A mixed-methods study on the pharmacological management of pain in Australian and Japanese nursing homes.

BACKGROUND: Understanding how analgesics are used in different countries can inform initiatives to improve the pharmacological management of pain in nursing homes. AIMS: To compare patterns of analgesic use among Australian and Japanese nursing home residents; and explore Australian and Japanese healthcare professionals' perspectives on analgesic use. METHODS: Part one involved a cross-sectional comparison among residents from 12 nursing homes in South Australia (N = 550) in 2019 and four nursing homes in Tokyo (N = 333) in 2020. Part two involved three focus groups with Australian and Japanese healthcare professionals (N = 16) in 2023. Qualitative data were deductively content analysed using the World Health Organization six-step Guide to Good Prescribing. RESULTS: Australian and Japanese residents were similar in age (median: 89 vs 87) and sex (female: 73% vs 73%). Overall, 74% of Australian and 11% of Japanese residents used regular oral acetaminophen, non-steroidal anti-inflammatory drugs or opioids. Australian and Japanese healthcare professionals described individualising pain management and the first-line use of acetaminophen. Australian participants described their therapeutic goal was to alleviate pain and reported analgesics were often prescribed on a regular basis. Japanese participants described their therapeutic goal was to minimise impacts of pain on daily activities and reported analgesics were often prescribed for short-term durations, corresponding to episodes of pain. Japanese participants described regulations that limit opioid use for non-cancer pain in nursing homes. CONCLUSION: Analgesic use is more prevalent in Australian than Japanese nursing homes. Differences in therapeutic goals, culture, analgesic regulations and treatment durations may contribute to this apparent difference.

Deprescribing Opportunities for Frail Residents of Nursing Homes: A Multicenter Study in Australia, China, Japan, and Spain.

OBJECTIVE: Deprescribing opportunities may differ across health care systems, nursing home settings, and prescribing cultures. The objective of this study was to compare the prevalence of STOPPFrail medications according to frailty status among residents of nursing homes in Australia, China, Japan, and Spain. DESIGN: Secondary cross-sectional analyses of data from 4 cohort studies. SETTING AND PARTICIPANTS: A total of 1142 residents in 31 nursing homes. METHODS: Medication data were extracted from resident records. Frailty was assessed using the FRAIL-NH scale (non-frail 0-2; frail 3-6; most-frail 7-14). Chi-square tests and prevalence ratios (PRs) were used to compare STOPPFrail medication use across cohorts. RESULTS: In total, 84.7% of non-frail, 95.6% of frail, and 90.6% of most-frail residents received ≥1 STOPPFrail medication. Overall, the most prevalent STOPPFrail medications were antihypertensives (53.0% in China to 73.3% in Australia, P < .001), vitamin D (nil in China to 52.7% in Australia, P < .001), lipid-lowering therapies (11.1% in Japan to 38.9% in Australia, P < .001), aspirin (13.5% in Japan to 26.2% in China, P < .001), proton pump inhibitors (2.1% in Japan to 32.0% in Australia, P < .001), and antidiabetic medications (12.3% in Japan to 23.5% in China, P = .010). Overall use of antihypertensives (PR, 1.15; 95% CI, 1.06-1.25), lipid-lowering therapies (PR, 1.78; 95% CI, 1.45-2.18), aspirin (PR, 1.31; 95% CI, 1.04-1.64), and antidiabetic medications (PR, 1.31; 95% CI, 1.00-1.72) were more prevalent among non-frail and frail residents compared with most-frail residents. Antihypertensive use was more prevalent with increasing frailty in China and Japan, but less prevalent with increasing frailty in Australia. Antidiabetic medication use was less prevalent with increasing frailty in China and Spain but was consistent across frailty groups in Australia and Japan. CONCLUSIONS AND IMPLICATIONS: There were overall and frailty-specific variations in prevalence of different STOPPFrail medications across cohorts. This may reflect differences in prescribing cultures, application of clinical practice guidelines in the nursing home setting, and clinician or resident attitudes toward deprescribing.

Symptomatic and preventive medication use according to age and frailty in Australian and Japanese nursing homes.

