sequence data were uploaded to the MG-RAST metagenomics analysis pipeline (version 4.03) for quality processing and basic taxonomic analysis.2, or Fisher exact test where appropriate, was used to compare participant groups. The statistical significance level was set at .05 using 2-sided tests. Spearman ρ correlation was used to assess for associations.Descriptive statistics were used for the analysis of demographic and periodontal characteristics. For continuous data, median and interquartile range (IQR) were presented and Kruskal-Wallis tests were used to compare demographic and periodontal parameters (including PISA) between participant groups. Where significant differences were found for a periodontal parameter, post hoc testing was then performed using Bonferroni correction with a Dunn test. For categorical data , χP gingivalis, A actinomycetemcomitans, and F alocis) were normalized and compared between different groups using the DESeq2 package  in R statistical software version 3.5.1 .22P values were adjusted for multiple testing using false-discovery rate correction. A pseudocount of 0.5 was added to the normalized count to allow for log scale plotting. Effect sizes and standard errors were calculated using DESeq2 based on the negative binomial (gamma-Poisson) distribution.For the analysis of the 3 periodontal pathogens, raw count data of each species , 26 patients with ERA , and 32 healthy individuals  were included. Groups were balanced for age, sex, and smoking status . Of 26 pAll CCP+ at-risk individuals and patients with ERA had tested positive for serum IgG anti-CCP2 antibodies using the Bioplex 2200 kit (BioRad)  when they were recruited to the study. Of these 83 participants, 5 (6%) had a negative serum IgG anti-CCP2 test result using the Immunocap kit (Phadia)  at the tThe percentage of periodontal sites with CAL 2 mm or greater, PPD 4 mm or greater, BOP, PDD, and active PDD were all greater in anti-CCP+ at-risk individuals compared with healthy individuals . In contP = .02). The median (interquartile range) percentage of periodontal sites with disease was greater in CCP+ at-risk individuals compared with healthy individuals (3.3% [0%-11.3%] vs 0% [0%-0.7%]). Rates of periodontitis did not show difference in prevalence between CCP+ at-risk participants and patients with ERA (1.1% [0%-13.1%]) .Overall clinical periodontal status of all participants is shown in 2 (81-504 mm2) compared with 40 mm2 (12-205 mm2) in healthy individuals and 116 mm2 (25-269 mm2) in patients with ERA , indicating higher total periodontal inflammation in CCP+ at-risk individuals compared with healthy participants (P = .002)  PISA in CCP+ at-risk individuals was 221 mmAll CCP+ at-risk individuals underwent US assessment. Synovitis was defined as the presence of gray scale synovial hypertrophy greater than or equal to 1 and PD signal greater than or equal to 1  at the same joint. Of 48 CCP+ at-risk individuals, 46 (96%) had no US synovitis, suggesting the absence of both clinical and subclinical joint inflammation in these subjects. Of the 2 patients who had US synovitis, 1 had synovitis in both first metatarsophalangeal joints and the other in both wrist joints.P gingivalis, A actinomycetemcomitans, and F alocis at healthy and diseased periodontal sites was compared according to RA status. Read counts of P gingivalis, A actinomycetemcomitans, and F alocis were generated from MG-RAST (RefSeq database with 95% identity) and normalized by DESeq2. At healthy periodontal sites, there was a greater abundance of P gingivalis in CCP+ at-risk individuals compared with healthy participants  and patients with ERA  . There were no differences in the abundance of A actinomycetemcomitans or F alocis according to RA status at either healthy or diseased periodontal sites.The abundance of d test]) . InteresP gingivalis at the healthy periodontal sites of CCP+ at-risk participants compared with control individuals and patients with ERA.In this exploratory cross-sectional study, we have identified an increased prevalence of periodontal disease and periodontal inflammation in CCP+ at-risk individuals without joint inflammation. Furthermore, we identified an increased abundance of the periodontopathic bacterium 30 and a recent meta-analysis6 have confirmed an association between periodontal disease and established RA. However, whether or not periodontal disease is truly a trigger for inflammatory arthritis cannot be assessed in patients who have already developed RA. In the current study, we performed comprehensive periodontal examination, using quantitative periodontal measurement, overall clinical periodontal status, and total periodontal inflammatory burden (by PISA) in at-risk participants with anti-CCP antibodies but no clinical or subclinical synovitis. We found an increased prevalence of periodontal disease sites and overall clinical periodontitis as well as an increased PISA in CCP+ at-risk individuals compared with control individuals matched for age and smoking status. The presence of high-titer serum ACPA in these individuals indicates a break in tolerance and the development of RA-related systemic autoimmunity. Prospective data from the CCP+ at-risk cohort suggests that while not all these individuals can be considered to be pre-RA, some will go on to develop clinical arthritis and RA.15 This suggests periodontal inflammation may precede joint inflammation in the development of RA and supports the concept of the periodontium as a mucosal site of disease initiation. These findings are in line with a recent study31 showing a higher frequency and greater severity of periodontal disease in first-degree relatives of patients with RA, although in that study, only 9% of first-degree relatives with periodontitis were anti-CCP positive.Several observational studies33Porphyromonas gingivalis is a key component of the so-called red complex of gram-negative bacteria associated with periodontal disease.9 It has been hypothesized that P gingivalis, through posttranslational citrullination of periodontal mucosal proteins, may provide an antigenic source for ACPA in RA.11 A recent study34 showed oral priming with P gingivalis can trigger an erosive ACPA-positive arthritis in an in vivo animal model. In the current study, we found an increased abundance of P gingivalis in subgingival plaque from the healthy periodontal sites of CCP+ at-risk participants compared with healthy individuals and patients with ERA. This association was not seen at diseased periodontal sites where P gingivalis was identified at similar levels in CCP+ at-risk and healthy participants. The reason for the lower abundance of P gingivalis at diseased sites in patients with ERA compared with the other groups is not fully clear, but this may be due to an early effect of therapy; 62% of these patients were receiving DMARDs and most had also received corticosteroids. It is possible that these treatments may have had an early influence on the periodontal microbiome. As periodontitis may be caused by a dysregulated inflammatory response initiated by the biofilm, it is possible that immunomodulatory therapies could have a direct effect on periodontal inflammation, with consequent effects on the microbiome. Also, DMARDs are believed to have antimicrobial properties.37 It is possible that P gingivalis may be affected in this way, and this would be an interesting area to explore in future work.The microbiome in periodontal disease is distinct from that seen in periodontal