PCR in Clinical Biochemistry - TPMT genotyping

Rachel Marrington, Craig Webster and Jane Starczynski

Birmingham Heart of England NHS Foundation Trust

Thiopurine S-Methyl Transferase (TPMT) is involved in the metabolism of thiopurine drugs which are commonly used in the treatment of a range of conditions such as autoimmune diseases.  Point mutations in the TPMT gene can cause deficiency of the enzyme which can result in enhanced bone marrow toxicity.  Prospective measurement of either TPMT enzyme activity or the determination of TPMT genotype is recommended prior to therapy to avoid potentially fatal haematopoietic toxicity.  Birmingham Heartlands Hospital (BHH) have measured TPMT enzyme activity in whole blood by HPLC with fluorescence detection for over a year, and have recently set up a TPMT genotype assay to compliment the service.

Multiplexed amplification refractory mutation system (ARMS) based on the method by Roberts et al. is used for the genotyping analysis.  The PCR is multiplexed to include primers for both TPMT*2 and TPMT*3.  The method is described along with a comparison to the results obtained to that from phenotyping.  The advantages and limitations of TPMT genotyping is discussed, for example, TPMT activity is subject to sample degradation and drug interference.  A summary is also given of possible methods for the future in this area.