OBJECTIVE: To investigate symptomatic and preventive medication use according to age and frailty in Australian and Japanese nursing homes (NHs). METHODS: Secondary cross-sectional analyses of two prospective cohort studies involving 12 Australian NHs and four Japanese NHs. Frailty was measured using the FRAIL-NH scale (non-frail 0-2; frail 3-6; most-frail 7-14). Regular medications were classified as symptomatic or preventive based on published lists and expert consensus. Descriptive statistics were used to compare the prevalence and ratio of symptomatic to preventive medications. RESULTS: Overall, 550 Australian residents (87.7 ± 7.3 years; 73.3% females) and 333 Japanese residents (86.5 ± 7.0 years; 73.3% females) were included. Australian residents used a higher mean number of medications than Japanese residents (9.8 ± 4.0 vs 7.7 ± 3.7, p < 0.0001). Australian residents used more preventive than symptomatic medications (5.5 ± 2.5 vs 4.3 ± 2.6, p < 0.0001), while Japanese residents used more symptomatic than preventive medications (4.7 ± 2.6 vs 3.0 ± 2.2, p < 0.0001). In Australia, symptomatic medications were more prevalent with increasing frailty (non-frail 3.4 ± 2.6; frail 4.0 ± 2.6; most-frail 4.8 ± 2.6, p < 0.0001) but less prevalent with age (< 80 years 5.0 ± 2.9; 80-89 years 4.4 ± 2.6; ≥ 90 years 3.9 ± 2.5, p = 0.0042); while preventive medications remained similar across age and frailty groups. In Japan, there was no significant difference in the mean number of symptomatic and preventive medications irrespective of age and frailty. CONCLUSIONS: The ratio of symptomatic to preventive medications was higher with increasing frailty but lower with age in Australia; whereas in Japan, the ratio remained consistent across age and frailty groups. Preventive medications remained prevalent in most-frail residents in both cohorts, albeit at lower levels in Japan.

Changes in Polypharmacy and Potentially Inappropriate Medications in Homebound Older Adults in Japan, 2015-2019: a Nationwide Study.

BACKGROUND: With rising worldwide population aging, the number of homebound individuals with multimorbidity is increasing. Improvement in the quality of home medical care (HMC), including medications, contributes to meeting older adults' preference for "aging in place" and securing healthcare resources. OBJECTIVE: To evaluate the changes in drug prescriptions, particularly potentially inappropriate medications (PIMs), among older adults receiving HMC in recent years, during which measures addressing inappropriate polypharmacy were implemented, including the introduction of clinical practice guidelines and medical fees for deprescribing. DESIGN: A cross-sectional study. PARTICIPANTS: Using data from the national claims database in Japan, this study included older adults aged ≥ 75 years who received HMC in October 2015 (N = 499,850) and October 2019 (N = 657,051). MAIN MEASURES: Number of drugs, prevalence of polypharmacy (≥ 5 regular drugs), major drug categories/classes, and PIMs according to Japanese guidelines were analyzed. Random effects logistic regression models were used to evaluate the differences in medications between 2015 and 2019, considering the correlation within individuals who contributed to the analysis in both years. KEY RESULTS: The number of drugs remained unchanged from 2015 to 2019 (median: 6; interquartile range: 4, 9). The prevalence of polypharmacy also remained unchanged at 70.0% in both years (P = 0.93). However, the prescription of some drugs (e.g., direct oral anticoagulants, new types of hypnotics, acetaminophen, proton pump inhibitors, and ß-blockers) increased, whereas others (e.g., warfarin, vasodilators, H2 blockers, acetylcholinesterase inhibitors, and benzodiazepines) decreased. Among the frequently prescribed PIMs, benzodiazepines/Z-drugs (25.6% in 2015 to 21.1% in 2019; adjusted odds ratio: 0.52) and H2 blockers (11.2 to 7.3%; 0.45) decreased, whereas diuretics (23.8 to 23.6%; 0.90) and antipsychotics (9.7 to 10.5%; 1.11) remained unchanged. CONCLUSIONS: We observed some favorable changes but identified some continuous and new challenges. This study suggests that continued attention to medication optimization is required to achieve safe and effective HMC.

Dispensing by Family Pharmacists as a Potential Factor in Pharmacist-initiated Prescription Change: A Retrospective Observational Study.

