A Frameshift Variant of GluN2A Identified in an Epilepsy Patient Results in NMDA Receptor Mistargeting.

N-methyl-D-aspartate receptors (NMDARs) are crucial for neuronal development and synaptic plasticity. Dysfunction of NMDARs is associated with multiple neurodevelopmental disorders, including epilepsy, autism spectrum disorder, and intellectual disability. Understanding the impact of genetic variants of NMDAR subunits can shed light on the mechanisms of disease. Here, we characterized the functional implications of a de novo mutation of the GluN2A subunit (P1199Rfs*32) resulting in the truncation of the C-terminal domain. The variant was identified in a male patient with epileptic encephalopathy, multiple seizure types, severe aphasia, and neurobehavioral changes. Given the known role of the CTD in NMDAR trafficking, we examined changes in receptor localization and abundance at the postsynaptic membrane using a combination of molecular assays in heterologous cells and rat primary neuronal cultures. We observed that the GluN2A P1199Rfs*32-containing receptors traffic efficiently to the postsynaptic membrane but have increased extra-synaptic expression relative to WT GluN2A-containing NMDARs. Using in silico predictions, we hypothesized that the mutant would lose all PDZ interactions, except for the recycling protein Scribble1. Indeed, we observed impaired binding to the scaffolding protein postsynaptic protein-95 (PSD-95); however, we found the mutant interacts with Scribble1, which facilitates the recycling of both the mutant and the WT GluN2A. Finally, we found that neurons expressing GluN2A P1199Rfs*32 have fewer synapses and decreased spine density, indicating compromised synaptic transmission in these neurons. Overall, our data show that GluN2A P1199Rfs*32 is a loss-of-function variant with altered membrane localization in neurons and provide mechanistic insight into disease etiology.

Assessing Left Ventricular Systolic Function in Patients with Pregnancy-Induced Hypertension Using Three-Dimensional Speckle Tracking Technology.

Objective: This study investigates the clinical utility of three-dimensional speckle tracking technology in assessing left ventricular systolic function in pregnancy-induced hypertension syndrome (PIH). Methods: We retrospectively enrolled 70 patients with diagnosed PIH treated at our institution between July 2019 and August 2021 as the study group. A total of 70 healthy pregnant women undergoing routine antenatal examinations at the same institution during the same period were included in the control group. Two-dimensional conventional echocardiography measured left ventricular parameters in both groups. Three-dimensional speckle tracking technology analyzed Left Ventricular Global Longitudinal Peak Strain (LVGLS), Left Ventricular Global Radial Peak Strain (LVGRS), and Left Ventricular Global Circumferential Peak Strain (LVGCS). Differences in left ventricular systolic function and pregnancy outcomes were compared. Results: In the study group, LVEDD, LVPWTd, and IVSTd (47.67±4.88, 10.68±1.21, 11.24±1.03) exceeded those in the control group (45.21±5.65, 8.17±0.98, 8.91±0.37). LVEF (62.12±5.63) was lower than the control group (65.25±5.17) (all P < .05). LVGLS, LVGCS, and LVGAS in the study group (-15.66±1.07, -20.17±2.89, -23.17±3.43) were higher than the control group (-20.14±1.27, -25.17±1.36, -37.68±3.29), while LVGRS (30.29±3.61) was lower than the control group (34.18±4.08) (all P < .05). The study group had 72.86% natural deliveries and 27.14% cesarean sections; the control group had 31.43% natural deliveries and 68.57% cesarean sections (all P < .05). Weeks of delivery and birth weight in the study group (36.87±1.23, 2.71±0.41) were lower than the control group (38.96±1.54, 3.41±0.78) (both P < .05). Conclusions: Compared to traditional methods, three-dimensional speckle tracking technology more sensitively detects left ventricular strain and rotation in PIH patients. It holds clinical relevance in early left ventricular dysfunction detection, effectively mitigating adverse pregnancy outcomes and warranting clinical adoption and application.

Physiologically Based Pharmacokinetic Modeling To Guide Management of Drug Interactions between Elexacaftor-Tezacaftor-Ivacaftor and Antibiotics for the Treatment of Nontuberculous Mycobacteria.

