Binding affinities of human IgG1 and chimerized pig and rabbit derivatives to human, pig and rabbit Fc gamma receptor IIIA.

While pigs and rabbits are used as models for human immune diseases, FcγR binding is poorly characterized in both test species. To evaluate antibody binding to FcγRIIIA, a receptor involved in antibody-dependent cellular cytotoxicity, chimerized antibodies were generated by grafting the variable regions of a human IgG1 onto scaffolds from both species. The affinities of the parent and chimeric antibodies to the FcγRIIIA proteins from all three species were determined. While the human IgG1 and rabbit IgG had similar affinities for each FcγRIIIA with notable differences across species, pig IgG1 only bound pig FcγRIIIA with appreciable affinity. Also, the functional pig and rabbit proteins described here can be used in future experiments, such as pharmacology and mechanism of action studies.

Dying Too Soon: Excess Mortality in Severe Mental Illness.

Aims: We aimed to identify baseline predictors of mortality in patients with a severe mental illness (SMI) over a 6-year period and to describe mortality rates as standardised mortality ratios (SMRs). We hypothesised that cardiovascular diseases, older age, cigarette smoking, more severe psychiatric symptoms and more severe psychotropic side effects, and alcohol or drug use were independent risk factors for mortality. Method: Medical examinations were conducted at baseline in a cohort of 322 SMI patients. SMRs were estimated after 6 years and an evaluation was made of the impact of a wide range of variables on survival time. Results: Almost 11% of the SMI patients had died at the end of the study period. All-cause SMRs were 4.51 (95% CI 3.07-5.95) for all SMI patients (4.89, 95% CI 2.97-6.80 for men, and 3.94, 95% CI 1.78-6.10 for women). Natural causes accounted for 86% of excess mortality and unnatural causes for 14%. Cardiovascular disease was a major contributor to this excess mortality. Multivariate Cox regression analyses showed that premature death was associated with a longer history of tobacco use (HR: 1.03, 95% CI 1.02-1.03) and more severe symptoms of disorganisation (HR: 2.36, 95% CI 2.21-2.52). Conclusions: The high SMR and the incidence of cardiovascular disease-related death in SMI patients in our study justify concern. This study underscores the urgent need for interventions to reduce excess mortality in patients with SMI.

Use of outcome measurements in clinical practice: How specific should one be?

OBJECTIVE: Progress feedback is often measured with generic instruments that measure common symptoms and generic aspects of functioning. The current study aims to explore the relative usefulness of disorder-specific measures. We hypothesized that disorder-specific instruments reveal more improvement than generic instruments and that the addition of disorder-specific instruments results in better treatment outcomes. METHOD: We used a cohort of 3419 patients with a depression. As generic measures, we used the BSI or the symptoms distress subscale of the OQ-45. In 946 patients, a specific instrument, the IDS-SR, was added. We compared mean change scores and percentages of clinical significant change. In a matched case control design, we analyzed whether the additional use of the IDS-SR resulted in better treatment outcomes. RESULTS: Mean change scores of both types of instruments were comparable. When comparing clinical significant change, agreement was moderate. We found better outcomes on the generic instruments when both a generic and a disorder-specific instrument were used. CONCLUSION: In individual treatment of depression, generic and disorder-specific instruments are not interchangeable. The additional use of disorder-specific instruments provides a more complete picture of the patient's progress than the use of a generic instrument alone. Clinical or methodological significance of this article: In outcome management often rather generic instruments are used, that do not address the specific symptoms of the primary diagnosis of patients. In daily practice clinicians do not always use the feedback on treatment progress, when they perceive the feedback as not specific or relevant enough. The current study aims to explore the relative usefulness of measures that focus on symptoms that characterize the primary diagnosis of patients with a depression compared to the generic measures. We used a large cohort of existing data of patients of several mental health care organizations that share an application for outcome measurement. First, we compared outcomes of generic instruments and a disorder-specific instrument of a subsample of patients with a depressive disorder that completed both kinds of instruments. Next, we applied a matched case control design to control for differences between patients and analyzed whether the additional use of disorder-specific instruments predicted better outcomes. With this methodology, we tried to optimize both the methodological quality as well as the clinical significance of our research.

