Assessing rapid microbial detection systems

by Dr. Ray Nims

With each passing year, it seems that there are more options available for rapid microbial detection. These rapid systems come in a variety of “flavors”, that is - they differ with respect to a set of key attributes. For instance, how rapid is rapid? What is the sensitivity? What is the maximum sample volume that may be tested? Is it quantitative or qualitative? What units are the results given in? Is it destructive or non-destructive (i.e., can the organism, once detected, be identified)? When one considers the variety of applications for which rapid methods may potentially replace existing culture methods, it rapidly becomes clear that there may not be “one shoe that fits all”.

In order to select an appropriate rapid method for use in one of the many microbial detection applications, one must first assess the available rapid systems for the key attributes mentioned above. This then provides the opportunity to rule out systems which for one reason or the other will not suit the application. There may be some applications for which no rapid system currently meets all requirements. Those rapid systems which do appear to possess the attributes required may be further evaluated for cost and for performance capabilities using specific sample matrices.

In the table below, we have listed some of the currently available rapid microbial detection systems. These include only systems which are 48 hours in duration or less, and therefore some of the sterility replacement assays involving reduced incubation durations (e.g., BacT/ALERT®, Growth Direct™) are not listed.

The key attributes of these rapid systems are displayed in the table below. The systems are arranged by principle of detection, as in the table above. For certain methods (e.g., Micro Pro™) increased sensitivity can be gained through increasing the duration of the incubation time. For non-destructive methods, the ability to identify the organism(s) detected is facilitated by an additional incubation post-detection.

What is the regulatory position on rapid microbial detection methods? The U.S. FDA Guidance for Industry: Sterile Drug Products Produced by Aseptic Processing states that other suitable microbiological tests (e.g., rapid methods) may be considered for environmental monitoring, in-process control testing, and finished product release testing after it has been demonstrated that these new methods are equivalent or better than conventional (e.g., USP) methods. Additionally, the FDA Process Analytical Technology (PAT) initiative encourages the voluntary development and implementation of innovative approaches in pharmaceutical development, manufacturing, and quality assurance (from MJ Miller, PDA Journal 45: 1-5, 2002).

Are rapid methods being used in the pharmaceutical industry? ScanRDI was approved by the FDA for water testing at GSK and for sterility testing at Alcon; Pallchek has been approved by the FDA for bioburden testing at GSK; and Wyeth received approval for use of Celsis for microbial limits testing.

Like all methods proposed to replace existing “gold standards”, these rapid microbial detection systems must be demonstrated through comparability protocols to be equivalent to or better than the existing methods. The effort required should pay dividends in terms of shortened turnaround times and reduced costs.

FDA regulation of “combination” products

by Dr. Ray Nims

The FDA’s Office of Combination Products (OCP) was established in 2002 to shepherd combination products, those comprised of a combination of drug, biological, and/or device, through the review and regulation process. An example of a combination product is the drug-releasing stent (see figure below). The OCP does not conduct the reviews, but is responsible for: assigning the product to the appropriate FDA center; coordinating reviews involving more than one center; and working with agency centers to develop guidance and regulations to make the regulation of combination products “as clear, consistent and predictable as possible”. 

An example of a drug-releasing stent.

From Wikipedia

The complexity of combination products arises because the efficacy and safety of the individual constituent components (i.e., the drug, biologic, or device) must be considered alone, as well as within the context of the combination product. Because of this complexity, there is no single development paradigm for all combination products. The guidance recommends that combination product developers consider any prior approval/clearance of the constituent parts, as well as how their testing may be influenced by the interaction of the components. Factors that should be considered (from the guidance) include:

• Are the constituent parts already approved for an indication?
• Is the indication for a given constituent part similar to that proposed for the combination product?
• Does the combination product broaden the indication or intended target population beyond that of the approved constituent part?
• Does the combination product expose the patient to a new route of administration or a new local or systemic exposure profile for an existing indication?
• Is the drug formulation different than that used in the already approved drug?
• Does the device design need to be modified for the new use?
• Is the device constituent used in an area of the body that is different than its existing approval?
• Are the device and drug constituents chemically, physically, or otherwise combined into a single entity?
• Does the device function as a delivery system, a method to prepare a final dosage form, and/or does it provide active therapeutic benefit?
• Is there any other change in design or formulation that may affect the safety/effectiveness of any existing constituent part or the combination product as a whole?
• Is a marketed device being proposed for use with a drug constituent that is a new molecular entity?
• Is a marketed drug being proposed for use with a complex new device?

