Review: Proteomics and the creation and development of pharmaceutical drugs

17 min readMar 25, 2023

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Abstract

The pharmaceutical is extremely integral to the healthcare industry and also a highly competitive, highly funded industry. The reason for this is that drugs can either save someone’s life or negatively impact it for the worse. Because of this, it is necessary for companies to both find drug targets for diseases and come up with safe, well-developed drugs to put into the market.

Using proteomic technology, pharmaceutical companies do both of these things. Technology like protein microarrays and 2D-Gel Electrophoresis is essential in finding potential drug targets at any given condition. Meanwhile, technology like mass spectrometry is great in understanding the effect of the drug on the body.

This review article covers an overview of proteins, proteomics, and the pharmaceutical industry as well as the most prominent proteomic technologies used in early and late-stage drug development. It also discusses the current challenges and limitations in applying proteomics to drug discovery and development.

1. Proteins
1.1 Introduction to proteins
1.2 How proteins are made
2. Introduction to Proteomics
2.1 Definition of Proteomics
2.2 Comparison to genomics
2.4 Different uses of proteomics
3. Pharmaceutics and proteomics
3.2 Challenges in the Pharmaceutical Industry and the Solution Proteomics Provides
4. Proteomic techniques with pharmaceutics
4.1 Mass Spectrometry
4.2 2D Gel Electrophoresis
4.3 Protein Microarrays
4.4 Immunoprecipitation
4.5 Limitations of these technologies
5. Proteomics in early-stage drug discovery
5.1 Main uses in this stage
5.2 Case Study: Herceptin
5.3 Other Cases of proteomic analysis
6. Proteomics in late-stage drug development
6.1 Safety Assessment
6.2 Case study: Avandia
6.3 Limitations with Proteomics and Late-stage Drug Development
7. Non proteomic targeted drugs
8. Conclusion
9. Sources

1. Proteins

1.1 Introduction to proteins

Proteins are molecules that play many crucial roles in the body. They do most of the work in cells and are needed for the structure, function, and regulation of the body’s tissues and organs. At any given time, there are approximately at least 10,000 proteins working to perform different functions.

There are many different types of proteins; from things like hormones to coordinate the body’s functions to antibodies that support the immune system’s functions (1).

1.2 How proteins are made

Proteins are made through a process called protein synthesis. Protein synthesis is done through two steps: transcription and translocation. (3)

Transcription: The DNA sequence in a gene is copied to a messenger RNA (mRNA) inside the nucleus. After this is done, the mRNA molecule then leaves the nucleus and enters the cytoplasm.
Translation: In this step, the code on the mRNA is used to create the protein. This process takes place on ribosomes. The ribosome reads the sequence of codons (groups of three nucleotides) on the mRNA molecule and matches each codon to the appropriate amino acid, which is brought to the ribosome by a molecule called transfer RNA (tRNA). The ribosome then links the amino acids together in the correct order to form a protein.

Once the protein is complete, it may undergo additional processing such as folding or modification to become fully functional (3). The final folded protein is then transported to its proper location in the cell, where it can perform its specific functions. This protein can also change in shape and bind to other molecules later on.

1.3 Types of proteins

There is various type of proteins. Here are the most common ones in the body (2):

Digestive enzymes: helping convert nutrients to more simple forms of cells
Transporting proteins: carrying substances in the blood or lymph system
Structural: constructing different structures in the body such as the skeleton and bones
Hormones: coordinating activity in the body
Defense: working with the immune system to keep the body protected from pathogens
Contractile: allows the muscle to contract
Storage: provide nourishment to the embryo and during the early stages of the pregnancy

Each protein is crucial for the body to perform specific functions and can be targeted as they relate to different medical and diseased conditions.

2. Introduction to Proteomics

2.1 Definition of Proteomics

The term proteomics is defined as the analysis of proteins in the human body. Rather than just relying on the genes — which do not contain the ‘code’ for all proteins that are produced in the body — proteomics focuses on the outcome of these genes for analysis. Further, proteomics allows for a larger-scale analysis of human proteins and allows researchers to analyze multiple proteins rather than only just one individually.