Treating Tissue Factor-Positive Cancers with Antibody-Drug Conjugates That Do Not Affect Blood Clotting.

The primary function of tissue factor (TF) resides in the vasculature as a cofactor of blood clotting; however, multiple solid tumors aberrantly express this transmembrane receptor on the cell surface. Here, we developed anti-TF antibody-drug conjugates (ADC) that did not interfere with the coagulation cascade and benchmarked them against previously developed anti-TF ADCs. After screening an affinity-matured antibody panel of diverse paratopes and affinities, we identified one primary paratope family that did not inhibit conversion of Factor X (FX) to activated Factor X (FXa) and did not affect conversion of prothrombin to thrombin. The rest of the antibody panel and previously developed anti-TF antibodies were found to perturb coagulation to varying degrees. To compare the anticancer activity of coagulation-inert and -inhibitory antibodies as ADCs, a selection of antibodies was conjugated to the prototypic cytotoxic agent monomethyl auristatin E (MMAE) through a protease-cleavable linker. The coagulation-inert and -inhibitory anti-TF ADCs both killed cancer cells effectively. Importantly, the coagulation-inert ADCs were as efficacious as tisotumab vedotin, a clinical stage ADC that affected blood clotting, including in patient-derived xenografts from three solid tumor indications with a need for new therapeutic treatments-squamous cell carcinoma of the head and neck (SCCHN), ovarian, and gastric adenocarcinoma. Furthermore, a subset of the anti-TF antibodies could also be considered for the treatment of other diseases associated with upregulation of membranous TF expression, such as macular degeneration. Mol Cancer Ther; 17(11); 2412-26. ©2018 AACR.

Treatment results for severe psychiatric illness: which method is best suited to denote the outcome of mental health care?

BACKGROUND: The present study investigates the suitability of various treatment outcome indicators to evaluate performance of mental health institutions that provide care to patients with severe mental illness. Several categorical approaches are compared to a reference indicator (continuous outcome) using pretest-posttest data of the Health of Nation Outcome Scales (HoNOS). METHODS: Data from 10 institutions and 3189 patients were used, comprising outcomes of the first year of treatment by teams providing long-term care. RESULTS: Findings revealed differences between continuous indicators (standardized pre-post difference score ES and ΔT) and categorical indicators (SEM, JTRCI, JTCS, JTRCI&CS, JTrevised) on their ranking of institutions, as well as substantial differences among categorical indicators; the outcome according to the traditional JT approach was most concordant with the continuous outcome indicators. CONCLUSIONS: For research comparing group averages, a continuous outcome indicator such as ES or ΔT is preferred, as this best preserves information from the original variable. Categorical outcomes can be used to illustrate what is accomplished in clinical terms. For categorical outcome, the classical Jacobson-Truax approach is preferred over the more complex method of Parabiaghi et al. with eight outcome categories. The latter may be valuable in clinical practice as it allows for a more detailed characterization of individual patients.

Burden on caregivers of older patients with bipolar disorder.

OBJECTIVES: Available data suggest high burden on caregivers of patientswith bipolar disorder (BD), yet the well-being of patients with BD increasingly depends on family members, partners and close friends. Aspatients with BD get older, the need for informal care may shift. We aimed to describe the caregivers of older adults with BD (OABD) and explore what patients' and caregivers' characteristics are associated with caregiver burden. METHOD: Forty-seven caregivers of OABD were questioned about their perceived burden and depressive symptoms. Linear regression analyses were performed to examine the influence of various patients' and caregivers' characteristics on caregiver burden or depression. RESULTS: More than half of all caregivers experienced some degree of burden,and 6.4% reported depressive symptoms. The number of psychiatric admissions and social functioning were the only patients' characteristics associated with higher burden. Caregiver burden was significantly associated with caregiver's other obligations. None of the patient or caregiver characteristics was significantly associated with depression in caregivers of OABD. CONCLUSION: In OABD, even with few residual symptoms, more than half of all caregivers experience substantial burden. Future studies are needed to confirm if improving social functioning and preventing psychiatric hospitalizations decrease the burden on the caregivers of OABD.

