DIAGNOSIS OF MELANOMA BY NUCLEIC ACID ANALYSIS

• US 20080274908A1
• Filed: 05/02/2008
• Published: 11/06/2008
• Est. Priority Date: 05/04/2007
• Status: Abandoned Application

• Current Assignee: DermTech International
• Original Assignee: DermTech International
• Inventors: Chang, Sherman
• Application Number:
  US12/114,669
• Publication Number:
  US 20080274908A1
• Time in Patent Office: Days
• US Class Current: 506/8
• CPC Class Codes: C12Q 1/6886 for cancer immunoassay for cancer G01N33/574
• CPC Class Codes: C12Q 2600/158 Expression markers
• CPC Class Codes: G16B 25/00 ICT specially adapted for hybridisation; ICT specially adapted for gene or protein expression
• CPC Class Codes: G16B 25/10 Gene or protein expression profiling; Expression-ratio estimation or normalisation
• CPC Class Codes: G16B 40/00 ICT specially adapted for biostatistics; ICT specially adapted for bioinformatics-related machine learning or data mining, e.g. knowledge discovery or pattern finding
• CPC Class Codes: G16B 40/30 Unsupervised data analysis

Abstract

The present invention provides methods for diagnosing melanoma in a subject by analyzing nucleic acid molecules obtained from the subject. The present invention also provides methods for distinguishing melanoma from dysplastic nevi and/or normal pigmented skin. The methods include analyzing expression or mutations in epidermal samples, of one or more skin markers. The methods can include the use of a microarray to analyze gene or protein profiles from a sample.

Claims

1. A method for characterizing a skin lesion in a subject comprising analyzing a nucleic acid molecule from one or more genes listed in any of Tables 1-6, or any combination thereof, in a sample of the skin lesion, thereby characterizing a skin lesion of the subject.

2. The method of claim 1, wherein the nucleic acid molecule is RNA.

3. The method of claim 1, wherein analyzing the nucleic acid molecule comprises detecting one or more mutations in the nucleic acid sequence of the nucleic acid molecule.

4. The method of claim 3, wherein the one or more mutations are selected from the group consisting of a substitution, a deletion, and an insertion.

5. The method of claim 1, further comprising amplifying the nucleic acid molecule obtained from the sample prior to analyzing.

6. The method of claim 1, wherein the sample is obtained by applying an adhesive tape to a target area of skin in a manner sufficient to isolate the sample adhering to the adhesive tape.

7. The method of claim 1, further comprising using the characterizing to determine a treatment regimen.

8. The method of claim 2, wherein the isolated nucleic acid molecule or an amplification product thereof, is applied to a microarray.

9. The method of claim 8, wherein an expression profile is detected using a microarray.

10. The method of claim 1, wherein the sample is obtained from a biopsy taken at the site of the skin lesion or surrounding margin.

11. The method of claim 6, wherein the tape comprises a rubber adhesive on a polyurethane film.

12. The method of claim 6, wherein about one to ten adhesive tapes or one to ten applications of a tape are applied and removed from the skin.

13. The method of claim 6, wherein about one to eight adhesive tapes or one to eight applications of a tape are applied and removed from the skin.

14. The method of claim 6, wherein about one to five adhesive tapes or one to five applications of a tape are applied and removed from the skin.

15. The method of claim 6, wherein the method further comprises taking a biopsy of the target area of the skin.

16. The method of claim 2, wherein the analyzing is performed in situ.

17. The method of claim 1, wherein the nucleic acid molecule is from one or more genes listed in Table 6.

60. The method of any one of claims 1, 18, 33, or 47, wherein the subject is human.

61. The method of any one of claims 1, 18, 33, or 47, wherein the sample is an epidermal sample.

18. A method of distinguishing melanoma from dysplastic nevi or normal pigmented skin in a subject comprising analyzing a nucleic acid molecule from one or more genes listed in any of Tables 1-6, or any combination thereof, in a sample from the subject, thereby distinguishing melanoma from dysplastic nevi or normal pigmented skin in a subject.

19. The method of claim 18, wherein the nucleic acid molecule is RNA.

20. The method of claim 18, wherein analyzing the nucleic acid molecule comprises detecting one or more mutations in the nucleic acid sequence of the nucleic acid molecule.

21. The method of claim 20, wherein the one or more mutations are selected from the group consisting of a substitution, a deletion, and an insertion.

22. The method of claim 18, further comprising amplifying the nucleic acid molecule obtained from the sample prior to analyzing.

23. The method of claim 18, wherein the sample is obtained by applying an adhesive tape to a target area of skin in a manner sufficient to isolate the sample adhering to the adhesive tape, wherein the sample comprises nucleic acid molecules.

24. The method of claim 19, wherein the isolated nucleic acid molecule or an amplification product thereof, is applied to a microarray.

25. The method of claim 22, wherein an expression profile is detected using a microarray.

26. The method of claim 18, wherein the sample is obtained from a biopsy taken from a target area of skin from the subject.

27. The method of claim 23, wherein the tape comprises a rubber adhesive on a polyurethane film.

28. The method of claim 23, wherein about one to ten adhesive tapes or one to ten applications of a tape are applied and removed from the skin.

29. The method of claim 23, wherein about one to eight adhesive tapes or one to eight applications of a tape are applied and removed from the skin.

30. The method of claim 23, wherein about one to five adhesive tapes or one to five applications of a tape are applied and removed from the skin.

