I’m always looking for examples of the distinction between pharmacokinetics (PK) and pharmacodynamics (PD), partly to blog about and partly as new material for the undergrads I lecture to in the Pharmaceutical Product Development program at West Chester University.  I came across an interesting one recently, although maybe not exactly PK vs. PD…more of an indirect connection.

In a paper in the December 15, 2011 issue of The Journal of Infectious Diseases (http://jid.oxfordjournals.org/content/204/12/1951.full.pdf+html), researchers torpedoed a widely held and highly influential assumption in the field of tuberculosis (TB) therapy.  The assumption is that poor compliance by TB patients is responsible for the rise of multidrug-resistant TB.  This has led the World Health Organization (WHO) to require that healthcare workers directly witness each patient taking every dose of a combination of drugs over several months of therapy, a practice that is more expensive than the drugs themselves in many parts of the world.  However, multidrug resistant strains of TB continue to emerge in spite of >98% adherence to the direct-observation regimen.

The researchers used in vitro, hollow-fiber systems inoculated with M. tuberculosis, including a drug-resistant strain in some cases.  The cultures were maintained for 4 or 8 weeks, with antibiotics (isoniazid, rifampin, and pyrazinamide) administered daily by programmable syringe pumps (and gradual dilution with fresh culture medium to model PK clearance).  Antibiotic non-adherence (varying degrees and different patterns) was modeled by skipping the drug infusion on some days.  The emergence of drug-resistant bacteria was monitored by sampling the cultures throughout the study.  Therapeutic failure (defined as incomplete killing of bacteria) occurred only with >70% non-adherence, but the multidrug-resistant strain never reached 1% of the total bacterial population under any conditions.  Thus, non-adherence was not a sufficient condition for emergence of multidrug-resistant TB.

To test an alternative hypothesis, the researchers performed Monte Carlo simulations to determine the effect of inter-subject PK variability on efficacy (bacterial killing) and the emergence of drug resistance.  Based on previous reports of the variability in the PK of rifampin and isoniazid in a large population of patients, the authors concluded that a small percentage of patients clear rifampin rapidly enough that they are effectively treated with isoniazid monotherapy, which is known to promote the emergence of multidrug-resistant TB.  Thus, they hypothesize that PK variability, not non-adherence, is to blame for multidrug resistance among TB patients.  Furthermore, they propose that monitoring the blood levels of antibiotics in patients (enabling appropriate adjustment of doses) would be more effective than direct observation of therapy.  I’m not sure whether monitoring blood levels would cost less than direct observation or even be feasible in many remote areas, but it was interesting to see the importance of PK recognized in an unexpected circumstance.

Back to PK vs. PD: in the sense that any drug that you don’t take can’t be effective, one could consider non-adherence an aspect of PD.  As for PK, rest assured that Absorption Systems (www.absorption.com) knows it inside and out…PK is our business, and our portfolio of preclinical ADME services is both broad and deep.  Contact us for help with your ADME challenges.

Leadership in the field of pharmaceutical and medical device development depends on innovation.  Scientists and engineers with a new idea, who can turn their idea into something tangible, help shape future medical products.  Many people mistakenly think such innovation takes place only in universities or in within start-up companies themselves.  But this misses the important part played by contract research organizations (CROs) in facilitating innovation.

At a recent conference sponsored by the California Biomedical Research Association (CBRA), it was abundantly clear that many CROs are actively participating in innovative research.  Many of them, including Absorption Systems, attend investor forums and establish working relationships with academic institutions and non-profit organizations that foster the creation of start-up companies. 

What do CROs bring to the table?  They bring years of experience in how to design a testing program to maximize the likelihood of success in in-vivo trials, experience that can trim months to years from development timelines.  An effective study director can give a client insights into current regulatory demands and design studies that yield the maximum amount of information in the shortest time possible.   To further enhance the relationship, sponsor personnel should be able to participate directly in the study execution as well.  This is done frequently for preclinical medical device testing, where the sponsor may include leading authorities in a particular field but needs the expertise and infrastructure of a CRO to perform a study that will be acceptable to regulatory authorities.  Many CROs are known for their familiarity with and adherence to the Good Laboratory Practices (GLP) and animal welfare regulations, thus freeing a startup company to focus on its product’s development.