Homesick: residential and care patterns in patients with severe mental illness.

BACKGROUND: Changes in the residential and care settings of patients with severe mental illness (SMI) are a concern because of the large variety of possible negative consequences. This study describes patterns of changes in the residential and care settings of SMI patients and explores associations between these changes, sociodemographics, and clinical characteristics. METHODS: From January 2006 to January 2012, all data relating to changes in residential and/or care setting by SMI patients (N = 262) were collected from electronic case files. Data covering psychopathology, substance use, and medication adherence were assessed in 2006. RESULTS: There were more changes in the residential than in the care setting. In 6 years, only 22% of our sample did not move, 23% changed residence once, 19% twice, 10% three times, and 26% four or more times. Substance use predicted changes of care and/or residential setting and rehospitalisation. The severity of negative symptoms predicted rehospitalisation and duration of hospitalisation. Disorganisation symptoms predicted the duration of hospitalisation. CONCLUSIONS: A majority of patients with SMI changed residential and/or care settings several times in 6 years. Patients with substance use or severe negative and disorganisation symptoms may need more intensive and customised treatment. Further research is needed to investigate prevention programmes for highly-frequent movers.

Pathways through care of severely mentally ill individuals experiencing multiple public crisis events: a qualitative description.

BACKGROUND: Patients experiencing severe mental illnesses (SMI) need continuing support and remain vulnerable in many domains. Crisis interventions and compulsory admissions are common, causing a huge burden on police, health workers, the community and patients. The aim of this retrospective case-file study is to determine profiles of SMI-patients and their pathways through care among those experiencing multiple public crisis events. METHODS: Data from a larger study of 323 SMI-patients in Amsterdam were used. These data were linked to data of the public mental health care (PMHC) in order to identify persons that experienced crisis interventions (CI's) between January 2004 and November 2012. The cut-off point for inclusion in the study population was set on three CI's, resulting in a group of 47 SMI-patients. PMHC and mental health care (MHC) data were linked in order to identify profiles in patterns of care. Qualitative content analysis was used to gather and analyze chronological timelines. RESULTS: Three profiles were identified: SMI-patients with CI's during continuous MHC, SMI-patients with CI's after discharge and SMI-patients with CI's during unstable MHC. For each profile events prior to, during and after a CI were identified. CONCLUSIONS: PMHC and MHC can possibly identify cases with a high risk of CI's and predict these events based on the results of this study. CI's seem inevitable for a group of SMI-patients in care but they do not only require acute psychiatric care. The collaboration between MHC, PMHC and police could be further developed in a quick and effective triage in order to tackle the complexity of problems of the SMI-patients.

Victimisation in adults with severe mental illness: prevalence and risk factors.

BACKGROUND: Patients with a severe mental illness (SMI) are more likely to experience victimisation than the general population. AIMS: To examine the prevalence of victimisation in people with SMI, and the relationship between symptoms, treatment facility and indices of substance use/misuse and perpetration, in comparison with the general population. METHOD: Victimisation was assessed among both randomly selected patients with SMI (n = 216) and the general population (n = 10 865). RESULTS: Compared with the general population, a high prevalence of violent victimisation was found among the SMI group (22.7% v. 8.5%). Compared with out-patients and patients in a sheltered housing facility, in-patients were most often victimised (violent crimes: 35.3%; property crimes: 47.1%). Risk factors among the SMI group for violent victimisation included young age and disorganisation, and risk factors for property crimes included being an in-patient, disorganisation and cannabis use. The SMI group were most often assaulted by someone they knew. CONCLUSIONS: Caregivers should be aware that patients with SMI are at risk of violent victimisation. Interventions need to be developed to reduce this vulnerability.