Nontuberculous mycobacteria (NTM) are the pathogens of concern in people with cystic fibrosis (pwCF) due to their association with deterioration of lung function. Treatment requires the use of a multidrug combination regimen, creating the potential for drug-drug interactions (DDIs) with cystic fibrosis transmembrane conductance regulator (CFTR)-modulating therapies, including elexacaftor, tezacaftor, and ivacaftor (ETI), which are eliminated mainly through cytochrome P450 (CYP) 3A-mediated metabolism. An assessment of the DDI risk for ETI coadministered with NTM treatments, including rifabutin, clofazimine, and clarithromycin, is needed to provide appropriate guidance on dosing. The CYP3A-mediated DDIs between ETI and the NTM therapies rifabutin, clarithromycin, and clofazimine were evaluated using physiologically based pharmacokinetic (PBPK) modeling by incorporating demographic and physiological "system" data with drug physicochemical and in vitro parameters. Models were verified and then applied to predict untested scenarios to guide continuation of ETI during antibiotic treatment, using ivacaftor as the most sensitive CYP3A4 substrate. The predicted area under the concentration-time curve (AUC) ratios of ivacaftor when coadministered with rifabutin, clofazimine, or clarithromycin were 0.31, 2.98, and 9.64, respectively, suggesting moderate and strong interactions. The simulation predicted adjusted dosing regimens of ETI administered concomitantly with NTM treatments, which required delayed resumption of the standard dose of ETI once the NTM treatments were completed. The dosing transitions were determined based on the characteristics of the perpetrator drugs, including the mechanism of CYP3A modulation and their elimination half-lives. This study suggests increased doses of elexacaftor/tezacaftor/ivacaftor 200/100/450 mg in the morning and 100/50/375 mg in the evening when ETI is coadministered with rifabutin and reduced doses of elexacaftor/tezacaftor 200/100 mg every 48 h (q48h) and ivacaftor 150 mg daily or a dose of elexacaftor/tezacaftor/ivacaftor 200/100/150 mg q72h when coadministered with clofazimine or clarithromycin, respectively. Importantly, the PBPK simulations provide evidence in support of the use of treatments for NTM in pwCF receiving concomitant dose-adjusted ETI therapy.

Treatment of early-stage mycosis fungoides: results from the PROspective Cutaneous Lymphoma International Prognostic Index (PROCLIPI) study.

BACKGROUND: The PROspective Cutaneous Lymphoma International Prognostic Index (PROCLIPI) study is a prospective analysis of an international database. Here we examine front-line treatments and quality of life (QoL) in patients with newly diagnosed mycosis fungoides (MF). OBJECTIVES: To identify (i) differences in first-line approaches according to tumour-nodes-metastasis-blood (TNMB) staging; (ii) parameters related to a first-line systemic approach and (iii) response rates and QoL measures. METHODS: In total, 395 newly diagnosed patients with early-stage MF (stage IA-IIA) were recruited from 41 centres in 17 countries between 1 January 2015 and 31 December 2018 following central clinicopathological review. RESULTS: The most common first-line therapy was skin-directed therapy (SDT) (322 cases, 81·5%), while a smaller percentage (44 cases, 11·1%) received systemic therapy. Expectant observation was used in 7·3%. In univariate analysis, the use of systemic therapy was significantly associated with higher clinical stage (IA, 6%; IB, 14%; IIA, 20%; IA-IB vs. IIA, P < 0·001), presence of plaques (T1a/T2a, 5%; T1b/T2b, 17%; P < 0·001), higher modified Severity Weighted Assessment Tool (> 10, 15%; ≤ 10, 7%; P = 0·01) and folliculotropic MF (FMF) (24% vs. 12%, P = 0·001). Multivariate analysis demonstrated significant associations with the presence of plaques (T1b/T2b vs. T1a/T2a, odds ratio 3·07) and FMF (odds ratio 2·83). The overall response rate (ORR) to first-line SDT was 73%, while the ORR to first-line systemic treatments was lower (57%) (P = 0·027). Health-related QoL improved significantly both in patients with responsive disease and in those with stable disease. CONCLUSIONS: Disease characteristics such as presence of plaques and FMF influence physician treatment choices, and SDT was superior to systemic therapy even in patients with such disease characteristics. Consequently, future treatment guidelines for early-stage MF need to address these issues.

Should we be imaging lymph nodes at initial diagnosis of early-stage mycosis fungoides? Results from the PROspective Cutaneous Lymphoma International Prognostic Index (PROCLIPI) international study.

