WHO Expert Committee on Specifications for Pharmaceutical Preparations: fifty-fourth report

The World Health Organisation has published ‘WHO Expert Committee on Specifications for Pharmaceutical Preparations: fifty-fourth report’.

The WHO Technical Report Series makes available the findings of various international groups of experts that provide WHO with the latest scientific and technical advice on a broad range of medical and public health subjects. Members of such expert groups serve without remuneration in their personal capacities rather than as representatives of governments or other bodies; their views do not necessarily reflect the decisions or the stated policy of WHO.

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There is a new Annex 3 – ‘Production of water for injection by means other than distillation’.

An important section for microbiologists reads:

“The system should be monitored for its ongoing performance within defined parameters, including but not limited to, conductivity, total organic carbon (TOC) and microbial contamination.

A combination of online and offline monitoring of WFI should be done, to ensure that the appropriate water specification is maintained. TOC and conductivity should be monitored with online instruments. Use of rapid microbiological methods is encouraged for timely monitoring, and aids with rapid responses to prevent deterioration of the system.”

The report can be found here: https://www.who.int/publications-detail/978-92-4-000182-4

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

Human Drug Compounding Outsourcing Facilities

The U.S. FDA has issued a new draft guidance document: “Current Good Manufacturing Practice—Guidance for Human Drug Compounding Outsourcing Facilities Under Section 503B of the FD&C Act Guidance for Industry.”

The guidance reflects further consideration of how CGMP requirements should be applied in light of the size and scope of an outsourcing facility's operations.

The document can be found here: https://www.fda.gov/regulatory-information/search-fda-guidance-documents/current-good-manufacturing-practice-guidance-human-drug-compounding-outsourcing-facilities-under

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

Best practices for airflow visualization

Air is of fundamental importance to cleanrooms and clean air devices, either as a contamination source (microorganisms carried in the airstream) or as a control measure to minimise contamination (through the supply of clean air). Therefore, controlling a cleanroom requires careful attention to air filtration and airflow.

Assessing airflow is undertaken through airflow visualisation studies (or what are colloquially termed ‘smoke studies’). The purpose of flow visualisation is to confirm the smoothness, flow patterns and other spatial and temporal characteristics of airflow in an installation.

For this, the airflow is examined through airflow visualisation mapping whereby smoke is generated, and the behaviour of the smoke is studied and then captured by a video camera.

Air flow visualisation is one of the less clear-cut activities required of aseptic manufacturers. Regulations refer to such studies, yet little in the way of guidance is provided. However, judging by warning letters and other regulatory trends, many industries are running behind in terms of compliance expectations. The regulator findings suggest that evaluation, presentation and demonstration of airflow is not well understood. In this article, we will discuss the main regulatory findings, as evidenced from recent USA Food and Drug Administration (FDA) 483s and assess these for the main areas for concern and offer best practices for addressing the regulatory observations.

In relation to this topic, a new article of interest has been published in GMP Review. The reference is:

Saghee, M. R., Sandle, T., and Das, P. (2019) Regulatory inspections of sterile facilities – the focal points Part 2: Airflow visualisation, GMP Review, 18 (3): 15-21

For further details, please contact Tim Sandle.

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

Cleaning validation Q&A

Recently Pharmig ran a session on cleaning validation. Here are some of the questions posed, together with the answers.

How can irregular shaped objects (such as pipes) be swabbed to an exact figure e.g. 25cm². Is the surface area calculated and a depth decided upon to swab?

This is obviously not easy. If the surface area is smaller, then the count is still assessed as if it was 25cm² (to indicate what a ‘norm’ is and to allow a comparison to be made). With other areas, all you can do is gain an approximate assessment in terms of the number and length of strokes. It is generally accepted that where a template cannot be used, the swabbing is approximate.

How can a swab template be used for the irregular surfaces? If not, what justification may be used?

A swab template cannot be used on an irregular surface, all you can do is gain an approximate assessment in terms of the number and length of strokes. It is generally accepted that where a template cannot be used, the swabbing is approximate.

Where you can, it is best to use a contact plate – it gives a better recovery and if there are cleaning residues present, the agar can contain an appropriate neutralisers.