Antibody-Drug Conjugates (ADCs) Derived from Interchain Cysteine Cross-Linking Demonstrate Improved Homogeneity and Other Pharmacological Properties over Conventional Heterogeneous ADCs.

Conventional antibody-drug conjugates (ADCs) are heterogeneous mixtures of chemically distinct molecules that vary in both drugs/antibody (DAR) and conjugation sites. Suboptimal properties of heterogeneous ADCs have led to new site-specific conjugation methods for improving ADC homogeneity. Most site-specific methods require extensive antibody engineering to identify optimal conjugation sites and introduce unique functional groups for conjugation with appropriately modified linkers. Alternative nonrecombinant methods have emerged in which bifunctional linkers are utilized to cross-link antibody interchain cysteines and afford ADCs containing four drugs/antibody. Although these methods have been shown to improve ADC homogeneity and stability in vitro, their effect on the pharmacological properties of ADCs in vivo is unknown. In order to determine the relative impact of interchain cysteine cross-linking on the therapeutic window and other properties of ADCs in vivo, we synthesized a derivative of the known ADC payload, MC-MMAF, that contains a bifunctional dibromomaleimide (DBM) linker instead of a conventional maleimide (MC) linker. The DBM-MMAF derivative was conjugated to trastuzumab and a novel anti-CD98 antibody to afford ADCs containing predominantly four drugs/antibody. The pharmacological properties of the resulting cross-linked ADCs were compared with analogous heterogeneous ADCs derived from conventional linkers. The results demonstrate that DBM linkers can be applied directly to native antibodies, without antibody engineering, to yield highly homogeneous ADCs via cysteine cross-linking. The resulting ADCs demonstrate improved pharmacokinetics, superior efficacy, and reduced toxicity in vivo compared to analogous conventional heterogeneous ADCs.

CD39 is a promising therapeutic antibody target for the treatment of soft tissue sarcoma.

Soft tissue sarcoma (STS) is a heterogenous tumor arising from the embryonic mesoderm represented by approximately 50 histological subtypes. Effective therapeutic intervention is lacking for recurrent, late stage and metastatic disease. CD39, a cell-surface ectonucleotidase, has previously been shown to be upregulated in hematological malignancies and various epithelial tumors, but not in STS. Here, we show by mass spectrometry and immunohistochemistry that CD39 is highly expressed in primary patient sarcoma samples. Moreover, CD39 nucleotidase activity is enhanced in fibrosarcoma compared with normal control cells. We demonstrate that an inhibitory monoclonal anti-CD39 antibody, abrogates CD39 enzymatic activity significantly and prolongs survival in a lethal metastatic patient-derived sarcoma model. Taken together, the data suggest CD39 is a novel therapeutic target for the treatment of STS.

Quantitative assessment of antibody internalization with novel monoclonal antibodies against Alexa fluorophores.

Antibodies against cell surface antigens may be internalized through their specific interactions with these proteins and in some cases may induce or perturb antigen internalization. The anti-cancer efficacy of antibody-drug conjugates is thought to rely on their uptake by cancer cells expressing the surface antigen. Numerous techniques, including microscopy and flow cytometry, have been used to identify antibodies with desired cellular uptake rates. To enable quantitative measurements of internalization of labeled antibodies, an assay based on internalized and quenched fluorescence was developed. For this approach, we generated novel anti-Alexa Fluor monoclonal antibodies (mAbs) that effectively and specifically quench cell surface-bound Alexa Fluor 488 or Alexa Fluor 594 fluorescence. Utilizing Alexa Fluor-labeled mAbs against the EphA2 receptor tyrosine kinase, we showed that the anti-Alexa Fluor reagents could be used to monitor internalization quantitatively over time. The anti-Alexa Fluor mAbs were also validated in a proof of concept dual-label internalization assay with simultaneous exposure of cells to two different mAbs. Importantly, the unique anti-Alexa Fluor mAbs described here may also enable other single- and dual-label experiments, including label detection and signal enhancement in macromolecules, trafficking of proteins and microorganisms, and cell migration and morphology.

