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Many prostate cancer patients also suffer from depression, which can decrease their life satisfaction and also impede recovery from their cancer. This study described the network structure of depressive symptomatology in prostate cancer patients, with a view to providing suggestions for clinical interventions for depressed patients.Using a cross-sectional design, 555 prostate cancer patients completed the Patient Health Questionnaire-9 (PHQ-9).Network analysis and multidimensional scaling indicated that anhedonia was the most central symptom for these men, and that several sets of depression symptoms were closely associated with each other. These included anhedonia-depressed mood; sleeping problems-fatigue/lethargy; and suicidal ideation-low self-worth-depressed mood. Other depression symptoms such as appetite problems, concentration problems, and motor problems, were less well-related with the remainder of the network. Patients receiving treatment for reocurring prostate cancer (PCa) had significantly higher PHQ9 scores than patients undergoing their initial treatment, but no major differences in their network structures. Implications for clinical practice were derived from the relationships between individual depression symptoms and the overall depression network by examining node predictability.The use of total depression scores on an inventory does not reflect the underlying network structure of depression in PCa patients. Identification and treatment of the central symptom of anhedonia in PCa patients suggests the need to adopt specific therapies that are focussed upon this symptom.

Supplementary Figure S1. Analytical characterization of INT016-ADCs; Supplementary Figure S2. Validation of B7-H4 antibodies for use in IHC; Supplementary Figure S3. Relationship between B7-H4 expression level and heterogeneity in the studied human triple-negative cohort; Supplementary Figure S4. Binding of antibody INT016 to HEK 293 cells stably expressing human, mouse, or cynomolgus monkey B7-H4; Supplementary Figure S5. Binding of antibody INT016 to human breast cancer cell lines and to HT29 cells stably expressing human B7-H4; Supplementary Figure S6. Internalization and lysosomal trafficking of INT016; Supplementary Figure S7. In vitro stability of INT016-ADCs in mouse and cynomolgus monkey serum; Supplementary Figure S8. In vitro cytotoxicity of INT016-ADCs in HT29 and HT29-huB7-H4 isogenic cell lines; Supplementary Figure S9. In vivo efficacy of INT016-ADCs in the MX-1 xenograft model; Supplementary Figure S10. Growth rate analysis of MX-1 in vivo efficacy study comparing INT016-ADCs from days 14–40.

Surface plasmon resonance; In vitro cytotoxicity assays; In vitro proliferation assays; Internalization assays; Subcellular localization microscopy; Immunoblotting; Statistical analysis

Supplementary Table S1. Primary antibodies used for immunoblotting; Supplementary Table S2. Characterization of ADCs evaluated in this study; Supplementary Table S3. B7-H4 expression in human normal tissue; Supplementary Table S4. Binding affinity of anti-B7-H4 antibody INT016 Fab to human, cynomolgus monkey, and mouse B7-H4, measured by surface plasmon resonance.

Purpose:We evaluated the activity of AZD8205, a B7-H4–directed antibody–drug conjugate (ADC) bearing a novel topoisomerase I inhibitor (TOP1i) payload, alone and in combination with the PARP1-selective inhibitor AZD5305, in preclinical models.Experimental Design:IHC and deep-learning–based image analysis algorithms were used to assess prevalence and intratumoral heterogeneity of B7-H4 expression in human tumors. Several TOP1i-ADCs, prepared with Val-Ala or Gly–Gly–Phe–Gly peptide linkers, with or without a PEG8 spacer, were compared in biophysical, in vivo efficacy, and rat toxicology studies. AZD8205 mechanism of action and efficacy studies were conducted in human cancer cell line and patient-derived xenograft (PDX) models.Results:Evaluation of IHC-staining density on a per-cell basis revealed a range of heterogeneous B7-H4 expression across patient tumors. This informed selection of bystander-capable Val-Ala–PEG8–TOP1i payload AZ14170133 and development of AZD8205, which demonstrated improved stability, efficacy, and safety compared with other linker–payload ADCs. In a study of 26 PDX tumors, single administration of 3.5 mg/kg AZD8205 provided a 69% overall response rate, according to modified RECIST criteria, which correlated with homologous recombination repair (HRR) deficiency (HRD) and elevated levels of B7-H4 in HRR-proficient models. Addition of AZD5305 sensitized very low B7-H4–expressing tumors to AZD8205 treatment, independent of HRD status and in models representing clinically relevant mechanisms of PARPi resistance.Conclusions:These data provide evidence for the potential utility of AZD8205 for treatment of B7-H4–expressing tumors and support the rationale for an ongoing phase 1 clinical study (NCT05123482).See related commentary by Pommier and Thomas, p. 991

The management of Hodgkin Lymphoma (HL) has undergone significant changes in recent years. Due to the predilection of HL to affect younger patients, balancing cure and treatment related morbidity is a constant source of concern, for physicians and patients alike. PET adapted therapy has been developed for both early and advanced stage HL to try and improve the outcome of treatment, whilst minimising toxicities. The aim of this review is to digest the plethora of studies recently conducted and provide some clear, evidence-based practice statements to simplify the management HL. This article is protected by copyright. All rights reserved.

