An Open-Label Phase I Study of Metformin and Nelfinavir in Combination With Bortezomib in Patients With Relapsed and Refractory Multiple Myeloma.

BACKGROUND: In preclinical models, combining a GLUT4 inhibitor with an oxidative phosphorylation inhibitor shows synergistic therapeutic potential against multiple myeloma (MM). Thus, this study evaluated the safety and tolerability of repurposing metformin, a complex I inhibitor, and nelfinavir, a GLUT4 inhibitor, in combination with bortezomib for the treatment of relapsed/refractory MM that had progressed on all standard of care therapies. MATERIALS AND METHODS: This trial utilized a 3 + 3 dose escalation design with 3 dose levels planned for up to a maximum of 6 (21-day) cycles. Metformin and nelfinavir were administered for 14 of 21 days, and subQ bortezomib was administered to a portion of patients on days 1, 8, and 15. The primary objective was to determine the maximal tolerated dose, and the secondary objective was to evaluate the safety and overall response rate (ORR) of this combination. RESULTS: Nine patients were accrued with a median age of 65 (range: 42-81) and received a median of 7 prior lines of therapy (Range: 5-12). The first 3 patients received only metformin (500 mg BID) and nelfinavir (1250 mg BID) at the first dose level, with 1 patient experiencing an unconfirmed minimal response (MR) in the first cycle, 1 experiencing progressive disease after 1 cycle of treatment and 1 patient going off treatment prior to assessing response but with signs of progressive disease. Given the limited therapeutic activity, the upfront addition of bortezomib (1.3 mg/m2) was utilized for the subsequent 6 patients accrued. Three of these 6 patients went off study due to progressive disease, 1 patient achieved an unconfirmed partial response after 1 cycle of treatment but reported progressive disease in the subsequent cycle, 1 patient went off study to enter hospice, and the remaining patient experienced stable disease (SD) after receiving 6 cycles of clinical trial treatment. The study was closed before accrual to the next dose level was started. CONCLUSION: This is the first study to evaluate the safety and efficacy of this repurposed drug combination in this very difficult-to-treat population of relapsed and refractory MM. This was an overall negative study with no ORR observed. Fortunately, 1 patient experienced an SD response, allowing this combination to stabilize their disease until another novel therapy on a clinical trial was available.

PRESTO: A Phase III, Open-Label Study of Intensification of Androgen Blockade in Patients With High-Risk Biochemically Relapsed Castration-Sensitive Prostate Cancer (AFT-19).

PURPOSE: Patients with biochemically recurrent prostate cancer (BRPC) after radical prostatectomy and a short PSA doubling time are at risk for distant metastases. Apalutamide, an androgen receptor antagonist, and abiraterone acetate plus prednisone (AAP) prolong survival in the metastatic setting. We evaluated whether intensification of androgen-deprivation therapy (ADT) improves outcomes in BRPC. PATIENTS AND METHODS: PRESTO is a randomized phase III, open-label trial in patients with BRPC and PSA doubling time ≤9 months (ClinicalTrials.gov identifier: NCT03009981). Patients were randomly assigned 1:1:1 to receive a finite 52-week treatment course with ADT control, ADT + apalutamide, or ADT + apalutamide + AAP. The primary end point was PSA progression-free survival (PSA-PFS), defined as serum PSA >0.2 ng/mL after treatment completion. RESULTS: Five hundred three patients were enrolled. The median PSA was 1.8 ng/mL (IQR, 1.0-3.6). At the first planned interim analysis, both experimental arms significantly prolonged PSA-PFS compared with the control arm (median, 24.9 months for ADT + apalutamide v 20.3 months for ADT; hazard ratio [HR], 0.52 [95% CI, 0.35 to 0.77]; P = .00047; median, 26.0 months for ADT + apalutamide + AAP v 20.0 months for ADT; HR, 0.48 [95% CI, 0.32 to 0.71]; P = .00008). Median time to testosterone recovery did not differ across treatment arms. The most common grade ≥3 adverse event was hypertension (7.5%, 7.4%, and 18% in ADT, ADT + apalutamide, and ADT + apalutamide + AAP arms, respectively). CONCLUSION: Intensified AR blockade for a finite duration prolongs PSA-PFS with a manageable safety profile, without adversely affecting time to testosterone recovery. The addition of apalutamide to ADT should be considered in patients with high-risk BRPC.