Are there any regulations or guidelines for swabbing in this manner?

Not for microbiological swabbing. There are a number of good guidance texts, although these are not ‘standards’. The microbiological aspects are invariably overlooked, which was the reason for putting the webinar together.

I’d recommend the work of Andrew Walsh, for example: https://pbe-expert.com/wp-content/uploads/2018/04/CleaningValidationforthe21stCentury-AcceptanceLimitsforCleaningAgents.pdf

How can training be provided for personnel to encompass this irregular swabbing?

Pharmig have run too training courses run by Gordon Farquharson in relation to this.

The best way is to use old items of equipment and allow personnel to practice on the lab bench, assessing their technique e.g. number of swab strokes, twisting the swab head etc.

Also when swabbing a surface do companies use cut out templates to get an accurate surface area? Or do companies just go on samplers judgement of surface area- between different samplers this could introduce a lot of variability?

Some companies use templates, which are sterilised. These will only work on relatively flat, regular shaped surfaces.

Others use judgement, and this has to be applied to irregular shaped surfaces. This can be supported by training.

As cannot contaminate equipment with organisms like can with chemicals. What does he think to the suggestion of supplementing cleaning validation study with lab based studies on samples of surfaces representative of the equipment? E.g. samples of SS 316L in lab – could contaminate with organism(s) representative of those found in facility and demonstrate effectiveness of cleaning regime/agents etc. is possible.  Overkill & unnecessary or good to do if have the time & resource?

This is useful if there is sufficient resource. The method outlined is similar to disinfectant efficacy testing in terms of phase 2 surface evaluation. However, it is not (as yet) a regulatory requirement. The standard approach is to evaluate ‘cleanliness’ through direct (swabbing) and indirect (rinse) samples.

The reference to facility isolates is interesting, although these are difficult to standardise.

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

'Designer virus' is first new oral polio vaccine in 50 years

Phase 1 trial shows promise for completion of stalled eradication effort; offers lessons for COVID-19 vaccine development.

Virologists report promising Phase 1 clinical results for the first new oral polio vaccine in 50 years, which they have designed to be incapable of evolving the ability to cause disease in humans.

Before being halted due to the COVID-19 pandemic, a relentless vaccination campaign had nearly succeeded in eradicating polio from the world. Between 2000 and 2017, the World Health Organization (WHO) estimated that its campaign had reduced the burden of the disease by 99 percent, preventing more than 13 million children from becoming infected and risking potentially debilitating paralysis.

But in recent years, the eradication effort has been plagued by outbreaks of vaccine-derived polio -- in which the weakened virus used in oral polio vaccines evolved the ability to escape from vaccinated individuals and spread in communities with poor vaccination rates.

Now, with support from the Bill and Melinda Gates Foundation, UC San Francisco virologist Raul Andino, PhD and Andrew Macadam, PhD, of the UK's National Institute for Biological Standards and Control (NIBSC) report promising Phase 1 clinical results for the first new oral polio vaccine in 50 years, which they have designed to be incapable of evolving the ability to cause disease in humans.

In a 2017 study, Andino and colleagues discovered that in every vaccine-derived polio outbreak they studied, the virus had used the same three evolutionary steps to mutate from harmless vaccine into a regional menace.

In their new study, Andino, Macadam, and colleagues at the Gates Foundation, the Center for Vaccine Innovation and Access in Seattle, and the Centre for the Evaluation of Vaccination at the University of Antwerp have employed clever genetic wizardry based on decades of study of the poliovirus's biology to redesign the vaccine to ensure that is incapable of following this three-step pathway to re-evolve virulence. Specifically, they stabilized a region of the viral genome that is required for it to re-evolve the ability to infect humans, and ensured that the virus could not get rid of this modification even by exchanging genetic material with related viruses.

The trial found that the new designer polio vaccine was both more stable and more effective than the 50-year old Sabin vaccine from which it was derived. Specifically, the new vaccine caused participants to generate plentiful antibodies against the poliovirus, and despite shedding viral particles in their stool, those particles were unable to infect or cause paralysis in mice. In contrast, previous studies have found that when mice are exposed to viral samples shed by people vaccinated with the standard Sabin oral polio vaccine, as many as 90 percent develop paralysis.

