What are some of the main challenges in the development of mAbs against cancer?
There are many challenges to take into consideration when developing mAbs.2-5 Briefly, challenges exist all the way from the design of mAbs to manufacturing processing steps, to formulation efforts to delivery, as well as concerns on stability issues, bioavailability and immunological engagement. Some of these issues are difficult to address in a short time. For example, producing mAbs using a mammalian cell line will always be expensive, time consuming and several critical aspects such as post-translational modifications (PTMs) (especially glycosylation) are dependent on both intrinsic and external factors that are not easy to control. Current industrial production of mAbs mainly involves using Chinese hamster ovary (CHO) cell lines, which introduces challenges like making mAbs with potentially immunogenic glycosylation profiles.
The cost of mAbs is directly related to manufacturing costs, so even with stable cell lines that the industry relies on, most mAbs are too expensive to be used commonly”
The cost of mAbs is directly related to manufacturing costs, so even with stable cell lines that the industry relies on, most mAbs are too expensive to be used commonly. This means that many are not first line therapies, but perhaps a last resort after traditional methods such as chemotherapy or surgery, or else they may be used in combination with these approaches. In developing countries this is especially true, where several thousands of dollars per dose (or tens of thousands of dollars per patient per year) is not a realistic expectation. The latest example of this was mAbs that were developed against SARS-CoV-2. These products are successful for neutralising the virus; however, they were not available for many patients even in developed countries due mainly to their cost. Luckily, we were not reliant on them due to the availability of vaccines. Unfortunately, a similar situation exists for mAbs that are used for cancer too: around the globe, even if we have different mAbs available for a particular type of cancer, they may not be the first choice of therapy in many countries and in particular, countries where a national subsidy scheme exists. Hence, in many cases, a patient will be offered a mAb only after undergoing chemotherapy treatment, surgery, radiation therapy or a combination of these, although sometimes a biologic might be offered as a combination therapy alongside traditional approaches.6
Another major issue is protein aggregation which can be observed at any stage during the manufacturing, transport or long-term storage conditions, making it one of the main degradation routes of protein‑based medicines. Considering that proteins are marginally stable in these conditions, preventing and predicting protein aggregation is an ongoing challenge.
Rational development of mAbs requires predictive and sensitive spectroscopy-based methods during development and manufacture. However, most of the current processing steps are still empirical and rely on trial-and-error approaches. There are examples of such methods7-12 but they are not yet widely adapted.
A cancer-specific challenge is the target, ie, biomarker, for a specific cancer type. Existing targets such as HER2 and EGFR are good but not all cancer types have such a specific biomarker or have not yet been determined. For example, while there are excellent mAbs targeting HER2, this biomarker is overexpressed only in 20 to 30 percent of breast cancers; thus developing mAbs for other cancer types requires other targets to be identified.
Lastly, we do not know why some cancer patients respond well to a certain mAb while others do not due to the lack of mechanistic understanding on how some therapeutic mAbs work and how our bodies respond to the treatment. An example of this is immune checkpoint inhibitors; cancers that have many mutations such as melanoma respond well to immune checkpoint inhibitors, but only a small percentage of people with this type of cancer are able to benefit from these mAbs and we do not why other patients do not show improvement. Clearly, more research is needed to provide better understanding so that more patients can benefit.
How could these challenges be overcome?
Solubility and formulation challenges such as high viscosity or aggregation might be addressed by using alternative excipients. There are some new developments in this field, eg, ionic liquids as co-solvents or additives, hyperglycosylation or PEGylation.
Bispecifics are extremely promising when dealing with several different diseases as it involves utilising immune cells too”
New sensitive analytical methods that can be used in situ could also be beneficial to address some of the concerns such as detection of protein aggregates early on. Using computational techniques with experimental approaches could also be helpful,13 specifically when molecular-level information is required; for instance, when aggregation-prone regions on the protein surface need to be determined.