Absorption Systems takes seriously its commitment to supporting client innovation.  We work with clients closely through all stages of product development to help ensure the best study designs and outcomes.  We also keep on top of regulatory changes and guidances that will affect our clients’ business.  You will definitely feel our support when doing business with us.
 

What makes a scientific conference great? New, cutting-edge data from experts in the field, to be sure. To me, it’s equally important to have lively, stimulating, insightful discussion after the oral presentations; this is extremely rare, but it really helps illuminate the presentations and put them in perspective. I’ve come to expect both from the annual International Conference on Drug-Drug Interactions (DDIs) in Seattle, presented by the Institute for Scientific Communications, and this year’s edition (the 14th) was no exception. It’s not a large meeting by any means (~125 attendees), but apparently the optimal size for productive give-and-take.

DDI-2011 brought together leading experts from industry and academia in various aspects of drug-drug interactions. Many of the faces were familiar, as they attend this conference every year. Others were new, and they presented what were for me some of the most interesting results. Several people who were supposed to be at the conference were at an invitation-only meeting at the FDA to finalize the draft guidance on DDI testing. One recommendation will reportedly be the use of cryopreserved, rather than fresh, hepatocytes (the advantages presumably being pre-characterization and the ability to use the same batch over time with multiple test compounds).

If a clinical perspective on DDIs is your thing, you would have enjoyed the annual review of the DDI literature. Two clinical observations of note: the AUC of aliskiren is elevated 6.5-fold when co-administered with itraconazole, apparently due to inhibition of P-gp in the gut; a component of apple juice and orange juice inhibits gut OATPs, which decreases the exposure of many drugs, including atenolol, celiprolol, ciprofloxacin, fexofenadine, and pravastatin. Two different speakers discussed the fact that circulating metabolites of drugs frequently contribute to DDIs but are seldom evaluated in pre-clinical studies.

If you’re into preclinical prediction of DDI risk, there was plenty of information for you as well. Rheem Totah (University of Washington) thoroughly covered CYP2J2, an enzyme that is expressed in liver, intestine, and kidney, but at especially high levels in cardiac myocytes. Most CYP2J2 substrates are also metabolized by CYP3A4, often at the same position. And all of the most potent inhibitors of CYP2J2 cause QT prolongation and inhibit hERG, which raises the question of the role of CYP2J2 in QT prolongation. N.L. Simmons (University of Newcastle upon Tyne, UK) presented some compelling data on P-gp induction in human intestinal T84 cells. I was evidently not alone in my ignorance of the fact that digoxin, a P-gp substrate, also induces expression of the transporter.

If you’ve ever evaluated CYP induction in human hepatocytes, you know that sometimes a compound will be scored positive in some donors and negative in others. According to the results of a PhRMA survey of CYP induction practices in the industry, (i) if a compound is a CYP inducer in even one donor (but not in others) it should be considered an inducer, and (ii) if you only measure one endpoint, it should be mRNA.

Finally, have you ever observed apparent stimulation of transporter activity at low concentrations of a compound that is an inhibitor at higher concentrations? We have, although we didn’t know what to make of it. Similar observations (apparent allosteric modulation) were reported by two speakers, including a possible connection to cholestasis in the clinic for modulators of MRP2 in the liver.

You can be sure that Absorption Systems will continue to stay on top of cutting-edge research in DDIs and related fields, both by actively participating in conferences like this one and by presenting the results of our own research, as we often do.
 

The IQPC conference Clinically Relevant Drug Transporters, held in London June 28-30, was a forum for presentation of case studies and exchange of scientific thought related to drug transporters. Presentations pertaining to last year’s ITC white paper and forthcoming EMA and FDA guidelines sought to address the question on everyone’s mind, “What does the future hold?” Participants shared experiences on strategies to reduce compound attrition due to transporter-mediated toxicity and lack of efficacy. Most importantly, it was established that DDI prediction from early preclinical in vitro studies can significantly impact drug development and time to market. As Constance Hoefer, who co-hosted a pre-conference workshop on transporters with Absorption Systems, said, ”It’s never too early to start thinking about transporters.”
 