BACKGROUND: Early-stage mycosis fungoides (MF) includes involvement of dermatopathic lymph nodes (LNs) or early lymphomatous LNs. There is a lack of unanimity among current guidelines regarding the indications for initial staging imaging in early-stage presentation of MF in the absence of enlarged palpable LNs. OBJECTIVES: To investigate how often imaging is performed in patients with early-stage presentation of MF, to assess the yield of LN imaging, and to determine what disease characteristics promoted imaging. METHODS: A review of clinicopathologically confirmed newly diagnosed patients with cutaneous patch/plaque (T1/T2) MF from PROspective Cutaneous Lymphoma International Prognostic Index (PROCLIPI) data. RESULTS: PROCLIPI enrolled 375 patients with stage T1/T2 MF: 304 with classical MF and 71 with folliculotropic MF. Imaging was performed in 169 patients (45%): 83 with computed tomography, 18 with positron emission tomography-computed tomography and 68 with ultrasound. Only nine of these (5%) had palpable enlarged (≥ 15 mm) LNs, with an over-representation of plaques, irrespectively of the 10% body surface area cutoff that distinguishes T1 from T2. Folliculotropic MF was not more frequently imaged than classical MF. Radiologically enlarged LNs (≥ 15 mm) were detected in 30 patients (18%); only seven had clinical lymphadenopathy. On multivariate analysis, plaque presentation was the sole parameter significantly associated with radiologically enlarged LNs. Imaging of only clinically enlarged LNs upstaged 4% of patients (seven of 169) to at least IIA, whereas nonselective imaging upstaged another 14% (24 of 169). LN biopsy, performed in eight of 30 patients, identified N3 (extensive lymphomatous involvement) in two and N1 (dermatopathic changes) in six. CONCLUSIONS: Physical examination was a poor determinant of LN enlargement or involvement. Presence of plaques was associated with a significant increase in identification of enlarged or involved LNs in patients with early-stage presentation of MF, which may be important when deciding who to image. Imaging increases the detection rate of stage IIA MF, and identifies rare cases of extensive lymphomatous nodes, upstaging them to advanced-stage IVA2.

Improving clinical management of colon cancer through CONNECTION, a nation-wide colon cancer registry and stratification effort (CONNECTION II trial): rationale and protocol of a single arm intervention study.

BACKGROUND: It is estimated that around 15-30% of patients with early stage colon cancer benefit from adjuvant chemotherapy. We are currently not capable of upfront selection of patients who benefit from chemotherapy, which indicates the need for additional predictive markers for response to chemotherapy. It has been shown that the consensus molecular subtypes (CMSs), defined by RNA-profiling, have prognostic and/or predictive value. Due to postoperative timing of chemotherapy in current guidelines, tumor response to chemotherapy per CMS is not known, which makes the differentiation between the prognostic and predictive value impossible. Therefore, we propose to assess the tumor response per CMS in the neoadjuvant chemotherapy setting. This will provide us with clear data on the predictive value for chemotherapy response of the CMSs. METHODS: In this prospective, single arm, multicenter intervention study, 262 patients with resectable microsatellite stable cT3-4NxM0 colon cancer will be treated with two courses of neoadjuvant and two courses of adjuvant capecitabine and oxaliplatin. The primary endpoint is the pathological tumor response to neoadjuvant chemotherapy per CMS. Secondary endpoints are radiological tumor response, the prognostic value of these responses for recurrence free survival and overall survival and the differences in CMS classification of the same tumor before and after neoadjuvant chemotherapy. The study is scheduled to be performed in 8-10 Dutch hospitals. The first patient was included in February 2020. DISCUSSION: Patient selection for adjuvant chemotherapy in early stage colon cancer is far from optimal. The CMS classification is a promising new biomarker, but a solid chemotherapy response assessment per subtype is lacking. In this study we will investigate whether CMS classification can be of added value in clinical decision making by analyzing the predictive value for chemotherapy response. This study can provide the results necessary to proceed to future studies in which (neo) adjuvant chemotherapy may be withhold in patients with a specific CMS subtype, who show no benefit from chemotherapy and for whom possible new treatments can be investigated. TRIAL REGISTRATION: This study has been registered in the Netherlands Trial Register (NL8177) at 11-26-2019, https://www.trialregister.nl/trial/8177 . The study has been approved by the medical ethics committee Utrecht (MEC18/712).

The incidence and survival of melanoma and nonmelanoma skin cancer in patients with vitiligo: a nationwide population-based matched cohort study in Korea.

