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We Support each other to Develop and Grow at Novartis
No full textThe culture of Novartis starts at the very top with our dynamic CEO Vas Narasimhan. He publicly announced in 2018 the importance of unleashing the power of our people as a strategic priority to reimagine medicine. Vas planted the seeds of change in the company to ensure all employees feel inspired, curious and unbossed. In turn, it has been up to every single one of us to take those words and turn them into actions. Several years since the initial commitment, I personally have seen the growth of our organization at all levels. Gone are the days of traditional leadership centered around hierarchy, competition to get ahead, and micromanagement. In its place are servant leaders, peer mentoring circles, and empowered teams. I truly see Novartis today as a collaborative group of innovators working toward the same goal to best serve our patients. I myself have benefited from the investment we have made into the leadership and to each other. In this article, I will share with you my experiences within Novartis on how we take this culture to heart and support each other to develop and grow every day. The opportunities to grow and gain exposure to new development experiences at Novartis are endless and are only restricted now by our own self-induced boundaries
Species-dependent hepatic and intestinal metabolism of selective oestrogen receptor degrader LSZ102 by sulphation and glucuronidation.
No full textLSZ102 is an orally bioavailable selective oestrogen receptor degrader in clinical development for the treatment of breast cancer. Preclinical studies showed efficacy in xenograft models on oral dosing. However, oral bioavailability was relatively low in several preclinical species (7-33%), and was associated with first-pass metabolism, particularly intestinal first-pass.To investigate metabolism and first-pass effects, metabolites were analysed in human plasma samples after oral dosing of LSZ102 to patients, rat plasma samples after oral dosing of [14C]LSZ102, and in vitro incubations of [14C]LSZ102 with human and rat hepatocytes and intestinal S9 fractions. The kinetics of human sulfotransferase (SULT) enzymes potentially involved in metabolism of LSZ102 was characterised.Sulphate metabolites were found to be the major components in human plasma, as well as in human hepatocytes and intestinal S9 fractions. Contrastingly, glucuronidation was predominant in rat plasma, hepatocytes and intestinal S9. LSZ102 was found to be metabolised by several human SULTs expressed in liver and intestine. The combined metabolism data in rat and human provide supporting evidence for an extensive intestinal first-pass metabolism effect via sulphation in human but glucuronidation in rat.As LSZ102 is metabolised by a number of different SULTs, drug-drug interactions resulting from the inhibition of one SULT are unlikely.Despite the observed species difference in metabolism, the major human metabolites of LSZ102, sulphate M5, glucuronide M4, and secondary glucuronide/sulphate metabolite M12, have no or weak pharmacological activity and are not considered a toxicity risk as they are phase II conjugative metabolites
Donor-Acceptor Pyridinium Salts for Photo-Induced Electron-Transfer-Driven Modification of Tryptophan in Peptides, Proteins, and Proteomes Using Visible Light.
No full textTryptophan (Trp) plays a variety of critical functional roles in protein biochemistry; however, owing to its low natural frequency and poor nucleophilicity, the design of effective methods for both single protein bioconjugation at Trp as well as for in situ chemoproteomic profiling remains a challenge. Here, we report a method for covalent Trp modification that is suitable for both scenarios by invoking photo-induced electron transfer (PET) as a means of driving efficient reactivity. We have engineered biaryl N-carbamoyl pyridinium salts that possess a donor-acceptor relationship that enables optical triggering with visible light whilst simultaneously attenuating the probe's photo-oxidation potential in order to prevent photodegradation. This probe was assayed against a small bank of eight peptides and proteins, where it was found that micromolar concentrations of the probe and short irradiation times (10-60 min) with violet light enabled efficient reactivity toward surface exposed Trp residues. The carbamate transferring group can be used to transfer useful functional groups to proteins including affinity tags and click handles. DFT calculations and other mechanistic analyses reveal correlations between excited state lifetimes, relative fluorescence quantum yields, and chemical reactivity. Biotinylated and azide-functionalized pyridinium salts were used for Trp profiling in HEK293T lysates and in situ in HEK293T cells using 440 nm LED irradiation. Peptide-level enrichment from live cell labeling experiments identified 290 Trp modifications, with 82% selectivity for Trp modification over other π-amino acids, demonstrating the ability of this method to identify and quantify reactive Trp residues from live cells
High-content cellular screen image analysis benchmark study
No full textRecent development of novel methods based on deep neural networks has transformed how high-content microscopy cellular images are analyzed. Nonetheless, it is still a challenge to identify cellular phenotypic changes caused by chemical or genetic treatments and to elucidate the relationships among treatments in an unsupervised manner, due to the large data volume, high phenotypic complexity and the presence of a priori unknown phenotypes. Here we benchmarked five deep neural network methods and two feature engineering methods on a well-characterized public data set. In contrast to previous benchmarking efforts, the manual annotations were not provided to the methods, but rather used as evaluation criteria afterwards. The seven methods individually performed feature extraction or representation learning from cellular images, and were consistently evaluated for downstream phenotype prediction and clustering tasks. We identified the strengths of individual methods across evaluation metrics, and further examined the biological concepts of features automatically learned by deep neural networks
The proteomic landscape of glioblastoma recurrence reveals novel and targetable immunoregulatory drivers.