Furthermore, closer collaboration between academia and industry is also needed to address some of the challenges that the industry is facing. Lack of funding, adequate amounts of protein and long-term data are some of the major roadblocks in academia for progress on therapeutic mAbs. The industry could benefit from the new developments and expertise that research institutes could offer to overcome some of these challenges.
Can you outline your current research into mAbs?
In addition to our ongoing and established research efforts in the development of mAbs, formulations, vaccines and predictive methods, in recent years our research has diversified and now includes ADCs, bispecifics, nanoparticle-mAb complexes and new methods and additives to enhance structural and formulation stability of biologics. To this end, we have recently applied for a patent for a novel bispecific platform that is highly versatile. We also developed a multi-payload ADC approach in the last several years that shows better therapeutic outcomes compared to antibodies alone or a single-payload ADC. Additionally, we enhanced the formulation stability of mAbs and vaccines with a new class of additives using ionic liquids.
What exciting results have you seen?
We observed that, using new additives, we can almost stop protein aggregation while also enhancing vaccine formulation stability. Most biologics including mAbs and vaccines are highly thermolabile and thus being able to prepare a highly stable formulation would be very useful for many products including influenza and COVID-19 vaccines. We are currently testing these ideas with the ultimate goal being to realise a vaccine formulation that is stable at room temperature.
Further, we achieved aggregation-resistant mAbs via hyperglycosylation.14,15 For this, we introduced additional glycosylation sites on constant regions of mAbs. Although slight glycosylation heterogeneity is introduced, mAbs aggregate to a lesser extent and their solubility is increased greatly. This will allow us to prepare higher concentrations of mAb formulations with no concerns about protein aggregation.
We also engineered a dual-payload ADC and expect that this approach will be highly adaptable for different mAbs and payloads, aiding the preparation of novel ADCs. Having two different payloads is very useful in destroying cancer cells rapidly and safely before they have a chance to develop resistance.
Bispecifics are extremely promising when dealing with several different diseases as it involves utilising immune cells too. However, current bispecifics have several limitations, so we recently developed a platform that overcomes these limitations. A patent application has been filed by the University of Sydney for this technology.
New formulation approaches are also quite promising”
Lastly, our efforts on developing new formulation approaches for biologics using new additives such as ionic liquids yielded very exciting results. To realise the full potential of this approach, we are furthering our understanding about molecular-level interactions of protein additives, eg, mAb-ionic liquid,16 using a variety of different experimental and computational approaches.
What emerging trends are we seeing in the development of mAbs for cancer?
Each year we see more papers published in the literature about mAbs for cancer, more patents emerging, regulatory agencies approving more biologics including various formats of mAbs for a variety of different indications and more mAbs and antibody-based products are entering clinical trials; so, there seems to be an overall shift towards biologics with mAbs being the leading class by far.
We also observe that along with full-size mAbs and biosimilars, there is an increased number of publications on other mAb-based therapeutic candidates such as ADCs and bi- or multi-specifics and nanoparticle-mAb complexes. Along with other relatively recent developments in the immunotherapy field, such as immune checkpoint inhibitors and CAR T-cell therapies, the trend seems to be shifting towards these novel developments. The number of clinical trials is also indicative of this tendency.
In the long-run, perhaps cancer vaccines and other immunotherapies might be combined. In the near future, I expect to see more combination therapies involving immune therapies and chemotherapies as well as different types of immunotherapies.
Any other comments?
This is a rapidly evolving space and the mAb market seems to be growing exponentially. Additionally, there is an urgent need for predictive and sensitive biophysical methods for the rational development of any biologic, including mAbs and vaccines. We and others have been developing simple but elegant spectroscopy-based techniques that could facilitate the development of mAbs, from early phases of development such as candidate selection to formulation screening to the prediction of aggregation propensity in the long-term storage conditions from accelerated studies.7-12
New formulation approaches are also quite promising. I expect that these efforts will come to fruition soon and more stable formulations will be available.