In recent years, proteomics has rapidly grown due to advances in technology such as mass Spectrometry, 2D Gel Electrophoresis, protein microarrays, and immunoprecipitation. These will be discussed later in this article.

Proteomics has many applications in areas such as medicine, forensic sciences, and the food development industry (4). Specifically, in medicine, proteomics is used to identify potential biomarkers for disease diagnosis and to develop new drugs that target specific proteins involved in disease processes.

2.2 Comparison to genomics

In comparison to proteomics, one heavily used tool for analysis is genomics. Genomics involves sequencing and understanding the human genome as well as its functions (5). The key difference between proteomics and genomics is that proteomics focuses on the outcome of the genome, rather than the genome itself. Unlike genomics, proteomics shows internal and external conditions.

For example, something that proteomics would be able to detect but genomics would not be able to be a difference of protein expression. A specific gene may be expressed at the same level in two different cells, but the protein produced by that gene may be modified or present at different levels in those cells, resulting in different cellular functions or responses. Another example of a change in protein function that genomics would not be able to detect is cancer. Rather then just a mutation of the genome sequence, it also produces different proteins (or too many proteins) not usually produced by the body (Ex. syndecan-1 (FTC-133) is only produced by thyroid cancer cells).

2.3 Different uses of proteomics

Here are the four main ways proteomics is used. (6)

Understanding the functions of proteins:

Using proteomics, there is a better understanding of proteins and how they link to specific diseases. This would help as we would better understand the functions based on the data collected after testing.

Another use of this is the 3D rendering of proteins for an understanding of their function. Some proteins are only understood after a 3D rendering of their model these proteins. Technologies such as Mass Spectrometry help scientists model the protein.

2. Protein modifications

Proteins are known to modify as time progresses. By using proteomics, it is easier to understand the change in the protein after a modification has occurred due to external conditions and/or because of a change in genetic code.

3. Protein localization

Proteins being in the correct place in the body helps the body keep homeostasis. However, when the protein is mislocated a disease occurs. For example, Amyloidosis occurs when a protein called amyloid is in the organs leading the organ to not work properly (7).

Some proteomic technologies are able to map out the human proteome and aid in better understanding where these proteins are traveling to in the human body.

4. Protein-protein interactions

At all times, proteins are working together with other proteins to perform specific functions in the human body. Understanding how protein-protein interactions help have a better understanding of the use and purpose of these proteins. For example, BAD BCL2 is a protein that only performs cell death (8).

This is a type of interaction only proteomics can perform as other technologies such as genomics would not be able to detect these interactions, however, proteomics would be able to do this.

3. Pharmaceutics and proteomics

3.1 Introduction to the use of proteomics in the pharmaceutical industry

The pharmaceutical industry is a necessary component of the healthcare system, involved in the research, development, manufacturing, and marketing of drugs and medical devices.

Proteomics plays a key part in this is through understanding the correlation between the proteins released and physiological changes related to a healthy or diseased condition. Further, this way of analyzing proteins helps describe drug mechanisms and side effects. As a result, over 70% of new drugs approved by the US Food and Drug Administration (FDA) between 2010 and 2019 were targeted at proteins (9).

Along with the formulation of drugs, the pharmaceutical industry also looks for cost-effective drugs that can also be easily absorbed by the body. Consistent proteomic testing can allow the manufacturers to stay in competition with others while still having high-quality drugs that meet regulatory standards.

However, the development of the drugs to go into the market is a long process that can take up to 20 years to develop. These technologies make this process easier and aid in shortening this process for the companies and making it easier. Further, when the companies can prove the drug to be safe, it would take less time to verify it to the regulators such as the FDA or HPFB.