Introduction:. Prostate-specific antigen (PSA) kinetic patterns during neoadjuvant androgen deprivation therapy have been shown to predict unfavorable long-term outcomes. Objective:. To investigate the effect of testosterone escape (TE) on these kinetic patterns, as this had not been previously reported. Methods:. There were 50 consecutive prostate cancer patients who received 6 months of triptorelin prior to definitive radiotherapy (RT). Testosterone and PSA levels were measured at baseline and every 6 weeks. Clinical factors were tested for their ability to predict for TE and unfavorable PSA kinetic patterns. The effects of TE, at both 1.7 and 0.7 nmol/L levels, were analyzed. Results:. TE occurred in at least one reading for 14% and 34% of the patients at the 1.7 and 0.7 nmol/L levels, respectively. No baseline factors predicted TE. The median PSA halving time was 25 days and the median pre-RT PSA level was 0.55 ng/mL. The only factor significantly associated with a higher pre-RT PSA level was a higher baseline PSA level. The only factor that significantly predicted a longer PSA halving time was TE at the 1.7 nmol/L level. Conclusions:. TE and higher baseline PSA levels may adversely affect PSA kinetics and other outcomes for patients undergoing neoadjuvant hormone therapy prior to radiotherapy. Studies investigating the tailoring of neoadjuvant therapy by extending the duration in those patients with a higher baseline PSA level or by the addition of anti-androgens in those demonstrating TE, should be considered.

Aims Perineural invasion (PNI) by prostatic adenocarcinoma is debated as a prognostic parameter. This study investigates the prognostic predictive value of PNI in a series of patients with locally advanced prostate cancer treated with radiotherapy and androgen deprivation using 10 years outcome data from the TROG 03.04 RADAR trial. Methods Diagnostic prostate biopsies from 976 patients were reviewed and the presence of PNI noted. Patients were followed for 10 years according to the trial protocol or until death. The primary endpoint for the study was time to bone metastasis. Secondary endpoints included time to soft tissue metastasis, transition to castration resistance, prostate cancer-specific mortality and all-cause mortality. Results PNI was detected in 449 cases (46%), with 234 cases (24%) having PNI in more than one core. The presence of PNI was significantly associated with higher ISUP grade, clinical T staging category, National Comprehensive Cancer Network risk group, and percent positive biopsy cores. The cumulative probability of bone metastases according to PNI status was significant over the 10 years follow-up interval of the study (log-rank testP < 0.0001). PNI was associated with all endpoints on univariable analysis. After adjusting for baseline clinicopathological and treatment factors, bone metastasis was the only endpoint in which PNI retained its prognostic significance (hazard ratio 1.42, 95% confidence interval 1.05-1.92,P = 0.021). Conclusions The association between PNI and the development of bone metastases supports the inclusion of this parameter as a component of the routine histology report. Further this association suggests that evaluation of PNI may assist in selecting those patients who should be monitored more closely during follow-up.

While alpha-blockers are commonly used to reduce lower urinary tract symptoms in prostate cancer patients receiving radiotherapy, their impact on response to radiotherapy remains unknown. Therefore, this pilot study aimed to retrospectively determine if alpha-blockers use, influenced response to radiotherapy for localised prostate cancer. In total, 303 prostate cancer patients were included, consisting of 84 control (alpha-blocker naïve), 72 tamsulosin and 147 prazosin patients. The main outcomes measured were relapse rates (%), time to biochemical relapse (months) and PSA velocity (ng/mL/year). Recurrence free survival was calculated using Kaplan-Meier analysis. Prazosin significantly reduced biochemical relapse at both two and five-years (2.72%, 8.84%) compared to control (22.61%, 34.52%). Recurrence free survival was also significantly higher in the prazosin group. This remained after multivariable analysis (HR: 0.09, 95% CI: 0.04–0.26, p

In 2016, during a visit to the Chemical Heritage Foundation in Philadelphia (now the Science History Institute), I and Carin Berkowitz, then director of the Beckman Center for the History of Chemis...

To clarify the relative effects of duration of androgen suppression (AS) and radiation dose escalation (RDE) on distant progression (DP) in men with locally advanced prostate cancer.Participants with locally advanced prostate cancer in the TROG 03.04 RADAR trial were randomized to 6 or 18 months AS ± 18 months zoledronic acid (Z). The trial incorporated a RDE program by stratification at randomization and dosing options were 66, 70, or 74 Gy external beam radiation therapy (EBRT), or 46 Gy EBRT plus high-dose-rate brachytherapy boost (HDRB). The primary endpoint for this study was distant progression (DP). Secondary endpoints included local progression, bone progression, prostate cancer-specific mortality and all-cause mortality. Effect estimates for AS duration and RDE were derived using Fine and Gray competing risk models adjusting for use of Z, age, tumor stage, Gleason grade group, prostate-specific antigen, and treatment center. Cumulative incidence at 10 years was estimated for each RDE group.A total of 1051 out of 1071 randomized subjects were eligible for inclusion in this analysis. Compared with 6 months AS, 18 months AS significantly reduced DP independently of radiation dose (subhazard ratio 0.70; 95% confidence interval [CI], 0.56-0.87; P = .002). No statistically significant interaction between effect of AS duration and RT dose was observed (Wald test P = .76). In subgroup analyses, DP was significantly reduced by the longer duration of AS in the 70 Gy and HDRB groups but not in the 66 Gy and 74 Gy. Compared with 70 Gy, HDRB significantly reduced DP (subhazard ratio 0.68 [95% CI, 0.57-0.80]; P.0001) independently of AS duration. At 10 years, adjusted cumulative incidences were 26.1% (95% CI, 18.9%-33.2%), 26.7% (22.9%-30.6%), 24.9% (20.0%-29.8%) and 19.7% (15.5%-23.8%) for DPs in the respective radiation dose groups.Compared with 6 months AS, 18 months AS reduced DP independently of radiation dose. Men treated with HDRB gained a significant benefit from a longer duration of AS. Evidence of improved oncologic outcomes for HDRB compared with dose-escalated EBRT needs to be confirmed in a randomized trial.