BACKGROUND: No studies in Japan have examined whether dispensing by family pharmacists, who are incentivized by reimbursement to provide continuous and exclusive medication management, results in prescription changes. Our primary objective was to identify the variables affecting prescription changes, particularly to investigate dispensing by family pharmacists as a possible factor. METHODS: We identified 333,503 records of pharmacy claims data from patients aged 65 years or older who received medication instructions at outpatient pharmacies at Tsukuba, a medium-sized city near Tokyo, between April 2018 and March 2019. We extracted data on dispensing by family pharmacists, number of medicines, patient sex, patient age, and pharmacy category. A multilevel modified Poisson regression analysis was performed to analyze the correlation between dispensing by family pharmacists and pharmacist-initiated prescription change. RESULTS: Dispensing by family pharmacists was 1.37 times more likely to involve a record of prescription change than dispensing by non-family pharmacists. Older age, female sex, polypharmacy, and small-scale pharmacies were also found to be factors. CONCLUSION: This study indicated that dispensing by family pharmacists was a potential factor for pharmacist-initiated prescription changes that may prevent excessive medication and limit pharmacological interactions. Since the likelihood of inappropriate prescriptions being issued varies from hospital to hospital, subsequent studies should take into account the quality of each institution.

Drug prescribing changes in the last year of life among homebound older adults: national retrospective cohort study.

OBJECTIVES: This study aimed to evaluate the prescription patterns of drugs during the last year of life in homebound older adults who received home medical care. METHODS: We used a nationwide claims database in Japan and selected older adults aged ≥75 years who received home medical care services from ≥12 months before their death. We evaluated medications prescribed 12 months before death (month 12), 3 months before death (month 3) and in the last month of life (month 1). We explored the factors associated with the decreased number of cardiovascular preventive drugs from month 12 to both month 3 and month 1. RESULTS: A total of 118 661 participants were included, and the majority were aged ≥90 years and women. The prevalence of cardiovascular preventive drugs decreased but remained common in month 1, which included antihypertensives (34.7%), antiplatelets (15.9%), oral anticoagulants (7.6%), antidiabetic drugs (7.3%) and lipid-lowering drugs (6.1%). The relative decrease from month 12 to month 1 was the largest for lipid-lowering drugs (44.8%) and the smallest for oral anticoagulants (13.6%). Among other drugs, laxatives (enema), antiemetics, oral corticosteroids, analgesics, expectorants, bronchodilators and antibiotics showed a large relative increase. Older age, duration of home medical care services for <1 year and diagnoses of cancer, dementia and Parkinson's disease were associated with a greater likelihood of a decreased number of cardiovascular preventive drugs. CONCLUSIONS: There is room for deprescribing to avoid inappropriate polypharmacy by balancing preventive and symptom management drugs in those receiving home medical care with a limited life expectancy.

Development and validation of a risk score to predict the frequent emergency house calls among older people who receive regular home visits.

BACKGROUND: The demand for home healthcare is increasing in Japan, and a 24-hour on-call system could be a burden for primary care physicians. Identifying high-risk patients who need frequent emergency house calls could help physicians prepare and allocate medical resources. The aim of the present study was to develop a risk score to predict the frequent emergency house calls in patients who receive regular home visits. METHODS: We conducted a retrospective cohort study with linked medical and long-term care claims data from two Japanese cities. Participants were ≥ 65 years of age and had newly started regular home visits between July 2014 and March 2018 in Tsukuba city and between July 2012 and March 2017 in Kashiwa city. We followed up with patients a year after they began the regular home visits or until the month following the end of the regular home visits if this was completed within 1 year. We calculated the average number of emergency house calls per month by dividing the total number of emergency house calls by the number of months that each person received regular home visits (1-13 months). The primary outcome was the "frequent" emergency house calls, defined as its use once per month or more, on average, during the observation period. We used the least absolute shrinkage and selection operator (LASSO) logistic regression with 10-fold cross-validation to build the model from 19 candidate variables. The predictive performance was assessed with the area under the curve (AUC). RESULTS: Among 4888 eligible patients, frequent emergency house calls were observed in 13.0% of participants (634/4888). The risk score included three variables with the following point assignments: home oxygen therapy (3 points); long-term care need level 4-5 (1 point); cancer (4 points). While the AUC of a model that included all candidate variables was 0.734, the AUC of the 3-risk score model was 0.707, suggesting good discrimination. CONCLUSIONS: This easy-to-use risk score would be useful for assessing high-risk patients and would allow the burden on primary care physicians to be reduced through measures such as clustering high-risk patients in well-equipped medical facilities.

