Feliza Mirasol is the science editor for 

Feliza Mirasol

On March 10, 2023, Pfizer announced that FDA approved ZAVZPRET (zavegepant), which, according to a company press release, is the first and only calcitonin gene-related peptide receptor antagonist nasal spray for migraine. The drug is approved for the acute treatment in adults of migraine with or without aura.

FDA’s approval is based on two pivotal randomized, double-blind, placebo-controlled Phase III studies that established the drug’s efficacy, tolerability, and safety profiles. The results showed that ZAVZPRET was statistically superior to placebo on the co-primary endpoints of pain freedom and freedom from most bothersome symptom at two hours post-dose. Study results also demonstrated that pain relief was achieved as early as 15 minutes in a prespecified secondary endpoint versus placebo. Following approval, the drug is expected to be available in pharmacies in July 2023.

“The FDA approval of ZAVZPRET marks a significant breakthrough for people with migraine who need freedom from pain and prefer alternative options to oral medications,” said Angela Hwang, chief commercial officer, president, Global Biopharmaceuticals Business, Pfizer, in the press release.

“Among my migraine patients, one of the most important attributes of an acute treatment option is how quickly it works,” said Kathleen Mullin, associate medical director at New England Institute for Neurology & Headache. “As a nasal spray with rapid drug absorption, ZAVZPRET offers an alternative treatment option for people who need pain relief or cannot take oral medications due to nausea or vomiting, so they can get back to normal function quickly.”

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FDA has approved Pfizer’s ZAVZPRET (zavegepant), a new nasal spray treatment for migraine. 

FDA Approves Pfizer’s New Migraine Nasal Spray

Bioassays play a crucial role in biologic drug development as a means to determine potency, toxicity effects, pharmacological activity, stability, purity, and other product qualities. The complexity of biological molecules, however, requires a wide range of analytical procedures to adequately characterize the product. In particular, the concern over immunogenicity in the use of biologics is driving the need to develop more highly sensitive assays. Because of this concern, there is a continual need to refine biological assays to increase accuracy and reproducibility. There is a specific need to replace 

 bioassays are important tools employed during drug discovery/drug development, according to Weihong Wang, PhD, director of Technology Development, Eurofins BioPharma Product Testing. 

 bioassays use cell culture and offer the benefit of low cost and high throughput, both of which are essential during early development. An 

 bioassay also has the necessary robustness needed for use as a lot-release assay for product quality control later in the drug development life cycle, Wang explains.

 bioassay is a more efficient method that allows researchers to clearly characterize product mechanism of action in a controlled environment, says Catherine Liloia, director, Cell & Microbiology, PPD clinical research, Thermo Fisher Scientific. “This approach is scalable and reproducible across production and can assess product stability,” she states.

 bioassays (non-clinical) use animal models and are considered more relevant to potential clinical outcome, says Wang. 

 bioassays are often costly, however, and are less robust and subjected to strict regulations and compliance standards. “Regulatory agencies across the world have been encouraging sponsors to replace, reduce, and refine (the three Rs) dependence on animal studies by advancing development of, and evaluating new, fit-for-purpose methodologies,” Wang explains.

 bioassays are more representative of the full biological complexity and pharmacokinetics of the human body; they can, therefore, more broadly assess potential patient safety impacts stemming from metabolic changes and other side effects. The downside, however, is that 

 bioassays require high investment, strict animal care protocols, and are not as conducive to high-level analytics, according to Liloia.

, generate valuable data needed for regulatory filings, such as investigational new drug (IND) applications. 

 bioassays support determination of product potency, for instance, says Liloia. “When included in analytical panels during release and stability testing, potency data can demonstrate lot-to-lot consistency and long-term stability of the product,” she says.

In addition, bioassays provide data to better understand the impact of manufacturing process changes through process development. “Early 

 methods may be needed to help discern true product mechanism of action in biological pathways that are complex or not well characterized. These methods can sometimes be consolidated or replaced on a product release panel as data [are] collected to determine a single method’s capability to reliably determine product potency across production samples,” Liloia explains.

 method cannot by itself model the biological response in a reliable manner, as is the case with immunogenicity and other safety testing, Liloia adds.

