Xenograft models have long been the gold standard method for preclinically assessing the relationship between PK/PD and efficacy. However, technological advances have unlocked an alternative complementary approach!
Drug development, preclinical studies and PK
In drug development, preclinical studies are crucial to informing decisions before clinical trials. Whilst there may be differences in the experimental approach, preclinical studies are required to define drug pharmacokinetics (what the body does to the drug), pharmacodynamics (what the drug does to the body) and toxicology/safety profiles1.
Initial studies are often performed in vitro to screen compounds. These convenient, quick and reproducible screening approaches are valuable; however, their value is limited by two important factors. Firstly, the simplistic nature of 2D cell cultures, which poorly replicate human biology. Secondly, fixed concentration drug dosing poorly replicates the concentration changes that occur within the body after a drug is administered (PK profiles), when the concentration in the bloodstream initially rises as it is absorbed and distributed, then falls as it is metabolised and excreted. The dynamic nature of this process means that the concentration of a drug at the target is difficult to estimate and continuously changing over time. The problem becomes even more complex for combination therapies, a cornerstone of cancer treatment where two, or more, drugs are administered together. Therefore, animal xenograft models are often utilized to enable preclinical PK/PD/efficacy studies.
The use of xenografts in oncology
Using animal models for the preclinical testing of cancer drugs has long been the gold standard. These studies typically use immunodeficient mice as hosts for tumor xenografts. Alternatively, cancer cell line-derived xenograft models (CDX) involve grafted immortalized cell lines developed in vitro to mimic tumors for anti-cancer drug development2. Additional models use segments of an actual patient’s excised tumor that are grafted into the mouse (patient-derived xenografts, PDX)3. The latter has the added benefit of containing tissue from an actual patient, with the aim that the preclinical data derived using the model would better predict outcomes in the patient.
These xenograft methodologies are widely used in preclinical cancer drug development; however, they all suffer from similar limitations – they don’t replicate human pharmacokinetics! The in vivo concentration changes of a drug (it’s PK profile), generally differ significantly between the animal model used to replicate the human response and an actual human. Conventional fixed dosing in 2D cultures, is far worse in terms of physiological relevance as these models offer no/little concentration change in drug dose over time – see image below.
Technological advance & in vitro preclinical studies
There is hope on the horizon as technological advances have now unlocked the capability to perform more complex studies in vitro to address the poor replication of human biology within 2D cell culture. Microphysiological systems (MPS), otherwise known as organ-on-a-chip, improve the replication of cellular microenvironments by enabling the formation of more human-relevant tissues for use in drug testing4. Similarly, it is postulated that more human-relevant responses to drug dosing can be studied through better in vitro replication of the concentration changes that occur following a drug’s administration.
Studying the response of a lab-grown human organ to actual concentration change profiles with time in a preclinical in vitro setting would be incredibly valuable to drug development programs. It would enable the study of known PK profiles, but also the study of theoretical profiles for potential adaptations of drugs, or the study of the concentration and time effects not possible in animal models – all of which will positively contribute to a more accurate, efficient and successful drug discovery process.
Replicated PK dose vs traditional fixed dose and in vivo studies
A recent study in PLOS Biology introduced the Microformulator, a microfluidic device capable of replicating programmable PK profiles in vitro5, that has since been renamed the PhysioMimix™ PK by CN Bio. The Microformulator controls the movement of media and drug-containing media within the well of a standard well plate to dilute the concentration of drug over time in a programmable manner.
Many different drugs can be diluted within the well at any one time, enabling the study of combination therapies, which are prevalent in oncology. By adjusting the timing of combination dose therapies, it is possible to interrogate the effect of different dosing strategies on established in vitro cellular models.
The publication set out to demonstrate the system’s utility through a series of studies. The first study highlighted the device’s ability to identify cell lines with different sensitivities to compounds, known to be different in vivo but with little/no difference observed in traditional fixed concentration dosing. The results of these experiments highlighted the improved capability of the device over traditional fixed concentration dosing.
A further study within the publication focussed on whether the system was able to recreate data from ‘gold standard’ xenograft models. The device can mimic most PK profiles; therefore, murine PK profiles were replicated to simulate the concentration changes in the xenograft. The study focussed on combinations of PARP inhibitors and DNA-PK inhibitors, which had been demonstrated in the literature to have increased effectiveness when used in collaboration. Combinations were dosed once a day and twice a day, respectively. The in vitro results using the Microformulator demonstrated regression of the tumor cell model in the combination therapy group from day 10 and reduced growth in the monotherapy groups compared to the vehicle control.
Interestingly a similar result was observed in the xenograft models, whereby the relative tumor volume showed signs of regression from day 7 and increased regression on day 10 in the combination therapy group. Also, the same rank order effectiveness was observed in the other treatment groups when comparing the Microformulator data to the xenograft data. Together, these data demonstrate the unique ability of the device to replicate PK profiles that stimulate in vivo like responses within an in vitro setting.
Oncology services
At CN Bio, we provide cutting-edge technologies that enable our customers to perform preclinical studies that replicate the human body and its responses to drugs. We have licenced use of the Microformulator from Vanderbilt and Northwestern Universities and utilize it within our Oncology Services program to apply the PK profiles of known/customer compounds to tumor models. Many oncology therapeutics are more effective when dosed together; therefore, we also offer the ability to apply multiple PK profiles to ascertain optimal dose combinations. Additionally, the effects of differing dosing schedules can also be evaluated. Furthermore, the technological capability of the device enables us to replicate known PK profiles from any species or to perform concentration change profiles that are completely made up to test specific scientific questions. By improving the human relevance of drug dosing, this technology unlocks the door to many interesting studies and brings us ever closer to bridging the gap between in vitro preclinical studies and actual drug responses in vivo.
Contact us to understand how we can help study your compound’s responses to PK profile replicated doses today!
Steinmetz, K.L. and Spack, E.G., 2009. The basics of preclinical drug development for neurodegenerative disease indications.BMC neurology,9(1), pp.1-13.
Richmond, A. and Su, Y., 2008. Mouse xenograft models vs GEM models for human cancer therapeutics.Disease models & mechanisms,1(2-3), pp.78-82.
Lai, Y., Wei, X., Lin, S., Qin, L., Cheng, L. and Li, P., 2017. Current status and perspectives of patient-derived xenograft models in cancer research.Journal of hematology & oncology,10(1), pp.1-14.
Rubiano, A., Indapurkar, A., Yokosawa, R., Miedzik, A., Rosenzweig, B., Arefin, A., Moulin, C.M., Dame, K., Hartman, N., Volpe, D.A. and Matta, M.K., 2021. Characterizing the reproducibility in using a liver microphysiological system for assaying drug toxicity, metabolism, and accumulation.Clinical and translational science,14(3), pp.1049-1061.
Singh, D., Deosarkar, S.P., Cadogan, E., Flemington, V., Bray, A., Zhang, J., Reiserer, R.S., Schaffer, D.K., Gerken, G.B., Britt, C.M. and Werner, E.M., 2022. A microfluidic system that replicates pharmacokinetic (PK) profiles in vitro improves prediction of in vivo efficacy in preclinical models.PLoS Biology,20(5), p.e3001624.