health, with an increased abundance of pathogenic bacterial communities.38 Abundance of P gingivalis was no different between groups in that study, which may be expected, as the patients did not have periodontitis. In contrast, in the current study, P gingivalis was enriched at the periodontally healthy sites of CCP+ at-risk participants, the majority of whom also had periodontal disease sites. It is possible that the dysbiosis related to periodontitis has a broader effect specifically in CCP+ at-risk individuals, leading to dysbiosis at distant healthy periodontal sites. Of note, this association was not seen for A actinomycetemcomitans or F alocis, suggesting P gingivalis may be especially significant in CCP+ at risk. While A actinomycetemcomitans has been shown to be capable of citrullination,12 it is particularly associated with aggressive forms of periodontitis that were not seen in our participants.39Interestingly, a recent study found evidence of dysbiosis of the subgingival microbiome in periodontally healthy patients with RA compared with controls, suggesting RA may be associated with changes to the periodontal microbiome independently of periodontal inflammation.40 We were not aware of our participants being affected by these conditions, but they were not specifically excluded. These limitations mean that the findings of this study must be considered as exploratory. These data should be validated in other cohorts, and longitudinal follow-up will be important to assess whether periodontal disease predicts the onset of clinical arthritis in CCP+ at-risk individuals.This study is limited by a relatively small sample size; CCP+ at-risk individuals are difficult to identify and our patients have been recruited from a national primary care study. Owing to frequency matching during the recruitment period, the final group numbers were unequal; despite the planned number of participants being exceeded, fewer patients were identified and recruited in the healthy control group and ERA group compared with the CCP+ at-risk group. Some individuals from all groups declined study participation. Unfortunately, details of eligible participants who declined in each group are not available. It is possible that this self-selection may have introduced bias. However, a participant survey conducted prior to periodontal assessment suggests no difference in access to dental care or self-reported oral symptoms between CCP+ at-risk and healthy control groups (eAppendix 3 and the eTable in the P gingivalis in anti-CCP–positive individuals at risk of RA. These data suggest periodontal inflammation and the enrichment of P gingivalis may precede joint inflammation in RA and support an association between these risk factors and disease initiation. This study adds to an emerging evidence base linking periodontal and systemic disease and, therefore, further highlights the potential importance of improving dental health and reducing the burden of periodontal disease on the risk of chronic systemic diseases such as RA. Importantly, these findings suggest periodontal inflammation may be a legitimate target to explore for preventive intervention in RA.This study is the first, to our knowledge, to demonstrate an increased prevalence of periodontal disease together with an increased abundance of


Although breast cancer mortality is decreasing, morbidity following treatment remains a significant issue, as patients face symptoms such as cancer-related fatigue (CRF). The aim of the present study is to develop a classification system that monitors fatigue via integration of an objective clinical assessment with patient self-report. Forty-three women participated in this research. Participants were post-treatment breast cancer survivors who had been surgically treated for their primary tumour with no evidence of neoplastic disease at the time of recruitment. Self-perceived fatigue was assessed with the Spanish version of the Piper Fatigue Scale-Revised (R-PFS). Objective fatigue was assessed by the 30 second Sit-to-Stand (30-STS) test. Confirmatory factor analysis was done with Maximum Likelihood Extraction (MLE). Internal consistency was obtained by Cronbach's α coefficients. Bivariate correlation showed that 30-STS performance was negatively-inversely associated with R-PFS. The MANOVA model explained 54.3% of 30-STS performance variance. Using normalized scores from the MLE, a classification system was developed based on the quartiles. This study integrated objective and subjective measures of fatigue to better allow classification of patient CRF experience. Results allowed development of a classification index to classify CRF severity in breast cancer survivors using the relationship between 30-STS and R-PFS scores. Future research must consider the patient-perceived and clinically measurable components of CRF to better understand this multidimensional issue. While breast cancer is the most commonly diagnosed type of cancer in women, accounting for 30% of all new diagnoses, mortality rates have decreased by 38% in recent years . HoweverExperience of CRF is essential to monitor for, to enable healthcare practitioners to provide treatment and monitor its effectiveness . FurtherFatigue has been defined as a multidimensional concept involving psychological and physiological dimensions . Given tIn self-reported fatigue, patient-reported outcomes (PRO) are the primary assessment tool. PROs are used to assess a patient's symptoms or functional status at a specific time ,17. AlthWhile PRO tools offer ease in self-monitoring and assessment  and are Given the high prevalence, variability and multidimensionality of CRF in breast cancer survivors, work is needed to develop a more complete objective and subjective assessment of CRF. Hence, the aim of the present study is to develop a classification system that improves clinical assessment and monitoring of fatigue via integration of an objective clinical assessment with patient self-report, using the 30-STS and R-PFS, respectively.Forty-three women aged between 32 and 69 years old volunteered to participate in this cross-sectional study. They were recruited by Medical Oncology faculty specialists from the University Clinical Hospital Virgen de la Victoria . Participants were post-surgery breast cancer survivors who had been surgically treated for their primary tumour with no evidence of neoplastic disease at the time of recruitment. Patients undergoing hormonal treatment were allowed to participate. Exclusion criteria included suffering any cardiovascular event defined as: stable or unstable angina; acute pulmonary oedema; cardiac rhythm disorders; and syncope of cause not affiliated in the year prior to inclusion. All participants signed an informed consent prior to participation. Ethical clearance for the study, following the Helsinki declaration, was obtained from Ethics Committee of the Provincial Research of Málaga, Andalusian Health Service .Participants completed testing at the hospital. In addition to providing demographic and clinical history, they completed the following assessments:Self-perceived fatigue: The Spanish version of the Piper Fatigue Scale-Revised (R-PFS) was used to obtain a subjective PRO of fatigue. This self-administered questionnaire contains 22 items whose scores range from 0 to 10 and includes four domains of subjective fatigue: behavioral/severity (6 items); affective meaning (5 items); sensory (5 items); and cognitive/mood (6 items). The scale has high