BACKGROUND: Exome sequencing is emerging as a first-line diagnostic method in some clinical disciplines, but its usefulness has yet to be examined for most constitutional disorders in adults, including chronic kidney disease, which affects more than 1 in 10 persons globally. METHODS: We conducted exome sequencing and diagnostic analysis in two cohorts totaling 3315 patients with chronic kidney disease. We assessed the diagnostic yield and, among the patients for whom detailed clinical data were available, the clinical implications of diagnostic and other medically relevant findings. RESULTS: In all, 3037 patients (91.6%) were over 21 years of age, and 1179 (35.6%) were of self-identified non-European ancestry. We detected diagnostic variants in 307 of the 3315 patients (9.3%), encompassing 66 different monogenic disorders. Of the disorders detected, 39 (59%) were found in only a single patient. Diagnostic variants were detected across all clinically defined categories, including congenital or cystic renal disease (127 of 531 patients [23.9%]) and nephropathy of unknown origin (48 of 281 patients [17.1%]). Of the 2187 patients assessed, 34 (1.6%) had genetic findings for medically actionable disorders that, although unrelated to their nephropathy, would also lead to subspecialty referral and inform renal management. CONCLUSIONS: Exome sequencing in a combined cohort of more than 3000 patients with chronic kidney disease yielded a genetic diagnosis in just under 10% of cases. (Funded by the National Institutes of Health and others.)

The cell-surface glycoprotein B7-H4 is overexpressed in a range of solid tumors including breast cancer, ovarian serous carcinoma, endometrial carcinoma, and cholangiocarcinoma, yet has limited expression in normal tissue, making it an attractive target for an antibody-drug conjugate (ADC). This presentation describes for the first time the development of AZD8205, a B7-H4 targeted ADC incorporating a novel topoisomerase 1 inhibitor (TOP1i) linker-warhead, AZ’0133 which was designed to exploit the full potential of B7-H4 as an ADC target. Initially, we investigated a series of more than 35 TOP1i compounds as warheads and achieved activity in a clinically relevant nM range. We further optimized the conjugation site and chemistry to reduce the potential for aggregation while maintaining potency, overcoming major synthetic challenges to deliver a robust synthetic route amenable to scale-up. Finally, with a series of optimized linker-warheads, we explored the impact of linker-warhead design on ADC hydrophobicity, stability, efficacy, pharmacokinetics and tolerability culminating in the development of AZD8205. The primary mechanism of action of AZD8205 is intracellular delivery of the TOP1i warhead to B7-H4 positive cells, leading to DNA damage and apoptotic cell death. AZD8205 drove bystander killing of target negative cells in mixed cultures in vitro, which is further supported by robust antitumor activity observed in in vivo studies with patient-derived xenograft (PDX) tumors with heterogeneous target expression, representing multiple tumor indications. In a study of 26 human TNBC PDX tumors, a single IV administration of 3.5 mg/kg AZD8205 provided an overall response rate of 69% (tumor regression of 30% or greater from baseline) and complete responses observed in 9/26 (36%) of models. To understand the biology underlying antitumor response, we conducted a multiparametric analysis including genomics, proteomics and computational pathology and found that deeper antitumor activity was observed in models with elevated B7-H4 expression as well as in models with defects in DNA damage repair (DDR). To further exploit the DNA damage elicited by the TOP1i warhead, we examined combinations of AZD8205 with small molecules, including a novel PARP1 selective inhibitor, in a BRCA wild type MDA-MB-468 model. These data suggest that AZD8205 is a promising therapeutic candidate for the treatment of B7-H4 positive solid tumors. A first in human phase 1 study in patients with advanced solid tumors is currently ongoing (NCT05123482). Citation Format: Krista Kinneer, Niall J. Dickinson, Luke Masterson, Thais Cailleau, Ian Hutchinson, Balakumar Vijayakrishnan, Nazzareno Dimasi, R. James Christie, Mary McFarlane, Kathryn Ball, Arthur Lewis, Sofia Koch, Lee Brown, Yue Huang, Anton I. Rosenbaum, Jiaqi Yuan, Si Mou, Noel R. Monks, Jon Chesebrough, Ravinder Tammali, Judith Anderton, Darrin Sabol, Frances Anne Tosto, Philipp Wortmann, Zachary A. Cooper, Pauline Ryan, John Hood, Carlos Fernandez Teruel, Carlos Serra Traynor, Andy Pike, Michael Davies, Elisabetta Leo, Kimberly Cook, Nadia Luheshi, Philip W. Howard, Puja Sapra. Discovery and first disclosure of AZD8205, a B7-H4-targeted antibody-drug conjugate utilizing a novel topoisomerase I linker-warhead [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1765.

