Pharmacogenomics Tests And Methodologies

The majority of the current testing might be classified as pharmacogenetics (PGx). In attempting to guide the possible planning of clinical pharmacogenomics testing, AACC conducted a survey of the top 10 tests in 2005, showing the following.

1. CYP 2D6

2. TPMT

3. CYP 2C9

4. CYP 2C19

6. CYP 3A5

7. UGT1A1

8. MDR1

9. CYP 2B6 10. MTHFR

With recent pharmacogenomics findings for warfarin therapy, it would be appropriate to add the VKORC1 gene to this list (35). Within the list, CYP and other phase II enzymes such as UGT1A1 accounted for the majority of drug/substrate metabolism for drugs approved in the United States, about 75% involving CYP 3A4 and CYP 2D6 enzymes. According to the draft of LMPG by NACB, listed in the website http://www.nacb.org/lmpg/LMPG_Pharmacogenetics.pdf, the proposed alleles to be initially included for clinical pharmacogenetics were CYP 2D6*1 to CYP 2D6*12, CYP 2D6*17 and CYP 2D6*2A, 2C9*1 to 2C9*6, and 2C19*1 to 2C19*8 and 2C19*17. The final recommendations are pending. During the third FDA-DIA workshop, O'Kane presented an assessment of possible routine pharmacogenomics testing for medical care (36). He summarized some of the barriers including the current findings of more than 160 alterations for CYP 2D6 genes. Assay problems would include allelic drop-out, intra-allelic recombination, the need for specific assays not affected by pseudogenes CYP 2D7 and CYP 2D8 and to address gene conversion of CYP 2D6 from CYP 2D7. Cautions were recommended that CYP 2D6 genotyping might not be that routine.

Some of the AACC top 10 pharmacogenomic tests are readily performed either by home-brew assay or by some commercially available, FDA-approved test/platform. Payne (37) recently reviewed how to choose a method; Jannetto et al. (38), Weber (39),

Table 1

Methodologies for Pharmacogenetics Testing (37)

FDA Cleared or

Table 1

Methodologies for Pharmacogenetics Testing (37)

FDA Cleared or

Method

Company

Approved

Sequencing3

Abbott (Abbott Park, IL)

Yes

Real-time PCR

Applied Biosystems (Foster City, CA)

-

PCR Arrays

Autogenomics (Carlsbad, CA)

-

Sequencinga

Bayer Healthcare (Tarrytown, NY)

Yes

Pyrosequencing

Biotage AB (Uppsala, Sweden)

-

Real-time PCR

Celera Diagnostics (Alemeda, CA)

-

Real-time, allele-specific

DxS Genotyping (Manchester, UK)

-

PCR

PCR

Gentris (Morrisville, NC)

-

User-developed PCR

Nanogen (San Diego, CA)

-

arrays

Nanoparticles

Nanosphere (Northbrook, IL)

-

PCR arrays

Roche Diagnostics (Indianapolis, IN)

Yes

Invader assay

Third Wave Technologies (Madison, WI)

Yes

PCR bead-based

Tm Biosciences Corp (Toronto, Ontario)

-

detection

FISH

Vysis (Des Plaines, IL)b

Yes

FDA, Food Drug Administration; FISH, fluorescent in-situ hybridization; PCR, polymerase chain reaction.

a Sequencing for HIV drug resistance. b Vysis is now Abbott Molecular Diagnostics.

FDA, Food Drug Administration; FISH, fluorescent in-situ hybridization; PCR, polymerase chain reaction.

a Sequencing for HIV drug resistance. b Vysis is now Abbott Molecular Diagnostics.

and other recently published chapters (6) also summarized the currently available technologies. The approaches included non-amplification, for example, fluorescent in-situ hybridization, target and signal amplification methods including end-point polymerase chain reaction (PCR) detection, allele-specific primers, length analysis using restiction fragment length polymorphism (RFLP) and Oligomer Ligase Assays (OLA), real-time PCR, signal amplifications, and new methods including solid-phase microarray and fluorescent-based bead assay (liquid microarray). Other technologies reviewed included Pyrosequencing and, in the future, nanotechnology. The manufacturers and the status of FDA approval are listed in Table 1.

From personal communications, some labs performing genotyping has adapted the PCR liquid bead-based detection. The choice of the platform and assays seem to reflect, similar to the selection of clinical chemistry analyzers and tests, on the ease of "home-brew" assay development and the cost of the instrument and reagents.

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