Targeting the differential addiction to anti-apoptotic BCL-2 family for cancer therapy

AbstractBCL-2 family proteins are central regulators of mitochondrial apoptosis and validated anti-cancer targets. Using small cell lung cancer (SCLC) as a model, we demonstrated the presence of differential addiction of cancer cells to anti-apoptotic BCL-2, BCL-XL or MCL-1, which correlated with the respective protein expression ratio. ABT-263 (navitoclax), a BCL-2/BCL-XL inhibitor, prevented BCL-XL from sequestering activator BH3-only molecules (BH3s) and BAX but not BAK. Consequently, ABT-263 failed to kill BCL-XL-addicted cells with low activator BH3s and BCL-XL overabundance conferred resistance to ABT-263. High-throughput screening identified anthracyclines including doxorubicin and CDK9 inhibitors including dinaciclib that synergized with ABT-263 through downregulation of MCL-1. As doxorubicin and dinaciclib also reduced BCL-XL, the combinations of BCL-2 inhibitor ABT-199 (venetoclax) with doxorubicin or dinaciclib provided effective therapeutic strategies for SCLC. Altogether, our study highlights the need for mechanism-guided targeting of anti-apoptotic BCL-2 proteins to effectively activate the mitochondrial cell death programme to kill cancer cells.</jats:p

N-terminal acetylation levels are maintained during acetyl-CoA deficiency in Saccharomyces cerevisiae

N-terminal acetylation (Nt-acetylation) is a highly abundant protein modification in eukaryotes and impacts a wide range of cellular processes, including protein quality control and stress tolerance. Despite its prevalence, the mechanisms regulating Nt-acetylation are still nebulous. Here, we present the first global study of Nt-acetylation in yeast cells as they progress to stationary phase in response to nutrient starvation. Surprisingly, we found that yeast cells maintain their global Nt-acetylation levels upon nutrient depletion, despite a marked decrease in acetyl-CoA levels. We further observed two distinct sets of protein N termini that display differential and opposing Nt-acetylation behavior upon nutrient starvation, indicating a dynamic process. The first protein cluster was enriched for annotated N termini showing increased Nt-acetylation in stationary phase compared with exponential growth phase. The second protein cluster was conversely enriched for alternative nonannotated N termini (i.e. N termini indicative of shorter N-terminal proteoforms) and, like histones, showed reduced acetylation levels in stationary phase when acetyl-CoA levels were low. Notably, the degree of Nt-acetylation of Pcl8, a negative regulator of glycogen biosynthesis and two components of the pre-ribosome complex (Rsa3 and Rpl7a) increased during starvation. Moreover, the steady-state levels of these proteins were regulated both by starvation and NatA activity. In summary, this study represents the first comprehensive analysis of metabolic regulation of Nt-acetylation and reveals that specific, rather than global, Nt-acetylation events are subject to metabolic regulation

Food Science Challenge: Translating the Dietary Guidelines for Americans to Bring About Real Behavior Change

Food scientists and nutrition scientists (dietitians and nutrition communicators) are tasked with creating strategies to more closely align the American food supply and the public's diet with the Dietary Guidelines for Americans (DGA). This paper is the result of 2 expert dialogues to address this mandate, which were held in Chicago, Illinois, and Washington, D.C., in early October 2010 between these 2 key scientific audiences. It is an objective that has largely eluded public health experts over the past several decades. This document takes the perspective of food scientists who are tasked with making positive modifications to the food supply, both in innovating and reformulating food products, to respond to both the DGA recommendations, and to consumer desires, needs, and choices. The paper is one of two to emerge from those October 2010 discussions; the other article focuses on the work of dietitians and nutrition communicators in effecting positive dietary change

NAA80 is actin’s N-terminal acetyltransferase and regulates cytoskeleton assembly and cell motility