Comparison of quartz and sapphire optical chambers for infrared laser sealing of vascular tissues using a reciprocating, side-firing optical fiber: Simulations and experiments.

INTRODUCTION: Infrared (IR) lasers are being tested as an alternative to radiofrequency (RF) and ultrasonic (US) surgical devices for hemostatic sealing of vascular tissues. In previous studies, a side-firing optical fiber with elliptical IR beam output was reciprocated, producing a linear IR laser beam pattern for uniform sealing of blood vessels. Technical challenges include limited field-of-view of vessel position within the metallic device jaws, and matching fiber scan length to variable vessel sizes. A transparent jaw may improve visibility and enable custom treatment. METHODS: Quartz and sapphire square optical chambers (2.7 × 2.7 × 25 [mm3 ] outer dimensions) were tested, capable of fitting into a 5-mm-OD laparoscopic device. A 1470 nm laser was used for optical transmission studies. Razor blade scans and an IR beam profiler acquired fiber (550-µm-core/0.22NA) output beam profiles. Thermocouples recorded peak temperatures and cooling times on internal and external chamber surfaces. Optical fibers with angle polished distal tips delivered 94% of light at a 90° angle. Porcine renal arteries with diameters of 3.4 ± 0.7 mm (n = 13) for quartz and 3.2 ± 0.7 mm (n = 14) for sapphire chambers (p > 0.05), were sealed using 30 W for 5 s. RESULTS: Reflection losses at material/air interfaces were 3.3% and 7.4% for quartz and sapphire. Peak temperatures on the external chamber surface averaged 74 ± 8°C and 73 ± 10°C (p > 0.05). Times to cool down to 37°C measured 13 ± 4 s and 27 ± 7 s (p < 0.05). Vessel burst pressures (BP) averaged 883 ± 393 mmHg and 412 ± 330 mmHg (p < 0.05). For quartz, 13/13 (100%) vessels were sealed (BP > 360 mmHg), versus 9/14 (64%) for sapphire. Computer simulations for the quartz chamber yielded peak temperatures (78°C) and cooling times (16 s) similar to experiments. CONCLUSIONS: Quartz is an inexpensive material for use in a laparoscopic device jaw, providing more consistent vessel seals and faster cooling times than sapphire and current RF and US devices.

Combined Bilateral Salpingo-oophorectomy and Cesarean Delivery in BRCA1/2 Alteration Carriers : A Case Series.

BACKGROUND: The cumulative lifetime risk of ovarian cancer is 16-68% and 11-30% in female BRCA1 and BRCA2 gene alteration carriers, respectively. Risk-reducing bilateral salpingo-oophorectomy (RRSO) is the only proven way to reduce ovarian cancer mortality. We report a series of patients who underwent risk-reducing surgery at the time of planned obstetric-indicated cesarean delivery. CASES: This is a case series of four women carrying a pathogenic germline BRCA1 or BRCA2 gene alteration who underwent RRSO at the time of cesarean delivery between March 1, 2018, and March 31, 2022. All women were referred during pregnancy to the University College London Hospitals Familial Cancer Clinic for consideration of RRSO at the time of obstetric-indicated cesarean delivery. Women were considered eligible for RRSO if they had a proven pathogenic germline alteration, would have completed childbearing after the cesarean delivery, and were older than age 35 or 40 years with BRCA1 or BRCA2 alterations, respectively. Operating time, blood loss, transfusion requirements, length of hospital stay, complications, and ability to breastfeed were assessed and, where possible, compared with the institutional means for similar patients who underwent cesarean delivery only, to determine whether RRSO was associated with increased morbidity. Women were contacted 11-59 months postprocedure to assess satisfaction. The mean blood loss was 687 mL (range 400-1,000 mL), mean operating time was 68 minutes, mean length of hospital stay was 3 days, and mean change in hemoglobin was -1 g/dL. No patient required a transfusion, had internal organ damage, returned to the operating room, or was readmitted. One of two women with intact breast tissue successfully breastfed, and the other chose to bottle feed. The mean contemporaneous institutional blood loss for cesarean delivery was not significantly different at 681 mL for singleton pregnancies and 872 mL for twin pregnancies. All four women reported a high level of satisfaction with the combined procedure. CONCLUSION: Our results show that RRSO can be performed at the time of cesarean delivery with high patient satisfaction. This approach can be offered to appropriately counseled individuals, with the benefit of avoiding the need for two separate procedures, with potentially reduced patient morbidity and health care costs.