Criminal victimisation in people with severe mental illness: a multi-site prevalence and incidence survey in the Netherlands.

BACKGROUND: Although crime victimisation is as prevalent in psychiatric patients as crime perpetration (and possibly more so), few European figures for it are available. We therefore assessed its one-year prevalence and incident rates in Dutch severely mentally ill outpatients, and compared the results with victimisation rates in the general population. METHOD: This multisite epidemiological survey included a random sample of 956 adult severely mentally ill outpatients. Data on victimisation were obtained using the victimisation scale of the Dutch Crime and Victimisation Survey, which assesses crime victimisation over the preceding 12 months. Comparison data were derived from the nationwide survey on safety and victimisation in the Netherlands. Prevalence and incident rates were weighted for sex, age, ethnicity and socioeconomic status, and compared with a general population sample matched by region (N = 38,227). RESULTS: In the past year, almost half of the severely mentally ill outpatients (47%) had been victim of a crime. After control for demographic differences, prevalence rates of overall and specific victimisation measures were significantly higher in severely mentally ill outpatients than in the general population. The relative rates were especially high for personal crimes such as violent threats (RR = 2.12, 95% CI: 1.72-2.61), physical assaults (RR = 4.85, 95% CI: 3.69-6.39) and sexual harassment and assaults (RR = 3.94, 95% CI: 3.05-5.09). In concordance, severely mentally ill outpatients reported almost 14 times more personal crime incidents than persons from the general population (IRR = 13.68, 95% CI: 12.85-14.56). CONCLUSION: Crime victimisation is a serious problem in Dutch severely mentally ill outpatients. Mental-healthcare institutions and clinicians should become aware of their patients' victimisation risk, and should implement structural measures to detect and prevent (re-)victimisation.

Monoclonal antibody binding-site diversity assessment with a cell-based clustering assay.

The diversity of a panel of antibodies that target a specific antigen can be established in various assay formats. In conventional epitope binning assays purified antibodies are tested in a pairwise manner: two antibodies that compete with each other for binding to an antigen are grouped into the same cluster or bin, while they are assigned to two different clusters when they do not compete. Here we present a high through put assay that enables grouping of crude hybridoma supernatants without a need for antibody purification. In addition, the assay does not require recombinant protein, because it is conducted on cells that express the antigen of interest. Hence, one can use the antibody-clustering assay for cell surface proteins that are not amenable to purification. Heavy chain variable region (VH) sequencing shows that VH composition within clusters is conserved. Finally, the assay is in good agreement with a conventional epitope binning assay with purified antigen.

Psychometric properties of the Dutch version of the self-sufficiency matrix (SSM-D).

Measuring treatment outcomes can be challenging in patients who experience multiple interlinked problems, as is the case in public mental health care (PMHC). This study describes the development and psychometric properties of a Dutch version of the self-sufficiency matrix (SSM-D), an instrument that measures outcomes and originates from the US. In two different settings, clients were rated using the SSM-D in combination with the Health of the Nation Outcome Scales (HoNOS) and the Camberwell assessment of need short appraisal schedule (CANSAS). The results provided support for adequate psychometric properties of the SSM-D. The SSM-D had a solid single factor structure and internal consistency of the scale was excellent. In addition, convergent validity of the SSM-D was indicated by strong correlations between HoNOS and CANSAS, as well as between several subdomains. Further research is needed to establish whether the results presented here can be obtained in other PMHC settings.

Victimization of patients with severe psychiatric disorders: prevalence, risk factors, protective factors and consequences for mental health. A longitudinal study.