Background Biliary atresia (BA), a fibrosing disorder of the developing biliary tract leading to liver failure in infancy, has an elevated incidence in indigenous New Zealand (NZ) Māori. We investigated a high rate of BA in a group of children (n = 12) belonging to a single Māori iwi (or ‘tribe’, related through a remote ancestor). Methods Population and geographical data was used to estimate the rate of BA in Māori sub-groups, and a pedigree linking most of the affected children was constructed from oral and documented history. Array genotyping was used to examine hypotheses about the inheritance of a possible genetic risk factor, and the history of the affected population, and Exome Sequencing to search for candidate genes. Results Most of these affected children (n = 7) link to a self-reported pedigree and carry a 50-fold increase in BA risk over unrelated Māori (χ2 = 296P 0.63). Genome-wide quantitation of intervals of contiguous, homozygous-by-state markers reached a similar conclusion (F (2,399) = 1.99, P = 0.138). Principal component analysis and investigation with STRUCTURE found no evidence of increased allele frequency of either a recessive variant, or additive, low-risk variants due to reproductive isolation. To identify candidate causal factors, Exome Sequencing datasets were scrutinised for shared rare coding variants across 8 affected individuals. No rare, non-synonymous, phylogenetically conserved variants were common to 6 or more affected children. Conclusion The substantially elevated risk for development of BA in this subgroup could be mediated by genetic factors, but the iwi exhibits no properties indicative of recent or remote reproductive isolation. Resolution of any risk loci may rely on extensive genomic sequencing studies in this iwi or investigation of other mechnaisms such as copy number variation. Electronic supplementary material The online version of this article (10.1186/s12920-018-0440-0) contains supplementary material, which is available to authorized users.

Men who suffer from prostate cancer (PCa) need to make important decisions regarding their treatment options. There is some evidence that these men may suffer from sleep difficulties due to their cancer or its diagnosis and treatment. Although sleep difficulties have been associated with cognitive depression in other samples of men, they have not been examined in PCa patients, despite the importance of decision-making for these men. This study was designed to investigate the association between sleep difficulties and cognitive depression in PCa patients. A sample of 96 PCa patients completed a background questionnaire, the Zung Self-Rating Depression Scale, and the Insomnia Severity Index. Comparison was made between sleep difficulty scores from before the patients received their diagnosis of PCa to the time of survey, allowing use of a “retrospective pretest” methodology. Just over 61% of the sample reported a deterioration in sleep quality, and this was significantly associated with cognitive depression ( r = .346, p = .007). At the specific symptom level, having a clear mind significantly contributed to the variance in difficulty falling asleep (R2 change = .140, F for change = 9.298, p = .003). Sleeping difficulties, particularly falling asleep, are common and associated with depression-related to ability to think clearly in PCa patients. This has potentially adverse effects upon the ability of men with PCa to understand their treatment options and make decisions about them.

IntroductionAsthma risk is a complex interplay between genetic susceptibility and environment. Despite many significantly-associated common variants, the contribution of rarer variants with potentially greater effect sizes has not been as extensively studied. We present an exome-based study adopting 24,576 cases and 120,530 controls to assess the contribution of rare protein-coding variants to the risk of early-onset or all-comer asthma.MethodsWe performed case-control analyses on three genetic units: variant-, gene- and pathway-level, using sequence data from the Scandinavian Asthma Genetic Study and UK Biobank participants with asthma. Cases were defined as all-comer asthma (n=24,576) and early-onset asthma (n=5,962). Controls were 120,530 UK Biobank participants without reported history of respiratory illness.ResultsVariant-level analyses identified statistically significant variants at moderate-to-common allele frequency, including protein-truncating variants in FLG and IL33. Asthma risk was significantly increased not only by individual, common FLG protein-truncating variants, but also among the collection of rare-to-private FLG protein-truncating variants (p=6.8×10−7). This signal was driven by early-onset asthma and did not correlate with circulating eosinophil levels. In contrast, a single splice variant in IL33 was significantly protective (p=8.0×10−10), while the collection of remaining IL33 protein-truncating variants showed no class effect (p=0.54). A pathway-based analysis identified that protein-truncating variants in loss-of-function intolerant genes were significantly enriched among individuals with asthma.ConclusionsAccess to the full allele frequency spectrum of protein-coding variants provides additional clarity about the potential mechanisms of action for FLG and IL33. Beyond these two significant drivers, we detected a significant enrichment of protein-truncating variants in loss-of-function intolerant genes.

Despite recent advances in treatment, breast cancer remains the second-most common cause of cancer death among American women. A greater understanding of the molecular characteristics of breast tumors could ultimately lead to improved tumor-targeted treatment options, particularly for subsets of breast cancer patients with unmet needs. Using an unbiased genomics approach to uncover membrane-localized tumor-associated antigens (TAAs), we have identified glial cell line derived neurotrophic factor (GDNF) family receptor α 1 (GFRA1) as a breast cancer TAA. Immunohistochemistry (IHC) revealed that GFRA1 displays a limited normal tissue expression profile coupled with overexpression in specific breast cancer subsets. The cell surface localization as determined by fluorescence-activated cell sorting (FACS) and the rapid internalization kinetics of GFRA1 makes it an ideal target for therapeutic exploitation as an antibody-drug conjugate (ADC). Here, we describe the development of a pyrrolobenzodiazepine (PBD)-armed, GFRA1-targeted ADC that demonstrates cytotoxicity in GFRA1-positive cell lines and patient-derived xenograft (PDX) models. The safety profile of the rat cross-reactive GFRA1-PBD was assessed in a rat toxicology study to find transient cellularity reductions in the bone marrow and peripheral blood, consistent with known off-target effects of PBD ADC's. These studies reveal no evidence of on-target toxicity and support further evaluation of GFRA1-PBD in GFRA1-positive tumors.

