Given the evolving requirements from regulatory bodies and insurers for establishing the analytic and clinical validity of genetic tests, Invitrogen is marketing a procedure to lower the false positive rates for tests by preventing sample contamination.
Invitrogen and Greiner Bio-One recently announced they had finalized a licensing agreement that would grant Greiner non-exclusive rights to use Invitrogen’s dUTP (deoxyuridine triphosphate) to improve the reliability of two of its diagnostic tests, the HPV DNA test PapilloCheck and the DNA array CytoCheck.
"The overall benefit of using the Invitrogen dUTP during the PCR is that false-positive results are reliably prevented at the beginning of the procedure, while the genuine sample DNA remains intact," Heinz Schmid, Greiner Bio-One managing director, said in a release.
The announcement comes at a time when the US Food and Drug Administration, the Centers for Medicare and Medicaid Services, Congress, insurance companies, and patient groups are all trying to grapple with the level of regulatory oversight, and the amount of clinical and analytical data diagnostic firms should provide to ensure the safety and efficacy of genetic tests [see PGx Reporter 06-27-2007, 06-20-2007].
An Invitrogen spokesperson told Pharmacogenomics Reporter in an e-mail that as many as a dozen diagnostics companies have licensed the dUPT product, but would not identify the companies.
“The dUTP technology can be used to decrease false positives across a majority of PCR-based technologies,” the spokesperson said. “Greiner Bio-One is just one of multiple diagnostic companies who have recognized the value of this technology and have licensed it from Invitrogen.”
Although the financial details of the deal between Invitrogen and Greiner were not disclosed, Invitrogen’s agreement with Greiner covers the rights for the use of dUTP in PCR-based methods to “prevent sample contamination by amplification products in aerosols.”
Invitrogen’s patented procedure adds dUTP as the base to the primer during the PCR reaction, instead of dTTP (deoxythymidine triphosphate). Invitrogen’s process further removes the risk of amplifying a contaminated product, and thus reduces the chance of a false-positive result, by adding the enzyme uracil-DNA-glycosylase prior to starting the reaction, which selectively degrades potential carry over of dUTP-containing amplification products into a subsequent PCR reaction.
“The unamplified target sample, which contains dTTP, will not be degraded by the UDG and will amplify normally,” the Invitrogen spokesperson said.
In a release announcing the Invitrogen/Greiner agreement, the companies noted that Greiner began delivering PapilloCheck with dUTP in the MasterMix in June. Greiner will include dUTP in the product kits for CytoCheck starting this fall.
According to scientific literature, the most common source of DNA contamination causing false positive results is previously amplified PCR products, called amplicons, which are copies of the original DNA or RNA target region. The risk of amplicon contamination increases when a diagnostic PCR is performed multiple times for one DNA sequence, and when the process is set to detect a small number of DNA molecules.
Although Griener would not reveal the false positive rate of its products with and without the use of dUTP, the company said that good laboratory practices can reduce contamination even without Invitrogen’s dUTP procedure.
“The dUTP technology can be used to decrease false positives across a majority of PCR-based technologies. Greiner Bio-One is just one of multiple diagnostic companies who have recognized the value of this technology and have licensed it from Invitrogen.”
“The rate of false positives depends strongly on the careful handling, separation of working areas, etc. Also, without dUTP, false positives can successfully be prevented by sterile working conditions and careful handling,” a Greiner spokesperson told Pharmacogenomics Reporter in an e-mail this week. However, the company decided to include dUTP to make the handling of its products “easier and safer for customers.”
Good laboratory practices cannot completely wipe out the risk of false positives due to amplicon contamination, according to scientific literature. As such, investment in negative controls and repeating assays even if the minutest possibility of contamination is suspected adds to the cost of performing diagnostic tests.
The incorporation of dUTP, followed by the eradication of the amplicon with uracil-DNA-glycosylase to avoid contamination in subsequent reactions, is a method widely used by laboratories to manage amplicon contamination issues, according to scientific literature.
"Greiner Bio-One's HPV test will now deliver more reliable results for women who may need to be treated for HPV, which is the leading cause of cervical cancer," said Amy Butler, Invitrogen vice president of gene expression profiling, in a statement. "Licensing patented technologies like dUTP is allowing Invitrogen to assist diagnostic companies around the world to improve the sensitivity and specificity of their offerings."
Some pharmaceutical companies have cited the risk of false positive results with certain diagnostics as a point of apprehension when deciding to develop companion tests for their therapeutic products. In comments to FDA’s draft guidance for in vitro diagnostic multivariate index assays, Genentech expressed concern about the possibility of false positive or negative results with IVDMIAs that are not approved by the FDA [see PGx Repoter 06-13-2007].
Greiner’s spokesperson said that the use of dUTP is expected to increase customer confidence in the reliability of its products, which may fuel greater market acceptance.
“It is definitely an advantage over other PCR-based methods not using dUTP,” Greiner’s spokesperson said. “The acceptance for these kits including this additional security feature will of course be increased.”