Cell Biology

Cytochrome P450 Inhibition: Old Drug, New Tricks

Posted on by Darcia Schweitzer
multiwell screening plate and various pills on a table

Cytochrome P450 (CYP) inhibitors are often used as boosting agents in combination with other drugs. This drug development strategy is front and center for Paxlovid, the new anti-SARS-CoV-2 treatment from Pfizer. Paxlovid is a combination therapy, comprised of two protease inhibitors, nirmatrelvir and ritonavir. It significantly reduces the risk of COVID-19 hospitalization in high-risk adults and is ingested orally rather than injected, which is an advantage over other SARS-CoV-2 treatments, such as Remdesivir.

Nirmatrelvir was originally developed by Pfizer almost 20 years ago to treat HIV and works by blocking enzymes that help viruses replicate. Pfizer created another version of this drug to combat SARS in 2003, but, once that outbreak ended, further development was put on pause until the advent of the COVID-19 pandemic. After developing an intravenous form of nirmatrelvir early in the pandemic, Pfizer created another version that can be taken orally and combined it with ritonavir.

When ritonavir was originally developed, it wasn’t considered particularly useful because it metabolized so quickly in the body. Now it is recognized as a pharmacokinetic enhancer in combination with other drugs. Ritonivir inhibits CYP3A4, an enzyme which plays a key role in the metabolism of drugs and xenobiotics. By inhibiting CYP3A4, ritonivir slows the metabolism of other drugs. In the case of Paxlovid, this allows nirmatrelvir to stay in the body longer at a high enough concentration to be effective against the virus. This ultimately means that patients can be given lower doses of the drug with reducing efficacy.

Diagram of Nirmaltrelvir mechanism of action.
Nirmatrelvir inhibits the viral 3CL protease, so that functional, smaller viral proteins cannot be produced.
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Cultured Meat Viability Increases in Biotech

Posted on by Riley Bell
Cultured meat grows in a plastic dish in laboratory conditions.

The biotechnology industry has been powering through barriers standing between the lab and the dinner plate as cultured meat advances toward the market. Challenges like scaling up the technology and getting products to the market are significant, but future food demands are an even bigger obstacle. Earth’s population is projected to reach 10 billion people by 2050. Our current agricultural practices will not be able to meet the food demands. Therefore, we need to find alternative ways to produce food–like “growing” it in the lab.

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GPCRs and PROTACs: New Approaches for Designing More Effective Drug Candidates

Posted on by Ken Doyle
NanoBRET target engagement assay

G protein-coupled receptors (GPCRs) comprise a large group of cell surface receptors, characterized by the unique structural property of crossing the cell membrane seven times. They respond to a diverse group of signaling molecules, such as peptides, neurotransmitters, cytokines, hormones and other small molecules (1). Upon activation, GPCRs interact with GTP-binding (G) proteins and arrestins to regulate a wide variety of signaling pathways. This broad range of functions makes GPCRs attractive targets for drug discovery. The importance of GPCR research was highlighted in 2012, with the Nobel Prize in chemistry being awarded to Robert Lefkowitz and Brian Kobilka “for studies of G-protein–coupled receptors”.

Based on structure and function, GPCRs are categorized into six classes, A–F. The class A GPCRs, or rhodopsin-like receptors, have been studied extensively due to their association with many types of diseases (2). Within the class A GPCRs is a group that share a highly conserved structural motif (3) and respond to chemokines—small “chemotactic cytokines” that stimulate cell migration, especially that of white blood cells (4). A subfamily of class A GPCRs respond to chemokines that have two cysteine residues near the N-terminus, known as CC chemokines. GPCRs activated by CC chemokines are called CC chemokine receptors or CCRs, and these interactions have been implicated in both pro- and anti-cancer pathways (5).

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New Study Suggests Long Mononucleotide Repeat Markers Offer Advantages for Detecting Microsatellite Instability in Multiple Cancers

Posted on by Kelly Grooms

A new study, published in the Journal of Molecular Diagnostics (1), highlights the potential of using long mononucleotide repeat (LMR) markers for characterizing microsatellite instability (MSI) in several tumor types. The paper is a result of a collaborative effort between researchers from Johns Hopkins University and Promega to evaluate the performance of a panel of novel LMR markers for determining MSI status of colorectal, endometrial and prostate tumor samples.

Microsatellite instability (MSI) is the accumulation of insertion or deletion errors at microsatellites, which are short tandem repeats of DNA sequences found throughout the genome. MSI in cancerous cells is the result of a functional deficiency within one or more major DNA mismatch repair proteins (dMMR). PCR-based MSI testing is a commonly used method that can help understand a tumor’s genomic profile as it relates to MMR protein function.