Actin, one of the most abundant proteins in nature, participates in countless cellular functions ranging from organelle trafficking and pathogen motility to cell migration and regulation of gene transcription. Actin's cellular activities depend on the dynamic transition between its monomeric and filamentous forms, a process exquisitely regulated in cells by a large number of actin-binding and signaling proteins. Additionally, several posttranslational modifications control the cellular functions of actin, including most notably N-terminal (Nt)-acetylation, a prevalent modification throughout the animal kingdom. However, the biological role and mechanism of actin Nt-acetylation are poorly understood, and the identity of actin's N-terminal acetyltransferase (NAT) has remained a mystery. Here, we reveal that NAA80, a suggested NAT enzyme whose substrate specificity had not been characterized, is Nt-acetylating actin. We further show that actin Nt-acetylation plays crucial roles in cytoskeletal assembly in vitro and in cells. The absence of Nt-acetylation leads to significant differences in the rates of actin filament depolymerization and elongation, including elongation driven by formins, whereas filament nucleation by the Arp2/3 complex is mostly unaffected. NAA80-knockout cells display severely altered cytoskeletal organization, including an increase in the ratio of filamentous to globular actin, increased filopodia and lamellipodia formation, and accelerated cell motility. Together, the results demonstrate NAA80's role as actin's NAT and reveal a crucial role for actin Nt-acetylation in the control of cytoskeleton structure and dynamics

RHAMM/HMMR (CD168) is not an ideal target antigen for immunotherapy of acute myeloid leukemia

Background : Criteria for good candidate antigens for immunotherapy of acute myeloid leukemia are high expression on leukemic stem cells in the majority of patients with acute myeloid leukemia and low or no expression in vital tissues. It was shown in vaccination trials that Receptor for Hyaluronic Acid Mediated Motility (RHAMM/HMMR) generates cellular immune responses in patients with acute myeloid leukemia and that these responses correlate with clinical benefit. It is not clear however whether this response actually targets the leukemic stem cell, especially since it was reported that RHAMM is expressed maximally during the G2/M phase of the cell cycle. In addition, tumor specificity of RHAMM expression remains relatively unexplored. Design and Methods : Blood, leukapheresis and bone marrow samples were collected from both acute myeloid leukemia patients and healthy controls. RHAMM expression was assessed at protein and mRNA levels on various sorted populations, either fresh or after manipulation. Results : High levels of RHAMM were expressed by CD34+CD38+ and CD34- acute myeloid leukemia blasts. However, only baseline expression of RHAMM was measured in CD34+CD38- leukemic stem cells, and was not different from that in CD34+CD38- hematopoietic stem cells from healthy controls. RHAMM was significantly up-regulated in CD34+ cells from healthy donors during in vitro expansion and during in vivo engraftment. Finally, we demonstrated an explicit increase in the expression level of RHAMM after in vitro activation of T cells. Conclusions : RHAMM does not fulfill the criteria of an ideal target antigen for immunotherapy of acute myeloid leukemia. RHAMM expression in leukemic stem cells does not differ significantly from the expression in hematopoietic stem cells from healthy controls. RHAMM expression in proliferating CD34+ cells of healthy donors and activated T cells further compromises RHAMM-specific T-cell-mediated immunotherapy

Parent–infant closeness after preterm birth and depressive symptoms : A longitudinal study