See: Ming Te Yeh, Erika Bujaki, Patrick T. Dolan, Matthew Smith, Rahnuma Wahid, John Konz, Amy J. Weiner, Ananda S. Bandyopadhyay, Pierre Van Damme, Ilse De Coster, Hilde Revets, Andrew Macadam, Raul Andino. Engineering the Live-Attenuated Polio Vaccine to Prevent Reversion to Virulence. Cell Host & Microbe, 2020; DOI: 10.1016/j.chom.2020.04.003

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

FDA - Q3D(R1) Elemental Impurities

FDA has now published this final guidance. Elemental impurities in drug products may arise from several sources; they may be residual catalysts that were added intentionally in synthesis or may be present as impurities (e.g., through interactions with processing equipment or container/closure systems or by being present in components of the drug product). Because elemental impurities do not provide any therapeutic benefit to the patient, their levels in the drug product should be controlled within acceptable limits.

See: https://www.fda.gov/media/135956/download

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

Communities of microbes found to have working memory

Biologists studying communities of bacteria have discovered that these so-called simple organisms feature a robust capacity for memory. Using light, researchers were able to encode complex memory patterns and visualize cells with memory. The discovery reveals surprising parallels between low-level single-celled organisms and sophisticated neurons that process memory in the human brain. The finding also provides a starting path for scientists to one day design basic computing systems with living organisms such as bacteria.

Following recent discoveries by the Süel lab that bacteria use ion channels to communicate with each other, new research suggested that bacteria might also have the ability to store information about their past states. In the new study, the researchers were able to encode complex memory patterns in bacterial biofilms with light-induced changes in the cell membrane potential of Bacillus subtilis bacteria. The optical imprints, they found, lasted for hours after the initial stimulus, leading to a direct, controllable single-cell resolution depiction of memory.

The ability to encode memory in bacterial communities, the researchers say, could enable future biological computation through the imprinting of complex spatial memory patterns in biofilms.

See:

Chih-Yu Yang, Maja Bialecka-Fornal, Colleen Weatherwax, Joseph W. Larkin, Arthur Prindle, Jintao Liu, Jordi Garcia-Ojalvo, Gürol M. Süel. Encoding Membrane-Potential-Based Memory within a Microbial Community. Cell Systems, 2020; DOI: 10.1016/j.cels.2020.04.002

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

FDA - Safety testing of drug metabolites

This guidance provides recommendations to industry on when and how to identify and characterize drug metabolites whose nonclinical toxicity needs to be evaluated. Drug metabolites may need to be determined in nonclinical studies when they are disproportionate drug metabolites, that is, metabolites identified only in humans or present at higher plasma concentrations in humans than in any of the animal species used during standard nonclinical toxicology testing.

See: https://www.fda.gov/media/72279/download

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

New insight into bacterial structure to help fight against superbugs

Scientists have produced the first high-resolution images of the structure of the cell wall of bacteria, in a study that could further understanding of antimicrobial resistance.

The findings set a new framework for understanding how bacteria grow and how antibiotics work, overturning previous theories about the structure of the outer bacterial layers.

The images give unprecedented insight into the composition of the bacterial cell wall and will inform new approaches to developing antibiotics in order to combat antibiotic resistance. There are no other examples of studies of the cell wall in any organism at comparable resolution, down to the molecular scale.

The team used an advanced microscopy technique called Atomic Force Microscopy (AFM), which works by using a sharp needle to feel the shape of a surface and build an image similar to a contour map, but at the scale of individual molecules.

See:

L. Pasquina-Lemonche, J. Burns, R. D. Turner, S. Kumar, R. Tank, N. Mullin, J. S. Wilson, B. Chakrabarti, P. A. Bullough, S. J. Foster, J. K. Hobbs. The architecture of the Gram-positive bacterial cell wall. Nature, 2020; DOI: 10.1038/s41586-020-2236-6 

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

Technological developments with wound treatment dressings

Dressings are long-established in the medical field for keeping wounds clean and preventing secondary infections. Dressings are essential for reducing infection risk; more recent advancements have seen dressings developed to promote wound healing.

