A NOVEL ASSAY TO DETECT TRANSLOCATION IN LEUKEMIAS

SEVERAL CHROMOSOMAL TRANSLOCATIONS ASSOCIATED WITH DIFFERENT TYPES OF LEUKEMIAS HAVE BEEN RECENTLY CHARACTERIZED. AS A RESULT OF THESE TRANSLOCATIONS, NEW MRNA SPECIES ARE PRODUCED, CODING FOR NOVEL FUSION PROTEINS. THE GOAL OF PHASE I IS TO DEVELOP A PANEL OF NON-ISOTOPIC ASSAYS TO DETECT T(1;19) (Q23;P13) ASSOCIATED WITH ACUTE LYMPHOBLASTIC LEUKEMIA WITH PREB PHENOTYPE; T(15;17) (Q21;Q11) ASSOCIATED WITH ACUTE PROMYELOCTIC LEUKEMIA; AND T(6;9) (P23;Q34) ASSOCIATED WITH ACUTE MYELOMONOCYTIC LEUKEMIA. THE ASSAYS WILL BE DEVELOPED BY CAPTURING MRNA FROM CELL LYSATES USING CAPTURE OLIGONUCLEOTIDES COUPLED TO POLYSTYRENE MICROBEADS, II) AMPLIFICATION OF THE CAPTURED MRNA USING POLYMERASE CHAIN REACTION (PCR), AND III) CAPTURE AND DETECTION OF PCR PRODUCTS USING ADDITIONAL INTERNAL CAPTURE OLIGONUCLEOTIDES ATTACHED TO MICROBEADS, BIOTINYLATED PROBES AND ALKALINE PHOSPHATASE AS A REPORTER. IN PHASE II, THESE ASSAYS, AS WELL AS OTHERS, WILL BE FORMATTED TO 96 WELL PLATES, IMPROVED TO ALLOW SEMI-QUANTITATION AND VALIDATED USING PATIENT SAMPLES. DURING PHASES II AND III, ASSAYS WILL BE COMMERCIALIZED AND MADE AVAILABLE AS RESEARCH REAGENTS. THESE ASSAYS WILL PERMIT THE DETECTION OF HYBRID MRNA IN ONE POSITIVE CELL PER MILLION CELLS, ALLOWING THEIR USE TO DETECT MINIMAL RESIDUAL DISEASE.