As with individual constituent drugs, biologics, and devices, the FDA will require that the combination products be manufactured according to current good manufacturing practices. In most cases, a single investigational application (IND or IDE) is submitted to enable the clinical trials planned for the combination product. The science and technology associated with the combination products should drive the selection of statistical approaches, sample sizes, study endpoints, and methods for active principle measurement and for evaluating possible interactions between components. It may be best to involve the FDA in these decisions.

Complexity for the regulation of combination products also stems from the fact that separate manufacturing processes may exist for the various constituent parts. Potential changes in any of the component manufacturing processes, subsequent to initiating clinical trials or post-market, will need to be evaluated for possible effects on the safety and efficacy of the combination product.

Combination products represent therapeutic modalities with great promise for advancing health care. We expect to see more and more pharmaceutical activity in this area going forward.

The Good Buffer

By Scott Rudge

“A Good Buffer” has a number of connotations in biochemistry and biochemical engineering. A “good buffer” would be one that has good buffering capacity at the desired pH. The best buffering capacity is at the pK of the buffer of course, although it seems buffer salts are rarely used at their pK.

Second, a good buffer would be one matched to the application. Or maybe that’s first. For example, the buffering ion in an ion exchange chromatography step should be the same charge as the resin (so as not to bind and take up resin capacity). For example, phosphate ion (negative) is a good choice for cation exchange resins (also negatively charged) like S and CM resins.

Another meaning of a “Good” buffer is a buffer described by Dr. Norman Good and colleagues in 1966 (N. E. Good, G. D. Winget, W. Winter, T. N. Connolly, S. Izawa and R. M. M. Singh (1966). "Hydrogen Ion Buffers for Biological Research". Biochemistry 5 (2): 467–477.). These twelve buffers have pK’s spanning the range 6.15 to 8.35, and are a mixture of organic acids, organic bases and zwitterions (having both an acidic and basic site). All twelve of Good’s buffers have pK’s that are fairly strongly temperature dependent, meaning that, in addition to the temperature correction required for the activity of hydrogen ion, there is an actual shift in pH that is temperature dependent. So, while a buffer can be matched to the desired pH approximately every 0.2 pH units across pH 7 ± 1, the buffers are expensive and not entirely suited to manufacturing applications.

In our view, a good buffer is one that is well understood and is designed for its intended purpose. To be designed for its intended purpose, it should be well matched to provide adequate buffering capacity at the desired pH and desired temperature. As shown in the figure, the buffering capacity of a buffer with a pK of 8 is nearly exhausted below pH 7 and above pH 9.

It’s easy to overshoot the desired pH at these “extremes”, but just such a mismatch between buffering ion and desired pH is often specified. Furthermore, buffers are frequently made by titrating to the desired pH from the pK of the base or the acid. This leads to batch to batch variation in the amount of titrant used, because of overshooting and retracing. In addition, the temperature dependence of the pK is not taken into account when specifying the temperature of the buffer. Tris has a pK of 8.06 at 20°C, so a Tris buffer used at pH 7.0 is already not a good idea at 20°C. The pK of Tris changes by -0.03 pH units for every 1°C in positive temperature change. So if the temperature specified for the pH 7.0 buffer is 5°C, the pK will have shifted to 8.51. Tris has 3% of its buffering capacity available at pH 7.0, 5°C, it’s not well matched at all.

A good buffer will have a known mass transfer rate in water, so that its mixing time can be predicted. Precise amounts of buffering acid or base and cosalt are added to give the exact pH required at the exact temperature specified. This actually reduces our reliance on measurements like pH and conductivity that can be inexact. Good buffers can be made with much more precision than ± 0.2 pH units and ± 10% of nominal conductivity, and when you start to make buffers this way, you will rely more on your balance and understanding of appropriate storage conditions for your raw materials, than making adjustments in the field with titrants, time consuming mixing and guessing whether variation in conductivity is going to upset the process.

Got animal-derived materials? Part 2

By Ray Nims

As part of a formal animal-derived materials program, biopharma companies must assess the viral and TSE risk associated with materials derived from animals or which have been in contact with animal-derived materials at some point. We have addressed the viral risk within a previous blog installment. Now let’s consider the TSE risk.

TSEs (transmissible spongiform encephalopathies) are fatal diseases believed to result from exposure of a few animal species (including various ruminants, cervids, ungulates, cats, minks, and humans) to “infectious” prion proteins. The infectious proteins (PrP^Sc) are capable of interacting with and altering the normal prion proteins (PrP^c) within the brain and spinal cord, changing the normal proteins to the abnormal form. Since humans have contracted prion disease as a result of consuming tissues from cattle with bovine spongiform encephalopathy (mad cow disease), there is concern about the use of bovine materials and materials from other “relevant species” in the manufacture of biopharmaceuticals.