Postoperative Pain Treatment in Patients with Dementia: A Retrospective Observational Study.

BACKGROUND: Inadequate postoperative analgesia has been noted in patients with dementia, but this topic has only been studied in hip fractures. This study aimed to examine whether the duration of postoperative analgesia associated with three surgical procedures was shorter in patients with dementia than in those without dementia. METHODS: This retrospective observational study was based on a nationwide discharge database of acute care hospitals in Japan and included patients aged ≥ 65 years diagnosed with lung cancer, rectal cancer, or hip fracture, who underwent surgery in 366 hospitals between April 2013 and September 2018. The primary outcome was the incidence of injectable analgesic use during the postoperative hospital stay. The number of person-days of opioid, non-steroidal anti-inflammatory drug, or acetaminophen use was calculated for patients with and without dementia after each surgery using generalized estimating equations to obtain the age-adjusted incidence per 100 person-days. RESULTS: Among the 32,379 patients included, 4828 (14.9%) had dementia. The duration of opioid administration per 100 person-days was 14% shorter in patients with dementia than in those without dementia after open rectal cancer surgery (incidence rate ratio [IRR] 0.86; 95% confidence interval [CI] 0.74-1.00) and 20% shorter in patients with dementia after open lung cancer surgery (IRR 0.80; 95% CI 0.70-0.91). In patients who underwent thoracoscopic lung cancer surgery, laparoscopic rectal cancer surgery, or hip replacement surgery, the duration of opioid administration was shorter in patients with dementia than in those without dementia, but the difference was not statistically significant. CONCLUSIONS: The length of postoperative opioid administration after lung and rectal cancer surgery was reduced in patients with dementia, suggesting that the duration of postoperative analgesia in patients with dementia may be insufficient. More careful pain assessment and management of postoperative analgesia in these patients may be necessary.

National trends in the proportion of in-hospital deaths by cause of death among older adults with long-term care: a nationwide observational study in Japan from 2007 to 2017.

BACKGROUND: Japan has promoted end-of-life care at home and in long-term care facilities, and the total proportion of in-hospital deaths has decreased recently. However, the difference in trends of in-hospital deaths by the cause of death remains unclear. We investigated the variation in trends of in-hospital deaths among older adults with long-term care from 2007 to 2017, by cause of death and place of care. METHODS: Using the national long-term care insurance registry, long-term care claims data, and national death records, we identified people aged 65 years or older who died between 2007 and 2017 and used long-term care services in the month before death. Using a joinpoint regression model, we evaluated time trends of the proportion of in-hospital deaths by cause of death (cancer, heart diseases, cerebrovascular diseases, pneumonia, and senility) and place of care (home, long-term care health facility, or long-term care welfare facility). RESULTS: Of the 3,261,839 participants, the mean age was 87.0 ± 8.0 years, and 59.2% were female. Overall, the proportion of in-hospital deaths decreased from 66.2% in 2007 to 55.3% in 2017. By cause of death, the proportion of in-hospital deaths remained the highest for pneumonia (81.6% in 2007 and 77.2% in 2017) and lowest for senility (25.5% in 2007 and 20.0% in 2017) in all types of places of care. The joinpoint regression analysis showed the steepest decline among those who died of senility, especially among long-term care health facility residents. CONCLUSIONS: The findings of this nationwide study suggest that there was a decreasing trend of in-hospital deaths among older adults, although the speed of decline and absolute values varied widely depending on the cause of death and place of care.

Dissociation of the AhR/ARNT complex by TGF-ß/Smad signaling represses CYP1A1 gene expression and inhibits benze[a]pyrene-mediated cytotoxicity.