GSK’s John Keogh provided an overview of the ITC white paper and its impact while addressing the role of transporters in clinical development. He presented examples of late-stage compounds (sometimes even post-approval) for which the FDA required in vitro transporter interaction studies.
 
Speaking on transporter strategies and the large pharma business model, Praveen Balimane of BMS made an interesting point highlighting differences in transporter phenotypes that he aptly called “different sites, different strokes”. For example, being a P-gp substrate may have little impact on a compound with good permeability at the gut wall but the BBB may be a different story. He also referred to “the transporter effects we didn’t know about,” citing the example of mibefradil, a calcium channel blocker that was withdrawn from the market in 1998 due to time-dependent inhibition of CYP3A4. Subsequently, it was discovered that mibrefadil is also a potent inhibitor of both P-gp and OCT2. His key message was to perform in vitro drug transporter evaluation studies early to dial out undesirable chemotypes.
 
Another important topic was the impact of transporters on drug safety and the disposition of endobiotics. Many transporters such as ATP, BSEP and MRP2 play a key role in homeostasis, disruption of which can lead to toxicity. Dubin-Johnson syndrome is a classic example, where MRP2 deficiency results in hyperbilirubinemia. Promising compounds can fail in human trials due to elevated liver enzymes, which could be caused by interaction with transporters (e.g., interference with biliary efflux of bile acids).

With all the focus on small molecules, we should not forget that biologics can also precipitate drug interactions (TPDI: therapeutic protein-drug interactions). More than thirty blockbuster drugs on the market today are biologics; BMS, for example, has doubled its portfolio of biologics. It was recently reported that the Roche rheumatoid arthritis drug tocilizumab (Actemra®) significantly reduces the exposure of simvastatin due to de-repression of CYP expression via reduction of circulating cytokine levels. Transporters may or may not also be involved…we don’t know.

Sanofi representative Katharina Mertsch discussed the reliability and validation that she expects from CROs performing transporter studies as well as the risks and advantages of outsourcing.  She reported that the EMA will not accept vesicle results, requiring cell-based in vitro transporter data. It appears that the EMA is also stringent about the qualification of probe substrates since substrate-dependent inhibition, a phenomenon long recognized with CYP3A4, is increasingly seen with transporters.  Substrate dependence complicates the extrapolation of DDI risk to the clinic because I/K_i depends on the in vitro probe substrate.

Finally, several speakers addressed the issue of species differences: can observed differences in exposure between rodents and non-rodent species be explained by transporters? The technology is not quite there yet to answer the question.

If the question is “Which transporters?” clearly the take-home message from this conference was, to truly reduce the risk of late-stage failure, include all available human transporters in early in vitro screening.  Download the Absorption Systems ebook Transporter Reference Guide to learn all you need to know.

On June 10th, the National Institutes of Health (NIH) issued a statement regarding their adoption of the new Guide for the Care and Use of Laboratory Animals as discussed in my May 9th post in this space. The comment period closed on May 24 and they received an overwhelming response. Currently, NIH’s Office of Laboratory Animal Welfare (OLAW) is reviewing the comments. After an undisclosed amount of time, they will issue a position statement with another sixty-day comment period. If they do decide to adopt the eighth edition of the Guide as a standard, they are prepared to provide an implementation plan.

The NIH statement doesn’t say much, except that we will have to wait some time for their position statement given the number of comments and the concerns that adoption of the new edition of the Guide may hinder the progress of biomedical research in the US (as discussed previously). Similar to what AAALAC International has done, it appears that NIH will not expect full compliance with the new edition immediately, if in fact it is adopted as the standard, but may perhaps implement it in a stepwise fashion with their own interpretations of the new guidelines.