reliability (Cronbach’s α = 0.96) in the breast cancer population . FurtherClinical assessment of fatigue: To obtain an objective, clinical assessment of fatigue, the 30 second Sit-to-Stand (30-STS) test was used. This test has been demonstrated to produce quadriceps fatigue , used toDescriptive analyses were used to present mean, standard deviation, minimum and maximum of anthropometric variables. Distribution and normality of variables were determined using one-sample Kolmogorov-Smirnov tests. Association between 30-STS performance and R-PFS scores, overall and for each domain, was reported using Pearson´s Correlation Coefficient (r). A multivariate analysis of variance (MANOVA) was completed to further examine relationships between 30-STS performance (number of repetitions) and both overall and individual domain scores on the R-PFS. A confirmatory factor analysis with Maximum Likelihood Extraction (MLE) was employed to determine if sample size was sufficient to construct a classification system with standard scores . InternaParticipants had undergone a lumpectomy (60%) or mastectomy (40%). Further, 93.3% of them had been treated with chemotherapy, radiotherapy and/or hormone therapy, with the vast majority (75%) still on hormone therapy. Additional demographic and clinical variables are presented in Bivariate correlation showed that 30-STS test was negatively-inversely associated with Total Piper Score and all its domains . HoweverTotal R-PFS and individual domain scores were used to develop a MANOVA model examining contribution of total and individual domain scores to 30-STS results. This model explained 54.3% of 30-STS result variance. Based on the decomposition of the model, the main contributing variable was Domain III (p = 0.247) followed by Domain II (p = 0.251). However, these variables did not reach the required value to explain the model by themselves .Kaiser–Meyer–Oklin values (0.865) and the Bartlett’s test of sphericity  (p<0.001) indicated the correlation matrix was adequate for the Maximum Likelihood Extraction. Maximum Likelihood Extraction detected one factor with Eigenvalues above 1, explaining 69.17% of the total variance See . Using tThis study attempted to integrate objective and subjective assessments of fatigue,allowing more thorough classification of patient experience. Study aims were achieved, as results allowed development of an ambulatory index to assess CRF in breast cancer survivors. Results of the 30-STS were correlated with scores from total and each domain of the R-PFS to examine associations and relationships. Based on an observed relationship between the objective (30-STS) and subjective (R-PFS) tests, a scale was developed classifying CRF as one of five levels: Subclinical, mild, moderate, severe or extreme. As highlighted, CRF is multidimensional and includes physical and psychosocial components, necessitating an assessment that captures all domains. While PRO outcome tools such as the R-PFS  and QuicResults from the Pearson correlations highlighted that 30-STS performance was inversely associated with both total and individual domain scores of the R-PFS. As expected, individuals with higher self-perceived fatigue demonstrated poorer performance on the objective test. That is to say, a higher score on R-PFS (suggesting greater perceived fatigue), the fewer repetitions achieved in the 30-STS test. Of note is that the association between 30-STS performance and the cognitive domain of fatigue (Domain III) did not reach statistical significance. This domain relates to fatigue contributing to things like decreased attention span and impaired perception, sometimes considered to be ‘attention fatigue’ . Given tBased on MANOVA results, 54.3% of 30-STS result variance was explained by R-PFS scores. Hence, the number of repetitions is explained 54.3% by self-perceived fatigue, while the remaining 45.7% of variability in the number of repetitions is explained by other factors. Future research could look to further examine other key variables associated with both 30-STS results and R-PFS. As breast cancer survivors with CRF often experience a range of other physical and psychosocial side effects , includiBased on the analyses and results, a classification system was devised using the observed relationship between 30-STS and R-PFS scores . Five raPrevious longitudinal studies have subjectively monitored the trajectory of CRF throughout breast cancer treatment. However, the focus has spanned from mean levels of fatigue to individual levels, making it difficult to get a full idea of overall experience. These studies have resulted in both differentiation of subgroups suffering from high versus low levels of fatigue ,42, as wThis approach to patient classification is used in other groups outside of cancer survivors. For example, the CODI index is an on-line classification tool used to rate severity of fibromyalgia symptoms . CliniciLimitations of this research must be acknowledged. These include a relatively small sample size, only post-treatment survivors and lack of assessment related to the influence of other clinical, psychosocial and socio-demographic factors on results. Further research should look to examine if the association between 30-STS performance and R-PFS remains similar in women still undergoing breast cancer treatment, as well as in other types of cancer. There is also a need to examine what other variables may impact 30-STS performance and correlation with self-reported fatigue levels, as well as greater understanding of the influence of the four fatigue dimensions. Study strengths included range in participant age, years since diagnosis and type of surgery undergone, as more extensive surgery, particularly when involved axillary lymph node dissection, has been linked to greater experience of CRF-related problems .In summary, the fatigue classification system developed in this study has promising clinical and research implications. It supports a more well-rounded, but easily accessible, assessment of CRF and treatment evaluation. By combining objective and subjective measures of fatigue and using these to classify the severity of an individual’s experience of the condition, management strategies can be better prescribed. Additionally, those conducting research on interventions to help with the management and treatment of CRF may use the scale for easier assessment of intervention effectiveness. Patients may also use the tool to track their fatigue and better understand their contributing factors and more effective self-management techniques. As highlighted, future research must consider the patient-perceived and clinically measurable components of CRF to better understand this multidimensional issue.This study integrated objective and subjective assessments of fatigue to better allow classification of patient experience. Results allowed development of an ambulatory index to assess CRF in breast cancer survivors using the relationship between 30-STS and R-PFS scores. By combining objective and subjective measures of fatigue and using these to classify the severity of an individual’s experience of the condition, management strategies can be better prescribed. Future research must consider the patient-perceived and clinically measurable components of CRF to better understand this multidimensional issue.S1 Appendix(DOCX)Click here for additional data file.S2 Appendix(DOCX)Click here for additional data file.S1 Database(SAV)Click here for additional data file.