Background: Preterm birth increases the risk for postpartum depression in both mothers and fathers, calling for strategies to alleviate and prevent depressive symptoms in parents of preterm infants. The aim of this study was to assess the association between early parent-infant closeness and later depressive symptoms among parents of preterm infants. We hypothesized that longer duration of closeness associate with fewer depressive symptoms in both parents. Methods: This prospective cohort study included 23 neonatal intensive care units (NICUs) from 15 countries in 2018 to 2020. Each unit recruited families with preterm infants aiming to 30 families. The total duration of parents’ presence in the NICU, and separately parent-infant skin-to-skin contact and holding, were measured using a Closeness Diary up to 14  days. The Edinburgh Postnatal Depression Scale (EPDS) was used at discharge and at 4  months corrected age of the infant. Results: The study included 684 mothers and 574 fathers. The median presence was 469   min (Q1 258 and Q3 1,087) per 24   h for the mothers and 259   min (Q1 100 and Q3 540) for the fathers; mean EPDS scores were 9.2 (SD 5.0) and 6.3 (SD 4.4) at discharge and 6.6 (4.7) and 4.3 (4.2) at 4  months, respectively. Parents’ presence and depressive symptoms varied greatly between the units. Parents’ presence as the total measure, or skin-to-skin contact and holding separately, did not associate with depressive symptoms in either mothers or fathers at either time point (adjusted). Conclusion: No association was found between the duration of parent-infant closeness in the neonatal unit and parents’ depressive symptoms. The beneficial effects of family-centered care on parents’ depression seem to be mediated by other elements than parent-infant physical closeness. More research is needed to identify the critical elements which are needed to alleviate parents’ depression after NICU stay.© 2022 Lehtonen, Lilliesköld, De Coen, Toome, Gimeno, Caballero, Tameliene, Laroche, Retpap, Grundt, Van Hoestenberghe, Skene, Pape and Axelin. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.fi=vertaisarvioitu|en=peerReviewed

Clinical evaluation of an evidence-based method based on food characteristics to adjust pancreatic enzyme supplements dose in cystic fibrosis

[EN] Background: Patients with cystic fibrosis (CF) and pancreatic insufficiency need pancreatic enzyme replacement therapy (PERT) for dietary lipids digestion. There is limited evidence for recommending the adequate PERT dose for every meal, and controlling steatorrhea remains a challenge. This study aimed to evaluate a new PERT dosing method supported by a self-management mobile-app. Methods: Children with CF recruited from 6 European centres were instructed to use the app, including an algorithm for optimal PERT dosing based on in vitro digestion studies for every type of food. At base-line, a 24h self-selected diet was registered in the app, and usual PERT doses were taken by the patient. After 1 month, the same diet was followed, but PERT doses were indicated by the app. Change in faecal fat and coefficient of fat absorption (CFA) were determined. Results: 58 patients (median age 8.1 years) participated. Baseline fat absorption was high: median CFA 96.9%, median 2.4g faecal fat). After intervention CFA did not significantly change, but range of PERT doses was reduced: interquartile ranges narrowing from 1447-3070 at baseline to 1783-2495 LU/g fat when using the app. Patients with a low baseline fat absorption (CFA<90%, n= 12) experienced significant improvement in CFA after adhering to the recommended PERT dose (from 86.3 to 94.0%, p=0.031). Conclusion: the use of a novel evidence-based PERT dosing method, based on in vitro fat digestion studies incorporating food characteristics, was effective in increasing CFA in patients with poor baseline fat absorption and could safely be implemented in clinical practice.We acknowledge the support of the MyCyFAPP Project consortium. We especially thank the participation and the effort of the patients involved in the study and their families. This work was fully funded by the European Union and the Horizon 2020 Research and Innovation Framework Programme (PHC-26-2014 call Self management of health and disease: citizen engagement and mHealth) under grant number 643806.Calvo-Lerma, J.; Boon, M.; Colombo, C.; De Koning, B.; Asseiceira, I.; Garriga, M.; Roca, M.... (2021). Clinical evaluation of an evidence-based method based on food characteristics to adjust pancreatic enzyme supplements dose in cystic fibrosis. Journal of Cystic Fibrosis. 20(5):e33-e39. https://doi.org/10.1016/j.jcf.2020.11.016Se33e3920

N-terminal acetylation shields proteins from degradation and promotes age-dependent motility and longevity