Copy of the NIH announcement:

Consideration of Public Comments on Proposed Adoption and Implementation of the Guide for the Care and Use of Laboratory Animals: Eighth Edition

The NIH has announced in Guide Notice NOT-OD-11-082 that the public comment period on the proposed adoption and implementation of the Guide for the Care and Use of Laboratory Animals: Eighth Edition closed May 24, 2011. OLAW is now reviewing comments submitted during the public comment period. When a thoughtful and in-depth analysis has been completed, comments will be posted on the OLAW website. Also at that time, OLAW will issue position statement(s) that will be posted on the OLAW website and a 60 day public comment period will commence to permit the public to provide feedback regarding their understanding of the position statement(s). If OLAW decides to adopt the eighth edition of the Guide, an updated implementation plan will be posted on the OLAW website. OLAW appreciates the effort of the public in providing numerous well-reasoned comments regarding the welfare of research animal subjects and wishes to reassure the community that all comments are being reviewed and considered.
 

Maybe the most important takeaway message for me from the third annual Ocular Diseases and Drug Discovery conference in Boston, MA at the end of May was the surprising amount of activity in drug development for diseases of the front of the eye, such as glaucoma and dry eye. No new classes of drugs to treat glaucoma have reached the market in more than 15 years, but many patients don’t respond to any given drug and most patients eventually need multiple drugs. One group of experts agreed that it is now difficult to find funding for age-related macular degeneration (AMD, a retinal degenerative disease) due to the fact that Lucentis and Avastin are so effective. Not that the back of the eye has been abandoned entirely: GSK, PanOptica, and Alcon are among the companies with drugs in the clinic to treat retinal diseases with topical administration.

There were presentations on all stages of ophthalmic drug development, including preclinical (PharmOptima and Absorption Systems for ocular pharmacokinetics; Sclera for efficacy with preclinical models of retinal and choroidal neovascularization), medical devices (a novel drug delivery device from iVeena that is implanted in the lens capsule and is showing promising results in rabbits), and clinical trials (the strategy and design of each stage of clinical trials of a Santen drug for dry eye disease that was launched last year in Japan), as well as reviews of dry eye disease and the use of non-human primates in ocular studies.

At least three companies (Altheos, Kowa and Aerie) have rho kinase inhibitors in the clinic for glaucoma, and Xcovery Vision is also developing kinase inhibitors for various ophthalmic indications.

GSK is ready to take a novel approach to AMD treatment into the clinic. The approach is to administer, systemically, an antibody vs. amyloid beta (Aβ), which accumulates in the retina in AMD and indirectly recruits activated complement. According to the “peripheral sink hypothesis,” the circulating therapeutic antibody should help clear Aβ from the retina. Preclinical results in mice are very encouraging, and they believe they have engineered the humanized antibody such that they will avoid the safety problems seen in previous clinical trials with anti-amyloid antibodies in Alzheimer’s disease.

I presented on stem cell-derived human corneal orbs, a tissue construct discovered by International Stem Cell Corporation. In characterizing corneal orbs as a model to predict the corneal permeability of drugs, Absorption Systems has obtained excellent correlations with excised human and rabbit cornea. Histologically, corneal orbs resemble corneal tissue, including stratified epithelium, Bowman’s membrane, stroma, and endothelium. Expression of several stem cell markers has been detected, along with tissue-appropriate drug transporters such as MRP1.

We are very excited about the future of human corneal orbs, not only as a corneal permeability marker but also for other potential applications and the potential for reducing the number of animals used in ocular drug testing. They exemplify the way we approach our business: cutting-edge research and innovative test systems, for all the right reasons. Stay tuned for the launch of this novel test system as part of Absorption Systems’ contract research portfolio. And contact us for all your ADME needs, ocular or otherwise.
 

Last month I attended and presented at the 37th annual conference of the Animal Transportation Association in Brussels, including a track on the transport of research-related materials, during which speakers outlined the current state of the shipping industry with regard to biomedical research. Routine procedures such as customs inspections and security screening of cargo by airlines and other shippers, as well as regulations regarding dry ice content, trans-national movement of biological materials, and labeling of containers can all be potential barriers to the movement of research materials to their destination on time and with integrity intact.

Biomedical research today is global; it is not feasible to complete all aspects of a new drug development program in a single location. Different analyses, procedures, and parts of studies must be performed in different facilities, domestically and worldwide, in order to take advantage of subject matter expertise. As a result, chemicals (test compounds) and biological specimens (e.g., plasma and other tissues) must be shipped from one facility to another. Shipping logistics then becomes paramount in importance when it comes to getting the materials to the right location at the right time. This is often much more complicated than it might appear.