Cytochrome P450 1A1 (CYP1A1) catalyzes the metabolic activation of polycyclic aromatic hydrocarbons (PAHs) such as benzo[a]pyrene (B[a]P) and is transcriptionally regulated by the aryl hydrocarbon receptor (AhR)/AhR nuclear translocator (ARNT) complex upon exposure to PAHs. Accordingly, inhibition of CYP1A1 expression reduces production of carcinogens from PAHs. Although transcription of the CYP1A1 gene is known to be repressed by transforming growth factor-ß (TGF-ß), how TGF-ß signaling is involved in the suppression of CYP1A1 gene expression has yet to be clarified. In this study, using mammalian cell lines, along with shRNA-mediated gene silencing, CRISPR/Cas9-based genome editing, and reporter gene and quantitative RT-PCR assays, we found that TGF-ß signaling dissociates the B[a]P-mediated AhR/ARNT heteromeric complex. Among the examined Smads, Smad family member 3 (Smad3) strongly interacted with both AhR and ARNT via its MH2 domain. Moreover, hypoxia-inducible factor 1α (HIF-1α), which is stabilized upon TGF-ß stimulation, also inhibited AhR/ARNT complex formation in the presence of B[a]P. Thus, TGF-ß signaling negatively regulated the transcription of the CYP1A1 gene in at least two different ways. Of note, TGF-ß abrogated DNA damage in B[a]P-exposed cells. We therefore conclude that TGF-ß may protect cells against carcinogenesis because it inhibits CYP1A1-mediated metabolic activation of PAHs as part of its anti-tumorigenic activities.

Intentional or unintentional drug poisoning in elderly people: retrospective observational study in a tertiary care hospital in Japan.

AIM: Intentional or unintentional acute drug poisoning occurs even in elderly people, but little is known about the factors influencing the intention to poisoning. A retrospective study was undertaken to describe the characteristics of acute drug poisoning in elderly people according to whether the poisoning was intentional or unintentional and the responsible agents. METHODS: The study was carried out in a single tertiary hospital in Japan. A total of 145 patients aged ≥65 years who were transferred by an ambulance service and were diagnosed with acute drug poisoning were included. Medical records were used to collect information on the intention behind poisoning and the responsible agents. Patients were divided into two groups according to whether they experienced intentional or unintentional poisoning and were further classified according to the responsible agent. Multivariable logistic regression models were used to estimate the association between hospitalization for acute drug poisoning and the use of benzodiazepine receptor agonists (BzRAs). RESULTS: Poisoning was unintentional in 102 (70.3%) patients and intentional in 43 (29.7%) patients. In total, 65 (44.8%) patients required hospitalization. Among patients in the unintentional poisoning group, those using non-BzRAs were more likely to be hospitalized than those using BzRAs (odds ratio, 6.64; 95% confidence interval, 2.56-17.22). The length of hospital stay was significantly longer in the unintentional poisoning group than in the intentional poisoning group (13.9 vs. 6.2 days; P = 0.013). CONCLUSIONS: The proportion of unintentional poisoning in the elderly is high, and particularly with respect to poisoning with non-BzRAs, the hospitalization rates are high.

PDZK1-interacting protein 1 (PDZK1IP1) traps Smad4 protein and suppresses transforming growth factor-ß (TGF-ß) signaling.

Transforming growth factor (TGF)-ß signaling in humans is stringently regulated to prevent excessive TGF-ß signaling. In tumors, TGF-ß signaling can both negatively and positively regulate tumorigenesis dependent on tumor type, but the reason for these opposite effects is unclear. TGF-ß signaling is mainly mediated via the Smad-dependent pathway, and herein we found that PDZK1-interacting protein 1 (PDZK1IP1) interacts with Smad4. PDZK1IP1 inhibited both the TGF-ß and the bone morphogenetic protein (BMP) pathways without affecting receptor-regulated Smad (R-Smad) phosphorylation. Rather than targeting R-Smad phosphorylation, PDZK1IP1 could interfere with TGF-ß- and BMP-induced R-Smad/Smad4 complex formation. Of note, PDZK1IP1 retained Smad4 in the cytoplasm of TGF-ß-stimulated cells. To pinpoint PDZK1IP1's functional domain, we created several PDZK1IP1 variants and found that its middle region, from Phe40 to Ala49, plays a key role in its Smad4-regulating activity. PDZK1IP1 knockdown enhanced the expression of the TGF-ß target genes Smad7 and prostate transmembrane protein androgen-induced (TMEPAI) upon TGF-ß stimulation. In contrast, PDZK1IP1 overexpression suppressed TGF-ß-induced reporter activities, cell migration, and cell growth inhibition. In a xenograft tumor model in which TGF-ß was previously shown to elicit tumor-promoting effects, PDZK1IP1 gain of function decreased tumor size and increased survival rates. Taken together, these findings indicate that PDZK1IP1 interacts with Smad4 and thereby suppresses the TGF-ß signaling pathway.