To determine incidence and outcome of biliary atresia (BA) between ethnic groups in New Zealand (NZ), a retrospective review was undertaken of children with BA born between 2002 and 2014. Prioritized ethnicity was used to determine ethnicity and was compared to population data. Uni- and multivariate analyses were undertaken to determine demographic and biochemical factors associated with outcome. Overall incidence was 1 in 9181 (Māori 1 in 5285; European 1 in 16,228; P < 0.0001). Overall and transplant-free survival rates at 1, 2, and 5 years were 92%, 86%, 82% and 70%, 49%, 30% respectively with Māori having improved transplant-free survival (P < 0.05) despite European children undergoing Kasai earlier (49 vs 63 days). BA is more common in NZ than Europe and North America, which is attributable to a higher incidence in Māori but overall outcome is poorer. Māori have improved transplant-free survival compared to NZ European children but the reason is unknown.

Classification, phylogeography and the testing of evolutionary hypotheses rely on correct estimation of species phylogeny. Early molecular phylogenies often relied on mtDNA alone, which acts as a single linkage group with one history. Over the last decade, the use of multiple nuclear sequences has often revealed conflict among gene trees. This observation can be attributed to hybridization, lineage sorting, paralogy or selection. Here, we use 54 groups of fishes from 48 studies to estimate the degree of concordance between mitochondrial and nuclear gene trees in two ecological grades of fishes: marine and freshwater. We test the hypothesis that freshwater fish phylogenies should, on average, show more discordance because of their higher propensity for hybridization in the past. In keeping with this idea, concordance between mitochondrial and nuclear gene trees (as measured by proportion of components shared) is on average 50% higher in marine fishes. We discuss why this difference almost certainly results from introgression caused by greater historical hybridization among lineages in freshwater groups, and further emphasize the need to use multiple nuclear genes, and identify conflict among them, in estimation of species phylogeny.

Introduction: A common treatment for localized prostate cancer (PCa) is radiotherapy; however, effectiveness is hampered because of toxicities and tumor resistance. Cyclooxygenase-2 (COX-2) inhibitors have been identified as potential agents that could improve treatment outcomes and have demonstrated ability to increase the radiosensitivity of many human carcinomas. This retrospective human study aims to investigate the ability of COX-2 inhibitors, celecoxib, and meloxicam, to improve treatment outcomes after radiotherapy. Methods: Prostate Specific Antigen (PSA) data of eligible patients were obtained from Genesis Cancer Care, Southport, Australia. The primary outcome was the percentage of patients in each group that had reached biochemical relapse at two and five years after treatment. Secondary outcomes included time to biochemical relapse and PSA velocity. Results: At two and five years after treatment, both the celecoxib (6.7%, 18.3%) and meloxicam (0.0%, 18.9%) showed lower relapse rates than the control (8.6%, 31.0%). Although not statistically significant, these results are clinically significant. In addition, the two treatment groups were found to increase the time to relapse, 46.20 months for celecoxib and 54.15 months for meloxicam, compared with the control group, 35.53 months. A similar trend was shown for PSA velocity with both treatment groups demonstrating lower PSA velocities compared with control. Conclusions: This study provides further evidence to the potential for COX-2 inhibitors to address gaps in localizedz PCa treatment by demonstrating high clinical significance for the use of celecoxib and meloxicam. Further research should be conducted including larger retrospective studies and prospective studies to fully evaluate the benefits of COX-2 inhibitors in combination with radiotherapy for PCa. Keywords: Biochemical relapse, Celecoxib, COX-2, Meloxicam, Prostate cancer

Here, we describe a diene-containing noncanonical amino acid (ncAA) capable of undergoing fast and selective normal electron-demand Diels–Alder (DA) reactions following its incorporation into antibodies. A cyclopentadiene derivative of lysine (CpHK) served as the reactive handle for DA transformations and the substrate for genetic incorporation. CpHK incorporated into antibodies with high efficiency and was available for maleimide conjugation or self-reaction depending on position in the amino acid sequence. CpHK at position K274 reacted with the maleimide drug-linker AZ1508 at a rate of ~79 M(−1)s(−1) to produce functional antibody–drug conjugates (ADCs) in a one-step process. Incorporation of CpHK at position S239 resulted in dimerization, which covalently linked antibody heavy chains together. The diene ncAA described here is capable of producing therapeutic protein conjugates with clinically validated and widely available maleimide compounds, while also enabling proximity-based stapling through a DA dimerization reaction.