IND-enabling studies, including pharmacology/pharmacodynamic, pharmacokinetics, and toxicology studies, are typically done through both 

 assays, states Wang. She adds that additional 

 assays to address product safety and quality, such as viral testing, and potency testing, are also necessary later on for product licensing.

Cell-based bioassays have also become an important tool in the analytics arsenal for biologics drug development. According to Wang, cell-based bioassays measure relevant physiological responses in cell cultures, but unlike 

 bioassays, cell-based bioassays do not involve the use of live animals and often have different types of assay readouts/endpoints. “Nevertheless, the physiological responses are reflective of mechanism of action, and, therefore, cell-based assays are considered necessary components of product characterization, in addition to other biophysical/physiochemical analytical methods,” she states.

Cell-based bioassays utilize biologically relevant cell lines to measure product potency, says Benjamin Ziehr, senior group leader, PPD clinical research, Thermo Fisher Scientific. He notes that there are several types of cell-based bioassays that vary depending on what is being measured. “The classic cell-based bioassay compares the response of cells treated with drug sample or reference standard. Typically, sample and standard are diluted in a manner suitable to produce a four-parameter response curve in the target cell line. Well-defined response curves are critical for maintaining and demonstrating control of the assay and drug product in these assays,” he explains.

Ziehr further explains that the relative activity of sample drug material to reference standard drug material is reported as the relative potency of the drug product. In comparing cell-based bioassays to other types of bioassays, the key distinction lays in the fact that cell-based bioassays allow mechanism-of-action-dependent measurements, whereas other bioassays measure physical characteristics of the drug product, such as epitope binding or receptor occupancy. “Therefore, cell-based bioassays allow measurement of the cellular processes responsible for a drug product’s effects on the patient,” says Ziehr.

Meanwhile, there are several other modes of cell-based bioassays that provide information about other attributes of the drug product. Ziehr points out two common examples of these other modes from cell and gene therapy product testing: infectivity and replication competency assays. “These assays measure the infectious potential of the drug product and the presence of replication-competent viral particles, respectively,” he states.

In other words, rather than measuring a cellular response, these infectivity and replication competency assays measure the accumulation of viral genetic material following infection of permissive cell lines. “While these assays don’t measure product potency, they do provide critical process and safety data and detect lot-to-lot variability of products,” says Ziehr.

Cell-based bioassays become a clear front runner in this movement, where applicable, remarks Wang, who points out that, in other circumstances, sponsors may be able to eliminate certain 

 assays based on risk analysis. For example, per the International Council for Harmonisation Q5A(R2) (2), 

 testing is no longer necessary for well-characterized cell lines such as Chinese hamster ovary, non-secreting murine myeloma, and SP2/0 (a standard mouse myeloma cell line), based on cell-line history, prior knowledge, and other risk-based considerations, notes Wang.

Despite their advantages, however, cell-based bioassays are limited in their ability to represent more complex, distal biological responses in a patient, cautions Liloia.

Meanwhile, the challenges in working with cell-based bioassays include not only the fickleness of working with certain cell lines, but also the fact that the day-to-day variability is much higher in terms of assay performance, observes Ziehr. The state in which cells enter the bioassay affects their response in the bioassay.

“Cellular state is dictated by many factors, notably metabolic stimuli, cellular density—both in culture leading up to a bioassay and in the bioassay itself—and cellular age. Stable transfections, optimized banking, and robust maintenance controls are needed to minimize cell variability,” Ziehr says.

Ziehr explains that, typically, robust maintenance controls involve the use of well-defined and controlled culture protocols, qualification of critical reagents, and substantial system suitability criteria in the bioassay. “Consistency of technique is challenging, even with clear procedure, so robust training with clear connection between biological processes and cell handling technique remains important,” he remarks.

