CASE STUDY

Plasmax: A physiologically relevant cell culture media

The future of Cancer Cell Culture

The Contributor: Dr. Saverio Tardito, Cancer Research UK Glasgow: The Beatson Institute

The Research Tool: Plasmax^TM

Plasmax^TM is a physiologically relevant cell culture medium that closely resembles the metabolic and nutritional profile of human plasma. Unlike traditional media designed to supply excessive levels of a few nutrients, it provides unmatched metabolic fidelity.

Cell culture media is a critical component of cell-based assays, but its contribution to results is often overlooked. The right cell culture is critical in order to comprise the correct energy and compounds to regulate and support the cell cycle. For in vitro experiments, scientists typically use media like Dulbecco’s Modified Eagle Medium (DMEM) - a mixture of vitamins, selected amino acids, sugars and salts which sustain cellular growth. Yet, such types of mediums typically focus on cell proliferation rather than the nutritional environment that cells withstand in tumours. Dr. Tardito, an oncometabolism expert from the CRUK Beatson Institute, required a cell culture medium which better reflected human physiological conditions in order to study cancer biology.

CancerTools.org spoke with Dr. Tardito about Plasmax^TM to explore the importance of its development and what its contribution to the CancerTools.org initiative can do for cancer research.

Overcoming in vitro and in vivo variance

Traditional cell culture media was originally produced to rapidly and successfully increase cell proliferation in an in vitro environment (Eagle, 1955). This was made possible by adding nutrients in excessive concentrations to avoid nutrient depletion and simultaneously promote cell growth. Such disproportionate nutrient composition, in comparison to in vivo conditions like human plasma, affects both phenotypic and genotypic behaviour of cells (Schug et al, 2015 and Tardito et al, 2015). Usage of traditional media for cell culture can therefore lead to unrepresentative in vitro conditions and variance between in vitro and in vivo cancer cell metabolism. This becomes particularly important in research relating to cancer cell biology and related metabolic pathways.

To address this challenge, the research team at the Beatson Institute for Cancer Research, Glasgow, UK, under the supervision of Dr. Saverio Tardito, developed a novel cell culture medium, Plasmax^TM, to study the cell metabolism in different tumour types.

The Development of PlasmaxTM

Dr. Tardito and his research team, optimised the concentrations of over 80 compounds typically found in human plasma to achieve the cell growing conditions. The cell culture medium contains all relevant elements to mimic human plasma, consisting of proteinogenic amino acids, vitamins, salts, and sugars, determined through Dr.Tardito’s optimisation experiments. The inclusion of metabolites enhances its physiological relevance and thereby mimics the in vivo environment. Trace elements, while essential for survival and proliferation, are often missing from traditional cell culture media and have to be supplemented before use. Plasmax^TM, is uniquely formulated with trace elements including vanadium, zinc, manganese, copper and selenium. The presence of these increase the antioxidant capacity of cells, which promotes colony growth by preventing ferroptosis-induced cell death (VandeVoorde et al., 2019).

To date Cancer Research UK’s researcher community:

Benefits of Plasmax^TM

It is critical for biomedical research to renew and refine models to improve their relevance to human physiology – which is exactly what the development of Plasmax^TM helps to execute.

Plasmax^TM has been successfully validated across primary cells of different tissue, species, and experimental conditions (see Table 1), and is suitable for both primary and established cell lines. Additional cell lines are successfully cultured using Plasmax^TMregularly, which makes Table 1 a running list of validated cell lines. Plasmax^TM is anticipated to work across a broad range of cancer cell culture models.

Using a physiological relevant medium significantly impacts the results obtained from common cellular assays, including colony formation and gene expression. This has the potential to improve results for cancer cell biology experiments associated with drug discovery and in vitro cancer models.

Physiologically relevant

Plasmax^TM is optimised to reflect the in vivo profiles of nutrients and metabolites found in human plasma, including essential and non-essential amino acids, amino acid derivatives, organic acids, and other polar metabolites.

Improves in vitro metabolic fidelity

Plasmax^TM can better approximate the overall metabolic phenotype of tumours, with both 2D and 3D cells cultured in Plasmax^TM, better recapitulating the tumours’ metabolic signatures.

By increasing the metabolic fidelity and biological relevance of in vitro cancer models, better drug discovery and improved understanding of cancer at a cellular level can ensue.

Better mimics tumour metabolism

Plasmax^TM better mimics tumour metabolism. Breast cancer spheroids grown in Plasmax^TM , have shown to better approximate the metabolic profile of mammary tumours (Vande Voorde et al., 2019).

Produces a faster proliferation

When compared with traditional mediums, Plasmax^TM produces a faster proliferation, even when aged up to 12 months, in comparison with DMEM when both are supplemented with 2.5% foetal bovine serum.