Breast cancer is the most frequently diagnosed malignancy among women, and triple-negative breast cancer (TNBC) is a highly aggressive subtype. Increasing evidence has shown that lncRNAs are involved in tumor growth, cell-cycle, and apoptosis through interactions with miRNAs or mRNAs. However, there is still limited data on ceRNAs involved in the molecular mechanisms underlying TNBC. In this study, we applied the weighted gene co-expression network analysis to the existing microarray mRNA and lncRNA expression data obtained from the breast tissues of TNBC patients to find the hub genes and lncRNAs involved in TNBC. Functional enrichment was performed on the module that correlated with Ki-67 status the most (Turquoise module). The hub genes in the Turquoise module were found to be associated with DNA repair, cell proliferation, and the p53 signaling pathway. We performed co-expression analysis of the protein-coding and lncRNA hub genes in the Turquoise module. Analysis of the RNA-seq data obtained from The Cancer Genome Atlas database revealed that the protein-coding genes and lncRNAs that were co-expressed were also differentially expressed in the TNBC tissues compared with the normal mammary tissues. On the basis of establishing the ceRNA network, two mRNAs (RAD51AP1 and TYMS) were found to be correlated with overall survival in TNBC. These results suggest that TNBC-specific mRNA and lncRNAs may participate in a complex ceRNA network, which represents a potential therapeutic target for the treatment of TNBC. It is characterized by at least four different clinically relevant molecular subtypes: Luminal A, Luminal B, her2-enriched type, and triple negative breast cancer (TNBC)2. TNBC is generally classified into basal-like and Claudin-low categories3. Due to the lack of expression of estrogen and progesterone receptors and HER2 amplification, there is no targeted therapy for this highly invasive breast cancer subtype4. In addition, TNBC presents with higher aggressiveness and poorer prognosis than the other subtypes as evidenced by the lower survival rate and increased risk of metastasis and recurrence in TNBC6. Therefore, the molecular mechanisms underlying TNBC should be further studied.Breast cancer is the fifth leading cause of death and most frequently diagnosed malignancy in women worldwide7. In recent years, increasing evidence has shown that lncRNAs are involved in tumor growth, cell-cycle, and apoptosis through interactions with miRNAs or mRNAs10. In 2011, Salmena et al. proposed the concept of competing endogenous RNAs (ceRNAs), a class of RNAs with miRNA binding sites with which miRNA-targeted RNAs compete for miRNAs, and thus elucidated a complex post-transcriptional regulatory network including mRNAs, lncRNAs, and other types of RNAs11. LncRNAs can regulate gene expression by interacting with miRNAs. Several studies have confirmed this concept13. However, there has been few studies on the involvement of ceRNA-mediated mechanisms in TNBC.Long noncoding RNAs (lncRNAs) are defined as RNA transcripts ≥200 nucleotides long that do not encode for a protein14. By constructing correlation networks, WGCNA can identify candidate biomarkers and therapeutic targets for different types of cancer18. For instance, a few of hub genes associated with the pathological stage of colon cancer have been obtained by using this method19. In addition, Liu et al.20 have identified gene modules that constitute a recurrence-associated network.Weighted gene co-expression network analysis (WGCNA) is a systematic in silico method for the analysis of complex gene regulatory networks and is based on gene expression data. WGCNA can be used to study biological networks based on genetic correlations. It identifies modules (clusters) of highly correlated geneshttps://tcga-data.nci.nih.gov/tcga/) database. As a result, 4,565 mRNAs, 427 miRNAs, and 4,852 lncRNAs were identified. Finally, 37 lncRNAs, 28 miRNAs, and 16 mRNAs were selected to construct a lncRNA-miRNA-mRNA ceRNA network. The major aim of this study is to discuss the molecular mechanisms underlying TNBC and provide novel prognostic factors for TNBC.In this study, we applied WGCNA in combination with functional enrichment analysis to the available TNBC mRNA and lncRNA expression data to identify the hub genes, including lncRNAs involved in TNBC. Verification and further analysis of the identified protein-coding genes and lncRNAs were conducted using The Cancer Genome Atlas  were screened out. The volcano plots of these mRNAs and lncRNAs are shown in Fig. The GSE76250 database includes 17,643 mRNAs from 198 samples, and GSE65216 database includes 2097 lncRNAs from 66 samples. In total, 9346 differentially expressed genes (DEGs) and 2097 lncRNAs . We filtered out the protein-coding genes and lncRNAs that had MM values > 0.8 and MM P-value < 0.05 in the modules. These hub genes are exhibited in Supplementary Tables The correlation between the tumor characteristics and Module eigengenes (MEs) was determined by the interaction analysis of the modules related to a clinical feature Fig. . The eigThe turquoise module was highly correlated with Ki-67 status. Therefore, the GO enrichment and KEGG pathway analyses were carried out to gain an insight into the biological characteristics of this module. For the turquoise module, the genes were mainly concentrated in GO: 0006281 (DNA repair) and GO: 0008283 (cell proliferation). The KEGG pathway analysis of the turquoise module revealed hsa04110 (Cell cycle), hsa05203 , and hsa04115 . The complete information is shown in Fig. p-value < 0.05, we identified 4,852 lncRNAs, 427 miRNAs, and 4,565 mRNAs differentially expressed in the TNBC tissues compared with the normal mammary tissues. Some of the differences were larger than 80-fold  were found to be correlated with overall survival in TNBC. These data indicate that the identified ceRNA network is involved in the formation and development of TNBC.27. Furthermore, hsa-mir-335 has been reported as the first selective tumor initiation and metastasis suppressor locus in breast cancer in humans28.Recently, miRNAs have extensively been studied. Mechanisms involving miRNAs have been shown to take part in various cancer types, and even the same