Because cell-based assays utilize live cells, they are subjected to intrinsic assay variability that are often larger than that of physiochemical methods, says Wang. Thus, the selection of an appropriate cell line that is not only relevant to product mechanism of action, but also able to support a robust response in a dose dependent manner, is the single most critical factor for successful method development, she emphasizes.

“Many details need to be considered and optimized to produce a robust assay suitable for quality control. In addition, for products that may have multiple mechanisms of action, a carefully selected matrix containing multiple cell-based assays may be necessary to fully characterize the product,” Wang says.

Wang also points out, however, that it may yet be difficult to develop a cell-based bioassay for some products that either have not known defined mechanisms of action or have complex mechanisms of action. Furthermore, some cell-based assays still require extended testing time, which can be a cause for concern regarding products that require rapid factory-to-patient delivery. In such cases, Wang suggests that development of surrogate assays and/or orthogonal assays may be beneficial, though subject to discussions with the relevant regulatory agency.

In terms of advantages conferred, compared to 

 bioassays, cell-based bioassays often offer the advantages of a more streamlined testing procedure, higher assay throughput, quicker turnaround time, and less assay variability, according to Wang. “It is generally easier to develop a cell-based assay with an accurate quantitative endpoint than an 

 assay, which is particularly essential when the method is intended to be used for quality control,” she states.

Liloia notes that cell-based methods are quite challenging in many areas. Solutions such as liquid handling automation helps in providing accurate and precise sample dilution pipetting and transfers to the low concentrations and small volumes often needed in a cell-based assay. She also notes, however, that analytical workflows have become more complex because of the diversity of analytical readouts needed to support expanding product areas.

The growing complexity of analytical workflows has resulted in a greater need for coordination across functional areas, broadened cross-training, and design changes to laboratory spaces to maintain efficiency while also maintaining process flows that consider biosafety and contamination prevention, Liloia says. “Data analysis expectations continue to evolve as laboratories and agencies gain experience with newer products, requiring strong statistical support and the ability to apply industry best practices in a new way,” she states.

Overall, cell-based bioassays are considered a critical tool to predict quality and safety of drug products in the patient. “When established early in the development process, cell-based bioassays can provide early indicators of lot-to-lot variability in manufacturing, drug potency stability, and impact from process changes,” states Ziehr.

“Bioassays have an essential role throughout the life cycle of product development of biologics.Many aspects of product discovery, development, characterization, and quality control require careful construction and implementation of bioassays in order to thoroughly assess product safety, efficacy, and stability,” Wang adds.

1. Mire-Sluis, A.R. Progress in the Use of Biological Assays During the Development of Biotechnology Products. 

, 18, 1239–1246. DOI: 10.1023/A:1013067424248.
2. ICH, Q5A(R2) 

Viral Safety Evaluation of Biotechnology Products Derived from Cell Lines of Human or Animal Origin

As the development of increasingly complex biologics grows, more robust bioassays are required to accurately characterize these molecules.

Robust Bioassays Needed for Increasingly Complex Analytical Workflows

An understanding of the pharmacokinetics (PK) of monoclonal antibodies (mAbs) is important so that potential patient responses to treatment can be predicted. Both PK and pharmacodynamic (PD) studies are necessary in the development of a new mAb drug candidate. 

PKPK/PD modelling is already widely used in pre-clinical and clinical drug development; these studies are important for characterizing drug candidates quantitatively as well as aiding go/no-go decisions and informing future trial design and optimal dosing regimens (1). However, what is less well known is that PK/PD modelling can also be an effective tool in the earliest stages of drug development, such as in target selection and lead candidate selection. The same modelling techniques can be used to predict and illuminate the drug candidates that have the necessary characteristics for potentially successful development, or, conversely, can help pinpoint which drug development programmes may be less successful, before too much time and cost are accrued on that specific programme.

In particular, mAbs, which are an important therapeutic class, have complex pharmacology and interdependent PK/PD properties. Understanding the PK and PD of mAbs and their biological activity as well as their mechanisms of action are crucial factors in enabling their design and selection. Understanding these factors allows for the design of appropriate studies to characterize the mAb’s efficacy and toxicity, the translation of PK/PD parameters to humans, and the optimization of a dose and regimen for maximizing success in clinical development (2).