Management of obesity across women's life course: FIGO Best Practice Advice.

Obesity is a chronic, progressive, relapsing, and treatable multifactorial, neurobehavioral disease. According to the World Health Organization, obesity affects 15% of women and has long-term effects on women's health. The focus of care in patients with obesity should be on optimizing health outcomes rather than on weight loss. Appropriate and common language, considering cultural sensitivity and trauma-informed care, is needed to discuss obesity. Pregnancy is a time of significant physiological change. Pre-, ante-, and postpartum clinical encounters provide opportunities for health optimization for parents with obesity in terms of, but not limited to, fertility and breastfeeding. Pre-existing conditions may also be identified and managed. Beyond pregnancy, women with obesity are at an increased risk for gastrointestinal and liver diseases, impaired kidney function, obstructive sleep apnea, and venous thromboembolism. Gynecological and reproductive health of women living with obesity cannot be dismissed, with accommodations needed for preventive health screenings and consideration of increased risk for gynecologic malignancies. Mental wellness, specifically depression, should be screened and managed appropriately. Obesity is a complex condition and is increasing in prevalence with failure of public health interventions to achieve significant decrease. Future research efforts should focus on interprofessional care and discovering effective interventions for health optimization.

Human DNA ligases I and III have stand-alone end-joining capability, but differ in ligation efficiency and specificity.

Double-strand DNA breaks (DSBs) are toxic to cells, and improper repair can cause chromosomal abnormalities that initiate and drive cancer progression. DNA ligases III and IV (LIG3, LIG4) have long been credited for repair of DSBs in mammals, but recent evidence suggests that DNA ligase I (LIG1) has intrinsic end-joining (EJ) activity that can compensate for their loss. To test this model, we employed in vitro biochemical assays to compare EJ by LIG1 and LIG3. The ligases join blunt-end and 3'-overhang-containing DNA substrates with similar catalytic efficiency, but LIG1 joins 5'-overhang-containing DNA substrates ∼20-fold less efficiently than LIG3 under optimal conditions. LIG1-catalyzed EJ is compromised at a physiological concentration of Mg2+, but its activity is restored by increased molecular crowding. In contrast to LIG1, LIG3 efficiently catalyzes EJ reactions at a physiological concentration of Mg2+ with or without molecular crowding. Under all tested conditions, LIG3 has greater affinity than LIG1 for DNA ends. Remarkably, LIG3 can ligate both strands of a DSB during a single binding encounter. The weaker DNA binding affinity of LIG1 causes significant abortive ligation that is sensitive to molecular crowding and DNA terminal structure. These results provide new insights into mechanisms of alternative nonhomologous EJ.

Synergy between inhibitors of two mitotic spindle assembly motors undermines an adaptive response.

Mitosis is the cellular process that ensures accurate segregation of the cell's genetic material into two daughter cells. Mitosis is often deregulated in cancer; thus drugs that target mitosis-specific proteins represent attractive targets for anticancer therapy. Numerous inhibitors have been developed against kinesin-5 Eg5, a kinesin essential for bipolar spindle assembly. Unfortunately, Eg5 inhibitors (K5Is) have been largely ineffective in the clinic, possibly due to the activity of a second kinesin, KIF15, that can suppress the cytotoxic effect of K5Is by driving spindle assembly through an Eg5-independent pathway. We hypothesized that pairing of K5Is with small molecule inhibitors of KIF15 will be more cytotoxic than either inhibitor alone. Here we present the results of a high-throughput screen from which we identified two inhibitors that inhibit the motor activity of KIF15 both in vitro and in cells. These inhibitors selectively inhibit KIF15 over other molecular motors and differentially affect the ability of KIF15 to bind microtubules. Finally, we find that chemical inhibition of KIF15 reduces the ability of cells to acquire resistance to K5Is, highlighting the centrality of KIF15 to K5I resistance and the value of these inhibitors as tools with which to study KIF15 in a physiological context.

Positional aphasia as a manifestation of syndrome of the trephined: illustrative case.