Uncover role of trace elements

Cancer cells seeded at low densities in the absence of the trace element selenium are unable to form colonies in traditional media due to lipid peroxidation and ferroptosis. The growth-enabling trace elements in addition to vitamins and inorganic salts in Plasmax^TM, prevent ferroptosis-induced cell death, and promote colony growth.

Table 1: A selected list of cultured cell lines successfully validated for growth and viability in Plasmax^TMunder standard conditions

Cell lines grown in Plasmax ^TM	Tissue of origin	Cell line status	Species
HepG2	Liver Cancer	Established line	Human
HuH7	Liver Cancer	Established line	Human
HuH6	Liver Cancer	Established line	Human
BT549	Breast Cancer	Established line	Human
MDA-MB-468	Breast Cancer	Established line	Human
Cal120	Breast Cancer	Established line	Human
A375	Melanoma	Established line	Human
Colo829	Melanoma	Established line	Human
LN18	Brain Cancer	Established line	Human
Naive glioblastoma cell line	Brain Cancer	Low passage lines	Human
Dermal fibroblasts	Epidermis	Primary	Human
Small intestine organoid	Small intestine	Primary	Mouse
Mammospheres	Mammary gland	Primary	Mouse
Mesenchimal stromal cell line	Bone marrow	Primary	Human
Embryonic stem cell line	Embryo	Primary	Human
Trophoblast stem cell line	Placenta	Primary	Human
A549	Lung cancer	Established line	Human
HCT116	Colon Cancer	Established line	Human
SaOS2	Bone tumour	Established line	Human
HT1080	Fibrosarcoma	Established line	Human

Plasmax ^TM Impact

Choosing an appropriate cell culture medium is a crucial step in in vitro cell biology research and finding the correct one for your cell type and experiment can be challenging. Such was experienced by Sunada Khadka, a PhD Candidate at MD Anderson, during her research on anaplerosis in glioma cells.

While using a traditional medium, Sunada’s initial results obtained in vitro were not reproduced in her in vivo experiments. This inconsistency in data led to a return to  in vitro  experimental conditions and a closer examination of the cell culture media used. Plasmax^TM was selected as a cell culture media that better reflected the  in vivo  nutrient profile. By comparing in vitro results from Plasmax^TM to DMEM, Sunada was able to understand the discrepancy, illuminating the importance of triaging cell culture media with physiologically relevant media like Plasmax^TM  in order to better recapitulate the  in vivo  environment.

"The clear cut differences between experiments performed with Plasmax^TM vs commercial media available at the time become obvious once you realise, they aren’t physiologically relevant”

– Dr. Tardito

“To my knowledge at least 4 labs are trying to re-create Plasmax^TM…it is like following a very complex cake recipe… which takes weeks, and isn’t going to guarantee you always get the same medium."

– Dr. Tardito

In the future, whatever metabolism related work I do, I'll make sure to compare DMEM to Plasmax^TM to ensure that the nutrient profile is not effecting the certain phenotype that I’m seeing."

“Using physiologically relevant media is a time saver and will make you more confident in your data.”

- Sunada Khadka

Make time for work that matters:

Optimising the relevant components in culture media and defining their physiological concentrations at scale, can be a challenging process. Having Plasmax^TMin a pre-prepared liquid form saves time, effort, initial investment of sourcing, and spares the process of optimising 80+ components to get their proportions accurate.

Plasmax^TM unique formula, maintains its effectiveness throughout its shelf life with no effect on cell growth from being aged. It is compatible across different cell types and is greatly beneficial to any cancer researcher interested in the study of cancer cell biology, in vitro cancer models and cell based assays.

Plasmax^TM is already being repeatedly purchased by various cancer researchers across different academic institutes worldwide.

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About CancerTools.org

CancerTools.org, the research tools arm of Cancer Research UK, is a non-profit, global community of cancer researchers, academic institutes and societies, with a shared mission to accelerate cancer research discoveries. In this collaborative, researchers contribute research tools and share knowledge to deepen our understanding of cancer, and drive innovation within cancer research.

About Dr. Saverio Tardito

Dr. Saverio Tardito is the group leader for the oncometabolism research group at the CRUK Glasgow Beatson Institute and senior lecturer for the School of Cancer Sciences at the University of Glasgow.

About Cancer Research UK Glasgow: The Beatson Institute

One of Cancer Research UK’s core-funded institutes, The Beatson Institute have built an excellent reputation for basic cancer research, including world-class metabolism studies and renowned in vivo modelling of tumour growth and metastasis. Learn more at: https://www.beatson.gla.ac.uk/About/about-beatson.html