The EMEA has provided a guidance document (EMEA/410/01 Rev. 2 October 2003; http://www.emea.europa.eu/pdfs/human/bwp/TSE%20NFG%20410-rev2.pdf) describing the requirements for the use of animal-derived materials from relevant animal species (cattle, sheep, goats, and other animals which are naturally susceptible to TSEs but not including humans or non-human primates) in the manufacture of medicinal or veterinary products. The guidance applies to active substances, excipients and adjuvants, raw and starting materials and reagents, and materials which come into contact with products or equipment used to make product. The major point of the guidance is that TSE risk can be minimized, but in many cases not entirely eliminated. Where TSE risk cannot be avoided through elimination of animal-derived materials completely, the guidance provides principles for the minimization of TSE risk which include: the use of low-risk (non-relevant) animal species, the geographical sourcing of relevant species from low-risk regions, the use of low-risk tissues, the use of appropriate slaughter techniques to reduce potential for contamination of low-risk tissues with high-risk tissues, the appropriate oversight of manufacture of the animal products through Quality Assurance and self and external auditing and Quality Control testing, and the implementation of process designs to remove or inactivate the abnormal prion proteins.

Biopharma companies using animal-derived materials are instructed to perform risk assessments on those materials, which take into account the factors described above. The risk assessments must be completed as part of a formalized animal-derived materials program, documented procedurally and executed by staff that are experienced and trained to conduct such assessments. EMEA inspectors will expect to see this formal program in place. Risk assessments for individual raw materials may then be rolled up into a risk assessment for the biopharma product. The rolled up risk assessment may also consider manufacturing steps at the biopharma which may remove or inactivate the abnormal prion proteins, though these, if cited, may need to be validated. Finally, it is expected that companies will conduct a benefit/risk evaluation to assure that any benefits realized by the patient taking the product will outweigh and justify the risk associated with the use of materials derived from relevant animal species.

Outsource it, and fuggedaboutit?

By Ray Nims

Much has been written about the rationales and advantages for outsourcing of manufacturing and/or testing services; about the selection of outsourcing partners; and about the optimization of the pharma/contractor relationship. In any pharma/contractor relationship, there are responsibilities associated with the pharma as well as contractor responsibilities. These include both business as well as compliance responsibilities. The business realities and regulatory expectations associated with the use, by a pharma company, of a contract testing organization must be considered when the decision is made to outsource. A contract testing organization desiring to provide services for a pharmaceutical must be aware of the expectations and responsibilities associated with such a partnership. The optimal and most defensible programs will be those in which the various practices to be described below are formalized within internal Quality Systems, policies, and/or standard operating procedures as well as Quality Agreements.


Responsibilities falling upon the pharmaceutical partner include: 1) the selection of the contract testing lab; 2) commissioning and providing test samples of raw materials and products for method verification (compendial methods) and method qualification (non-compendial methods); 3) instituting of a Quality Agreements, business agreement, and/or confidentiality agreement with the contractor; 4) scheduling and shipping of test samples in accordance with the requirements of the testing lab and the test system; 5) providing in-life guidance and oversight of investigations of unexpected and out of specification results; and 6) ongoing monitoring of the performance of the contract lab and its methods.

Responsibilities primarily falling upon the testing lab include: 1) attaining and maintaining GLP or GMP compliance as appropriate for the intended use of the method; 2) providing assurance that the methods offered will be available to the client over the long term; 3) responsiveness to the sponsoring pharma and adherence to the terms of the Quality and/or business agreements; 4) method validation, verification, and or qualification as appropriate for the intended use of the method; 5) control of reagent, raw material, control, and standard inventory and quality; 6) assuring secure and retrievable data archiving; and 7) retention of staff possessing the appropriate expertise for direction of operators and the methods.

Do You Use Risk Assessments in Auditing?

Audits are a critical component of quality systems, but are they guided by formal assessments of risk to your products? In this world of ICH Q9, can you offer even a semi-quantitative justification for your audit priorities? We have spoken to many people in the industry, and almost all mention a risk assessment being undertaken prior to an audit. But we have not found many people that formalize that risk assessment, or keep it updated from audit to audit. Even fewer communicate their scoring of risk to either their internal clients or the vendor that has been audited.