BACKGROUND: The authors report a case of a 66-year-old male who presented acutely with a subdural hematoma who was managed operatively with craniotomy. His course was complicated by a postoperative epidural hematoma, which, on the basis of intraoperative findings at the second surgery, was managed with evacuation of the hematoma and removal of the bone flap. OBSERVATIONS: The patient's subsequent recovery was remarkable for a reproducible positional aphasia in the early postoperative period with an ultimate diagnosis of syndrome of the trephined. The patient's cerebral edema permitted early autologous cranioplasty, which resulted in resolution of the patient's symptoms. LESSONS: The authors believe this case to be the first described of isolated positional aphasia as a manifestation of syndrome of the trephined. Recognition and treatment of the syndrome resulted in a positive patient outcome.

Phase 2 trial of ixazomib, cyclophosphamide, and dexamethasone for previously untreated light chain amyloidosis.

Bortezomib, a proteasome inhibitor (PI), has shown efficacy in the treatment of newly diagnosed and relapsed light chain (AL) amyloidosis, and is often used in combination with cyclophosphamide and dexamethasone. Ixazomib is the first oral PI to be approved in routine practice but has not yet been evaluated in the upfront treatment setting. Newly diagnosed AL amyloidosis patients with measurable disease and adequate organ function were enrolled. The primary objective was to determine the hematologic response rate of ixazomib in combination with cyclophosphamide and dexamethasone. Treatment was given for 12 cycles, followed by ixazomib maintenance until progression. Thirty-five patients were included; their median age was 67 years, and 69% were male. Major organ involvement included heart (66%) and kidneys (54%). A median of 4 induction cycles (range, 1-12) were administered. The overall hematologic response to induction was 63% and included complete response in 11.4% and very good partial response in 37.1% of patients. One patient was upstaged to complete response during maintenance. The most common reason for going off study was the institution of alternate therapy (61%). With a median follow-up of 29.7 months for the living patients, the 2-year progression-free survival and overall survival were 74% and 78%, respectively. The median time to alternate therapy was 7.5 months. Grade ≥3 hematologic and nonhematologic adverse events occurred in 23% and 49% of patients. Given ixazomib's favorable toxicity profile, which is an important advantage for the typically frail AL population, further evaluation of ixazomib in other combinations in the upfront setting is warranted. This trial was registered at www.clinicaltrials.gov as #NCT01864018.

Recrudescent infection after deep brain stimulator reimplantation.

OBJECTIVE: The efficacy of deep brain stimulation (DBS) in treating the symptoms of movement disorders can be life changing for patients. Thus, the 5%-15% incidence of stimulator-related infection requiring removal of the device can be particularly disheartening. Although DBS system reimplantation is generally successful, this is not always the case. The literature is replete with publications describing the incidence of infection and the associated features. However, the literature is sparse in terms of information on the incidence of recurrent or recrudescent infection after system reimplantation. The goal of this paper was to evaluate factors leading to unsuccessful reimplantation of a DBS system following initial infection. METHODS: Data were reviewed for all DBS procedures performed by one surgeon (K.L.H.) over 19 years including the infectious agent, location of infection, treatment regimen, and subsequent reimplantation of a DBS system and long-term outcome. RESULTS: In this series of 558 patients who had undergone DBS surgery, 37 (6.6%) subsequently developed an infection. Infections with methicillin-sensitive Staphylococcus aureus, Enterobacter species, or coagulase-negative staphylococci were predominant. Four patients had cerebritis, one had meningitis, and the rest had soft tissue infections of the pocket or scalp. All had their entire DBS system explanted, followed by 4-6 weeks of intravenous antibiotics and surveillance for recrudescence for an additional period of at least 30 days. Twenty-five patients subsequently underwent DBS system reimplantation, and the procedure was successful in 22. Three of the 4 patients with cerebritis developed a subsequent wound infection after system reimplantation. None of the other 22 patients developed a recurrence. The odds ratio for developing a recurrent infection after cerebritis was 28.5 (95% CI 1.931-420.5, p = 0.007). CONCLUSIONS: This study, the largest series of DBS system reimplantations following infection, demonstrated that most patients can have successful reimplantations without recurrent infection. However, patients who have had DBS-related cerebritis have a nearly 30-fold increased risk of developing reinfection after reimplantation. Alternative strategies for these patients are discussed.

UBCH5 Family Members Differentially Impact Stabilization of Mutant p53 via RNF128 Iso1 During Barrett's Progression to Esophageal Adenocarcinoma.