3.2 Challenges in the Pharmaceutical Industry and the Solution Proteomics Provides

The pharmaceutical industry also faces various challenges with bringing these new drugs into the market, including identifying new drug targets, developing drugs that are effective and safe, and reducing the time and cost of bringing a drug to market. (10)

Proteomics can help address these challenges in several ways:

Identify New Drug Targets: Based on information from protein expression levels, post-translational modifications, and interactions with other proteins. This information can be used to identify proteins that are involved in disease pathways and that may be potential drug targets. (4)
Drug Development: After proteomic analysis, different information on drug-protein interactions and potential off-target effects are shown through various graphs and data. This information can help identify potential drug candidates that are safe and effective and pharmaceutical companies are able to further develop and improve their drugs during later stages. (10)
Biomarker Discovery: This is able to discover various biomarkers that can be used to diagnose diseases, monitor disease progression, and evaluate the efficacy of drugs. This information can be used to develop diagnostic tests and to identify patient populations that are most likely to benefit from a particular drug. (4)

Another big factor that comes into play in this industry is the ability to compete with other big brands. Many companies want to develop the cheapest, safest, and most accurate drug. Various proteomic testing would allow the companies to reiterate their drugs to the point which is it better than the other companies.

4. Proteomics techniques with pharmaceutics

There are many proteomic techniques available for researchers to use, however, these are the most commonly used proteomic techniques in the pharmaceutical industry.

These techniques can be used for various stages and times during the development and drug life cycle. Each technique can be used for different needs the companies have and the length and mass of each protein targeted as well as the level of analysis needed.

Here are the most common ones:

4.1 Mass Spectrometry:

Image of a Mass Spectrometry Machine | Source

Mass spectrometry is a technique that can identify and quantify proteins and their post-translational modifications. It is widely used during the preclinical stages of drug discovery (11).

This technology measures the ‘mass-to-charge’ ratio of ions within the protein by using either magnetic or electric fields. Some of the sample’s molecules to break into charged fragments. These ions are then separated according to their mass-to-charge ratio, typically by accelerating them and subjecting them to an electric or magnetic field. detected by a mechanism capable of detecting charged particles, such as an electron multiplier (12)

Here are the steps to a mass spectrometry analysis: (13)

Sample preparation
Ionization: the sample is vaporized and passed through an ionization chamber. Here the collisions between the metal coil in the chamber and the particle give the sample a positive charge.
Acceleration: since the chamber is already a positive charge, the positively charged ions accelerate to the slits in the chamber.
Deflection: ions are deflected by a magnetic field where both the lighter mass and positively charged masses become deflected
Detection: finally, beams of ions are passed through the mass analyzer and detected based on the m/z (mass to charge) ratio.
The result of this analysis is a mass spectrum chart sent to a computer to show the different values of ions present with their abundance.

Mass spectrometry can be used to identify and quantify drugs and their metabolites in various biological matrices such as blood, urine, and tissues. Here are some reasons why mass spectrometry is ideal for drugs it has a high sensitivity to low concentrations of drugs, the ability to distinguish between drugs and metabolites, the ability to provide information on the 3D structure of the drug, and the precision of qualitative data (27).

4.2 2D Gel Electrophoresis

Image of the results of a 2D gel electrophoresis analysis | Source

2D gel electrophoresis is a widely used technique for the separation of proteins based on their charge and molecular weight (14). It is used for the identification of biomarkers and for the analysis of protein expression levels in the pharmaceutical industry (14).

Here are the steps to the technique: (14)

Sample preparation
Isoelectric focusing (IEF): here the protein is separated based on the pl (isoelectric points or the point when the protein has a net charge of zero). In this step, the machine has two ends (an anode and a cathode), and the more net positive or net negative charge will move toward the oppositely charged side
SDS-PAGE (SDS-polyacrylamide gel electrophoresis): this is a method for separating polypeptides according to their molecular weights (Mr). Here the larger the protein, the more resistance and slower migration.
The results of this analysis are a visualization of the proteins in the sample. There are some 2D gel analysis software available (such as PDQuest, REDFIN, and Progenesis)

2D gel electrophoresis is a great technique for separating and analyzing complex mixtures of proteins. For the use of drug development, 2D gel electrophoresis can be used to analyze the protein expression profiles of cells or tissues that have been treated with drugs. It is a great option to use as it provides high resolution even while separating high levels of proteins, the ability to identify drug targets and drug-induced changes, and is often used along with mass spectrometry to identify drug targets (14).