BACKGROUND: While studies report a lower incidence of skin cancer in white patients with vitiligo compared with controls, the skin cancer incidence in Asian patients with vitiligo is unknown. OBJECTIVES: To quantify the incidence of melanoma and nonmelanoma skin cancer (NMSC) in Korean patients with vitiligo and compare it with matched nonvitiligo controls. METHODS: A retrospective matched cohort study was performed with 131 245 incident vitiligo cases and 2 624 900 age- and sex-matched (1 : 20) controls at index date, who were selected from the Korean National Health Insurance database between January 2005 and December 2017. Stratified Cox proportional hazards regression (stratified by sex, birth year and index year) was used to calculate the hazard ratio (HR) of skin cancer in patients with vitiligo. RESULTS: Patients with vitiligo were followed up for a mean duration of 6·34 years compared with a follow-up period of 6·27 years for matched controls. Ultraviolet (UV) treatment-adjusted HR for melanoma in patients with vitiligo was 3·32 [95% confidence interval (CI) 2·29-4·81] and 1·29 (95% CI 1·06-1·56) for NMSC. The HRs for melanoma and NMSC in the vitiligo population without a history of UV treatment were 3·37 (95% CI 2·32-4·90) and 1·35 (95% CI 1·11-1·64), respectively. CONCLUSIONS: In contrast to white patients with vitiligo, the risk of skin cancer was increased in the Korean vitiligo population. However, it is noteworthy that the skin cancer incidence in Korean patients with vitiligo was lower than that of their white counterparts. Owing to possible ethnic differences in the susceptibility to skin cancer, skin cancer surveillance in the vitiligo population may be adjusted for race. What's already known about this topic? Prior studies have reported a lower incidence of melanoma and nonmelanoma skin cancer (NMSC) in white patients with vitiligo compared with nonvitiligo controls. The skin cancer incidence in Asian patients with vitiligo is unknown. What does this study add? In contrast to white patients, the risk of both melanoma and NMSC was increased in Korean patients with vitiligo compared with controls. Owing to possible ethnic differences in susceptibility to skin cancer, skin cancer surveillance in the vitiligo population should be adjusted for race.

Impact of immunogenicity on efficacy and tolerability of tumour necrosis factor inhibitors: pooled analysis of biosimilar studies in rheumatoid arthritis.

Objective: SB4, SB2, and SB5 are biosimilars of etanercept (ETN), infliximab (INF), and adalimumab (ADA), respectively. This pooled analysis evaluated the immunogenicity of these treatments across three phase III randomized controlled trials of patients with rheumatoid arthritis (RA). Methods: Patients had to have at least one anti-drug antibody (ADAb) assessment up to the time of the primary endpoint from each study (week 24 in SB4 and SB5 studies; week 30 in SB2 study). The effect of ADAbs on American College of Rheumatology 20% (ACR20) response and the incidences of injection-site reactions (ISRs)/infusion-related reactions (IRRs) were evaluated. Results: The study included 1709 patients. The cumulative incidences of ADAbs were 30.3% in the all-treatments-combined group, 29.1% in the biosimilars combined group, and 31.5% in the reference products combined group. ACR20 response rates were significantly lower in ADAb-positive patients in the all-treatments-combined [odds ratio (95% confidence interval) 1.77 (1.37, 2.27), p < 0.0001], biosimilars combined [2.24 (1.53, 3.30), p < 0.0001], and reference products combined [1.49 (1.06, 2.09), p = 0.0225] groups. ADAb-positive patients also had a higher likelihood of developing ISRs/IRRs in the all-treatments-combined group [0.56 (0.31, 1.01), p = 0.0550], predominantly due to the results observed with SB2 + INF combined rather than with SB4 + ETN or SB5 + ADA combined. Conclusion: In this pooled analysis, ADAbs were associated with reduced efficacy in patients with RA treated with biosimilars (SB4, SB2, and SB5) or their reference products (ETN, INF, and ADA). ADAbs were associated with an increased incidence of ISRs/IRRs in those treated with SB2 + INF. Clinical trial registration numbers: NCT01936181 (SB2 study), NCT01895309 (SB4 study), and NCT02167139 (SB5 study).

[Extraction and Separation of Sinapine from Rapeseed Cake and the Mode of Action of Melanin Production Inhibition].

