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Updated: May 2, 2026
Generation of Human Alloantigen-specific T Cells from Peripheral Blood
Published on: November 21, 2014
The human T-cell cloning assay: identifying genotypes susceptible to drug toxicity and somatic mutation
1Department of Biosciences, Karolinska Institute, CNT/NOVUM, 141 57, Huddinge, Sweden, saimei.hou@cnt.ki.se.
Abstract:
Humans exhibit marked genetic polymorphisms in drug metabolism that contribute to high incidence of adverse effects in susceptible individuals due to altered balance between metabolic activation and detoxification. The T-cell cloning assay, which detects mutations in the gene for hypoxanthine-guanine phosphoribosyl transferase (HPRT), is the most well-developed reporter system for studying specific locus mutation in human somatic cells. The assay is based on a mitogen- and growth factor-dependent clonal expansion of peripheral T-lymphocytes in which the 6-thioguanine-resistant HPRT mutants can be selected, enumerated, and collected for molecular analysis of the mutational nature. The assay provides a unique tool for studying in vivo and in vitro mutagenesis, for investigating the functional impact of common polymorphism in metabolism and repair genes, and for identifying risk genotypes for drug-induced toxicity and mutagenicity. This chapter presents a simple and reliable method for the enumeration of HPRT mutant frequency induced in vitro without using any source of recombinant interleukin-2. The other main feature is that only truly induced and unique mutants are collected for further analysis.
Insights
Genetic variations in drug metabolism affect how individuals respond to medications, leading to adverse effects. The hypoxanthine-guanine phosphoribosyl transferase (HPRT) T-cell assay helps identify these genetic risks for drug toxicity.
Area of Science:
- Pharmacogenomics
- Molecular Toxicology
- Human Genetics
Background:
- Human genetic polymorphisms significantly impact drug metabolism, leading to variable drug responses and adverse effects.
- The balance between metabolic activation and detoxification pathways is crucial for preventing drug-induced toxicity.
- The hypoxanthine-guanine phosphoribosyl transferase (HPRT) gene assay is a key tool for studying somatic cell mutations.
Purpose of the Study:
- To present a reliable method for enumerating HPRT mutant frequency in human T-lymphocytes induced in vitro.
- To facilitate the molecular analysis of induced mutations without recombinant interleukin-2.
- To identify individuals at risk for drug-induced toxicity based on genetic profiles.
Main Methods:
- Utilizing a T-cell cloning assay to detect mutations in the HPRT gene within human somatic cells.
- Employing a mitogen- and growth factor-dependent clonal expansion of peripheral T-lymphocytes.
- Selecting and enumerating 6-thioguanine-resistant HPRT mutants for molecular characterization.
Main Results:
- A simple and reliable method for enumerating in vitro induced HPRT mutant frequency was established.
- The assay allows for the collection of truly induced and unique mutants for further analysis.
- The HPRT assay serves as a reporter system for in vivo and in vitro mutagenesis studies.
Conclusions:
- The HPRT T-cell assay is a valuable tool for studying mutagenesis and identifying risk genotypes for drug toxicity.
- Understanding genetic polymorphisms in drug metabolism is essential for personalized medicine and reducing adverse drug reactions.
- This method provides a means to investigate the functional impact of genetic variations in metabolism and repair genes.