Since 1995, severe epidemics of cucumber mosaic virus (CMV) have occurred in select fields of tobacco (Nicotiana tabacum) and pepper (Capsicum annuum) in three counties in northern Florida. Yield losses greater than 50% have occurred in both crops. Baker and Zettler (1) identified the presence of CMV in one plant of tropical spiderwort (Commelina benghalensis) in an organic garden on the campus of the University of Florida 10 years ago. In addition, they infected tropical spiderwort and Asiatic dayflower (Commelina communis) with isolates of CMV. Since 1995, in one area of northern Alachua County, Asiatic dayflower has been found in abundance in and around some fields and found to be infected with CMV. Prior to this time, CMV had not been known to be epidemic in any crop in northern Florida. Also, commelinaceous weeds did not occur in such abundance in northern Florida. In Hamilton County, an epidemic of CMV occurred in one field of tobacco in 1997. Tropical spiderwort with viral-like symptoms was growing abundantly in that field. The symptoms in this weed included chlorotic ringspots and chevron-like line patterns. Light microscopy, with Azure A stain, revealed the presence of typical inclusions of CMV in pepper, tobacco, tropical spiderwort, and Asiatic dayflower. Symptomatic samples of the tobacco and the tropical spiderwort reacted in an immunodiffusion test with antiserum to a winged bean isolate of CMV (2). Extracts from tropical spiderwort (isolate 3603) were rubbed on squash. This isolate was thereafter maintained in squash (Cucurbita pepo cvs. Prelude II or Early Prolific Straightneck). Infected plants of both of these cultivars developed strong mosaic symptoms and were stunted. After passage through squash, the 3603 isolate induced mosaic in tobacco (cv. Burley 21). Some plants of the squash cultivars Destiny III and Liberator III, which have transgenic, coat protein-mediated resistance to CMV, developed restricted symptoms after inoculation with this isolate. CMV was recovered by back inoculation from symptomatic plants of these cultivars. Symptomless plants of tropical spiderwort transplanted from the field developed chlorotic ringspots and chevron-like line patterns following inoculation in the greenhouse with isolate 3603. Back inoculations to squash followed by immunodiffusion assays confirmed the presence of CMV in the inoculated tropical spiderwort plants but CMV was not detected in noninoculated control plants. This is the first report of tropical spiderwort being infected with CMV in a commercial situation in the United States. Because commelinaceous plants are well known to be excellent hosts of CMV (1), we believe that the increased presence of perennial, commelinaceous weeds is a factor contributing to the epidemics of CMV in northern Florida. References: (1) C. A. Baker and F. W. Zettler. Plant Dis. 72:513, 1988. (2) C. A. Ku-wite and D. E. Purcifull. Plant Dis. 66:1071, 1982.

Up to a quarter of all prostate cancer (PCa) patients suffer from clinically significant depression but treatments are inconsistent and short-lived in their efficacy. One possible reason could be that “male depression” is not adequately diagnosed by the criteria for major depressive disorder (MDD) used in many clinical settings. In response to this limitation, the Gotland Scale of Male Depression (GSMD) was developed to identify the extra symptoms of MDD in men. Although the factor structure of the GSMD has been reported in non-PCa samples, it has not been determined for this group of men. Two samples of PCa patients were recruited, 191 from Australia and 138 from the United Kingdom and all patients received the GSMD individually, plus a background questionnaire. Two-factor solutions were identified for each of the two samples. The Australian sample was characterized by changes in emotional and somatic function, followed by depressed mood. The U.K. sample exhibited the same two-factor solution but in reverse order of weighting. Targeted treatments for depression in PCa patients may benefit from identification of the loadings that individual patients have on these two GSMD factors so that specific clinical profiles and treatment needs may be based on this information about their depression.

Background Studies have identified many common genetic associations that influence renal function and all-cause CKD, but these explain only a small fraction of variance in these traits. The contribution of rare variants has not been systematically examined. Methods We performed exome sequencing of 3150 individuals, who collectively encompassed diverse CKD subtypes, and 9563 controls. To detect causal genes and evaluate the contribution of rare variants we used collapsing analysis, in which we compared the proportion of cases and controls carrying rare variants per gene. Results The analyses captured five established monogenic causes of CKD: variants in PKD1 , PKD2 , and COL4A5 achieved study-wide significance, and we observed suggestive case enrichment for COL4A4 and COL4A3 . Beyond known disease-associated genes, collapsing analyses incorporating regional variant intolerance identified suggestive dominant signals in CPT2 and several other candidate genes. Biallelic mutations in CPT2 cause carnitine palmitoyltransferase II deficiency, sometimes associated with rhabdomyolysis and acute renal injury. Genetic modifier analysis among cases with APOL1 risk genotypes identified a suggestive signal in AHDC1 , implicated in Xia–Gibbs syndrome, which involves intellectual disability and other features. On the basis of the observed distribution of rare variants, we estimate that a two- to three-fold larger cohort would provide 80% power to implicate new genes for all-cause CKD. Conclusions This study demonstrates that rare-variant collapsing analyses can validate known genes and identify candidate genes and modifiers for kidney disease. In so doing, these findings provide a motivation for larger-scale investigation of rare-variant risk contributions across major clinical CKD categories.

The thiol-maleimide linkage is widely used for antibody-drug conjugate (ADC) production; however, conjugation of maleimide-drugs could be improved by simplified procedures and reliable conjugate stability. Here, we report the evaluation of electron-rich and cyclic dienes that can be appended to antibodies and reacted with maleimide-containing drugs through the Diels-Alder (DA) reaction. Drug conjugation is fast and quantitative due to reaction acceleration in water, and the linkage is more stable in serum than in the corresponding thiol-maleimide adduct with the same drug. ADCs produced using the DA reaction (DAADCs) are effective in vitro and in vivo, demonstrating the utility of this reaction in producing effective biotherapeutics. Given the large number of commercially available maleimide compounds, this conjugation approach could be readily applied to the production of a wide range of antibody (or protein) conjugates.