BACKGROUND & AIMS: TP53 mutations underlie Barrett's esophagus (BE) progression to dysplasia and cancer. During BE progression, the ubiquitin ligase (E3) RNF128/GRAIL switches expression from isoform 2 (Iso2) to Iso1, stabilizing mutant p53. However, the ubiquitin-conjugating enzyme (E2) that partners with Iso1 to stabilize mutant p53 is unknown. METHODS: Single-cell RNA sequencing of paired normal esophagus and BE tissues identified candidate E2s, further investigated in expression data from BE to esophageal adenocarcinoma (EAC) progression samples. Biochemical and cellular studies helped clarify the role of RNF128-E2 on mutant p53 stability. RESULTS: The UBE2D family member 2D3 (UBCH5C) is the most abundant E2 in normal esophagus. However, during BE to EAC progression, loss of UBE2D3 copy number and reduced expression of RNF128 Iso2 were noted, 2 known p53 degraders. In contrast, expression of UBE2D1 (UBCH5A) and RNF128 Iso1 in dysplastic BE and EAC forms an inactive E2-E3 complex, stabilizing mutant p53. To destabilize mutant p53, we targeted RNF128 Iso1 either by mutating asparagine (N48, 59, and 101) residues to block glycosylation to facilitate ß-TrCP1-mediated degradation or by mutating proline (P54 and 105) residues to restore p53 polyubiquitinating ability. In addition, either loss of UBCH5A catalytic activity, or disruption of the Iso1-UBCH5A interaction promoted Iso1 loss. Consequently, overexpression of either catalytically dead or Iso1-binding-deficient UBCH5A mutants destabilized Iso1 to degrade mutant p53, thus compromising the clonogenic survival of mutant p53-dependent BE cells. CONCLUSIONS: Loss of RNF128 Iso2-UBCH5C and persistence of the Iso1-UBCH5A complex favors mutant p53 stability to promote BE cell survival. Therefore, targeting of Iso1-UBCH5A may provide a novel therapeutic strategy to prevent BE progression.

Association Between Psychiatric Comorbidities and Mortality in Epilepsy.

OBJECTIVE: To explore the impact of psychiatric comorbidities on all-cause mortality in adults with epilepsy from a cohort of patients admitted for video-EEG monitoring (VEM) over 2 decades. METHODS: A retrospective medical record audit was conducted on 2,709 adults admitted for VEM and diagnosed with epilepsy at 3 Victorian comprehensive epilepsy programs from 1995 to 2015. A total of 1,805 patients were identified in whom the record of a clinical evaluation by a neuropsychiatrist was available, excluding 27 patients who died of a malignant brain tumor known at the time of VEM admission. Epilepsy and lifetime psychiatric diagnoses were determined from consensus opinion of epileptologists and neuropsychiatrists involved in the care of each patient. Mortality and cause of death were determined by linkage to the Australian National Death Index and National Coronial Information System. RESULTS: Compared with the general population, mortality was higher in people with epilepsy (PWE) with a psychiatric illness (standardized mortality ratio [SMR] 3.6) and without a psychiatric illness (SMR 2.5). PWE with a psychiatric illness had greater mortality compared with PWE without (hazard ratio 1.41, 95% confidence interval 1.02-1.97) after adjusting for age and sex. No single psychiatric disorder by itself conferred increased mortality in PWE. The distribution of causes of death remained similar between PWE with psychiatric comorbidities and those without. CONCLUSION: The presence of comorbid psychiatric disorders in adults with epilepsy is associated with increased mortality, highlighting the importance of identifying and treating psychiatric comorbidities in these patients.

Neuroleptic malignant syndrome in a patient with moderate intellectual disability treated with olanzapine: A case report.

This case demonstrates the challenges encountered in a case of Neuroleptic Malignant Syndrome in a young woman with moderate Intellectual Disability.

Transient kinetic analysis of oxidative dealkylation by the direct reversal DNA repair enzyme AlkB.