Most eukaryotic proteins are N-terminally acetylated, but the functional impact on a global scale has remained obscure. Using genome-wide CRISPR knockout screens in human cells, we reveal a strong genetic dependency between a major N-terminal acetyltransferase and specific ubiquitin ligases. Biochemical analyses uncover that both the ubiquitin ligase complex UBR4-KCMF1 and the acetyltransferase NatC recognize proteins bearing an unacetylated N-terminal methionine followed by a hydrophobic residue. NatC KO-induced protein degradation and phenotypes are reversed by UBR knockdown, demonstrating the central cellular role of this interplay. We reveal that loss of Drosophila NatC is associated with male sterility, reduced longevity, and age-dependent loss of motility due to developmental muscle defects. Remarkably, muscle-specific overexpression of UbcE2M, one of the proteins targeted for NatC KO-mediated degradation, suppresses defects of NatC deletion. In conclusion, NatC-mediated N-terminal acetylation acts as a protective mechanism against protein degradation, which is relevant for increased longevity and motility. The most common protein modification in eukaryotes is N-terminal acetylation, but its functional impact has remained enigmatic. Here, the authors find that a key role for N-terminal acetylation is shielding proteins from ubiquitin ligase-mediated degradation, mediating motility and longevity.Association Francaise contre les Myopathies 261981, Canadian Institutes of Health Research (CIHR) 249843, United States Department of Health & Human Services National Institutes of Health (NIH) - USA F-12540, Portuguese national funding through Fundaco para a Ciencia e a Tecnologia (FCT) 171752-PR-2009-0222, National Funds through Fundaco para a Ciencia e a Tecnologia (FCT) G008018N, G002721N, University of Bergen MC_UU_00028/6, FDN-143264, FDN-143265, PJT-180285, PJT-463531, R01HG005853, R01HG005084, DL 57/2016/CP1361/CT0019, 2022.01782.PTDC,PTDC/BIA-BID/28441/2017,PTDC/BIA-BID/1606/2020, ALG-01-0145-FEDER-028441, PPBI-POCI-01-0145-FEDER-022122, LISBOA-01-0145-FEDER-022170info:eu-repo/semantics/publishedVersio

Assessing gastro-intestinal related quality of life in cystic fibrosis: Validation of PedsQL GI in children and their parents

Background: Most patients with cystic fibrosis (CF) suffer from pancreatic insufficiency, leading to fat malabsorption, malnutrition and abdominal discomfort. Until recently, no specific tool was available for assessing gastro-intestinal related quality of life (GI QOL) in patients with CF. As the Horizon2020 project MyCyFAPP aims to improve GI QOL by using a newly designed mobile application, a sensitive and reliable outcome measure was needed. We aimed to study the applicability of the existing child-specific Pediatric Quality of Life Inventory, Gastrointestinal Symptoms Scales and Module (PedsQL GI) in children with CF. Methods: A multicenter, prospective observational study was performed in 6 European centers to validate the PedsQL GI in children with CF during 3 months. Results: In total, 248 children and their parents were included. Within-patient variability of PedsQL GI was low (24.11), and there was reasonable agreement between children and parents (ICC 0.681). Nine of 14 subscales were informative (no ceiling effect). The PedsQL GI and the median scores for 4 subscales were significantly lower in patients compared to healthy controls. Positive associations were found between PedsQL GI and age (OR = 1.044, p = 0.004) and between PedsQL GI and BMI z-score (OR = 1.127, p = 0.036). PedsQL GI correlated with most CFQ-R subscales (r 0.268 to 0.623) and with a Visual Analogue Scale (r = 0.20). Conclusions: PedsQL GI is a valid and applicable instrument to assess GI QOL in children with CF. Future research efforts should examine the responsiveness of the CF PedsQL GI to change in the context of clinical interventions and trials

Protocol for Application, Standardization and Validation of the Forskolin-Induced Swelling Assay in Cystic Fibrosis Human Colon Organoids

This protocol describes the isolation, handling, culture of, and experiments with human colon stem cell organoids in the context of cystic fibrosis (CF). In human colon organoids, the function of cystic fibrosis transmembrane conductance regulator (CFTR) protein and its rescue by CFTR modulators can be quantified using the forskolin-induced swelling assay. Implementation procedures and validation experiments are described for six CF human colon organoid lines, and representative CFTR genotypes are tested for basal CFTR function and response to CFTR-modulating drugs. For complete details on the use and execution of this protocol, please refer to Dekkers et al (2016) and Berkers and van Mourik (2019)