The objective of the current study was to identify the patient-perceived “worst aspects” of their diagnostic and treatment processes for prostate cancer (PCa) so as to inform targeted interventions aimed at reducing patient anxiety and depression. Two hundred and fifty-two patients who had received their diagnoses less than 8 years ago answered a postal survey about (a) background information, (b) their own descriptions of the worst aspects of their diagnosis and treatment, and (c) their ratings of 13 aspects of that process for (i) how these aspects made them feel stressed, anxious, and depressed and (ii) how they affected their relationships with significant others. They also answered standardized scales of anxiety and depression. The worst aspects reported by patients were receiving the initial diagnosis of PCa, plus the unknown outcome of that diagnosis, because of the possibility of death, loss of quality of life and/or partner, and the shock of the diagnosis. The most common coping strategy was to “just deal with it,” but participants also thought that more information would help. Principal contributors to feeling stressed, anxious, and depressed were also the diagnosis itself, followed by surgery treatment effects. The aspects that most affected relationships were receiving the diagnosis and the side effects of hormone therapy. The identification of these specific worst aspects of the PCa experience provides a set of potential treatment and prevention “targets” for psychosocial care in PCa patients.

The Phoenicians emerged in the Northern Levant around 1800 BCE and by the 9th century BCE had spread their culture across the Mediterranean Basin, establishing trading posts, and settlements in various European Mediterranean and North African locations. Despite their widespread influence, what is known of the Phoenicians comes from what was written about them by the Greeks and Egyptians. In this study, we investigate the extent of Phoenician integration with the Sardinian communities they settled. We present 14 new ancient mitogenome sequences from pre-Phoenician (~1800 BCE) and Phoenician (~700–400 BCE) samples from Lebanon (n = 4) and Sardinia (n = 10) and compare these with 87 new complete mitogenomes from modern Lebanese and 21 recently published pre-Phoenician ancient mitogenomes from Sardinia to investigate the population dynamics of the Phoenician (Punic) site of Monte Sirai, in southern Sardinia. Our results indicate evidence of continuity of some lineages from pre-Phoenician populations suggesting integration of indigenous Sardinians in the Monte Sirai Phoenician community. We also find evidence of the arrival of new, unique mitochondrial lineages, indicating the movement of women from sites in the Near East or North Africa to Sardinia, but also possibly from non-Mediterranean populations and the likely movement of women from Europe to Phoenician sites in Lebanon. Combined, this evidence suggests female mobility and genetic diversity in Phoenician communities, reflecting the inclusive and multicultural nature of Phoenician society.

Contains fulltext : 153454.pdf (Publisher’s version ) (Open Access) Robinow syndrome (RS) is a phenotypically and genetically heterogeneous condition that can be caused by mutations in genes encoding components of the non-canonical Wnt signaling pathway. In contrast, germline mutations that act to increase canonical Wnt signaling lead to distinctive osteosclerotic phenotypes. Here, we identified de novo frameshift mutations in DVL1, a mediator of both canonical and non-canonical Wnt signaling, as the cause of RS-OS, an RS subtype involving osteosclerosis, in three unrelated individuals. The mutations all delete the DVL1 C terminus and replace it, in each instance, with a novel, highly basic sequence. We showed the presence of mutant transcript in fibroblasts from one individual with RS-OS and demonstrated unimpaired protein stability with transfected GFP-tagged constructs bearing a frameshift mutation. In vitro TOPFlash assays, in apparent contradiction to the osteosclerotic phenotype, revealed that the mutant allele was less active than the wild-type allele in the canonical Wnt signaling pathway. However, when the mutant and wild-type alleles were co-expressed, canonical Wnt activity was 2-fold higher than that in the wild-type construct alone. This work establishes that DVL1 mutations cause a specific RS subtype, RS-OS, and that the osteosclerosis associated with this subtype might be the result of an interaction between the wild-type and mutant alleles and thus lead to elevated canonical Wnt signaling.

Background: Due to the pressing need to understand the causal and associative factors of depression among prostate cancer (PCa) patients, a comprehensive research protocol for investigating depression in prostate cancer patients is suggested as a way of furthering the collection of data in consistent and informative ways. Methods: A detailed review of a range of predictors of, and buffers against, depression, plus methods of assessing depressive symptomatology and optimum time to collect data were used to develop a model for a comprehensive research protocol. Results: A model protocol is described that includes socioeconomic, genetic, endocrinal, immunological, physiological, psychological, relationship, and socioeconomic pathways to depression. In addition, methods of assessing depressive symptomatology are described, plus comorbidity of anxiety with depression, male depression, and the construct of Individual Burden of Illness for Depression. The need to collect multiple measures over time in o...