A new trend in auditing is to use a form of risk assessment both before and after the audit. A popular form is the Failure Modes and Effects Assessment, or FMEA (see, for example, http://www.sre.org/pubs/Mil-Std-1629A.pdf). In a traditional FMEA, risks of failure are identified in a detailed fashion, and scored in three categories related to the failure’s probability, detectability and severity. Scoring is done on a semi-quantitative or relative basis using an arbitrary scale such as 1-10. For an audit, you might use the same categories as they relate to a particular vendor's (or department's) ability to deliver a product or service, failure free. You could organize your FMEA according to the critical quality attributes of the product or service being delivered or according to a list of requirements from a guideline or the CFR's. Your FMEA should receive input from affected departments, and should be used for prioritization of audit items. You should have the FMEA in mind as you conduct your audit, and remember why various items received high prioritization. You may change ratings for probability or detectability based on what you observe. If instead, you confirm your evaluation, you should probe remediations that decrease your firm's primary concern. A remediation that addresses detectability, when the issue was probability, likely won’t mitigate the risk of failure.
When you return from your audit, rescore the FMEA with assessments based on your observations and data that you collected. Make sure that you share your analysis with the stakeholders. And monitor the performance of the vendor until the next audit; the data will help inform your next FMEA.

Do you already use FMEA's in audit preparation and reporting? Let us know your practices.

Why is Quality by Design so heavy with statistics?

Why is the literature on Quality by Design so laden with statistics and experimental design space jargon? After all, the definition of the term “design” doesn't seem to include the analysis of messy data leading to rough correlations with results that are valid only over a limited range. So what gives?

The idea behind QbD was to use mathematical, predictive models to predict process outcomes. This concept can be applied directly to simple unit operations, such as drying, distilling, heating and cooling. However, unlike in the petrochemical business, the thermodynamic properties of most active pharmaceutical ingredients are not known and are difficult to measure. The unit operations used to manufacture common biotechnology products, such as cell culture, chromatography and fermentation have been modeled, but the models are very sensitive to unknown or unmeasureable adjustable parameters. The batch nature of these operations also makes their control difficult, as classical control theory relies on the measurement of an output to make an adjustment to an input to correct the output back towards the design specification.

Since there does not appear to be a clear path to using models, an approach has been chosen that emphasizes getting as much phenomenological information from as few experiments as possible. This is the Design of Experiments approach, where input conditions or operating parameters are systematically varied over a range and the process outputs measured, with statistics used to deconvolute the results. The combined ranges tested become the “design space”, and the process performance outputs with the variations closest to the process failure limit become the critical performance parameters. The results are useful, but only within the design space, and only with the certainty that the statistics report. Also, since the results are phenomenological, the effect of scale is often unknown.

The statistical approach is acceptable, and for the immediate future it's probably the best that we can expect. But the focus on this approach seems to drown out the more pressing need for good process models and physical properties data. These are the elements that allowed the petrochemical and commodity chemicals industries to scale up processes with assurance that quality specifications would be met. There are countless models available for bioprocessing's more complicated unit operations, but they have parameters that we don't know and can't calculate from first principles. There is no question that we need to find ways to collect this data, and a commitment to publish or share it. There are also simpler unit operations that we can model, scale up and scale down with complete assurance. These include operations such as mixing and storing solutions, filtration, diafiltration, centrifugation and some reactions. We shouldn't let the more complicated operations that still require statistical DOE approaches prevent us from applying the true principles of QbD to our simpler unit operations.

Ten Steps to Choosing your Contract Manufacturer

For many young companies, choosing a contract manufacturer, or CMO, for their lead pharmaceutical candidate is critical. Choose the wrong contractor, and you could be faced with delays and cost overruns with which your investors and patients won’t be very sympathetic. While there is no guarantee that you will always make the right decision, here are some tips that can help you make your choice in an organized, thoughtful, meaningful and objective way:

1) Make a list of all the possible suppliers. Such lists may be purchased, but they are also easily assembled from internet searches. In fact, you can do a little pre-screening with your own internet search.

2) Screen potential suppliers with a phone call. You will probably speak with a business development or sales person representing the contractor, but usually these people are quite knowledgeable about their company’s capabilities, and common problems encountered in the industry. We recommend you not reveal too many details about your project, and be prepared mostly to listen. However, you should have three or four key capabilities or proficiencies that you are looking for in all of the potential vendors. If possible, try to rule out vendors who do not meet your “must-have” requirements at this stage. Stay tuned for a blog on how to establish your showstopper list, it’s a critical exercise, and may extend beyond key capabilities and proficiencies.