Before shipping items critical to a drug research and development program, you should select the shipping vendor with care, basing your decision on their experience with your type of shipment (e.g., ability to handle packages containing dry ice), capabilities (e.g., ability and willingness to add more dry ice in case of customs delays), reputation for reliability and on-time delivery, proper handling and storage, as well as cost. Obtain references if you’re not sure. Have a backup plan, as a single international incident can stop cargo shipments at any time. Shippers that specialize in time-sensitive shipments of drugs and biologics can help with customs paperwork and guarantee timelines as well as proper package handling by personally attending to the package from beginning to end.

Did you know that, whatever size box you use for a dry-ice shipment, it should be completely filled with dry ice? If you only need 5 lbs. of dry ice, don’t use a box that would hold 20 lbs. But if you use a larger box than you need, fill it with dry ice; the process of sublimation (change in phase from solid to gaseous CO2) takes place much more readily in a container with plenty of empty headspace.

Did you know that a separate CITES permit is required for each international shipment containing one or more specimens (e.g., plasma samples from a PK study) from an endangered species such as cynomolgus monkeys? If we are doing an “in-life only” non-human primate study and shipping samples overseas to you for bioanalysis, we will take care of the CITES permit, but if you are sending such specimens to us the permit is your responsibility.

Absorption Systems recognizes the critical nature of shipping, and we do not take it lightly! We work closely with the various shipping companies and track packages while in transit. We are also well aware of the regulatory requirements for shipping various types of research specimens. If your study requires shipping a unique type of specimen, one that we have not handled previously, we will work out the logistics well in advance to ensure the best outcome for the study. Post a comment to let me know if you have any other shipping “best practices” that might not be obvious to other readers.
 

On the 16th of June, the Therapeutic Products Directorate of Health Canada and the Canadian Society for Pharmaceutical Sciences organized a workshop on the Biopharmaceutics Classification System (BCS) to improve their understanding of how the US and EU apply biowaivers in practice. Experts from academia, industry (including Absorption Systems), the FDA, the EMA, and provincial formularies were brought in to share their viewpoints and provide information for use by Canadian regulators in drafting a biowaiver guidance by the fall of 2011, to be finalized by early 2012.

The first few talks established the rigorous science that forms the foundation of the BCS; discussed various non-clinical intestinal permeation techniques available for permeability classification; and analyzed case studies using in vitro test systems as a safe, suitable, and conservative approach for establishing the bioequivalence of pharmaceutically equivalent drug products. Emphasis was also placed on the applicability of this tool outside the regulatory context and the design of an improved drug development path that could save substantial time and cost. Speakers from innovator and generic drug developers underscored the utility of the BCS within their respective branch of the industry. Specific examples of biowaivers shared in the forum included an API with high risk of adverse drug reactions when administered to healthy volunteers, a highly variable drug that would require many more subjects for a clinical BE study, and an alternative to an in vivo fed/fasting study.

The afternoon session provided insights from the FDA and the EMA. Both agencies accept biowaivers for Class I drug substances, but the threshold for complete absorption is lower for the EMA (85% fraction absorbed) than the FDA (90%). The FDA speaker clearly outlined the BCS review process and presented several statistics on biowaiver requests. Two obvious trends were the greater number of biowaiver submissions over the past three years and the wider use of in vitro test systems for permeability. The EMA, however, currently prefers human PK data for absorption, a policy that seems counter-intuitive to the central BCS tenet of reduced human testing. This emphasis on in vivo testing ignores the intrinsic variability of human BE studies, in particular the difficulty of obtaining reliable measures of bioavailability or fraction absorbed for highly variable drugs, and overlooks the body of work that has repeatedly shown biowaivers to eliminate unnecessary human drug exposure while maintaining the high standard for therapeutic equivalence. Differences in solubility requirements between the agencies were also highlighted; in my opinion, a safe and practical compromise would be to require complete solubility of the highest single dose in 250 mL or less of aqueous medium between pH 1 and 6.8 (from the EMA) within a timeframe and for a duration consistent with the physiological context (e.g., three hours, previously accepted by the FDA per a risk/science-based assessment).