miRNA can be involved in multiple cancers. For example, in our ceRNA network, hsa-mir-335 and hsa-mir-942 have been shown to play key regulatory roles in a variety of cancers, including epithelial ovarian, lung, and colorectal cancers29, there is little known about the function of RAD51AP1 in TNBC. Additionally, CCNE2 and HIST1H2BF were found to have no significant correlation with overall survival in TNBC (p > 0.05). Nevertheless, CCNE2 may play an important mechanistic role in non-small cell lung cancer and breast cancer31. Unfortunately, there have been few studies about the roles of TYMS and HIST1H2BF in breast cancer. Therefore, further research is needed to clarify the role of these genes in TNBC.Regarding the correlation between the cancer-specific mRNAs and patient prognosis, the results we obtained are not entirely consistent with previous reports. Although high expression of RAD51AP1 has been reported as a biomarker for poor overall survival in lung cancerIn the cytoplasm, ceRNA-mediated regulatory mechanisms constitute an important pathway for lncRNAs to modulate post-transcriptional regulation. Previous studies have shown that lncRNAs can serve as miRNA “sponges,” and compete with miRNA-targeted mRNAs for miRNAs, thereby affecting the miRNA-mediated gene regulation. The ceRNA network includes the genes that we analyzed by the functional enrichment method, such as TYMS in cell proliferation and RAD51AP1 in DNA repair. In addition, we also found that genes were enriched in multiple TNBC-related pathways, such as CCNE2 in the p53 signaling pathway.There are several limitations to our study. First, not all the TNBC GEO data were analyzed in this study. Therefore, due to the two predictions, this study unavoidably suffers from selection bias. Moreover, this study lacks biological experimental confirmation. As the next step, we will validate and further investigate the lncRNA-miRNA-mRNA relationships of the ceRNA network alongside biological experiments.In summary, we have established a ceRNA network in TNBC. Our results suggest that the tumor-specific mRNAs and lncRNAs in TNBC may be involved in a complex ceRNA network, presenting them as potential therapeutic targets for the treatment of TNBC.https://tcga-data.nci.nih.gov/tcga/) data portal on March 29, 2019. Microarray mRNA and lncRNA expression data were downloaded from the NCBI Gene Expression Omnibus (GEO). The GSE76250 dataset contains mRNA expression data from 33 normal breast and 165 TNBC tissue samples, and the GSE65216 dataset contains lncRNA expression data from 11 normal breast and 55 TNBC tissue samples. The raw CEL files derived from each microarray dataset were normalized by using the R package “limma”. Next, we selected the probes for the mRNAs and lncRNAs from the corresponding microarray platforms. Finally, we obtained 17,644 unique protein-coding genes and 2097 annotated lncRNA transcripts that we used for the following analyses.The RNA-sequencing (RNA-Seq-HTSeq) and clinical data of The Cancer Genome Atlas (TCGA) BRCA dataset were downloaded from TCGA | > 1 were identified by Volcano plots by using the R package “ggplot2.”Differentially expressed protein-coding genes (DEGs) and lncRNAs (DE-lncRNAs) in TNBC and normal mammary samples were detected by “edgeR” with R package33. The processes followed to construct the lncRNA co-expression networks were similar to the gene co-expression networks.We used the R package “WGCNA” to construct the protein-coding gene and lncRNA co-expression networks according to the DEGs and DE-lncRNAs, respectively. Firstly, the outlier samples were removed and the hierarchical clustering analysis was performed with the “hclust” R function. Secondly, we used the integral function “pickSoftThreshold” to select the accurate cut-off point. Thirdly, a TOM was transformed from the adjacency. The gene “minModuleSize” was set at 30 to assure high reliability. As for the lncRNA network, since there were few DE-lncRNAs, the minModuleSize was set to 15. Then, to summarize the expression patterns of the module genes into a single characteristic expression profile, each module’s gene expression matrix component, called the module eigengene (ME), was obtained by the WGCNABased on the data source of the DEGs or DE-lncRNAs, the R package “caret” was used to generate two datasets named “train” and “test”. Afterward, we used the WGCNA (module preservation) analysis to calculate the Z scores. The Z scores ranging from 2–10 exhibit low-to-moderate preservation and those >10 exhibit high preservation. It should be noted that, regardless of the protein-coding gene or lncRNA modules, the grey module contains protein-coding genes or lncRNAs that are not part of any module, while the gold module is produced through the preservation function during the statistical analysis. Therefore, the preservation analysis results do not show these two modules.P-value. The MM of a gene was defined as the correlation between ME and the gene expression profile. We defined genes as hub genes in the modules if they had a value of MM > 0.8 and MM P-value < 0.05. The same method was applied to identify the hub lncRNAs.The WGCNA utilizes ME to evaluate the possible connections between the gene modules and clinical traits. In this study, genetic significance (GS) and modular significance (MS) were used to calculate the modular expression patterns associated with the clinical characteristics of the samples. The GS of a gene was defined as the correlation coefficients for different clinical characteristics, and MS was defined as the average GS of all the genes in the module. This analysis was only applied to the protein-coding gene dataset but not to the lncRNA dataset due to the lack of the necessary information about the samples. The hub protein-coding genes were determined by calculating the module membership (MM) and MM https://david.nciferf.gov/home.jsp). The threshold was set as P-value < 0.05. The DEGs were considered as hub genes if they were extracted from the GO functions or KEGG pathways.To explore the functions and pathways related to the hub genes and lncRNAs, we performed the gene ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways analyses by using the DAVID bioinformatics tool . The