Method of identifying myelodysplastic syndromes

Patent metadata

Field	Value
Publication number	US-9487776-B2
Application number	US-201213561687-A
Country	US
Kind code	B2
Filing date	Jul 30, 2012
Priority date	Jan 29, 2010
Publication date	Nov 8, 2016
Grant date	Nov 8, 2016

Abstract

Myelodysplastic syndromes display both hematological and biological heterogeneity with variable leukemia potential. To determine whether microRNAs expression offers diagnostic discrimination or influences malignant potential in MDS, bone marrow miRNA expression was investigated from prognostically distinct MDS subsets using a microarray platform. After background subtraction and normalization, data were analyzed indicating thirteen miRNA signature with statistically significant differential expression, including down-regulation of members of a leukemia associated miRNA family. A unique signature consisting of 10 miRNAs was closely associated with International Prognostic Scoring System risk category permitting discrimination between lower and higher risk disease. Selective overexpression of miRNA-181 family members was detected in higher risk MDS, indicating pathogenetic overlap with acute myeloid leukemia. Analysis of miRNA expression profile offers diagnostic utility, and provides pathogenetic and prognostic discimination in MDS.

First claim

What is claimed is:

1. A method of diagnosing and treating myelodysplastic disease syndrome in a patient, comprising:
collecting a sample suspected to be cancer from the patient;
detecting the existence of myelodysplastic disease syndrome, further comprising:
determining a quantified MDS-expression profile in a biological sample, wherein the quantified MDS-expression profile is obtained by:
generating a plurality of complementary DNA molecules for a plurality of MDS-marker genes from the biological sample;
contacting at least one complementary DNA molecule to at least one probe for at least one MDS-expression marker gene, wherein the MDS-expression marker gene is miR-222, miR-29a, miR-10a, miR-196a, miR-320, miR-100, miR124, miR-206, miR-146a, miR-150, miR-326, miR-7e, miR-197, miR-875-5p, let-7i, let-7c, let-7f, let-7d, or combinations thereof;
quantifying the at least one MDS-marker gene to generate the at least one MDS-expression profile;

comparing the quantified MDS-expression profile to a MDS-control profile obtained from normal donors to form a differential MDS-expression profile;
wherein the MDS-control profile is obtained by determining the expression of the at least one MDS-expression marker gene in the normal donors;
wherein the differential MDS-expression profile is indicative of presence of myelodysplastic disease syndrome when showing at least upregulation of miR-10a, miR-100, miR-196, miR-222, miR-320, let-7i, miR-320, let-7c, let-7f, let-7d, or miR-100, or downregulation of miR-7e, miR-29a, miR-124, miR-146a, miR-150, miR-197, miR-206, miR-326, or miR-875-5p, and

determining the risk level of myelodysplastic disease syndrome, further comprising:
determining a quantified risk-expression profile in a biological sample, wherein the quantified risk-expression profile is obtained by:
contacting at least one complementary DNA molecule to at least one probe for at least one risk-expression marker gene, wherein the risk-expression marker gene is miR126, miR125a-5p, miR21, miR125b, miR155, miR181a, miR181b, miR181c, miR181d, miR220, miR345, miR374b, miR18a, miR27a, miR424, miR376b, miR519d, miR138, miR494, miR451, miR486-5p, miR346, miR878-5p, miR498, miRN1221, miRN130a, or a combination thereof;
quantifying the at least one risk-expression marker gene to generate the quantified risk-expression profile;

comparing the quantified risk-expression profile of the biological sample to a quantified risk-control profile obtained from nucleic acids from age-matched control donors;
assigning a risk score based on variation between the risk-expression profile and the risk-control profile;
wherein the risk score is assigned by
assigning 1 point where the risk expression profile varies from the risk-control profile by 1-2 standard deviations and 2 points where the risk expression profile varies from the risk-control profile by over 2 standard deviations, or assigning 1 point where the risk expression profile varies from the risk-control profile by more than a threshold value, where the threshold value is a p value of 0.01 or a 1.5-fold change of the risk-control profile;
summing the points to obtain a risk score;

wherein a risk score of 2 or more is indicative of a high risk level of myelodysplastic disease syndrome; and

administering a treatment for myelodysplastic disease syndrome when the risk score is 2 or more, where the treatment for myelodysplastic disease syndrome is blood transfusion, platelet transfusion, non-myeloblative bone marrow transplant, administration of granulocyte colony-stimulating factor, erythropoiesis-stimulating agent, epogen, darbopoietin alpha, anti-thymocyte globulin, cyclosporine A, corticosteroid, lenalidomide, azacitidine, 5-azacytidine, low-dose chemotherapy, thalidomide, arsenic trioxide, or decitabine.