3) Keep a matrix, and record the date you first contacted the vendor, when they responded to you, the status of any confidentiality agreements, and all the contact information that you can gather (email addresses, cell phone numbers, main switchboards and extension numbers). Also note the responses that each contact had relative to your three or four show-stopper criteria.

4) Meet with your team, and select three to five potential vendors to request a proposal from. We don’t recommend more than five: getting good, comparable proposals is a lot of work, like 2n times the work, where n = the number of proposers. Not to mention the work that contract manufacturers go through to read your RFP and prepare a proposal. Your three or four showstopper criteria should help you limit the number of proposers; if necessary you can begin to narrow down based on “nice-to-have” criteria as well. You may also deliberately choose to look at a range of vendors that represent different strengths/weaknesses (for example, do you prefer a “one-stop shop” that is convenient, but maybe not the best at everything, or a “specialty” vendor that provides higher levels of expertise, but will require you to select and manage multiple vendors?).

5) Solicit proposals. Most contract manufacturing seekers have a Request for Proposals (RFP) process that includes a document. These RFP documents vary from one page requirements descriptions, to lengthy, legalistic documents that require a team, and a month, to respond to. You should do what is comfortable for your organization. There needs to be enough information so that the vendor is able to respond with a meaningful proposal. There is some legal danger, particularly with intellectual property, so it’s not a bad idea to get your RFP reviewed by your legal counsel. And you should only send an RFP to a vendor after they have signed a mutually agreed confidentiality agreement.

6) Score your proposals. Find some basis to make apples to apples comparisons. RMC uses a modified Kepner Tregoe analysis, but many forms of analysis will work. You should have determined how you will assess and weight qualitative data before you begin. And in doing so, you should not under-estimate intangible factors: the ability to communicate, time zone differences, good references (you’ve checked, right?) are some examples. At this stage you should be ranking and scoring on “nice-to-have” criteria as well as comparing cost/timeline, capabilities, capacity, and viability of the business. You might form your opinion of the viability of the business by reading annual reports and press releases, and by assessing how busy the manufacturing area and support labs look during your visit.

7) Visit the top two or three vendors in person. Vendors may not allow a formal quality audit prior to signing a contract, but be sure to bring your quality representatives even for an informal “technical visit”. If the project is large, you may take the resistance to a pre-use quality audit as a red flag. Again, spend as much time as possible listening, rather than talking. Get a tour, and copies of all presentations. Ensure that you have a meeting between the decision makers for both sides as well as aligning discussions between key technical and quality personnel. If there are disputes or further work to do, your decision makers must have a good working relationship.
Your visit is also your best opportunity to break past the business development group and take a true measure of the business. Chat with the people in the lab or production area if you can. Look over the state of the equipment, the cleanliness of the facility and the stock in the warehouse. Check their flexibility-- what can they make happen for you, vs. what will have to be run past someone in another building, or another city? Ultimately, you should think about hiring a contract manufacturer similarly to how you hire an employee, by hiring for expertise as well as fit with your team.

8) Consider entering contract negotiations with at least two vendors. Things can go wrong in negotiations, and your position is stronger if you can legitimately walk away. We typically don’t let any of the final three candidates off the hook until the ink is dry on the final contract. If your budget can justify it, having a second contractor performing development work and verifying the primary contractor’s results is an excellent idea. It may ultimately spread your risk in supply chain, and give you leverage in negotiating commercial supply agreements later on.

9) Revisit your analysis tool. You may learn new things in your contract negotiations that cause you to adjust your evaluation. Don’t be afraid to be frank if you feel like terms have been changed since the selection was made. This is a good reason to keep a back up vendor.

10) You must now manage the project according to the criteria by which you selected your supplier. Hold your supplier and yourself accountable to these criteria. For example, after selecting a vendor because they can meet a very aggressive timeline, do not put the project timeline at risk by failing to order back up critical path supplies, in case the primary order doesn’t arrive, or fails to meet specifications on arrival. We will have more to say about managing a contract manufacturer in a future blog.

Choosing the right manufacturing partner is critical for your success as a drug developer. Spend the time required to make a good decision. This time should be spent gleaning as much tangible and non-tangible information on all your options as possible, and then objectively comparing it. You should have an idea about how you’re going to evaluate and weight non-tangible factors into your decision. And once you have made the choice, manage according to your criteria. Although everyone has their own methods for vendor selection, these are some suggestions that have worked well for us and our clients. If you have questions or comments, please visit http://www.rmcpharma.com/ or email us at info@rmcpharma.com.