4.3 Protein Microarrays

Protein microarrays are used to study protein-protein interactions, protein-DNA interactions, and protein-small molecule interactions. They are useful for the identification of potential drug targets and for the screening of small molecule inhibitors. (15)

There are three main steps to this process: (16)

Design and fabrication: this step requires a microarray printer to put the antigens onto. The type of printer and surface depends on the type of antigen and its mechanisms (for example if it was a nucleic acid it would need a different substrate to attach to than a carbohydrate)
Array probing, detection, and scanning: this is a process consisting of three steps dilution, probing and finally washing by a fluorescent antibody. After this, a 12h incubation process must occur
Image processing and data analysis: this is done using microarray processing software and the mean fluorescent intensity (MFI) is converted to text.

Due to the protein microarrays' ability to scan through large protein samples obtained across an entire proteome using this approach, the specificity or side effects of a drug can be monitored. This technology can help aid the process of reiteration and improvement of how the drug works. (15)

4.4 Immunoprecipitation

The output of the Immunoprecipitation technique | Source

Immunoprecipitation is a technique that uses antibodies to isolate specific proteins from a complex mixture. It is useful for the identification of protein interactions and for the characterization of protein complexes. (17)

There are five main steps to this process: (17)

Sample preparation
Preclearing
Antibody incubation/formation of antibody-antigen complexes
Precipitation and washing
Analysis: This is done using software such as SDS-PAGE

In pharmaceutics, immunoprecipitation is mainly used to identify potential drug targets and test the efficacy of drugs targeting specific proteins (17). This tool is used along with a variety of techniques to identify protein targets.

4.5 Limitations to proteomics:

Although there are many positives to using this technology, there are also a few key limitations to proteomics that cannot be avoided. Here are the most significant limitations to using proteomics potential sample preparation errors, the sensitivity of the tests, and data analysis. (4)

A lot of the time is spent in sample preparation which is both time-consuming and can limit the throughput of proteomics experiments. The majority of the results depend on how well the sample has been prepared and can change depending on it.

A second limitation of the technology is the limitations in sensitivity and dynamic range. It can be difficult to detect low-abundance proteins or to accurately quantify proteins across a wide range of concentrations. Further, some proteins cannot be measured on some technologies making it hard to find the correct one per a study.

The outcome of proteomics analysis is large amounts of data that can be challenging to analyze and interpret. If researchers lack this skill it can be challenging for them to analyze and interpret this data. However, this also allows the intersection of bioinformatics and more job opportunities in this field. This could also be considered a downside as this is another cost for the companies, needing to hire more people for one project.

5. Proteomics in early-stage drug discovery

5.1 Main uses in this stage

As mentioned, many drugs target proteins in the body. Using technologies such as Mass spectrometry and 2D gel electrophoresis, companies are able to both find protein targets and verify existing protein targets.

In early-stage drug discovery, proteomics is heavily used for both target identification and validation (18). Here is a brief overview of these two uses.

Target identification:

This is a key factor that relates to the success of the drug; how well it targets the specific issue or root cause of the disease. A good target needs to be efficient, safe, and meet clinical standards.

Target Verification:

Once there is a target of the drug, it needs to be verified through many clinical studies as well as done in the lab. There are various ways for companies to go through with this like cell-based disease models or genetics and expression data

5.2 Case study: Herceptin

An example of a drug that was developed through the use of proteomics is Herceptin, used to treat breast cancer. Herceptin was developed to target a protein called HER2 (human epidermal growth factor receptor 2), which is overexpressed in some forms of breast cancer (19).

Through proteomic studies, it was shown that HER2 was overexpressed in a significant proportion of breast cancer cases and that HER2-positive breast cancer tended to not respond to chemotherapy treatment. Based on this information, researchers developed Herceptin as a targeted therapy for HER2-positive breast cancer.