Dementia and Risk of 30-Day Readmission in Older Adults After Discharge from Acute Care Hospitals.

OBJECTIVES: To assess the association between dementia and risk of hospital readmission and to evaluate whether the effect of dementia on hospital readmission varies according to primary diagnosis. DESIGN: Retrospective cohort study. SETTING: Nationwide discharge database of acute care hospitals in Japan. PARTICIPANTS: Individuals aged 65 and older diagnosed with one of the 30 most common diagnoses and discharged from 987 hospitals between April 2014 and September 2015 (N = 1,834,378). MEASUREMENTS: The primary outcome was unplanned hospital readmission within 30 days. Poisson generalized estimating equation models were fitted to assess the risks of readmission for individuals with and without dementia, using primary diagnosis as a possible effect modifier and clinical factors as potential confounders. RESULTS: The overall prevalence of dementia was 14.7% and varied according to primary diagnosis, ranging from 3.0% in individuals with prostate cancer to 69.4% in those with aspiration pneumonia. Overall, individuals with dementia had a higher risk of hospital readmission (8.3%) than those without (4.1%) (adjusted risk ratio (aRR])=1.46, 95% confidence interval (CI)=1.44-1.49), although diagnostic category substantially modified the relationship between dementia and hospital readmission. For hip fracture, dementia was associated with greater risk of hospital readmission (adjusted risk 11.5% vs 7.9%; aRR=1.46; 95% CI=1.28-1.68); this risk was attenuated for cholecystitis (adjusted risk 12.8% vs 12.4%; aRR=1.03; 95% CI=0.90-1.18). CONCLUSION: Risk of hospital readmission associated with dementia varied according to primary diagnosis. Healthcare providers could enforce interventions to minimize readmission by focusing on comorbid conditions in individuals with dementia and specific primary diagnoses that increase their risk of readmission.

TMED10 Protein Interferes with Transforming Growth Factor (TGF)-ß Signaling by Disrupting TGF-ß Receptor Complex Formation.

The intensity and duration of TGF-ß signaling determine the cellular biological response. How this is negatively regulated is not well understood. Here, we identified a novel negative regulator of TGF-ß signaling, transmembrane p24-trafficking protein 10 (TMED10). TMED10 disrupts the complex formation between TGF-ß type I (also termed ALK5) and type II receptors (TßRII). Misexpression studies revealed that TMED10 attenuated TGF-ß-mediated signaling. A 20-amino acid-long region from Thr91 to Glu110 within the extracellular region of TMED10 was found to be crucial for TMED10 interaction with both ALK5 and TßRII. Synthetic peptides corresponding to this region inhibit both TGF-ß-induced Smad2 phosphorylation and Smad-dependent transcriptional reporter activity. In a xenograft cancer model, where previously TGF-ß was shown to elicit tumor-promoting effects, gain-of-function and loss-of-function studies for TMED10 revealed a decrease and increase in the tumor size, respectively. Thus, we determined herein that TMED10 expression levels are the key determinant for efficiency of TGF-ß receptor complex formation and signaling.

A diagnostic marker for superficial urothelial bladder carcinoma: lack of nuclear ATBF1 (ZFHX3) by immunohistochemistry suggests malignant progression.