Brassica campestris L. is the important oil-bearing crop in China. Rapeseed cake is the main byproduct of rapeseed oil extraction. As the main active ingredient in rapeseed cake, sinapine has several important biological activities. Therefore, the inhibitory activity of sinapine on tyrosinase in vitro and its free radical-scavenging rate were determined. Tyrosinase activity in A-375 human melanocytes was also investigated and the effects of sinapine on the melanin content and its antioxidant effects on melanin biosynthesis were studied. The results showed that sinapine had significant antioxidant activity. Sinapine significantly inhibited A-375 human melanocytes in a dose-dependent manner. Sinapine inhibited melanin synthesis in A-375 cells by downregulating the mRNA and protein expression of TRP-1, TRP-2, and MITF factors. The results showed that rapeseed cake sinapine inhibited melanin production and could be used as a potential active ingredient in the development of whitening agents.

Egg Quality and Pregnancy Outcome in Young Infertile Women with Diminished Ovarian Reserve.

BACKGROUND This study investigated the quality and quantity of eggs and embryos as well as the clinical pregnancy outcome in young infertile women with diminished ovarian reserve (DOR) after in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI). MATERIAL AND METHODS We retrospectively reviewed records of 4285 infertile women and divided them into 3 groups according to age and ovarian reserve: young women with normal ovarian reserve (n=1695), young women with DOR (n=1121), and older women with DOR (n=1469). RESULTS In young women with DOR, the proportion of high-quality embryos was significantly higher than in older women with DOR and lower than in young women with normal ovarian reserve (P<0.01). The proportions of ovulation cancellation, ovulation without egg acquisition, and ovulation without available embryos in young women with DOR were significantly higher than in young women with normal ovarian reserve. The rates of biochemical pregnancy, clinical pregnancy, and embryo implantation in young women with DOR were significantly higher than in older women with DOR, and lower than in young women with normal ovarian reserve. The miscarriage rate was 19.17% in young women with DOR, significantly lower than in older women with DOR (33.90%), and higher than in young women with normal ovarian reserve. CONCLUSIONS Young women with DOR have ovarian hypo-response and low numbers of acquired eggs and embryos, but the possibilities of high-quality embryo and good clinical pregnancy are higher once eggs are acquired. The indications to IVF/ICSI can be widened and active treatments should be administered for these women.

Possible involvement of orphan receptors GPR88 and GPR124 in the development of hypertension in spontaneously hypertensive rat.

Hypertension (HBP) is a chronic disease characterized by increased blood pressure, which despite several treatments maintains a high morbi-mortality, which suggests that there are other mechanisms involved in this pathology, within which the orphan receptors could be candidates for the treatment of the HBP; these receptors are called orphan receptors because their ligand is unknown. These receptors have been suggested to participate in some pathologies because they are associated with various systems such as GPR88, which has been linked to the dopaminergic system, and GPR124 with angiogenesis, suggesting that these receptors could take part in HBP. Hence, the aim of this work was to study the expression of orphan receptors GPR88 and GPR124 in various tissues of normotensive and hypertensive rats. We used Wistar Kyoto (WKY) and spontaneously hypertensive rat (SHR) of 6-8 and 10-12 weeks of age and we determined systolic blood pressure (SBP), heart rate, as well as mRNA of GPR88 and GPR124 receptors by reverse transcription polymerase chain reaction (RT-PCR) in the aorta, heart, kidney, and brain. Our results showed that GPR88 and GPR124 were expressed in all analyzed tissues, but their expression is dependent on the age and development of HBP because their expression tends to be modified as HBP is established. Therefore, we conclude that GPR88 and GPR124 receptors may be involved in the development or maintenance of high blood pressure.

Quality of care in patients with diabetic kidney disease in Asia: The Joint Asia Diabetes Evaluation (JADE) Registry.

AIMS: Diabetic kidney disease independently predicts cardiovascular disease and premature death. We examined the burden of chronic kidney disease (CKD, defined as an estimated GFR < 60 ml/min/1.73 m(2) ) and quality of care in a cross-sectional survey of adults (age ≥ 18 years) with Type 2 diabetes across Asia. METHODS: The Joint Asia Diabetes Evaluation programme is a disease-management programme implemented using an electronic portal that systematically captures clinical characteristics of all patients enrolled. Between July 2007 and December 2012, data on 28 110 consecutively enrolled patients (China: 3415, Hong Kong: 15 196, India: 3714, Korea: 1651, Philippines: 3364, Vietnam: 692, Taiwan: 78) were analysed. RESULTS: In this survey, 15.9% of patients had CKD, 25.0% had microalbuminuria and 12.5% had macroalbuminuria. Patients with CKD were less likely to achieve HbA1c < 53 mmol/mol (7.0%) (36.0% vs. 42.3%) and blood pressure < 130/80 mmHg (20.8% vs. 35.3%), and were more likely to have retinopathy (26.2% vs. 8.7%), sensory neuropathy (29.0% vs. 7.7%), cardiovascular disease (26.6% vs. 8.7%) and self-reported hypoglycaemia (18.9% vs. 8.2%). Despite high frequencies of albuminuria (74.8%) and dyslipidaemia (93.0%) among CKD patients, only 49.0% were using renin-angiotensin system inhibitors and 53.6% were on statins. On logistic regression, old age, male gender, tobacco use, long disease duration, high HbA1c , blood pressure and BMI, and low LDL cholesterol were independently associated with CKD (all P < 0.05). CONCLUSIONS: The poor control of risk factors, suboptimal use of organ-protective drugs and high frequencies of hypoglycaemia highlight major treatment gaps in patients with diabetic kidney disease in Asia.