differentially expressed mRNAs, lncRNAs, and miRNAs between the TNBC and normal mammary tissues were analyzed by edgeR package in R language. A gene was defined as differentially expressed (DE) when the P-value was <0.05 and the fold change (FC) was ≥2-fold higher or lower (|log FC| > 1). The construction of the ceRNA network included three steps: (1) Co-expression analysis of the hub protein-coding genes (associated with Ki-67%) and lncRNAs in the turquoise module (more lncRNAs); (2) The putative mRNA targets of the miRNAs were predicted by using DIANA-microT-CDs (http://diana.imis.athena-innovation.gr/), and only the miRNAs predicted to bind to four genes were retained; (3) We used DIANA-LncBase v2 (http://diana.imis.athena-innovation.gr/) to predict the putative lncRNA targets of the miRNAs obtained in step (2). The intersection of the predicted lncRNA targets and those obtained in step (1) were used for further analyses. The lncRNA-miRNA-mRNA ceRNA network was established and visualized by using Cytoscape v3.034.For validation, RNA and miRNA sequencing data and clinical information were obtained from the TCGA database . The log-rank test was used.Supplementary tables


Few studies have examined melanoma incidence and survival rates among non-Hispanic black populations because melanoma risk is lower among this group than among non-Hispanic white populations. However, non-Hispanic black people are often diagnosed with melanoma at later stages, and the predominant histologic types of melanomas that occur in non-Hispanic black people have poorer survival rates than the most common types among non-Hispanic white people.We used the US Cancer Statistics 2001–2015 Public Use Research Database to examine melanoma incidence and 5-year survival among non-Hispanic black US populations.From 2011 through 2015, the overall incidence of melanoma among non-Hispanic black people was 1.0 per 100,000, and incidence increased with age. Although 63.8% of melanomas in non-Hispanic black people were of unspecified histology, the most commonly diagnosed defined histologic type was acral lentiginous melanoma (16.7%). From 2001 through 2014, the relative 5-year melanoma survival rate among non-Hispanic black people was 66.2%.Although incidence of melanoma is relatively rare among non-Hispanic black populations, survival rates lag behind rates for non-Hispanic white populations. Improved public education is needed about incidence of acral lentiginous melanoma among non-Hispanic black people along with increased awareness among health care providers. Non-Hispanic black people have lower incidence rates of melanoma than non-Hispanic white people; however, non-Hispanic black people are typically diagnosed at a later stage, have different proportions of histologic types of melanoma, and have poorer survival rates than non-Hispanic white people.From 2011 through 2015, the histology of most melanomas diagnosed among non-Hispanic black patients was acral lentiginous melanoma. Five-year relative survival rates for acral lentiginous melanoma are lower than for the predominant histologic type diagnosed among non-Hispanic white people.Improved education of health care providers about incidence of acral lentiginous melanoma among non-Hispanic black people is needed because of its atypical presentation and poor survival rates for this cancer.Melanoma is one of the most common cancers in the United States, and incidence is increasing . Most meSkin type or complexion is a key risk factor for skin cancer . MelaninPrevious analyses found that melanoma histologic type and the body site where it occurs differ by race ,9. AcralInternational Classification of Diseases for Oncology current at the time of diagnosis and later converted to codes in the International Classification of Diseases for Oncology, third edition (ICD-O-3) (We examined population-based cancer registry incidence data from the US Cancer Statistics (USCS) 2001–2015 Public Use Research Database . This daICD-O-3) . We defiP < .05. Survival data were taken from NPCR registries of the 39 participating states with survival data, which covered 81.1% of the US population for 2001 through 2014  to calculate age-adjusted rates per 100,000 population (based on the 2000 US standard population), rate ratios, 95% confidence intervals, average annual counts, and 5-year relative survival. Rates based on a count of fewer than 16 patients were not reported because of concerns for rate stability . We limiugh 2014 . SurvivaFrom 2011 through 2015, 1,795 non-Hispanic black people were diagnosed with melanomas in the United States, an average of 359 per year  . Rates wHalf (55%) of non-Hispanic black people and 78% of non-Hispanic white people were diagnosed with melanoma at a localized stage; 18% of non-Hispanic black people and 9% of non-Hispanic white people were diagnosed with regional-stage melanoma . More noWe examined data on 2,848 melanomas diagnosed from 2001 through 2014 . OverallRelative survival rates differed between non-Hispanic black populations and non-Hispanic white populations by sex and by stage at diagnosis . OverallOur analysis of survival data from 39 states (81.1% of the US population) showed lower survival rates among non-Hispanic black people with melanoma than among non-Hispanic white people ,18. Our Previous research indicated that most melanomas among non-Hispanic black people occur on the lower limbs and hip or lower extremities, including soles of the feet ,9. One sDark skin confers more protection against ultraviolet radiation than fair skin, but skin type and skin tone can vary widely among people of the same race/ethnicity . About 1ALM was the most common melanoma subtype diagnosed among non-Hispanic black populations in our study. Survival is poorer for ALM than for SSM, the most common subtype diagnosed among non-Hispanic white populations. Other studies have found that ALM is associated with lower survival rates than other subtypes and that the high proportion of ALM among non-Hispanic black people may be partially responsible for poor survival outcomes ,10,27. USurvival was also poor among non-Hispanic black people for nodular melanoma. As with ALM, nodular melanoma is associated with poor survival and is often