Finally, a representative from one of the provincial formularies concluded that the BCS is ‘scientifically supported,’ that its introduction into Canada ‘will be judged by the level of transparency associated with its implementation,’ and that the system is ‘consistent with the provincial risk-based review process’. In other words, the scientific framework is tested and valid, but educating the industry/regulators/health care professionals is a key for success. Toward this end, Absorption Systems is working on a BCS road show that will empower all stakeholders with the knowledge necessary to unlock the full potential of the BCS for biowaivers and beyond. Let us know if you are interested, and we’ll let you know when the road show will be in your area.

An article published in the May issue of the journal Drug Metabolism and Disposition reveals a seasonal variation in the serum concentrations of the CYP3A substrates sirolimus and tacrolimus, which is temporally and inversely correlated with UV-B light exposure and vitamin D. The investigators, from the Karolinska Institute in Stockholm, extracted clinical data by date for the two drugs (and two other immunosuppressants, cyclosporine A and mycophenolic acid) from the hospital’s therapeutic drug monitoring database. Dose-adjusted drug concentrations were compared with the average monthly UV light exposure in the Stockholm area and the monthly variation in serum vitamin D concentration in a separate cohort of Scandinavian patients.

As you would expect at high latitude, UV light is highest in the summer months (May-August) and much lower (nearly zero in some months) in the winter (November-February). 25-OH-Vitamin D, the biosynthesis of which requires UV light, followed a similar cycle, with a slight delay: highest in July and August, lowest in January-April. The concentrations of sirolimus and tacrolimus were the mirror image of the vitamin D concentrations: lowest in the summer and highest in the winter. There was no such relationship for mycophenolic acid, a negative control that is not metabolized by CYP3A. Surprisingly, there was also no significant seasonal variation for cyclosporine, which is apparently an artifact of the lack of specificity of the immunoassay that was used to quantify it, which also picks up cyclosporine metabolites.

In spite of some obvious shortcomings (i.e., the immunoassay problem and the fact that the vitamin D data came from a different patient cohort), this is a potentially groundbreaking study. It has been known for years that there is considerable intra- and inter-individual variability in CYP3A4 expression and activity, and the reason is still unclear. How much of that variation can be explained by variations in vitamin D? While the authors are careful to point out that no causal relationship has been proven, the temporal relationship is quite compelling.

According to Ken Thummel of the University of Washington, who has published extensively on induction of CYP3A4 by vitamin D, organ transplant surgeons handle such variations empirically, by frequent monitoring of immunosuppressant levels and adjustments on the fly as needed. So, while they might be interested in this phenomenon, it wouldn’t change the way they practice medicine.

Unfortunately, when the media gets hold of a story they often get it a little wrong. In this case, an online outlet called in-pharmatechnologist.com ran the story with the headline “Swedish Researchers Uncover Link between Sunlight and Drug Efficacy.” If you’ve read carefully up to this point, you’ve probably realized that the study only measured drug concentrations (pharmacokinetics), not drug efficacy (pharmacodynamics). That’s very fortunate in this case, because lack of immunosuppressant efficacy in the summer would presumably be associated with organ transplant rejection. It probably only bothers me because I’m a pharmacologist, but I still think a trade publication should know the difference. Rest assured that Absorption Systems does!

Let me know what you think about this study.

In a recent blog post, I discussed potential changes to the GLP regulations being proposed by the FDA.  Now, more change is on the way for the industry, including Absorption Systems’ AAALAC- accredited vivarium and any other institution with an AAALAC-accredited vivarium.

After soliciting and incorporating input from experts, the much-anticipated 8th edition of the Guide for the Care and Use of Laboratory Animals, first published in 1963 and most recently updated in 1996, was released in draft form in 2010.  The final version was published in April of this year.  It is internationally accepted as a primary reference on animal care and use.  This revision was produced primarily to address some of the significant changes occurring in animal husbandry and in-vivo research today, such as the use of ventilated rodent racks and genetically modified animals.  While the Association for Assessment and Accreditation of Laboratory Animal Care (AAALAC) International has adopted the new guidelines, other organizations have not been so quick to jump on board.