In relation to this, Tim Sandle has written an article for Infectious Disease Hub. Here is an extract:

Wound healing refers to a specific biological process related to the general phenomenon of growth and tissue regeneration. Wounds heal by the control of moisture and by a process of staving off infection; while dressings have addressed the former over centuries, with the latter specialized dressings are being developed. Dressings to aid wound healing have been used for several decades, such as the use of hydrocolloid dressings to help treat burns and with the application of hydrogels for wounds that are leaking little or no fluid. However, it is in more recent years that antimicrobial compounds have been incorporated.

This involves the addition of physical or chemical processes designed to kill any pathogenic microorganisms that might be present, and which pose a risk of triggering a disease such as sepsis (where the chemicals that body’s immune system releases into the bloodstream to combat an infection trigger inflammation throughout the entire body) or delay the rate of wound healing (one of the causes of delayed wound healing is the presence of microorganisms in the wound). This article examines some of these developments, and also considers an additional development in the form of color-changing dressings for signaling the presence of antimicrobial-resistant bacteria.

The article can be accessed here.

The reference is:

Sandle, T. (2020) Technological advances with dressings for wound treatment and detection, Infectious Disease Hub, published on 19^th February 2020. At: https://www.id-hub.com/2020/02/20/technological-advances-with-dressings-for-wound-treatment-and-detection/

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

Plastic eating bacterium offers pollution hope

German scientists have found a small solution to this big problem – plastic-eating bacteria.

This newly identified strain of Pseudomonas bacteria was discovered by a team consisting of researchers from Helmholtz Centre for Environmental Research (Leipzig, Germany), Freiberg University of Mining and Technology (Germany) and Helmholtz Centre for Infection Research (Braunschweig, Germany), who have demonstrated its ability to break down polyurethane, a toxic type of plastic that is particularly hard to recycle.

In 2015 alone, polyurethane accounted for 3.5 million tons of the plastic produced in Europe. Its lightweight, insulating and flexible properties make it suitable for use in many products, from shoes and refrigerators to kitchen sponges. However, it’s incredibly difficult and energy-intensive to recycle, so most ends up on landfill sites where it releases toxic, often carcinogenic, chemicals.

Most bacteria are unable to withstand the toxic fumes released by the plastic, but, as demonstrated in a recent study published in Frontiers in Microbiology, this newly discovered strain is able to not only withstand and survive in the harsh environment, but also degrade some of the chemical building blocks of polyurethane and use these for energy.

For further details, see Biotechniques: https://www.biotechniques.com/microbiology-virology/a-tiny-solution-for-plastic-pollution-scientists-discover-polyurethane-feeding-bacteria  

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

Ready for The Count? Back-To-Basics Review Of Microbial Colony Counting

Microbiological laboratories remain reliant on the accurate determination of the number of colony forming units (CFUs) on growth media; this is notwithstanding advances with rapid microbiological methods, at least for some applications. Mainstay methods include pour plates, spread plates, and membrane filtration. Counting of microbes is important as it enables a laboratory to estimate the microbial population in a variety of products (bioburden). Yet there are limitations with plate count methods, including the fact that they only count viable cells and culturable organisms.

Tim Sandle looks at the issue of colony counting from a new perspective, including limitations with the human eye, and with how data integrity can be improved.

The reference is:

Sandle, T. (2020) Ready for The Count? Back-To-Basics Review Of Microbial Colony Counting, Journal of GxP Compliance, 24 (1) : http://www.ivtnetwork.com/article/ready-count-back-basics-review-microbial-colony-counting

For details, please contact Tim Sandle

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

What type of cells does the novel coronavirus attack?

Scientists from the Berlin Institute of Health (BIH), Charité -- Universitätsmedizin Berlin and the Thorax Clinic at Heidelberg University Hospital, whose collaboration is taking place under the auspices of the German Center for Lung Research (DZL), have examined samples from non-virus infected patients to determine which cells of the lungs and bronchi are targets for novel coronavirus (SARS-CoV-2) infection. They discovered that the receptor for this coronavirus is abundantly expressed in certain progenitor cells. These cells normally develop into respiratory tract cells lined with hair-like projections called cilia that sweep mucus and bacteria out of the lungs.