Meanwhile, the practice of characterizing the relationship between PK and PD is an important tool in the drug discovery phase. PK/PD strategies should be implemented in the early phases of research during drug discovery projects because these studies can enable a successful transition from discovery phase to development phase (3). An effective PK/PD study design, as well as proper analysis and interpretation can help researchers characterize the PK–PD relationship. Understanding the relationship would enable understanding of the mechanism of drug action and help identify PK properties that can help improve and optimize the design of the drug compound (3).

To learn more about the necessity of PK/PD studies in early mAb development and the most reliable ways to conduct these studies to generate data compliant with investigational new drug application (IND) filings, Pharmaceutical Technology Europe spoke with Maotian Zhou, PhD, Principal Scientist—DMPK Services Department, at WuXi AppTec.

 In the early development of a new mAb, what role does PK play in the development lifecycle?

 PK plays an important role in the early development of mAb therapeutics. This is even more apparent when the antibody is designed against a novel target, or if the antibody aims to be a ‘best in class’ drug in an already competitive arena with well-studied targets (i.e., HER2, PD1 [programmed cell death protein 1]/PDL1 [programmed cell death ligand 1], etc.).

This may differ from past views on mAb drug development. It is usually assumed that, for humanized monoclonal antibodies, the 

 absorption, distribution, metabolism, and excretion (ADME) characteristics share more significant commonality than small-molecule drugs. Thus, the importance of PK studies in the early phase of mAb development is diminished. The efficacy (or pharmacodynamics) appears to be more significant in the leading-candidate mAb selection.

The importance of PK studies increases with a better understanding of mAb drug mechanisms. Many factors can affect mAb exposure–response relationships 

, the affinity between the mAb and its ligand, the association between the mAb and different Fc receptors, and the modifications and physicochemical characteristics of the mAb. These factors form the basis for the continuous refinement of current mAb drugs. The mAb PK properties act as a bridge between its design philosophy and ultimate efficacy and safety. Fully understanding candidate mAb PK parameters can provide support for leading-mAb drug selection.

In summary, the PK studies significantly impact whether a mAb moves forward, and should happen early in discovery. Pushing a mAb toward an IND submission without fully understanding its PK could waste time and resources and/or result in programme failure.

 I always see the terms PK and PD together when discussing drug candidate screening or in drug discovery. What is the importance of both in mAb development, and is it necessary to have both PK data and PD data (can you have one without the other)?

 Simply put, PK is ‘what the body does to the drug’, whereas PD is ‘what the drug does to the body’.Explicitly, PK quantitatively describes the process of absorption and disposition of the drug in the body. PD evaluates the time course of the pharmacological effects of drugs.

For mAb drug development, the PK and PD always communicate, because mAb drugs generally have long-acting PD effects, and PD will have a direct impact on PK. PK and PD are mechanistically linked (unlike small-molecule drugs). Having both PK and PD data for mAb development is strongly recommended. PK/PD studies serve distinct roles at different stages of mAb drug development.

 What is a ‘best practice’ approach to conducting PK/PD studies, and do these studies necessarily need to be compliant with regulatory standards?

 PK/PD studies should adapt to the different requirements of each research stage. In the discovery phase, the PK/PD assessments facilitate an understanding of the biology by validating and confirming target engagement. For the leading-mAb selection stage, PK/PD studies should help to select the most ‘potential’ mAb, along with efficacy and toxicity evaluations, to minimize the risk of off-targeting. At the IND stage, systematic PK/PD studies, along with safety studies, play a vital role in translating pre-clinical knowledge to first-in-human clinic studies. In the clinical phase, further in-depth PK/PD studies can identify the covariates and potential sources of population variability, establishing exposure–response relationships in different cohorts. And finally, these studies support proper dose selection, regimen optimization, and biomarker tools establishment.