AlkB is a bacterial Fe(II)- and 2-oxoglutarate-dependent dioxygenase that repairs a wide range of alkylated nucleobases in DNA and RNA as part of the adaptive response to exogenous nucleic acid-alkylating agents. Although there has been longstanding interest in the structure and specificity of Escherichia coli AlkB and its homologs, difficulties in assaying their repair activities have limited our understanding of their substrate specificities and kinetic mechanisms. Here, we used quantitative kinetic approaches to determine the transient kinetics of recognition and repair of alkylated DNA by AlkB. These experiments revealed that AlkB is a much faster alkylation repair enzyme than previously reported and that it is significantly faster than DNA repair glycosylases that recognize and excise some of the same base lesions. We observed that whereas 1,N6-ethenoadenine can be repaired by AlkB with similar efficiencies in both single- and double-stranded DNA, 1-methyladenine is preferentially repaired in single-stranded DNA. Our results lay the groundwork for future studies of AlkB and its human homologs ALKBH2 and ALKBH3.

Biallelic mutations in DNA ligase 1 underlie a spectrum of immune deficiencies.

We report the molecular, cellular, and clinical features of 5 patients from 3 kindreds with biallelic mutations in the autosomal LIG1 gene encoding DNA ligase 1. The patients exhibited hypogammaglobulinemia, lymphopenia, increased proportions of circulating γδT cells, and erythrocyte macrocytosis. Clinical severity ranged from a mild antibody deficiency to a combined immunodeficiency requiring hematopoietic stem cell transplantation. Using engineered LIG1-deficient cell lines, we demonstrated chemical and radiation defects associated with the mutant alleles, which variably impaired the DNA repair pathway. We further showed that these LIG1 mutant alleles are amorphic or hypomorphic, and exhibited variably decreased enzymatic activities, which lead to premature release of unligated adenylated DNA. The variability of the LIG1 genotypes in the patients was consistent with that of their immunological and clinical phenotypes. These data suggest that different forms of autosomal recessive, partial DNA ligase 1 deficiency underlie an immunodeficiency of variable severity.

Tackling overutilization of hospital tests and treatments: Lessons learned from a grassroots approach.

Recent data suggest unnecessary medical testing and treatment is relatively common in Canada. A number of harms to patients can arise as a result of unnecessary tests and treatments. In addition to patient harm, unnecessary tests and treatments add to the cost of medical care. Inspired by the Choosing Wisely campaign, St. Michael's Hospital in Toronto, Ontario, developed a hospital-wide program to address many different forms of overutilization at our hospital. The program prioritizes harm reduction over cost-containment and aims to create sustainable change through grassroots clinician engagement. This article will review important lessons learned from the St. Michael's experience.

Crystal structure of (2-{[(8-aminona-phthalen-1-yl)imino]-meth-yl}-4,6-di-tert-butyl-phenolato-κ3N,N',O)bromido-nickel(II).

The title compound, [NiBr(C25H29N2O)], contains an NiII atom with a slightly distorted square-planar coordination environment defined by one O and two N atoms from the 2-{[(8-aminona-phthalen-1-yl)imino]-meth-yl}-4,6-di-tert-butyl-phenolate ligand and a bromide anion. The Ni-O and Ni-N bond lengths are slightly longer than those observed in the phenyl backbone counterpart, which can be attributed to the larger steric hindrance of the naphthyl group in the structure of the title compound. The mol-ecule as a whole is substanti-ally distorted, with both the planar naphthalene-1,8-di-amine and imino-meth-yl-phenolate substitutents rotated against the NiN2OBr plane by 38.92 (7) and 37.22 (8)°, respectively, giving the mol-ecule a twisted appearance. N-H⋯Br hydrogen bonds and N-H⋯C(π) contacts connect the mol-ecules into dimers, and additional C-H⋯Br contacts, C-H⋯π inter-actions, and an offset stacking inter-action between naphthyl units inter-connect these dimers into a three-dimensional network.

Sugar sensor genes in the murine gastrointestinal tract display a cephalocaudal axis of expression and a diurnal rhythm.

