MONITORING METHOD FOR ADULT T-CELL LEUKEMIA/LYMPHOMA (ATL)

Abstract

publication date: 2018-10-11; filing date: 2017-04-06The present invention refers to a method for preparing a linear PCR product from genomic DNA derived from cells of a host subject infected with an retrovirus or a subject suffering from a disease associated with said retrovirus, wherein the PCR product contains a target sequence comprising an integration site of the retrovirus in the host genomic DNA of the cells, said integration site comprising at least the terminal end of 3'-LTR or 5'-LTR sequence of the retrovirus and the adjacent host genomic DNA sequence, wherein the PCR product comprises a first terminus and a second terminus and sequences in the following order: sequences specific for the first terminus, a sequence comprising at least6 consecutive random nucleotides followed by a linker sequence, host genomic DNA sequence, at least the terminal endof 3'-LTR or 5' -LTR sequence of the retrovirus, sequences specific for the second terminus; wherein the PCR product is prepared by specific steps. The present invention also refers to a method for determining and longitudinally monitor the dominant leukemic T lymphocyte clone in subjects suffering from Adult T-cell leukemia/lymphoma (ATL), wherein a linear PCR product is prepared by the method according to the first aspect of the present invention, said PCR product is subjected to multiplex sequencing thereby determining all insertion sites and all shearing sites, the shearing sites are correlated to the respective insertion site, followed by counting the number of different shear sites for each insertion site representing a specific T lymphocyte clone, removing any PCR duplicate from consideration by eliminating reads that have the same insertion site and the same random tag, and determining the abundance of each specific T lymphocyte clone therefrom