To combat the HER2 protein, Herceptin binds and blocks its activity which slows or stops the growth of cancer cells as a monoclonal antibody (19). Herceptin has been shown to improve survival rates in women with HER2-positive breast cancer and has become a part of the standard treatment for a type of breast cancer (20).

5.3 Other Cases of proteomic analysis

There are many other cases in which this technology has been used, as well as in the development of treatments for infectious diseases. By analyzing the proteins of pathogens, researchers can identify proteins that are essential for the survival or virulence of the pathogen

6. Proteomics in late-stage drug development:

Another big part of the pharmaceutical industry is how well it targets the specific protein in the body as well as how safe it is. Once the drug has been developed in the lab, researchers can further verify through safety assessments and understand how the drug can alter and change protein expression in the body (21).

6.1 Safety Assessment

Proteomics can also be used to study the effects of drugs on the proteome (the complete set of proteins expressed by a cell or organism) and to understand the mechanisms underlying the drug’s therapeutic or adverse effects. By analyzing changes in the proteome before and after drug treatment, researchers can identify proteins that are affected by the drug and gain insight into the drug’s mode of action. (21)

Further, this information can also be used to develop a personalized approach where the drug is tailored to the individual and maximize the benefits of the drug.

6.2 Case study: Avandia

An example of a drug that was studied to understand side effects is the antidiabetic drug rosiglitazone (Avandia). Rosiglitazone is a drug that is used to lower blood sugar levels in patients with type 2 diabetes.

However, concerns were raised about the cardiovascular safety of rosiglitazone as cases of the drug increasing the risk of heart attacks and strokes came up. So, researchers used proteomics to study the effects of rosiglitazone on heart cells.

One study, published in the Journal of Proteome Research in 2009, used quantitative proteomics to analyze changes in protein expression in heart cells treated with rosiglitazone (22). The study identified several proteins that were upregulated by the drug, including the enzyme 12/15-lipoxygenase (12/15-LOX), which is involved in inflammation and oxidative stress. Further, it was shown that the drug contributes to the drug’s cardiovascular side effects by promoting inflammation and endothelial dysfunction in blood vessels. (23)

Since then, Avandia has now been banned in multiple countries such as the European Union, and restricted in many more (24) as it was found that it is not a safe drug for patients to use.

6.3 Limitations with proteomics and late-stage drug development

However, there are some limitations to using proteomics to test for the safety and accuracy of the drug. The biggest one is that proteomics only provides a snapshot of protein expression at a given point in time and may not capture dynamic changes in protein expression or activity over time. (4)

Another limitation is the need for appropriate sample preparation and handling to ensure the reproducibility and accuracy of proteomic data. For example, variations in sample collection, storage, and processing can affect protein expression levels and introduce experimental bias (5).

7 Non-protein targeted drugs

Not all drugs are made to target proteins the body creates (25). There are various other types of drugs that target other parts of the body that also use other technologies to do this. Some other examples of drug targets are:

Nucleic acids: Some drugs work by targeting nucleic acids, such as DNA or RNA, and altering their expression or function.
Cell membranes: Some drugs work by targeting cell membranes and altering their structure or function.
Viruses or bacteria: Some drugs work by targeting viruses or bacteria directly.

8. Future of Proteomics in Drug Development

Proteins are a great target for drugs since there are a wide variety of proteins working in the body at any given time. Targeting these proteins through various drug treatments is a great way for the pharmaceutics industry to create drugs. Further, proteomics allows companies to research, develop and verify their findings through various proteomic processes.

As companies are creating more and more drugs, the testing of these drugs is necessary for us to ensure that there are safe, well-developed drugs in the market.

Proteomics will continue to be an integral part of the pharmaceutical industry.

9. Sources

Hey! Thanks for reading my article! It means a lot to me!!

My name is Anya Ishani Sharma, and I am a grade 11 student passionate about personalized medicine and tech! If you found this review interesting and would like to discuss it with me please feel free to reach out. You can contact me through my email (anya.ishani.sharma@gmail.com), and my Linkedin (click here).