PK/PD studies must comply with regulatory guidance, both during and after the IND phase of mAb drug development. It is also helpful to reference the regulatory standards in the early phase of drug development. Given that regulations are often adapted based on the lessons learned from many actual drug development cases, items rooted in regulatory guidance are often representative of key points in drug development. For example, a mAb PK bioanalysis in the discovery phase, using a partially validated method (validating specific parameters such as selectivity and stability) will make the mAb quantification more reliable in an unconventional matrix.

 What role or impact do PK/PD studies have on an IND filing for a new mAb?Are they even relevant at that stage?

 PK/PD studies are essential materials for an IND filing. PK/PD studies serve the following purpose in the IND phase for mAb development: integrate pharmacology and toxicology data and help correlate pre-clinical knowledge to first-in-human clinical study; bridge toxicology studies to define maximum tolerated dose (MTD) and select first-in-human dose; design a dose escalation scheme; and project efficacious dose in clinical.

 What currently are the major analytical/technical challenges in conducting PK/PD studies on new mAbs?

 Several challenges currently exist within PK/PD studies for mAb drugs. First, the assumptions used to simplify the exposure–response relationships in PK/PD modelling may not always be appropriate. For example, in the past, the PK/PD model assumed the drug concentration in tumor cells was related to the cell-killing effect. However, in the absence of an actual measurement of the distribution cascade, the validity of this assumption is uncertain.

Second, the translation from animals (pre-clinical) to humans (clinical) based on PK/PD modelling is also challenging. Furthermore, existing PK/PD modelling assumptions are usually based on prior experience, as such studies are needed to understand the applicability of new mAbs.

Third, the reliability of PK/PD data depends on the selection of a proper analytical method/platform, which may be a challenge in some instances. The bioanalysis should always follow a fit-for-purpose principle. Based on the ligand properties (such as solubility and endogenous concentrations), the availability of critical reagents, and desired sensitivity, a suitable assay format needs to be designed.

There is an ongoing effort regarding PK/PD analytical strategy optimization. The ability to accelerate research and develop safe and effective mAbs rely less on tools and more on an investigator’s skills, knowledge, and expertise. Drug developers and sponsors that do not have the time, capacity, or expertise to conduct these investigations in-house should consider working with a testing partner.

 77 (5) 740–745 (2014).
2. A.V. Kamath, 

 21–22, 75–83 (2016).
3. T. Tuntland, et al., 

PK/PD studies are a crucial tool in the successful development of biologics.

PK/PD Studies Prove Critical for mAb Development

In this exclusive Drug Digest video interview, Meg Rivers and Feliza Mirasol, Pharmaceutical Technology editors, discuss the benefits of automating entire workflows in analytics (laboratories) versus automating only specific pieces of equipment or only certain processes. What influence or impact would this have on workflows further downstream, such as clinical or commercial manufacturing?

They discuss additional topics, including:

Additional automation approaches and how modular approaches fit into this

Automation opportunities in pharma (i.e., small molecule) vs. biopharma (i.e., large molecule)

Whether the level of adoption of automation differs in the industry between pharma and biopharma

How this trend toward automation in laboratory workflows impacts clinical and commercial drug development and manufacturing


is the head of product at Automata, a London-based biotech company providing robotic automation solutions to the life sciences industry. He is a trained mechanical engineer and studied at Oxford University. Pattinson joined Automata to help make real, tangible change and help unlock the potential of scientists through automation in laboratories.

This episode of Drug Digest is sponsored by:

Drug Digest is a tech talk video series with the 

 editors, who interview industry experts to discuss the emerging opportunities, obstacles, and advances in the pharmaceutical and biopharmaceutical industry for the research, development, formulation, analysis, upstream and downstream processing, manufacturing, supply chain, and packaging of drug products.

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Pharmaceutical Technology editors interview an automation expert from Automata on the benefits of automating entire workflows in analytics versus automating only specific pieces of equipment or only certain processes.

Drug Digest: Automation in Biopharma and the Impact on Workflows


Vol. 34, No. 12
December 2022
Pages: 16–17

When referring to this article, please cite it as F. Mirasol, “The Power of PK/PD in mAb Development,” 

Although well known in R&D, PK/PD studies are an equally powerful tool for promoting successful biologic drug development.