Historically, MSI has been a biomarker associated with Lynch syndrome, the hereditary predisposition to colorectal and certain other cancers. In recent years, research interest in MSI has exploded, driven by the discovery that its presence in tumor tissue can be predictive of a positive response to anti-PD-1 immunotherapies (2,3).

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Real-Time Analysis for Cell Viability, Cytotoxicity and Apoptosis: What Would You Do with More Data from One Sample?

Posted on by Kari Kenefick

Originally posted May 25, 2017. Updated 2022

You are studying the effects of a compound(s) on your cells. You want to know how the compound affects cell health over a period of hours, or even days. Real-time assays allow you to monitor cell viability, cytotoxicity and apoptosis continuously, to detect changes over time.

Why use a real-time assay?
A real-time assay enables you to repeatedly measure specific events or conditions over time from the same sample or plate well. Repeated measurement is possible because the cells are not harmed by real-time assay reagents. Real-time assays allow you to collect data without lysing the cells.

Advantages of  Real-Time Measurement
Real-time assays allow you to:

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MISpheroID: A Knowledgebase to Improve Reproducibility in Spheroid Research

Posted on by Johanna Lee
Spheroid research is now a common component of cell biology and drug discovery science

Advantages of Spheroids

In the past decade, there has been a sharp rise in studies using spheroids as cell models for basic research and drug discovery. Spheroids are self-organized aggregation of cells that form a spherical mass, and they have become widely popular because they are much more physiologically relevant compared to flat 2D cell cultures.

In spheroids, the inner cells have less access to nutrients and oxygen compared to the outer layer, forming a natural gradient. As a result, metabolite concentration and cellular state such as proliferation and differentiation, can be very different at the periphery compared to the inner core. This phenomenon, known as “heterogeneity”, makes 3D tumor spheroids much more representative of actual tumors in the human body.

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Total Eclipse of the CAR T: How mRNA Vaccine Technology Can Be Used to Help Heal “Broken” Hearts

Posted on by Natalie Larsen

While you can rely on Taylor Swift and Adele to help heal emotional heartbreak, unfortunately treating a physically “broken” heart, a heart damaged by fibrosis, is a much more complicated process than putting on your favorite sad songs and wallowing in your feelings. In a recent study published in Science, researchers developed a therapeutic approach to treat damaged hearts in mice through the removal of scar tissue using genetically engineered immune cells (CAR T cells) and the mRNA technology used in the mRNA coronavirus vaccines.

Genetically engineered CAR T cells have been used l for repairing damaged hearts in mice
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How to Get Real-Time Kinetic Data With GloMax® Microplate Readers

Posted on by Johanna Lee

Understanding how a compound or drug affects cellular pathways often requires measuring kinetic changes over an extended period of time—from several hours to days. Live-cell kinetic cell-based assays that measure cell viability, cytotoxicity, apoptosis and other cellular pathways are great for collecting real-time data. You don’t necessarily need expensive equipment to run these types of assays. In the videos below, Dr. Sarah Mahan, a research scientist at Promega, demonstrates how you can easily get great 24-hour or multi-day kinetic data using a GloMax® Microplate Reader.

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An Ambitious Endeavor: The Human Proteoform Project

Posted on by Riley Bell

On November 15, 2021, Science Advances announced the launch of The Human Proteoform Project. The ambitious project, led by the Consortium for Top-Down Proteomics, aims to address a critical next step in disease research. This means developing new technologies to outline a complete set of protein forms based on the ~20,000 genes in the human genome.

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Save Precious Time with Same-Well Multiplexing

Posted on by AnnaKay Kruger
Scientist performing a multi-well assay. Same-well multiplexing enables you to look at one event from several perspectives.

A graduate student believes he has mastered the art of “the assay”. No need to run duplicates, he knows exactly which one will get him the answers he needs right away.  

To challenge this, his PI proposes an exercise. He asks of the graduate student, “What happens when you treat cells with doxorubicin?”

The graduate student raises his cells, treats them accordingly, and decides to run a cell viability assay to determine their fate. He returns to the PI with the final verdict: his cells are dead.

The PI takes a look at the data and asks the graduate student to repeat the experiment with an additional assay for cytotoxicity―but the cytotoxicity assay shows that the cell membranes are intact, which only puzzles the graduate student. The PI asks him to run a third assay for apoptosis, and when the student does so, it becomes clear that the cells are dying.

The PI uses this opportunity to make his point: “Now do you see why I ask for more than one assay?”

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