Managing and Mitigating Risk in Biologics Process Transfer

processes into clinical and commercial production facilities. However, the application of the full set of tactics for every process transfer is often not warranted. Product quality and process performance must fulfil expectations in every process transfer, but the tactics to achieve these goals are typically product/project-dependent. Two important factors to consider when allocating tactics (and the associated resources) to a particular process transfer are the probability of an initial process transfer delay, and the impact to patients and the company of such a delay. The probability of an initial process implementation problem is heighted under a number of circumstances: for example, when the transfer is to an unfamiliar plant, when the process is non-platformed in some aspect, or when there has been a change of equipment or automation software in the plant. The impact to patients and financial impact to the company of a process start-up delay is also project specific. Janssen Pharmaceuticals transfers bulk biologics processes to a variety of internal and external, clinical and commercial production facilities around the world. In this presentation, we will discuss our menu of process transfer tactics, and our business process for allocating tactics and resources to each process transfer. The presentation will be illustrated by examples from recent clinical and commercial process transfers

Akathisia and Newer Second‐Generation Antipsychotic Drugs: A Review of Current Evidence

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/155998/1/phar2404_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/155998/2/phar2404.pd

The identification of novel fragment leads by Biophysical methods for the inhibition of TbrPDEB1 as an anti-parasitic approach to Human African Trypanosomiasis

This drug design research project is based on identifying novel compound 'hits' against an established drug target by various screening methods such as NMR, X-Ray Crystallography and Biochemical Assays. Trypanosoma brucei cyclic nucleotide phosphodiesterase B1 (TbrPDEB1) is a validated drug target reported to be essential in the life cycle of the Trypanosoma brucei parasite - the causative agent of Human African Trypanosomiasis, one of several Neglected Parasitic Diseases which affects underdeveloped countries, and in some cases even First World countries. Otherwise known as Sleeping Sickness, the insect-borne parasitic disease occurs in 36 sub-Saharan African countries. The estimated mortality rate is up to 500,000 deaths per year and up to 80 million people are at risk of infection. With current methods of treatment being limited, ineffective and in some cases unsafe, there is an urgent need for new, more effective and safer medication. In this research project, the TbrPDEB1 catalytic domain construct plasmid (provided by the host laboratory at the University of Kent) was used to transform an established E. coli bacteria strain for the purpose of protein expression. A small fragment library put together by IOTA Pharmaceuticals consisting of 31 active human PDE inhibitors was initially validated by Proton NMR and used to soak TbrPDEB1 protein crystals. Ligand uptake was determined by X-Ray diffraction at the Diamond Light Source. Through the use of the three previously mentioned screening approaches, fragment binders to the TbrPDEB1 target enzyme were identified. Additionally, inhibition of the fragment binders to the target was also measured and assessed in comparison to NPD-008, a known TbrPDEB1 inhibitor of moderate potency. Out of the 31 fragments, one fragment was identified as an optimal binder and low potency inhibitor to TbrPDEB1, which underwent a similarity search follow-up thus to suggest similar chemotypes of interest for future work

Bortezomib‐Induced Sweet's Syndrome Confirmed by Rechallenge

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/106899/1/phar1383.pd

The identification of novel fragment leads by Biophysical methods for the inhibition of TbrPDEB1 as an anti-parasitic approach to Human African Trypanosomiasis

This drug design research project is based on identifying novel compound 'hits' against an established drug target by various screening methods such as NMR, X-Ray Crystallography and Biochemical Assays. Trypanosoma brucei cyclic nucleotide phosphodiesterase B1 (TbrPDEB1) is a validated drug target reported to be essential in the life cycle of the Trypanosoma brucei parasite - the causative agent of Human African Trypanosomiasis, one of several Neglected Parasitic Diseases which affects underdeveloped countries, and in some cases even First World countries. Otherwise known as Sleeping Sickness, the insect-borne parasitic disease occurs in 36 sub-Saharan African countries. The estimated mortality rate is up to 500,000 deaths per year and up to 80 million people are at risk of infection. With current methods of treatment being limited, ineffective and in some cases unsafe, there is an urgent need for new, more effective and safer medication. In this research project, the TbrPDEB1 catalytic domain construct plasmid (provided by the host laboratory at the University of Kent) was used to transform an established E. coli bacteria strain for the purpose of protein expression. A small fragment library put together by IOTA Pharmaceuticals consisting of 31 active human PDE inhibitors was initially validated by Proton NMR and used to soak TbrPDEB1 protein crystals. Ligand uptake was determined by X-Ray diffraction at the Diamond Light Source. Through the use of the three previously mentioned screening approaches, fragment binders to the TbrPDEB1 target enzyme were identified. Additionally, inhibition of the fragment binders to the target was also measured and assessed in comparison to NPD-008, a known TbrPDEB1 inhibitor of moderate potency. Out of the 31 fragments, one fragment was identified as an optimal binder and low potency inhibitor to TbrPDEB1, which underwent a similarity search follow-up thus to suggest similar chemotypes of interest for future work

Investigations Into Fragment Ligand Binding Using Quantitative STD and WaterLOGSY NMR Spectroscopy

