Quantification of Adaptive Immune Cell Genomes in a Complex Mixture of Cells

摘要

Compositions and methods are described for highly sensitive quantification of the relative representation of DNA from adaptive immune cells (e.g., T and/or B lymphocytes) in DNA extracted from complex mixtures of cells that include cells which are not adaptive immune cells. Included are methods for determining the relative presence in a tumor of tumor infiltrating lymphocytes (TIL), the relative presence of lymphocytes infiltrating a somatic tissue that is the target of an autoimmune disease, and the relative presence of lymphocytes infiltrating a transplanted organ.

主权项

1. A method for determining a relative quantity of tumor-infiltrating lymphocytes in a solid tumor, comprising:
obtaining a sample comprising a solid tumor tissue; contacting said sample with (1) a set of V-segment oligonucleotide primers and J-segment oligonucleotide primers, and (2) a pair of control sequence primers, wherein said control sequence primers are capable of amplifying a control sequence present in all cells in said sample; amplifying in a first PCR at least 80% of all rearranged TCR or Ig CDR3-encoding regions present in said sample using said set of V-segment oligonucleotide primers and J-segment oligonucleotide primers to produce a plurality of rearranged DNA amplicons; amplifying in a second PCR said control sequence present in said sample using said pair of control sequence primers; quantifying a number of adaptive immune receptor sequence reads in said sample generated from high-throughput sequencing (HTS) of said plurality of rearranged DNA amplicons; quantifying a number of control sequence reads in said sample using said amplified control sequence; and comparing said number of adaptive immune receptor sequence reads and said number of control sequence reads to estimate a relative quantity of tumor-infiltrating lymphocytes in said solid tumor.