Distributed along the length of the gastrointestinal (GI) tract are nutrient sensing cells that release numerous signaling peptides influencing GI function, nutrient homeostasis and energy balance. Recent studies have shown a diurnal rhythm in GI nutrient sensing, but the mechanisms responsible for rhythmicity are poorly understood. In this report we studied murine GI sugar sensor gene and protein expression levels in the morning (7 AM) and evening (7 PM). Sweet taste receptor ( tas1r2/tas1r3/gnat3/gnat1) sugar transporter ( slc5a1, slc2a2, slc2a5) and putative sugar sensor ( slc5a4a and slc5a4b) gene expression levels were highest in tongue and proximal and distal small intestine, respectively. Clock gene ( cry2/arntl) activity was detected in all regions studied. Slc5a4a and slc5a4b gene expression showed clear diurnal rhythmicity in the small intestine and stomach, respectively, although no rhythmicity was detected in SGLT3 protein expression. Tas1r2, tas1r3, gnat1, and gcg displayed a limited rhythm in gene expression in proximal small intestine. Microarray analysis revealed a diurnal rhythm in gut peptide gene expression in tongue (7 AM vs. 7 PM) and in silico promoter analysis indicated intestinal sugar sensors and transporters possessed the canonical E box elements necessary for clock gene control over gene transcription. In this report we present evidence of a diurnal rhythm in genes that are responsible for intestinal nutrient sensing that is most likely controlled by clock gene activity. Disturbances in clock gene/nutrient sensing interactions may be important in the development of diet-related diseases, such as obesity and diabetes.

Kinetic analyses of single-stranded break repair by human DNA ligase III isoforms reveal biochemical differences from DNA ligase I.

Humans have three genes encoding DNA ligases with conserved structural features and activities, but they also have notable differences. The LIG3 gene encodes a ubiquitous isoform in all tissues (LIG3α) and a germ line-specific splicing isoform (LIG3ß) that differs in the C-terminal domain. Both isoforms are found in the nucleus and the mitochondria. Here, we determined the kinetics and thermodynamics of single-stranded break ligation by LIG3α and LIG3ß and compared this framework to that of LIG1, the nuclear replicative ligase. The kinetic parameters of the LIG3 isoforms are nearly identical under all tested conditions, indicating that the BRCA1 C terminal (BRCT) domain specific to LIG3α does not alter ligation kinetics. Although LIG3 is only 22% identical to LIG1 across their conserved domains, the two enzymes had very similar maximal ligation rates. Comparison of the rate and equilibrium constants for LIG3 and LIG1 nevertheless revealed important differences. The LIG3 isoforms were seven times more efficient than LIG1 at ligating nicked DNA under optimal conditions, mainly because of their lower Km value for the DNA substrate. This could explain why LIG3 is less prone to abortive ligation than LIG1. Surprisingly, the affinity of LIG3 for Mg2+ was ten times weaker than that of LIG1, suggesting that Mg2+ availability regulates DNA ligation in vivo, because Mg2+ levels are higher in the mitochondria than in the nucleus. The biochemical differences between the LIG3 isoforms and LIG1 identified here will guide the understanding of both unique and overlapping biological roles of these critical enzymes.

Non-contraceptive oestrogen-containing preparations for controlling symptoms of premenstrual syndrome.