diagnosed in advanced stages ,28. BothOur study had limitations. One limitation was the exclusion from the incidence database of people with melanomas reported only on autopsy or death certificates; however, these patients constitute less than 5% of the database and therefore would not affect the incidence rates significantly if they were included . AnotherA strength of our analysis is its comprehensive coverage of USCS data, which contain cancer incidence rates for the entire US population. Furthermore, this study provides an in-depth look at melanoma incidence and survival rates among the non-Hispanic black population that have not been previously reported.Increased awareness is needed among the public and the medical community of the prevalence of the ALM histology type and the common anatomic sites where it occurs because of ALM’s atypical presentation. Opportunities exist to provide increased education and behavioral counseling among non-Hispanic black populations about minimizing exposure to ultraviolet radiation to reduce skin cancer risk and about the need for regular skin checks by medical professionals, particularly in non-sun–exposed areas such as the feet .


Heart failure (HF) is a chronic condition requiring continuous self-care. Health literacy is increasingly recognized as a key factor of self-care behaviors in patients with chronic diseases. Recently, frailty in chronic diseases has also been associated with self-care behaviors. However, relationships among health literacy, frailty, and self-care in the HF population are not well understood. Therefore, this cross-sectional study aimed to identify the impact of health literacy and frailty on self-care behaviors in patients with HF. Data were collected from 281 adults attending a cardiovascular outpatient clinic in Korea. Health literacy, frailty, and self-care behaviors were measured using Korean-validated instruments. The mean scores of health literacy and self-care behaviors were 8.89 (±3.44) and 31.49 (±5.38), respectively. The prevalence of frailty was around 26.3%. Health literacy was significantly associated with frailty and self-care behaviors. In a hierarchical linear regression analysis, health literacy was a significant determinant of self-care behaviors after adjusting for confounding variables, but frailty was not. Educational level was also a significant predictor of self-care behaviors. Our main findings showed that health literacy can facilitate improvements in HF self-care behaviors. Healthcare professionals should assess patients’ health literacy and educational backgrounds when designing self-management programs. Heart failure is the leading cause of mortality and healthcare costs worldwide . It is aSelf-care is a cornerstone in long-term management of chronic heart failure . A recenKnowing a patient’s level of comprehension of health information is particularly essential because health information allows patients to effectively apply treatment-related information to daily life. In this context, health literacy, which allows patients to assess and translate adequate health information to daily activities, is emerging as a facilitator of self-care behaviors for patients with heart failure ,15. HealFrailty is considered a biological syndrome associated with multisystem deterioration in physiological reserves . It is rTherefore, the relationships among health literacy, frailty, and self-care in patients with heart failure should be clearly identified. This study aimed to investigate whether health literacy and frailty are determinants of self-care behaviors in patients with heart failure after adjusting for the patients’ socio-demographic and clinical factors.This study adopted a cross-sectional research design. A convenience sample of patients with heart failure was recruited from the outpatient clinic of a general hospital in Pyeongtaek, South Korea. Convenience sampling in this study was chosen on the basis of availability, which was easily accessible within the defined study time period. From April to July 2018, a total of 281 patients who visited the outpatient clinic for post-discharge follow-up were recruited for this study.The inclusion criteria were (1) aged 20 years or older; (2) articulate in the Korean language; (3) diagnosed with heart failure by a cardiologist based on clinical history, echocardiographic findings, and the presence of symptoms and diagnoses for at least one year; and (4) willing to participate. The exclusion criteria were (1) previously diagnosed with dementia or major depressive disorder, (2) presence of paralysis or physical impairment, (3) a life expectancy of less than 6 months, and (4) categorized as New York Heart Association (NYHA) class IV, requiring hospice or palliative care.To determine the appropriate sample size for this study, a power analysis was conducted using G-power, version 3.1 . Based oSocio-demographic data  were obtained from standardized questions. The clinical information obtained from medical records included body mass index, duration of heart failure diagnosis, NYHA functional classification, physician-diagnosed comorbidities , prescribed medication, and ejection fraction .Health literacy was assessed using a 3-item set of brief screening questions developed by Chew, Bradley, and Boyko . This scTo assess the status of frailty, we adopted the Korean version of the FRAIL scale (K-FRAIL) . The K-FWe assessed participants’ self-care behaviors with the Korean version of the European Heart Failure Self-Care Behavior Scale 9 items (EHFScBS-9)  which waThis study protocol was reviewed and approved by the Institutional Review Boards of the authors’ institution (SBR-SUR-17-299). All patients were invited to participate in the study through telephone recruitment before visiting the outpatient clinic for follow-ups. A researcher and a trained research assistant confirmed patients as being eligible on the telephone recruitment list. If they were eligible, the researcher explained the study and the steps of agreement in detail. Written informed consent was obtained from patients who agreed to participate in the study at the outpatient clinic visit after fully understanding the study purpose and the rights and responsibilities of participants. The