BACKGROUND: Pathological stage and grade have limited ability to predict the outcomes of superficial urothelial bladder carcinoma at initial transurethral resection (TUR). AT-motif binding factor 1 (ATBF1) is a tumor suppressive transcription factor that is normally localized to the nucleus but has been detected in the cytoplasm in several cancers. Here, we examined the diagnostic value of the intracellular localization of ATBF1 as a marker for the identification of high risk urothelial bladder carcinoma. METHODS: Seven anti-ATBF1 antibodies were generated to cover the entire ATBF1 sequence. Four human influenza hemagglutinin-derived amino acid sequence-tagged expression vectors with truncated ATBF1 cDNA were constructed to map the functional domains of nuclear localization signals (NLSs) with the consensus sequence KR[X10-12]K. A total of 117 samples from initial TUR of human bladder carcinomas were analyzed. None of the patients had received chemotherapy or radiotherapy before pathological evaluation. RESULTS: ATBF1 nuclear localization was regulated synergistically by three NLSs on ATBF1. The cytoplasmic fragments of ATBF1 lacked NLSs. Patients were divided into two groups according to positive nuclear staining of ATBF1, and significant differences in overall survival (P = 0.021) and intravesical recurrence-free survival (P = 0.013) were detected between ATBF1+ (n = 110) and ATBF1- (n = 7) cases. Multivariate analysis revealed that ATBF1 staining was an independent prognostic factor for intravesical recurrence-free survival after adjusting for cellular grading and pathological staging (P = 0.008). CONCLUSIONS: Cleavage of ATBF1 leads to the cytoplasmic localization of ATBF1 fragments and downregulates nuclear ATBF1. Alterations in the subcellular localization of ATBF1 due to fragmentation of the protein are related to the malignant character of urothelial carcinoma. Pathological evaluation using anti-ATBF1 antibodies enabled the identification of highly malignant cases that had been overlooked at initial TUR. Nuclear localization of ATBF1 indicates better prognosis of urothelial carcinoma.

TGF- ß Signaling Cooperates with AT Motif-Binding Factor-1 for Repression of the α -Fetoprotein Promoter.

α-Fetoprotein (AFP) is known to be highly produced in fetal liver despite its barely detectable level in normal adult liver. On the other hand, hepatocellular carcinoma often shows high expression of AFP. Thus, AFP seems to be an oncogenic marker. In our present study, we investigated how TGF-ß signaling cooperates with AT motif-binding factor-1 (ATBF1) to inhibit AFP transcription. Indeed, the expression of AFP mRNA in HuH-7 cells was negatively regulated by TGF-ß signaling. To further understand how TGF-ß suppresses the transcription of the AFP gene, we analyzed the activity of the AFP promoter in the presence of TGF-ß. We found that the TGF-ß signaling and ATBF1 suppressed AFP transcription through two ATBF1 binding elements (AT-motifs). Using a heterologous reporter system, both AT-motifs were required for transcriptional repression upon TGF-ß stimulation. Furthermore, Smads were found to interact with ATBF1 at both its N-terminal and C-terminal regions. Since the N-terminal (ATBF1N) and C-terminal regions of ATBF1 (ATBF1C) lack the ability of DNA binding, both truncated mutants rescued the cooperative inhibitory action by the TGF-ß signaling and ATBF1 in a dose-dependent manner. Taken together, these findings indicate that TGF-ß signaling can act in concert with ATBF1 to suppress the activity of the AFP promoter through direct interaction of ATBF1 with Smads.

C18 ORF1, a novel negative regulator of transforming growth factor-ß signaling.

Transforming growth factor (TGF)-ß signaling is deliberately regulated at multiple steps in its pathway from the extracellular microenvironment to the nucleus. However, how TGF-ß signaling is activated or attenuated is not fully understood. We recently identified transmembrane prostate androgen-induced RNA (TMEPAI), which is involved in a negative feedback loop of TGF-ß signaling. When we searched for a family molecule(s) for TMEPAI, we found C18ORF1, which, like TMEPAI, possesses two PY motifs and one Smad-interacting motif (SIM) domain. As expected, C18ORF1 could block TGF-ß signaling but not bone morphogenetic protein signaling. C18ORF1 bound to Smad2/3 via its SIM and competed with the Smad anchor for receptor activation for Smad2/3 binding to attenuate recruitment of Smad2/3 to the TGF-ß type I receptor (also termed activin receptor-like kinase 5 (ALK5)), in a similar fashion to TMEPAI. Knockdown of C18ORF1 prolonged duration of TGF-ß-induced Smad2 phosphorylation and concomitantly potentiated the expression of JunB, p21, and TMEPAI mRNAs induced by TGF-ß. Consistently, TGF-ß-induced cell migration was enhanced by the knockdown of C18ORF1. These results indicate that the inhibitory function of C18ORF1 on TGF-ß signaling is similar to that of TMEPAI. However, in contrast to TMEPAI, C18ORF1 was not induced upon TGF-ß signaling. Thus, we defined C18ORF1 as a surveillant of steady state TGF-ß signaling, whereas TMEPAI might help C18ORF1 to inhibit TGF-ß signaling in a coordinated manner when cells are stimulated with high levels of TGF-ß.