The scientists knew, from studies by BIH Professor Christian Drosten, director of the Institute of Virology at Campus Charité; Mitte, and by others, that the virus's spike protein attaches to an ACE2 receptor on the cell surface. In addition, the virus needs one or more cofactors for it to be able to penetrate cells. But which cells are endowed with such receptors and cofactors? Which cells in which part of the respiratory system are particularly susceptible to SARS-CoV-2 infection? Eils and his colleagues at the BIH and Charité; now used single-cell sequencing technology to examine the cells in the samples from Heidelberg.

60,000 single cells were sequenced

The researchers discovered that certain progenitor cells in the bronchi are mainly responsible for producing the coronavirus receptors. These progenitor cells normally develop into respiratory tract cells lined with hair-like projections called cilia that sweep mucus and bacteria out of the lungs.

See:

Soeren Lukassen, Robert Lorenz Chua, Timo Trefzer, Nicolas C. Kahn, Marc A. Schneider, Thomas Muley, Hauke Winter, Michael Meister, Carmen Veith, Agnes W. Boots, Bianca P. Hennig, Michael Kreuter, Christian Conrad & Roland Eils. SARS-CoV-2 receptor ACE2 and TMPRSS2 are primarily expressed in bronchial transient secretory cells. EMBO Journal, 2020 DOI: 10.15252/embj.20105114

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

Nosopharm develops a first-in-class novel antibiotic

Nosopharm, specialized in exploring unconventional sources of antibiotics to discover new drugs to fight antimicrobial resistance, today announces it has been granted a US patent for NOSO-502. NOSO-502 is Nosopharm’s first clinical candidate in a new class of antibiotics called Odilhorhabdins, aimed at eradicating resistant bacteria. This patent secures Nosopharm’s market exclusivity rights in the US, the leading pharmaceutical market. It also covers numerous chemical analogs - the chemical space of NOSO-502 – thereby reinforcing its protection against exploitation by competitors.

Besides the US patent (US Patent No. 10,626,144), Nosopharm has been granted a patent in Russia, and a notice of acceptance has recently been issued in Australia. In addition, the company has filed patent applications in Canada, China, Europe (EPO), Japan, Hong Kong, Brazil, South Korea, India, Israel and Mexico. Apart from Canada and South Korea, which start examining patent applications later than other countries, decisions are pending in all these territories and are expected to be announced during the course of this year.

“The award of a US patent for our clinical candidate NOSO-502 is a key milestone in Nosopharm’s development,” said Philippe Villain-Guillot, co-founder and chief executive officer at Nosopharm. “The US market is the global leader in pharmaceuticals and we have now secured exclusive marketing rights there. Backed by this IP, there is significantly more incentive for both a potential industrial partner to in-license the NOSO-502 program and further developments to combat resistant bacteria.”

The growing resistance of pathogenic bacteria is a threat to the efficacy of antibiotics. In 2015 antibiotic-resistant bacteria were estimated to be responsible for 670,000 infections and 33,110 attributable deaths in the EU and the European Economic Area (EEA). From a global perspective, antimicrobial resistance could kill up to ten million people every year by 2050, which could cost up to €94 trillion ($100tn).

Nosopharm investigates a new class of antibiotics, Odilhorhabdins, that inhibit the bacterial ribosome with a new mechanism of action. It is intended for treating nosocomial infections caused by Enterobacteriaceae, including polymyxin– and carbapenem-resistant Enterobacteriaceae (CRE). In 2017, the WHO published a list of priority pathogens for the development of new antibiotics. Carbapenem-resistant Gram-negative bacteria (Enterobacteriaceae, Pseudomonas aeruginosa, Acinetobacter baumannii) were at the top of that list, with critical priority.



NOSO-502 has proven to be effective in vivo in Enterobacteriaceae infection models and demonstrated antibacterial activity in vitro against multi-drug resistant clinical isolates (KPC, NDM and OXA among others).

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

Coronavirus testing, antivirals, vaccines and other measures

Posted by Dr. Tim Sandle,
Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)