Most of these challenges require close collaboration with multi-stakeholders including industry, academia and consumer support groups. We are always eager to engage with industry at any level to enable the development of successful biologics.
Dr Veysel Kayser is an Associate Professor in the University of Sydney School of Pharmacy. He completed his PhD at the University of Leeds, UK, undertook post-doctoral fellowships at the Max-Planck Institute, Germany and MIT, US. He was a senior staff scientist at MIT prior to taking up his current position in mid-2013. His research interests focus on biologics, mAbs, protein aggregation, biosimilars, vaccines and their formulations. He currently supervises five PhD students and one honours student, has six patents and has published two edited books, four book chapters and over fifty research papers.
References
- Kayser V,Sen M. 2020. New Emerging Biotherapies: Cutting‐Edge Research to Experimental Therapies. Biologics, Biosimilars, and Biobetters: An Introduction for Pharmacists, Physicians, and Other Health Practitioners, p. 213-236.
- McKertish CM, Kayser V.2021. Advances and Limitations of Antibody Drug Conjugates for Cancer. Biomedicines. 9(8): p. 872.
- Sifniotis V, et al. 2019. Current Advancements in Addressing Key Challenges of Therapeutic Antibody Design, Manufacture, and Formulation. Antibodies (Basel), 8(2): p. 36.
- Cruz E, Kayser V. 2019. Monoclonal antibody therapy of solid tumors: clinical limitations and novel strategies to enhance treatment efficacy. Biologics, 13: p. 33-51.
- Elgundi Z, et al. 2017. The state-of-play and future of antibody therapeutics. Adv Drug Deliv Rev. 122(Supplement C): p. 2-19.
- Australia’s Pharmaceutical Benefits Scheme (PBS) Expenditure and Prescriptions Report for 2019-2020, indicates that 7 out of 10 PBS subsidised medicines by expenditure were biologics, mainly mAbs. In fact, this is a global trend. Obviously, this will continue to be the case in the future, especially for cancer treatment. https://www.pbs.gov.au/info/statistics/expenditure-prescriptions/pb-expenditure-and-prescriptions-report-1-july-2019
- Kayser V, et al. 2011. Evaluation of a non-Arrhenius model for therapeutic monoclonal antibody aggregation. J Pharm Sci, 100(7): p. 2526-2542.
- Kayser V, et al. 2011. Stability and aggregation of therapeutic monoclonal antibodies studied with ANS and Thioflavin T binding. mAbs. 4(3): p. 408-411.
- Sahin Z, Demir YK, Kayser V. 2016. Global kinetic analysis of seeded BSA aggregation. Eur J Pharm Sci. 86: p. 115-124.
- Chennamsetty N, et al.2010. Prediction of aggregation prone regions of therapeutic proteins. J Phys Chem B, 114(19): p. 6614-6624.
- Kayser V, et al. 2012. A screening tool for therapeutic monoclonal antibodies: Identifying the most stable protein and its best formulation based on thioflavin T binding. J. 7(1): p. 127-132.
- Sahin Z, et al. 2017. Nile Red fluorescence spectrum decomposition enables rapid screening of large protein aggregates in complex biopharmaceutical formulations like influenza vaccines. Vaccine, 35(23): p. 3026-3032.
- Kuyucak S, Kayser V. 2017. Biobetters From an Integrated Computational/Experimental Approach. Comput Struct Biotechnol J, 15: p. 138-145.
- Reslan M, et al. 2020. Enhancing the stability of adalimumab by engineering additional glycosylation motifs. Int J Biol Macromol, 158: p. 189-196.
- Cruz E, et al. 2021. Glycan profile analysis of engineered trastuzumab with ra-tionally added glycosylation sequons for enhanced physical stability.
- Reslan M, et al. 2018. Choline ionic liquid enhances the stability of Herceptin® (trastuzumab). Chem Commun, 54(75): p. 10622-10625.