Prevalence and prognosis of hypoglycaemia in patients receiving maintenance dialysis.

BACKGROUND: End-stage renal disease is a common predisposing condition for the development of hypoglycaemia. AIM: To determine the effect of hypoglycaemia on the mortality of patients undergoing maintenance dialysis. METHODS: Retrospective and descriptive analyses were performed in five dialysis centres in the Republic of Korea between June 2002 and August 2008. We enrolled 1685 patients who had undergone dialysis for at least 1 month. RESULTS: We identified 453 episodes of hypoglycaemia in 256 of 1685 patients (15.2%); 189 patients (73.8%) had diabetes, whereas the other patients did not. The occurrence of hypoglycaemia in patients receiving dialysis appeared to be a life-threatening complication because 27.0% of patients died within two days of the onset of a hypoglycaemic episode. Older age, low serum albumin levels and infections were independent risk factors for total mortality in these patients. Furthermore, the absence of diabetes, age and serum glucose levels were independent factors associated with early mortality within two days of the development of hypoglycaemia. CONCLUSION: Although several factors were associated with mortality, the degree of hypoglycaemia, absence of diabetes and old age were associated with early mortality. Elderly hypoglycaemic patients, especially those without diabetes, should be closely monitored.

Changes in protein and gene expression of angiotensin II receptors (AT1 and AT2) in aorta of diabetic and hypertensive rats.

Diabetes and hypertension have been associated with cardiovascular diseases and stroke. Some reports have related the coexistence of hypertension and diabetes with increase in the risk of developing vascular complications. Recently some studies have shown results suggesting that in the early stages of diabetes and hypertension exist a reduced functional response to vasopressor agents like angiotensin II (Ang II), which plays an important role in blood pressure regulation mechanism through the activation of its AT1 and AT2 receptors. For that reason, the aim of this work was to study the gene and protein expression of AT1 and AT2 receptors in aorta of diabetic SHR and WKY rats. Diabetes was induced by the administration of streptozotocin (60 mg/kg i.p.). After 4 weeks of the onset of diabetes, the protein expression was obtained by western blot and the mRNA expression by RT-PCR. Our results showed that the hypertensive rats have a higher mRNA and protein expression of AT1 receptors than normotensive rats while the AT2 expression remained unchanged. On the other hand, the combination of diabetes and hypertension increased the mRNA and protein expression of AT1 and AT2 receptors significantly. In conclusion, our results suggest that diabetes with hypertension modifies the mRNA and protein expression of AT1 and AT2 receptors. However, the overexpression of AT2 could be associated with the reduction in the response to Ang II in the early stage of diabetes.

Development of Anilino-Maytansinoid ADCs that Efficiently Release Cytotoxic Metabolites in Cancer Cells and Induce High Levels of Bystander Killing.

Antibody anilino maytansinoid conjugates (AaMCs) have been prepared in which a maytansinoid bearing an aniline group was linked through the aniline amine to a dipeptide, which in turn was covalently attached to a desired monoclonal antibody. Several such conjugates were prepared utilizing different dipeptides in the linkage including Gly-Gly, l-Val-l-Cit, and all four stereoisomers of the Ala-Ala dipeptide. The properties of AaMCs could be altered by the choice of dipeptide in the linker. Each of the AaMCs, except the AaMC bearing a d-Ala-d-Ala peptide linker, displayed more bystander killing in vitro than maytansinoid ADCs that utilize disulfide linkers. In mouse models, the anti-CanAg AaMC bearing a d-Ala-l-Ala dipeptide in the linker was shown to be more efficacious against heterogeneous HT-29 xenografts than maytansinoid ADCs that utilize disulfide linkers, while both types of the conjugates displayed similar tolerabilities.