BACKGROUND: Victimization among people with a Severe Mental Illness is a common phenomenon. The objectives of this study proposal are: to delineate the extent and kind of victimization in a representative sample of chronic psychiatric patients; to contribute to the development and validation of a set of instruments registering victimization of psychiatric patients; to determine risk factors and protective factors; and to gain insight into the possible consequences of victimization. METHODS/DESIGN: An extensive data set of 323 patients with Sever Mental Illness (assessed 4 years ago) is used. In 2010 a second measurement will be performed, enabling longitudinal research on the predictors and consequences of victimization. DISCUSSION: The consequences of (re)victimization have barely been subjected to analysis, partially due to the lack of a comprehensive, conceptual model for victimization. This research project will contribute significantly to the scientific development of the conceptual model of victimization in chronic psychiatric patients.

Paracrine Hedgehog signaling in cancer.

Ligand-dependent and ligand-independent activation of the Hedgehog (Hh) signaling pathway is involved in tumorigenesis. The importance of mutations in Hh pathway components leading to constitutive signaling has been well established in basal cell carcinoma and medulloblastoma. However, the role of ligand-driven Hh pathway activation in cancer remains to be established. Three recent articles support a model in which, in the absence of mutations in the Hh pathway, Hh ligands expressed by a subset of epithelial cancers, including colon, pancreatic, and ovarian cancer, promote tumor growth indirectly by activating Hh signaling in the surrounding stroma, which, in turn, provides a more favorable environment for tumor growth. These data have important implications for the use of Hh pathway inhibitors currently in development and for selection of tumors likely to respond to such inhibitors.

The mammalian Cos2 homolog Kif7 plays an essential role in modulating Hh signal transduction during development.

The Hedgehog (Hh) signaling pathway regulates development in animals ranging from flies to humans. Although its framework is conserved, differences in pathway components have been reported. A kinesin-like protein, Costal2 (Cos2), plays a central role in the Hh pathway in flies. Knockdown of a zebrafish homolog of Cos2, Kif7, results in ectopic Hh signaling, suggesting that Kif7 acts primarily as a negative regulator of Hh signal transduction. However, in vitro analysis of the function of mammalian Kif7 and the closely related Kif27 has led to the conclusion that neither protein has a role in Hh signaling. Using Kif7 knockout mice, we demonstrate that mouse Kif7, like its zebrafish and Drosophila homologs, plays a role in transducing the Hh signal. We show that Kif7 accumulates at the distal tip of the primary cilia in a Hh-dependent manner. We also demonstrate a requirement for Kif7 in the efficient localization of Gli3 to cilia in response to Hh and for the processing of Gli3 to its repressor form. These results suggest a role for Kif7 in coordinating Hh signal transduction at the tip of cilia and preventing Gli3 cleavage into a repressor form in the presence of Hh.

Artemis and nonhomologous end joining-independent influence of DNA-dependent protein kinase catalytic subunit on chromosome stability.

Deficiency in both ATM and the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) is synthetically lethal in developing mouse embryos. Using mice that phenocopy diverse aspects of Atm deficiency, we have analyzed the genetic requirements for embryonic lethality in the absence of functional DNA-PKcs. Similar to the loss of ATM, hypomorphic mutations of Mre11 (Mre11(ATLD1)) led to synthetic lethality when juxtaposed with DNA-PKcs deficiency (Prkdc(scid)). In contrast, the more moderate DNA double-strand break response defects associated with the Nbs1(DeltaB) allele permitted viability of some Nbs1(DeltaB/DeltaB) Prkdc(scid/scid) embryos. Cell cultures from Nbs1(DeltaB/DeltaB) Prkdc(scid/scid) embryos displayed severe defects, including premature senescence, mitotic aberrations, sensitivity to ionizing radiation, altered checkpoint responses, and increased chromosome instability. The known functions of DNA-PKcs in the regulation of Artemis nuclease activity or nonhomologous end joining-mediated repair do not appear to underlie the severe genetic interaction. Our results reveal a role for DNA-PKcs in the maintenance of S/G(2)-phase chromosome stability and in the induction of cell cycle checkpoint responses.