Thiosuccinimide-linked antibody-drug conjugates (ADCs) are susceptible to drug loss over time due to a retro-Michael reaction, which can be prevented by selecting stable conjugation positions or hydrolysis of the thiosuccinimide. Here, we investigate pyrrolobenzodiazepine (PBD) ADC drug-linkers equipped with N-phenyl maleimide functionality for stable thiol conjugation via thiosuccinimide hydrolysis. Two PBD drug-linker formats (enzyme-cleavable and non-cleavable) were evaluated following site-specific conjugation to an engineered cysteine incorporated at position T289, which is known to be unstable for N-alkyl maleimide conjugates. N-phenyl maleimide PBDs conjugated to antibodies with similar efficiencies as N-alkyl maleimide PBDs and enhanced thiosuccinimide hydrolysis for N-phenyl maleimide PBDs was confirmed by mass spectrometry, capillary isoelectric focusing, and a SYPRO Orange dye binding assay. All of the PBD ADCs were highly potent in vitro regardless of maleimide- or linker-type, exhibiting low pM EC50 values. Thiol conjugation to N-phenyl maleimide PBD minimized the retro-Michael reaction in both rat and mouse serum. However, cleavage of the valine-alanine dipeptide in mouse serum for ADCs containing cleavable drug-linker led to drug loss regardless of maleimide type, which impacted ADC potency in tumor growth inhibition studies that were conducted in mouse models. Therapeutic improvement in mouse tumor models was realized for ADCs prepared with non-cleavable PBD drug-linkers that were conjugated through N-phenyl maleimide, where a stronger tumor growth inhibition (TGI) response was achieved when compared to the analogous N-alkyl maleimide drug-linker ADC. Altogether, our findings highlight the stability and efficacy benefits of N-phenyl maleimide functionality for ADCs that are produced with thiol-maleimide conjugation chemistry.

Antibodies and antigen-binding fragments (Fabs) can be used to modify the surface of nanoparticles for enhanced target binding. In our previous work, site-specific conjugation of Fabs to polymeric micelles using conventional methods was limited to approximately 30% efficiency, possibly due to steric hindrance related to macromolecular reactants. Here, we report a new method that enables conjugation of Fabs onto a micelle surface in a controlled manner with up to quantitative conversion of nanoparticle reactive groups. Variation of (i) PEG spacer length in a heterofunctionalized cross-linker and (ii) Fab/polymer feed ratios resulted in production of nanoparticles with a range of Fab densities on the surface up to the theoretical maximum value. The biological impact of variable Fab density was evaluated in vitro with respect to cell uptake and cytotoxicity of a drug-loaded (SN38) targeted polymeric micelle bearing anti-EphA2 Fabs. Fab conjugation increased cell uptake and potency compared with non-targeted micelles, although a Fab density of 60% resulted in decreased uptake and potency of the targeted micelles. Altogether, our findings demonstrate that conjugation strategies can be optimized to allow control of Fab density on the surface of nanoparticles and also that Fab density may need to be optimized for a given cell-surface target to achieve the highest bioactivity.

Background: Previous anxiety and depression can influence current anxiety and depression, and psychological resilience may be a buffer against current anxiety and depression. However, despite the relevance of these two sets of predictors of anxiety and depression, and their potential in treatment planning, no reports have been published on their role in post-diagnosis anxiety and depression among prostate cancer (PCa) patients. Therefore, this study aimed to determine the roles of these predictors in a sample of men with prostate cancer. Method: Retrospective self-reports of anxiety and depression for how they were before receiving their diagnosis of PCa and at the time of testing after their diagnosis were collected from 425 PCa patients across two states of Australia. In addition, self-reported current psychological resilience was measured. Results: Although there was a significant increase in depression and a nonsignificant increase in anxiety from before their diagnosis, for the whole sample,...

Dr. Joseph Black had at one time, a house near us to the west. He was a striking and beautiful person; tall, very thin, and cadaverously pale; his hair carefully powdered, though there was little of it except what was collected in a long thin queue; his eyes dark, clear and large, like deep pools of pure water. He wore black speckless clothes, silk stockings, silver buckles, and either a slim green umbrella, or a genteel brown cane. The general frame and air were feeble and slender. The wildest boy respected Black. No lad could be irreverent toward a man so pale, so gentle, so elegant and so illustrious. So he glided, like a spirit, through our rather mischievous sportiveness, unharmed. He died seated, with a bowl of milk upon his knee, of which his ceasing to be did not spill a drop; a departure which it seemed, after the event, might have been foretold of this attenuated philosophical gentleman.

'Multifocal bone lymphoma' or 'polyostotic lymphoma' is a neoplasm with exclusive multifocal involvement of the skeleton, without affecting lymph nodes or other soft tissues. Knowledge on this uncommon condition is limited because the related literature is sparse and fragmentary. We reviewed cases of multifocal bone diffuse large B-cell lymphoma (MB-DLBCL) registered in a clinico-pathological database of the International Extranodal Lymphoma Study Group that includes 499 cases of bone lymphoma. Clinical features, management and prognosis of 37 MB-DLBCL patients and 63 'controls' (stage-IV DLBCL and skeletal involvement) were analysed. Presentation and treatment of MB-DLBCL and controls were identical. At a median follow-up of 52 months (10-189), MB-DLBCL patients exhibited a significantly better response rate (92% vs. 65%; P = 0·002), progression-free survival (5-year: 56 ± 9% vs. 34 ± 6%; P = 0·003) and overall survival (5-year: 74 ± 8% vs. 36 ± 7%; P = 0·002). Among MB-DLBCL patients, the use of post-chemo radiotherapy was associated with better overall survival (5-year: 83 ± 12% vs. 55 ± 16%; P = 0·003). Two MB-DLBCL patients (5·4%) with spine and skull involvement experienced central nervous system (CNS) relapse. Thus, MB-DLBCL patients exhibit a significantly better prognosis compared to patients with advanced-stage DLBCL, and should be treated with conventional anthracycline-based chemotherapy, keeping intensified treatment for relapsing cases, considering involved-field radiotherapy, and CNS prophylaxis in high-risk patients.