ZFHX4 interacts with the NuRD core member CHD4 and regulates the glioblastoma tumor-initiating cell state.

Glioblastoma (GBM) harbors subpopulations of therapy-resistant tumor-initiating cells (TICs) that are self-renewing and multipotent. To understand the regulation of the TIC state, we performed an image-based screen for genes regulating GBM TIC maintenance and identified ZFHX4, a 397 kDa transcription factor. ZFHX4 is required to maintain TIC-associated and normal human neural precursor cell phenotypes in vitro, suggesting that ZFHX4 regulates differentiation, and its suppression increases glioma-free survival in intracranial xenografts. ZFHX4 interacts with CHD4, a core member of the nucleosome remodeling and deacetylase (NuRD) complex. ZFHX4 and CHD4 bind to overlapping sets of genomic loci and control similar gene expression programs. Using expression data derived from GBM patients, we found that ZFHX4 significantly affects CHD4-mediated gene expression perturbations, which defines ZFHX4 as a master regulator of CHD4. These observations define ZFHX4 as a regulatory factor that links the chromatin-remodeling NuRD complex and the GBM TIC state.

A homeodomain-zinc finger protein, ZFHX4, is expressed in neuronal differentiation manner and suppressed in muscle differentiation manner.

Human ZFHX4 has recently been shown to be a candidate gene for congenital bilateral isolated ptosis. Here, we report molecular cloning of the human ZFHX4 cDNA and genomic organization of this gene. Human ZFHX4 is about 180 kb long, containing 12 exons that encodes a 3599-amino acid protein carrying four homeodomains and 22 zinc fingers. The 11th exon is 3.2 kb in length and encodes all the four homeodomains together with four of the 22 zinc fingers. ZFHX4 is 90% homologous to mouse Zfhx4, 52% to human ATBF1A and 24% to Drosophila ZFH-2. ZFHX4 was mapped to human chromosome 8q13.3-q21.11 by fluorescence in situ hybridization using BAC clone RP11-48D4 as a probe. RT-PCR analysis showed that ZFHX4 transcripts were expressed in adult human brain, liver and muscle. This, together with the finding that Zfhx4 was expressed transiently in differentiating P19 embryonal carcinoma cells and C2C12 myoblasts, suggests that ZFHX4/Zfhx4 is involved in neural and muscle differentiation.

ATBF1 enhances the suppression of STAT3 signaling by interaction with PIAS3.

ATBF1 was first discovered as a suppressor of AFP expression in hepatocytes. It is present in brain, adult liver, lung, and gastro-intestinal tract. Recently, it has been reported that ATBF1 regulates myoblastic differentiation and interacts with v-Myb in regulation of its transactivation. Using the yeast two-hybrid system, we searched for protein-protein interactions to uncover new functions for ATBF1. We present here experimental evidence that ATBF1 is a new regulatory factor for STAT3-mediated signal transduction through its interaction with PIAS3. PIAS3 was thus identified as an ATBF1-binding protein. In co-transfection experiments, the full-length ATBF1 was found to form complexes with PIAS3 in Hep G2 cells. In the luciferase assay, ATBF1 was found to have no influence on STAT3 signaling induced by IL-6 stimulation, but it did synergistically enhance PIAS3 inhibition of activated STAT3. In conclusion, ATBF1 can suppress the IL-6-mediated cellular response by acting together with PIAS3.