No full textGlioblastoma (GBM) is characterized by extensive cellular and genetic heterogeneity. Its initial presentation as primary disease (pGBM) has been subject to exhaustive molecular and cellular profiling. By contrast, our understanding of how GBM evolves to evade the selective pressure of therapy is starkly limited. The proteomic landscape of recurrent GBM (rGBM), which is refractory to most treatments used for pGBM, are poorly known. We, therefore, quantified the transcriptome and proteome of 134 patient-derived pGBM and rGBM samples, including 40 matched pGBM-rGBM pairs. GBM subtypes transition from pGBM to rGBM towards a preferentially mesenchymal state at recurrence, consistent with the increasingly invasive nature of rGBM. We identified immune regulatory/suppressive genes as important drivers of rGBM and in particular 2-5-oligoadenylate synthase 2 (OAS2) as an essential gene in recurrent disease. Our data identify a new class of therapeutic targets that emerge from the adaptive response of pGBM to therapy,
A comparison of the lowest effective concentration in culture media for detection of chromosomal damage in vitro and in blood or plasma for detection of micronuclei in vivo
No full textIt is often assumed that in vitro genotoxicity tests will be more effective than in vivo tests in detecting genotoxic substances since higher concentrations, more cytotoxicity and static exposures can be achieved. However, there is a paucity of data demonstrating whether genotoxic substances are detected at lower concentrations in cell culture in vitro than can be reached in the blood of animals treated in vivo. To investigate this issue, we compared the lowest concentration required for induction of chromosomal damage in vitro (lowest observed effective concentration, or LOEC) with the concentration of the test substance in blood at the lowest dose required for biologically relevant induction of micronuclei in vivo (lowest observed effective dose, or LOED). In total, 83 substances were found for which the LOED could be identified or estimated, where concentrations in blood and micronucleus data were available via the same route of administration in the same species, and in vitro chromosomal damage data were available. 39.8% of substances were positive in vivo at blood concentrations that were lower than the LOEC in vitro, 22.9% were positive at similar concentrations, and 37.3% of substances were positive in vivo at higher concentrations. Distribution analysis showed a very wide scatter of >6 orders of magnitude across these 3 categories. When mode of action was evaluated, the distribution of clastogens and aneugens across the 3 categories was very similar. Thus, the ability to detect induction of micronuclei in bone marrow in vivo regardless of the mechanism for micronucleus induction, is clearly not solely determined by the concentration of test substance which induced chromosomal damage in vitro
HistoNet: A Deep Learning-Based Model of Normal Histology
No full textWe introduce HistoNet, a deep neural network trained on normal tissue. On 1690 slides with rat tissue samples from 6 preclinical toxicology studies, tissue regions were outlined and annotated by pathologists into 46 different tissue classes. From these annotated regions, we sampled small 224 × 224 pixels images (patches) at 6 different levels of magnification. Using 4 studies as training set and 2 studies as test set, we trained VGG-16, ResNet-50, and Inception-v3 networks separately at each magnification level. Among these model architectures, Inception-v3 and ResNet-50 outperformed VGG-16. Inception-v3 identified the tissue from query images, with an accuracy up to 83.4%. Most misclassifications occurred between histologically similar tissues. Investigation of the features learned by the model (embedding layer) using Uniform Manifold Approximation and Projection revealed not only coherent clusters associated with the individual tissues but also subclusters corresponding to histologically meaningful structures that had not been annotated or trained for. This suggests that the histological representation learned by HistoNet could be useful as the basis of other machine learning algorithms and data mining. Finally, we found that models trained on rat tissues can be used on non-human primate and minipig tissues with minimal retraining
Full exome sequencing of 11 families with Hidradenitis suppurativa.
