Common consensus LNA probe for quantitative PCR assays in cancer: Vehicles for minimal residual disease detection in t(11;14) and t(14;18) positive malignant lymphomas

摘要

The use of locked nucleic acid (LNA) probes and primers potentially improves sensitivity and specificity of quantitative PCR (qPCR) assays. One area of application is that of minimal residual cancer where PCR techniques have proved to be highly relevant tools in patient follow-up. We present here sensitive and specific consensus qPCR assays for quantification of the malignant lymphoma translocations, t(ll;14) and t(14;18), by taking advantage of the thermodynamic properties of LNA. The assays were applied to genomic DNA from patients diagnosed with mantle cell lymphoma (MCL) and follicular lymphoma (FL), respectively. Two consensus forward primers targeting the BCU and BCL2 genes were designed together with a common consensus reverse primer and hydrolysis probe, the latter consisting exclusively of LNA, both targeting the J segments of the immunoglobulin heavy chain (IgH) gene. The quantitative range of both assays was 1 x 10° to 5 x 10~5, and the sensitivity was 10~-5 without the need for patient-specific primers. Peripheral blood (PB) and bone marrow (BM) samples from 36 patients diagnosed with MCL and nine patients diagnosed with FL were analysed using this novel qPCR approach. The level of minimal residual disease (MRD) using t(ll;14) and t(14;18) as genetic targets reflected the clinical status of the patients: low levels of MRD at clinical remission, and increasing levels at disease progression. The present assays could prove as useful tools in lymphoma therapy.