The Power of PK/PD in mAb Development

A look back at challenge and innovation highlights in 2022 for oral solid dosage forms, this podcast episode will review industry experts’ takes on factors driving the evolution of OSD forms as well as innovations in APIs, technology, and methodology. Highlights include commentary from experts at LGM Pharma, Hovione, Syntegon, and Colorcon.

, vice-president R&D and Regulatory Submission, LGM Pharma, joined LGM in 2020 to lead the company’s integrated R&D and regulatory submissions teams for CDMO clients. He has launched commercial products with several US and EU patents and handled submissions for NDAs, ANDAs, BLAs, and medical devices. Prior to joining Nexgen in 2015 to lead the R&D and Regulatory groups, Deepak was managing director at Actavis, chief scientific officer and vice-president of pharmaceutical development at PharmaNova, chief scientific officer and senior vice-president of pharma development at Holopack International, and associate director of Global Pharmaceutical Development and Technology at UCB Pharma. He earned his master’s in Pharmaceutics from Saugor University and his PhD and post-doctoral training from the University of Cincinnati in Pharmaceutical Sciences. He earned MBAs from Cornell University—in general business administration and finance—and Queen’s University.

, senior vice-president of Sales & Marketing, Hovione, graduated from the Chemical Engineering department at the Technical University of Lisbon, where he also obtained a PhD in chemistry in 2006. In 2007 he joined the Hovione R&D department as a scientist in the particle design discipline. The focus of his work was the application of particle engineering technologies to improve bioavailability of poorly water-soluble drugs. In May 2011 he was appointed the director of R&D Process Chemistry and was responsible for a group of dedicated scientists focused on the development and scale-up of chemical processes for the production of active ingredients. He has held several management responsibilities, including site general manager of Hovione operations in the United States and commercial-related functions. He is author of more than 15 scientific papers in peer-reviewed journals and book chapters, co-author in two patents, and an invited speaker in more than 10 international conferences.

, business development manager for the Huttlin portfolio line at Syntegon. He spent five years with Eli Lilly and Co. working on formulation and process development work for oral solid dosage forms before joining Syntegon in 2021. Trombley is experienced in early phase formulation development, process identification, scale-up, and technology transfer and has worked extensively with continuous manufacturing, high shear granulation, fluid bed granulation, tableting, and coating. At Syntegon, he is focused on technical support, customer trials, and market development for Huttlin products in North America. He studied chemical engineering with a minor in computer science at the University of Michigan.


 is the global lead authentication for Colorcon and is responsible for leading the overall strategy, business planning, and operational execution for Colorcon’s on-dose authentication business. Pond has more than 25 years of experience in product management, marketing, operations, and digital transformation, and he is an established leader in on-dose authentication. Prior to Colorcon, Pond held marketing and operational leadership roles at Sanofi, Merck & Co, Abraxis Bioscience, and IQVIA.


 is the vice president and chief innovation officer at Colorcon, based at the global headquarter in the United States. Rajabi-Siahboomi has held various academic positions in Nottingham and Liverpool JM Universities in the United Kingdom before joining Colorcon as technical director, responsible for Europe, the Middle East, and Africa in 2000. His main research interests are solid dosage form pharmaceutics, pharmaceutical technology with emphasis on oral drug delivery systems, solubility enhancement to improve bioavailbility, and consistency of drugs and bioequivalence formulations. He has published more than 300 articles, book chapters, abstracts, and patents and obtained his B.Pharm. and PhD in pharmacy from the University of Nottingham (UK).



 presents the Drug Solutions podcast, where the editors will chat with industry experts from across the pharmaceutical and biopharmaceutical supply chain. Join us as experts share insights into your biggest questions—from the technologies, to strategies, to regulations related to the development and manufacture of drug products.

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A look back at challenge and innovation highlights in 2022 for oral solid dosage forms, this podcast episode will review industry experts’ takes on factors driving the evolution of OSD forms as well as innovations in APIs, technology, and methodology. 