Design of Coltuximab Ravtansine, a CD19-Targeting Antibody-Drug Conjugate (ADC) for the Treatment of B-Cell Malignancies: Structure-Activity Relationships and Preclinical Evaluation.

Coltuximab ravtansine (SAR3419) is an antibody-drug conjugate (ADC) targeting CD19 created by conjugating a derivative of the potent microtubule-acting cytotoxic agent, maytansine, to a version of the anti-CD19 antibody, anti-B4, that was humanized as an IgG1 by variable domain resurfacing. Four different linker-maytansinoid constructs were synthesized (average ∼3.5 maytansinoids/antibody for each) to evaluate the impact of linker-payload design on the activity of the maytansinoid-ADCs targeting CD19. The ADC composed of DM4 (N(2')-deacetyl-N(2')-[4-mercapto-4-methyl-1-oxopentyl]maytansine) conjugated to antibody via the N-succinimidyl-4-(2-pyridyldithio)butyrate (SPDB) linker was selected for development as SAR3419. A molar ratio for DM4/antibody of between 3 and 5 was selected for the final design of SAR3419. Evaluation of SAR3419 in Ramos tumor xenograft models showed that the minimal effective single dose was about 50 µg/kg conjugated DM4 (∼2.5 mg/kg conjugated antibody), while twice this dose gave complete regressions in 100% of the mice. SAR3419 arrests cells in the G2/M phase of the cell cycle, ultimately leading to apoptosis after about 24 h. The results of in vitro and in vivo studies with SAR3419 made with DM4 that was [(3)H]-labeled at the C20 methoxy group of the maytansinoid suggest a mechanism of internalization and intracellular trafficking of SAR3419, ultimately to lysosomes, in which the antibody is fully degraded, releasing lysine-N(ε)-SPDB-DM4 as the initial metabolite. Subsequent intracellular reduction of the disulfide bond between linker and DM4 generates the free thiol species, which is then converted to S-methyl DM4 by cellular methyl transferase activity. We provide evidence to suggest that generation of S-methyl DM4 in tumor cells may contribute to in vivo tumor eradication via bystander killing of neighboring tumor cells. Furthermore, we show that S-methyl DM4 is converted to the sulfoxide and sulfone derivatives in the liver, suggesting that hepatic catabolism of the payload to less cytotoxic maytansinoid species contributes to the overall therapeutic window of SAR3419. This compound is currently in phase II clinical evaluation for the treatment of diffuse large B cell lymphoma.

Low immunogenicity of quadrivalent meningococcal vaccines in solid organ transplant recipients.

Immunization against meningococcal disease is recommended for solid organ transplant (SOT) recipients at high risk for meningococcal disease or travelling to an endemic country. However, the immunogenicity of meningococcal vaccines has not been studied in this population. We analyzed the immune response of quadrivalent (against Neisseria meningitidis serogroups A, C, Y, and W) polysaccharidic non-conjugate and conjugate meningococcal vaccines in kidney- and liver-transplant patients using bactericidal assays against the targeted serogroups. Upon vaccination with a non-conjugate (n = 5) or a conjugate vaccine (n = 10), respectively, 40% and 50% of patients were able to mount an immune response, achieving at least the threshold correlated with protection defined as human serum bactericidal antibody titers of ≥4. Responders showed only partial and low responses (titers ≤64), thus predicting a rapid decline in bactericidal response. Only 1 patient developed a booster response to preexisting immunity. Our data argue for the need of additional measures for SOT recipients, when they are at risk of meningococcal disease.

A cost-reducing extracorporeal membrane oxygenation (ECMO) program model: a single institution experience.

BACKGROUND: The worldwide demand for ECMO support has grown. Its provision remains limited due to several factors (high cost, complicated technology, lack of expertise) that increase healthcare cost. Our goal was to assess if an intensive care unit (ICU)-run ECMO model without continuous bedside perfusionists would decrease costs while maintaining patient safety and outcomes. METHOD: A new ECMO program was implemented in 2010, consisting of dedicated ICU multidisciplinary providers (ICU-registered nurses, mid-level providers and intensivists). In year one, we introduced an education platform, new technology and dedicated space. In year two, continuous bedside monitoring by perfusionists was removed and new management algorithms designating multidisciplinary providers as first responders were established. The patient safety and cost benefit from the removal of the continuous bedside monitoring of the perfusionists of this new ECMO program was retrospectively reviewed and compared. RESULTS: During the study period, 74 patients (28 patients in year 1 and 46 patients in year 2) were placed on ECMO (mean days: 8 ± 5.7). The total annual hospital expenditure for the ECMO program was significantly reduced in the new model ($234,000 in year 2 vs. $600,264 in year 1), showing a 61% decrease in cost. This cost decrease was attributed to a decreased utilization of perfusion services and the introduction of longer lasting and more efficient ECMO technology. We did not find any significant changes in registered nurse ratios or any differences in outcomes related to ICU safety events. CONCLUSION: We demonstrated that the ICU-run ECMO model managed to lower hospital cost by reducing the cost of continuous bedside perfusion support without a change in outcomes.