2. The method of claim 1 , wherein the biological sample is a bone marrow specimen collected from the patient.

3. The method of claim 1 , wherein the expression of miRNA in a biological sample is determined using a microarray platform.

4. The method of claim 1 , further comprising standardizing the expression profile by background subtraction and normalization using a set of housekeeping genes before comparing the expression profile of the at least one miRNA to those obtained from normal donors.

5. The method of claim 4 , wherein the expression profile is standardized using quantiles.

6. The method of claim 1 , further comprising isolating mature miRNA from the sample.

7. The method of claim 6 , further comprising obtaining complementary DNA from the mature miRNA by subjecting the mature miRNA to reverse transcription PCR.

8. The method of claim 1 , wherein the at least one miRNA used to diagnose the risk level of myelodysplastic disease syndrome is miR-181a, miR-181b, miR-181c, miR-181d, miR-1221, miR-376b, miR-125B, miR-155, miR130a, and miR-486-5p.

9. The method of claim 1 , wherein the miRNA has at least a 1.5 fold change in expression compared to the miRNA expression from normal donors, indicating the presence of myelodysplastic disease syndrome.

10. The method of claim 1 , further comprising standardizing the expression profile by background subtraction and normalization using a set of housekeeping genes before comparing the expression profile of the at least one miRNA to those obtained from normal donors.

11. The method of claim 1 , wherein the at least one miRNA used to detect the existence of myelodysplastic disease syndrome is miR-222, miR-10a, miR-196a, miR-320, miR-100, miR124, miR-206, miR-146a, miR-150, miR-326, miR-7e, miR-197, and miR-875-5p.

12. The method of claim 1 , wherein a risk score of 3 or more is indicative of the risk level of myelodysplastic disease syndrome.

13. The method of claim 1 , wherein the treatment for myelodysplastic disease syndrome is platelet transfusion, blood transfusion, non-myeloblative bone marrow transplant, low-dose chemotherapy, 5-azacytidine, thalidomide, arsenic trioxide, or azacitidine.

14. A method of predicting myelodysplastic disease syndrome in a patient, comprising
collecting a biological sample suspected to be cancer from the patient;
detecting the existence of myelodysplastic disease syndrome, further comprising:
determining a quantified MDS-expression profile in the biological sample, wherein the quantified MDS-expression profile is obtained by:
generating a plurality of complementary DNA molecules for a plurality of MDS-marker genes from the biological sample;
contacting at least one complementary DNA molecule to at least one probe for at least one MDS-expression marker gene, wherein the MDS-expression marker gene is miR-222, miR-29a, miR-10a, miR-196a, miR-320, miR100, miR124, miR-206, miR-146a, miR-150, miR-326, miR-7e, miR-197, miR-875-5p, let-7i, let-7c, let-7f, let-7d, or combinations thereof;
quantifying the at least one MDS-marker gene to generate the quantified MDS-expression profile;

comparing the quantified MDS-expression profile to a MDS-control profile obtained from normal donors to form a differential MDS-expression profile using class signatures or Significance Analysis of Microarrays;
wherein the MDS-control profile is obtained by determining the expression of the at least one MDS-expression marker gene in the normal donors;
wherein the differential MDS-expression profile is indicative of presence of myelodysplastic disease syndrome when showing at least upregulation miR-10a, miR-100, miR-196, miR-222, miR-320, let-7i, miR-320, let-7c, let-7f, let-7d, or miR-100, or downregulation of miR-7e, miR-29a, miR-124, miR-146a, miR-150, miR-197, miR-206, miR-326, or miR-875-5p; and

determining the risk level o

Assignees

• List Alan F
• Sokol Lubomir
• H Lee Moffitt Cancer Ct & Res

Inventors

• List Alan F
• Sokol Lubomir

Classifications

• C12Q1/6886

  for cancer (immunoassay for cancer G01N33/575) · CPC title

• C12Q2600/178

  miRNA, siRNA or ncRNA · CPC title

• C12N2310/141

  MicroRNAs, miRNAs · CPC title

• C12Q2600/118

  Prognosis of disease development · CPC title

• C12N2320/10

  in screening processes · CPC title