survey took approximately 15 min to complete. The consent information included details about the study aim, the confidentiality and anonymity of patient information, and the voluntary nature of participation.t-test, one-way analysis of variance, and the post hoc test (Scheffe test) were performed to identify differences between variables by characteristics. The correlations among variables were analyzed using Pearson’s correlation coefficient. Hierarchical regression was used to identify the impact of health literacy and frailty on self-care behaviors after adjusting for patient characteristics. All tests were performed at a p = 0.05 statistical significance level.The collected data were analyzed using IBM SPSS Statistics version 23 . Descriptive statistics were used to summarize data for presenting the patients’ characteristics. Continuous data were reported as means with standard deviations. Categorical data were expressed as numbers with percentages. The independent n = 171) were men, and the mean age was 68.7 (±11.1) with 37.4% (n = 108) of participants aged less than 60. Of the 281 patients, only 38.1% (n = 107) were well educated (above high school level), and 20.3% (n = 57) lived alone. The majority were unemployed  and had low monthly incomes   .n = 170), had a relatively short duration after heart failure diagnosis (1–2 years), and had a relatively normal LVEF (≥50%). The mean durations of heart failure and mean LVEF were 3.89 ± 2.89 years and 53.54 ± 11.39%, respectively , men , and those with a higher educational level . Employed individuals  and those with higher monthly incomes  also had higher health literacy levels. In terms of clinical characteristics, the health literacy level was higher for NYHA I than for NYHA II and III . There were no differences in health literacy levels by heart failure duration, comorbidity, LVEF level, or medications except diuretics .In n = 154). The prevalence of frailty was 26.3% (n = 74). Health literacy had a significantly negative correlation with frailty . In contrast, health literacy had a significantly positive correlation with self-care behaviors . Frailty had a significantly negative association with self-care behaviors .For the hierarchical regression analysis, socio-demographic  and clinical variables (NYHA class and the status of taking diuretics) were first entered into Model 1 to determine the predictors of self-care behaviors, and then both health literacy and frailty were entered into Model 2 .β = −0.17, p = 0.017), those with a higher educational level , and those in the lower NYHA class  engaged more in self-care behaviors. In Model 2, health literacy  was a significant determinant of self-care behaviors after adjusting for socio-demographic and clinical variables, but frailty  was not. Educational level  remained significant after adding health literacy and frailty.In Model 1, younger individuals (The importance of self-care in patients with heart failure is well documented ,11. In rIn this study, the mean health literacy score was 8.89, which was below the 11 to 12 score that indicated adequate health literacy . It was The average score of self-care behaviors in our study was 31.49, which was higher than the number reported in a previous review . This stA notable finding of this study was that health literacy had a significant impact on self-care behaviors after adjusting for patient characteristics and frailty. This finding was consistent with findings from previous studies ,15,35. SIn addition, our study found that educational level was also a significant predictor of self-care behaviors. This result was in line with previous findings that educational level was related to health literacy ,15 and sIn this study’s univariate analysis, frailty was significantly associated with self-care behaviors even though frailty was not a significant predictor of self-care behaviors in the hierarchical linear regression. This finding might be related to the samples’ characteristics. Namely, the majority of the patients in our study belonged to NYHA class I with relatively normal EF and were therefore different from the frail heart failure populations of previous studies ,40. FraiThere were several limitations to the present study. First, our study was not designed to evaluate longitudinal associations between health literacy, frailty, and self-care behaviors in patients with heart failure. The severity of frailty in the heart failure population may change over time, especially in elderly patients. Therefore, future studies are needed to determine the optimal screening intervals for these three factors to prevent adverse health outcomes including cognitive decline during the heart failure disease trajectory. Secondly, the study participants were limited to outpatients from a single tertiary care referral center using a convenience sampling; therefore, the results might not generalize to other clinical settings. Third, the present study included a small portion of the NYHA functional classes III and IV. The higher NYHA classes were positively associated with cognitive dysfunction in patients with heart failure. Finally, health literacy, frailty, and self-care behaviors in this study were self-reported measures which could lead to bias and overestimation of these variables.The most critical components of managing heart failure are to plan optimal self-care interventions that consider the patients’ characteristics, share the plan with patients, and ultimately support them in performing adequate self-care. Our findings highlighted that high or adequate health literacy can lead to better compliance of self-care behaviors in patients with heart failure. Therefore, we recommend assessing health literacy in patients with heart failure using a screening tool. The purposes of the assessment are the monitoring of the level of health literacy and the identification of data for empirical and individual intervention. Through this, healthcare providers should be aware of the importance of early assessment for health literacy before patient education. Finally, offering tailored education based on the level of health literacy and educational background may help patients with heart failure improve self-care behaviors.