Growth performance, carcass yield, and quality and chemical traits of meat from commercial korean native ducks with 2-way crossbreeding.

This work was conducted to investigate the performance and meat characteristics of commercial Korean native duck (KND). A total of 180 1-d-old ducklings of 2-way crossbreds from A and B lines (from National Institute of Animal Science) were used in this work and divided into 4 groups (3 replicates/group, 15 birds/replicate). The four groups were 4 crossbreds as AA (A line [♀]×A line [♂]), AB (A line [♀]×B line [♂]), BB (Pure line B strains) and BA (B strains [♀]×A strain [♂]). Ducks were fed diets based on corn-soybean meal for 0 to 3 wk (22.4% crude protein [CP], 2,945 kcal/kg metabolizable energy [ME]) and 3 to 8 wk (18.4% CP, 3,047 kcal/kg ME). As a result of this study, average body weight of 4 crossbreds were 625, 1,617, 2,466, and 2,836 g at 2, 4, 6, and 8 weeks, respectively, and significantly increased over the period of time (p<0.05). Body weight of BB group was greater than other crossbreds at the age of 6 weeks (p<0.05). There was a significant difference in weekly body weight gains (p<0.05), which were 573, 991, 850, and 371 g at 2, 4, 6, and 8 weeks old, respectively. Uniformity of 4 crossbreds was 84.9%, 80.5%, and 72.5% at 6, 7, and 8 weeks, respectively, and there was no difference among crossbreds. Body weight gain of BB crossbred was highest among crossbreds (p<0.05). Weekly feed intake significantly increased with weeks as 669, 1,839, 2,812, and 3,381 g at 2, 4, 6, and 8 weeks respectively (p<0.05). Feed intakes of AA and BB crossbreds were higher at 2 to 4 weeks old than others and that of BB crossbred was highest at 4 to 6 weeks old (p<0.05). Weekly feed conversion ratios were 1.17, 1.86, 3.32, and 9.37 at 0 to 2, 2 to 4, 4 to 6, and 6 to 8 weeks old, respectively, and it increased with age (p<0.05). There was no significant difference in feed conversion ratio among crossbreds. Carcass yields of 4 crossbreds were 73.6%, 71.6%, 73.5%, and 71.7%, respectively, so there was no significant difference among crossbreds. There was no difference in wing, neck, breast and leg ratios among crossbreds. However, back ratios of 4 crossbreds were 17.6%, 18.0%, 15.8%, and 17.6%, respectively, and back ratio of BB was the highest among crossbreds. Finally, these results may provide the basic data on the production, carcass quality, fatty acid and amino acid composition of commercial KND with 2-way crossbreeding.

Effect of lithium chloride on endoplasmic reticulum stress-related PERK/ROCK signaling in a rat model of glaucoma.

Elevated intraocular pressure (IOP) is considered as the major risk factor for the loss of retinal ganglion cells (RGCs) and their axons in glaucoma. Lithium chloride (LiCl) inhibits glycogen synthase kinase-3 beta (GSK-3ß) and attends PERK-induced endoplasmic reticulum stress (ERs) transition. PERK is a type I transmembrane protein located in the endoplasmic reticulum. PERK pathway activation takes place in ERs early inhibiting protein synthnesis to protect cell and promote cell survival. Here, we firstly evaluate that LiCl reduced IOP when administered intraperitoneally. After 6 weeks, IOP dropped by around 21.9% in LiCl treated rats. Then we investigated the effects of LiCI on PERK-mediated signaling pathways. LiCl treatment activated PERK and inhibited the expression of ROCK-1 and ROCK-2 in a rat model of glaucoma. Collectively, these results suggest that LiCl reduced the IOP through the phosphorylation of PERK by the regulation of PERK/ROCK signaling in glaucoma rat model.