BACKGROUND: Premenstrual syndrome (PMS) is a psychological and somatic disorder of unknown aetiology, with symptoms typically including irritability, depression, mood swings, bloating, breast tenderness and sleep disturbances. About 3% to 10% of women who experience these symptoms may also meet criteria for premenstrual dysphoric disorder (PMDD). PMS symptoms recur during the luteal phase of the menstrual cycle and reduce by the end of menstruation. PMS results from ovulation and may be due to ovarian steroid interactions relating to neurotransmitter dysfunction. Premenstrual disorders have a devastating effect on women, their families and their work.Several treatment options have been suggested for PMS, including pharmacological and surgical interventions. The treatments thought to be most effective tend to fall into one of two categories: suppressing ovulation or correcting a speculated neuroendocrine anomaly.Transdermal oestradiol by patch, gel or implant effectively stops ovulation and the cyclical hormonal changes which produce the cyclical symptoms. These preparations are normally used for hormone therapy and contain lower doses of oestrogen than found in oral contraceptive pills. A shortened seven-day course of a progestogen is required each month for endometrial protection but can reproduce premenstrual syndrome-type symptoms in these women. OBJECTIVES: To determine the effectiveness and safety of non-contraceptive oestrogen-containing preparations in the management of PMS. SEARCH METHODS: On 14 March 2016, we searched the following databases: the Cochrane Gynaecology and Fertility Group (CGF) Specialised Register; Cochrane Central Register of Studies (CRSO); MEDLINE; Embase; PsycINFO; CINAHL; ClinicalTrials.gov; metaRegister of Controlled trials (mRCT); and the World Health Organization (WHO) International Clinical Trials Registry Platform (ICTRP) Search Portal. In addition, we checked the reference lists of articles retrieved. SELECTION CRITERIA: We included published and unpublished randomized placebo or active controlled trials on the efficacy of the use of non-contraceptive oestrogen-containing preparations in the management of premenstrual syndrome in women of reproductive age with PMS diagnosed by at least two prospective cycles without current psychiatric disorder. DATA COLLECTION AND ANALYSIS: Two review authors independently selected studies, assessed risk of bias, extracted data on premenstrual symptoms and adverse effects and entered data into Review Manager 5 software. Where possible, intention-to-treat or modified intention-to-treat analysis was used. Studies were pooled using a fixed-effect model, analysing cross-over trials as parallel trials. Standardised mean differences (SMDs) with 95% confidence intervals (CIs) were calculated for premenstrual symptom scores. Risk ratios (RRs) with 95% confidence intervals (CIs) were calculated for dichotomous outcomes. The overall quality of the evidence was assessed using the GRADE working group methods. MAIN RESULTS: The search resulted in 524 potentially relevant articles. Five eligible randomized controlled trials (RCTs) were identified (305 women). Trials using oral tablets, transdermal patches and implants were identified. No trial used gels.One small cross-over trial (11 women, effective sample size 22 women considering cross-over trials) compared oral luteal-phase oestrogen versus placebo. Data were very low quality and unsuitable for analysis, but study authors reported that the intervention was ineffective and might aggravate the symptoms of PMS. They also reported that there were no adverse events.Three studies compared continuous oestrogen with progestogen versus placebo (with or without progestogen). These trials were of reasonable quality, although with a high risk of attrition bias and an unclear risk of bias due to potential carry-over effects in two cross-over trials. Continuous oestrogen had a small to moderate positive effect on global symptom scores (SMD -0.34, 95% CI -0.59 to -0.10, P = 0.005, 3 RCTs, 158 women, effective sample size 267 women, I² = 63%, very low quality evidence). The evidence was too imprecise to determine if the groups differed in withdrawal rates due to adverse effects (RR 0.64, 95% CI 0.26 to 1.58, P = 0.33, 3 RCTs, 196 women, effective sample size 284 women, I² = 0%, very low quality evidence). Similarly, the evidence was very imprecise in measures of specific adverse events, with large uncertainties around the true value of the relative risk. None of the studies reported on long-term risks such as endometrial cancer or breast cancer.One study compared patch dosage (100 vs 200 µg oestrogen, with progestogen in both arms) and had a high risk of performance bias, detection bias and attrition bias. The study did not find evidence that dosage affects global symptoms but there was much uncertainty around the effect estimate (SMD -1.55, 95% CI -8.88 to 5.78, P = 0.68, 1 RCT, 98 women, very low quality evidence). The evidence on rates of withdrawal for adverse events was too imprecise to draw any conclusions (RR 0.70, 95% CI 0.34 to 1.46, P = 0.34, 1 RCT, 107 women, low-quality evidence). However, it appeared that the 100 µg dose might be associated with a lower overall risk of adverse events attributed to oestrogen (RR 0.51, 95% Cl 0.26 to 0.99, P = 0.05, 1 RCT, 107 women, very low quality evidence) with a large uncertainty around the effect estimate.The overall quality of the evidence for all comparisons was very low, mainly due to risk of bias (specifically attrition), imprecision, and statistical and clinical heterogeneity. AUTHORS' CONCLUSIONS: We found very low quality evidence to support the effectiveness of continuous oestrogen (transdermal patches or subcutaneous implants) plus progestogen, with a small to moderate effect size. We found very low quality evidence from a study based on 11 women to suggest that luteal-phase oral unopposed oestrogen is probably ineffective and possibly detrimental for controlling the symptoms of PMS. A comparison between 200 µg and 100 µg doses of continuous oestrogen was inconclusive with regard to effectiveness, but suggested that the lower dose was less likely to cause side effects. Uncertainty remains regarding safety, as the identified studies were too small to provide definite answers. Moreover, no included trial addressed adverse effects that might occur beyond the typical trial duration of 2-8 months. This suggests the choice of oestrogen dose and mode of administration could be based on an individual woman's preference and modified according to the effectiveness and tolerability of the chosen regimen.