Ligand-observed NMR spectroscopy is frequently employed in early-stage drug discovery, often as an initial screen to narrow the field of potential drug-like molecules. However, its use is limited to this early stage. More information regarding binding mode can be extracted from these experiments via quantification, and this should help extend the remit of these experiments beyond simple screening functions. Initially, it was shown that the amount of signal that could be produced from an STD NMR experiment could be dramatically increased by careful consideration of the selective saturation pulse. By systematically shortening the Gaussian pulse and positioning it at specific offset positions, it was shown that these dramatic increases in signal are genuine and need not result in false positives. Quantitative STD NMR spectroscopy as applied to Hsp90 and a series of small fragment ligands provided evidence to suggest that the precise inter-atomic distances between a protein and ligand within a crystal structure correlate with both initial rates of STD build up, and T1-adjusted STD values. This precise correlation has implications for chemotype clustering and initial binding mode selection, something which should be useful in the absence of a crystal structure. Taking the same quantitative principles and applying to LOGSY experiments elucidated another, discrete property of protein-ligand binding. Examining the ‘LOGSY difference’ signal for protons of a ligand allows us to see what protons are in close proximity to conserved, bound water at the protein-ligand binding interface. This is fundamentally different to the information gained from STD experiments. Applying the insights to a protein of a different nature, Ras, it was shown that quantitative STD can be applied to proteins of both different size and structure. Furthermore, more evidence was acquired to suggest that conserved, bound water in the binding site really is responsible for generating LOGSY signal. In the absence of these molecules, as in Ras, proximity of a proton to an exchangeable tends to dominate. In addition we were able to show that these quantitative methods can be used together to help eliminate incorrect computationally generated docking poses. The work presented in this thesis provides evidence for the advantages of STD and LOGSY NMR spectroscopy in fragment-based drug discovery. The information that can be extracted from relatively simple ligand-observed NMR experiments should be used to provide more evidence at an earlier stage of the drug discovery process, hopefully reducing late-stage attrition and helping us get to the therapeutic drug molecules we need a little more quickly

Vedolizumab: an α4β7 integrin antagonist for ulcerative colitis and Crohn’s disease

Ulcerative colitis (UC) and Crohn’s disease (CD) are chronic, relapsing inflammatory bowel diseases associated with significant morbidity. Conventional therapies for these diseases include corticosteroids, aminosalicylates, immunomodulators, and monoclonal antibodies. Over the years tumor necrosis factor (TNF)-α antagonists alone or in combination with other therapies have emerged as the cornerstone of treatment for induction and maintenance of remission of moderate to severe UC and CD. Unfortunately, some patients with moderate to severe UC and CD are unable to attain or maintain remission with TNF-α antagonist treatment. Vedolizumab, a humanized monoclonal antibody, is the first integrin receptor antagonist approved that selectively antagonizes α4β7 gastrointestinal integrin receptors. US Food and Drug Administration approval is for treatment of patients with moderate to severe active UC and CD who have inadequate response with, lost response to, or are intolerant to a TNF-α antagonist or an immunomodulator; or have inadequate response with, are intolerant to, or demonstrate dependence on corticosteroids. When administered according to approved dosing in patients with moderate to severe CD and UC, vedolizumab induces clinical response rates up to 31.4% and 47.1% at week 6, and clinical remission rates up to 39% and 41.8% at week 52, respectively. Serious adverse events reported with vedolizumab include serious infections, malignancies, and anaphylaxis. Since vedolizumab is gastrointestinal selective, to date, it has not shown evidence of causing progressive multifocal leukoencephalopathy; however, postmarketing studies monitoring for this adverse effect are ongoing. Further assessment of vedolizumab earlier in the course of these diseases and in combination with other therapies is warranted

Restoration of CFTR function in patients with cystic fibrosis carrying the F508del-CFTR mutation

<div><p>Restoration of BECN1/Beclin 1-dependent autophagy and depletion of SQSTM1/p62 by genetic manipulation or autophagy-stimulatory proteostasis regulators, such as cystamine, have positive effects on mouse models of human cystic fibrosis (CF). These measures rescue the functional expression of the most frequent pathogenic CFTR mutant, F508del, at the respiratory epithelial surface and reduce lung inflammation in <i>Cftr^F508del</i> homozygous mice. Cysteamine, the reduced form of cystamine, is an FDA-approved drug. Here, we report that oral treatment with cysteamine greatly reduces the mortality rate and improves the phenotype of newborn mice bearing the <i>F508del-CFTR</i> mutation. Cysteamine was also able to increase the plasma membrane expression of the F508del-CFTR protein in nasal epithelial cells from <i>F508del</i> homozygous CF patients, and these effects persisted for 24 h after cysteamine withdrawal. Importantly, this cysteamine effect after washout was further sustained by the sequential administration of epigallocatechin gallate (EGCG), a green tea flavonoid, both <i>in vivo</i>, in mice, and <i>in vitro</i>, in primary epithelial cells from CF patients. In a pilot clinical trial involving 10 <i>F508del-CFTR</i> homozygous CF patients, the combination of cysteamine and EGCG restored BECN1, reduced SQSTM1 levels and improved CFTR function from nasal epithelial cells <i>in vivo</i>, correlating with a decrease of chloride concentrations in sweat, as well as with a reduction of the abundance of <i>TNF/TNF-alpha (tumor necrosis factor)</i> and <i>CXCL8</i> (<i>chemokine [C-X-C motif] ligand 8</i>) transcripts in nasal brushing and TNF and CXCL8 protein levels in the sputum. Altogether, these results suggest that optimal schedules of cysteamine plus EGCG might be used for the treatment of CF caused by the <i>F508del-CFTR</i> mutation.</p></div