Rare clonotypes and uses thereof
The invention is directed to a method of selecting disease-correlated clonotypes that have a reduced likelihood of producing a false positive signal of relapse when, used to monitor minimal residual disease. In accordance with the invention, candidate correlating clonotype are obtained from a patient, the rarity of each is determined either by comparison with a clonotype database or a clonotype model, and one or more of the rarest of such clonotypes are used to monitor the minimal residual disease.
1. A method of selecting one or more patient-specific clonotypes correlated with a lymphoid neoplasm for monitoring a minimal residual disease thereof, the method comprising the steps of:
(a) amplifying molecules of nucleic acid from T-cells and/or B-cells in a sample from a patient, the molecules of nucleic acid comprising recombined DNA sequences from T-cell receptor genes or immunoglobulin genes;
(b) sequencing the amplified molecules of nucleic acid to form a clonotype profile containing the one or more patient-specific clonotypes;
(c) comparing nucleotide sequences of the one or more patient-specific clonotypes from the clonotype profile to nucleotide sequences of clonotypes in a clonotype database, wherein the nucleotide sequences of clonotypes in a clonotype database are from at least one individual other than the patient to determine a presence, absence and/or level in the clonotype database of each of the nucleotide sequences of the one or more patient-specific clonotypes from the clonotype profile; and
(d) selecting the one or more patient-specific clonotypes for monitoring the minimal residual disease which are each correlated with the lymphoid neoplasm and which are each absent from the clonotype database or at a level in the clonotype database that is below a predetermined frequency.
2. The method of claim 1 , wherein the clonotype database contains clonotype profile data from each individual from a population of individuals.
3. The method of claim 2 , wherein the population comprises at least 10 individuals.
4. The method of claim 2 , wherein the clonotype profile data from each individual comprises nucleotide sequences of at least 10 4 clonotypes.
5. The method of claim 1 , wherein the clonotype profile comprises at least 10 4 clonotypes.
6. The method of claim 1 , wherein the nucleotide sequences of the one or more patient-specific clonotypes or the nucleotide sequences of clonotypes in the clonotype database are each 25 to 400 nucleotides in length encoding a segment of an immune receptor or immune receptor component selected from the group consisting of a VDJ rearrangement of IgH, a DJ rearrangement of IgH, a VJ rearrangement, of IgK, a VJ rearrangement of IgL, a VDJ rearrangement of TCRβ, a DJ rearrangement of TCRβ, a VJ rearrangement of TCRα, a VJ rearrangement of TCRγ, a VDJ rearrangement of TCRδ, and a VD rearrangement of TCRδ.
7. The method of claim 1 , wherein the predetermined frequency of the clonotypes in the clonotype database is less than 10 −6 .
8. The method of claim 1 , wherein the step of comparing includes counting a clonotype from the one or more patient-specific clonotypes in the clonotype profile as present in the clonotype database if a clonotype from the clonotypes in the clonotype database is a member of an identical clan as the clonotype from the one or more patient-specific in the clonotype profile.
9. The method of claim 8 , wherein the clonotype from the clonotypes in the clonotype database is in the identical clan as the clonotype the clonotype profile if (a) the clonotype from the clonotypes in the clonotype database is at least ninety percent identical to the clonotype from the one or more patient-specific clonotypes in the clonotype profile; (b) the clonotype from the clonotypes in the clonotype database and the clonotype from the one or more patient-specific clonotypes in the clonotype profile each comprise recombined sequences from immunoglobulin heavy chain and are related by a VH replacement; (c) the clonotype from the clonotypes in the clonotype database and the clonotype from the one or more patient-specific clonotypes in the clonotype profile have identically mutated V region and J region but have a different NDN region, or (d) the clonotype from the clonotypes in the clonotype database and the clonotype from the one or more patient-specific clonotypes in the clonotype profile each comprise recombined sequences from immunoglobulin heavy chain and are related by hypermutation.
10. A method of selecting one or more patient-specific clonotypes correlated with a lymphoid neoplasm for monitoring a minimal residual disease thereof, the method comprising the steps of:
(a) amplifying molecules of nucleic acid from T-cells and/or B-cells in a sample from a patient, the molecules of nucleic acid comprising recombined DNA sequences from T-cell receptor genes or immunoglobulin genes;
(b) sequencing the amplified molecules of nucleic acid to form a clonotype profile containing the one or more patient-specific clonotypes;
(c) comparing nucleotide sequences of constituent regions of each of the one or more patient-specific clonotypes from the clonotype profile to corresponding nucleotide sequences of constituent regions of a clonotype model to determine an expected frequency of occurrence for each of the constituent regions; and
(d) selecting at least one clonotype from the one or more patient-specific clonotypes for monitoring the minimal residual disease, wherein the at least one clonotype is correlated with the lymphoid neoplasm, and wherein a product of the expected frequencies of occurrence of each of the constituent regions of the at least one clonotype is below a predetermined frequency.