No full textImportance: Hidradenitis Suppurativa (HS) is not a well-studied or easily treated disease. Genetic information is essential for advances in the understanding and treatment of HS.
Objective: This study aims to examine mutations in the gamma-secretase complex, the Notch signaling pathway and to perform a Mendelian analysis of genetic variants that segregated with disease in a full exome sequencing of 11 families with HS.
Design: Whole exome sequencing and Mendelian analysis of 11 families with HS from Denmark.
Setting: Single-centre tertiary level clinic.
Participants: Patients with a clinical diagnosis of active HS and a positive family history of HS, were recruited. Consenting family members were enrolled and examined for HS as well. We included 11 families, with a total of 51 participants, 24 with HS and 27 without.
Intervention(s) (for clinical trials) or Exposure(s) (for observational studies): Whole exome sequencing using HiSeq platform as paired-end 2 x 150 bases, targeting 80x coverage.
Main Outcome(s) and Measure(s): We report mutations in genes in gamma-secretase and Notch pathway. As well as mutations perfectly segregated with the disease.
Results: We found mutations in the Notch pathway for all families, but only mutations in the PSENEN and APH1B of the gamma-secretase genes. We also report 161 variants of unknown significance that segregated with the disease within these families.
Conclusions and Relevance: We did not find causative mutation for each family in this study, supporting the theory that HS is rarely caused by single-gene mutations. We suggest that future genetic studies should be focused on genome-wide association with thousands of cases, as this technique is much better suited for suspected polygenic disease
Design of thioether cyclic peptide scaffolds with passive permeability and oral exposure
No full textAdvances in the design of permeable peptides and in the synthesis of large arrays of macrocyclic peptides with diverse amino acids have evolved on parallel, but independent tracks. Less precedence combines their respective attributes, in turn limiting potential to identify permeable peptide ligands for key protein targets. Herein, we present one strategy for focusing the powerful ligand-finding capability of DNA- or RNA-templated peptide synthesis within permeability-biased property space. Despite higher than standard molecular weights (from 774 to 1076 g·mol-1), the 6-, 7-, and 8-mer cyclic peptides of the present contribution are partially N-methylated to achieve low energy conformations with low desolvation penalties. The N-methylation patterns were selected using in silico methods, then experimentally validat-ed with high passive permeability and oral exposure. Further, the present work shows that chemical structures that overlap the synthetic capabilities of DNA- or RNA-templated peptide syntheses in water, can be both permeable and orally exposed. We envision that, by retaining the backbone N-methylation pattern and consequent bias toward per-meability, one can generate large peptide arrays with sufficient side chain diversity to identify permeability-biased lig-ands to a variety of protein targets
Ebi2 is temporarily upregulated in mo3.13 oligodendrocytes during maturation and regulates remyelination in the organotypic cerebellar slice model
No full textThe EBI2 receptor regulates the immune system and is expressed in various immune cells including B and T lymphocytes. It is also expressed in astrocytes in the central nervous system (CNS) where it regulates pro-inflammatory cytokine release, cell migration and protects from chem-ically induced demyelination. Its signaling and expression are implicated in various diseases including multiple sclerosis, where its expression is increased in infiltrating immune cells in the white matter lesions. Here, for the first time, the EBI2 protein in the CNS cells in the human brain was examined. The function of the receptor in MO3.13 oligodendrocytes, as well as its role in remye-lination in organotypic cerebellar slices, were investigated. Human brain sections were co-stained for EBI2 receptor and various markers of CNS-specific cells and the human oligodendrocyte cell line MO3.13 was used to investigate changes in EBI2 expression and cellular migration. Organotypic cerebellar slices prepared from wild-type and cholesterol 25-hydroxylase knock-out mice were used to study remyelination following lysophosphatidylcholine (LPC)-induced demyelination. The data showed that EBI2 receptor is present in OPCs but not in myelinating oligodendrocytes in the human brain and that EBI2 expression is temporarily upregulated in maturing MO3.13 oligodendrocytes. Moreover, we show that migration of MO3.13 cells is directly regulated by EBI2 and that its signaling is necessary for remyelination in cerebellar slices post-LPC-induced demyelination. The work reported here provides new information on the expression and role of EBI2 in oligodendrocytes and myelination and provides new tools for modulation of oligodendrocyte biology and therapeutic approaches for demyelinating diseases