31. The method of claim 23, wherein the method further comprises taking a biopsy of the target area of the skin.

32. The method of claim 19, wherein the analyzing is performed in situ.

33. A method for diagnosing melanoma in a subject comprising detecting an altered level of a target protein in a sample from the subject, as compared to the level of the target protein in a corresponding sample from a subject that does not have melanoma, wherein the protein is an expression product of a gene listed in any of Tables 1-6, or any combination thereof, thereby diagnosing melanoma in the subject.

34. The method of claim 33, wherein the sample is obtained by applying an adhesive tape to a target area of skin in a manner sufficient to isolate the sample adhering to the adhesive tape, wherein the sample comprises cells, and further comprising lysing the cells to extract the target protein.

35. The method of claim 34, wherein the tape comprises a rubber adhesive on a polyurethane film.

36. The method of claim 34, wherein between one and ten adhesive tapes are applied to the skin and removed from the skin.

37. The method of claim 34, wherein about one to eight adhesive tapes are applied and removed from the skin.

38. The method of claim 34, wherein about one to five adhesive tapes are applied and removed from the skin.

39. The method of claim 34, wherein the method further comprises taking a biopsy of the target area of the skin.

40. The method of claim 39, wherein protein is extracted from the biopsy sample, and the level of protein in the biopsy and the level of protein in the tape sample are analyzed.

41. The method of claim 33, wherein the sample is obtained by from a biopsy of a target area of skin.

42. The method of claim 33, further comprising obtaining a sample from uninvolved tissue of the subject.

43. The method of claim 42, wherein the sample from uninvolved tissue is obtained by taking a biopsy of the uninvolved skin.

44. The method of claim 42, wherein the sample from uninvolved tissue is obtained by:

a) applying an adhesive tape to skin of the subject in a manner sufficient to isolate a skin sample adhering to the adhesive tape, wherein the skin sample comprises cells from the stratum corneum and wherein the skin is unaffected by a disease to be tested;

b) lysing the cells to extract a protein; and

c) quantitating the extracted protein.

45. The method of claim 42, wherein the uninvolved skin is from the mastoid process or the upper back.

46. The method of claim 33, wherein the protein is an expression product of a gene listed in Table 6.

47. A method for diagnosing melanoma in a subject, comprising:

a) providing a gene expression profile of a target area suspected of being melanoma on the skin of the subject, wherein the target area of the skin simultaneously expresses a plurality of genes at the protein level that are markers for melanoma; and

b) comparing the subject'"'"'s gene expression profile to a reference gene expression profile obtained from a corresponding normal skin sample, wherein the reference gene expression profile comprises an expression value of a target gene selected from the group consisting of human mRNA for tyrosinase-related protein 1, LIM homeobox 2, melan-A, spermidine/spermine N1-acetyltransferase, NDRG family member 2, carbohydrate (N-acetylglucosamine-6-O) sulfotransferase 2, syndecan binding protein (syntenin), crystallin, alpha B, endothelin receptor type B, tetratricopeptide repeat domain 3, WAP four-disulfide core domain 3, cytochrome P450 (family 1, subfamily B, polypeptide 1), p8 protein (candidate of metastasis 1), interferon gamma-inducible protein 16, dopachrome tautomerase (dopachrome delta-isomerase, tyrosine-related protein 2), solute carrier family 39 (zinc transporter), member 6, heat shock 70 kDa protein 2, vesicle-associated membrane protein 2 (synaptobrevin 2), Similar to AI661453 protein, and transient receptor potential cation channel (subfamily M, member 1), tyrosinase (oculocutaneous albinism IA), endothelin receptor type B, tetratricopeptide repeat domain 3, syndecan binding protein (syntenin), v-kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog, OTU domain, ubiquitin aldehyde binding 1, protein phosphatase 1 regulatory (inhibitor) subunit 12A, calponin 2, dishevelled dsh homolog 3 (Drosophila), tribbles homolog 2 (Drosophila), mucolipin 3, actinin alpha 4, ribosomal protein S15, CDC37 cell division cycle 37 homolog (S. cerevisiae), jumonji domain containing 3, v-maf musculoaponeurotic fibrosarcoma oncogene homolog B (avian), c-Maf-inducing protein, myosin heavy polypeptide 14, Hypothetical protein MGC40222, or any combination thereof.

48. The method of claim 47, wherein the reference gene expression profile further comprises an expression value of any target gene listed in Tables 1-6.

49. The method of claim 47 or 48, wherein the reference gene expression profile is contained within a database.

50. The method of claim 47, wherein said comparing is carried out using a computer algorithm.

51. A kit for characterizing a skin lesion in a subject comprising a skin sample collection device and one or more probes or primers that selectively bind to one or more nucleic acid molecules in any of Tables 1-6, or to a nucleic acid or protein expression product of a nucleic acid molecule in any of Tables 1-6.

52. The kit of claim 51, wherein the kit provides a probe which binds to a portion of a nucleic acid molecule in any of Tables 1-6.

53. The kit of claim 51, wherein the kit provides one or more primer pairs comprising a forward primer that selectively binds upstream of a gene on one strand and a reverse primer that selectively binds upstream of a gene on a complementary strand, wherein the gene is listed in any of Tables 1-6.

54. The kit of claim 51, wherein the skin sample collection device is a biopsy needle.

55. The kit of claim 51, wherein the skin sample collection device is an adhesive tape.

56. The kit of claim 55, wherein the adhesive tape comprises a rubber adhesive on a polyurethane film.

57. The kit of claim 51, further comprising a microarray containing at least a fragment of a gene or a nucleic acid or protein product of a gene identified in any of Tables 1-6 or any combination thereof.

58. A kit for characterizing a skin lesion in a subject comprising an applicator and one or more probes or primers that selectively bind to one or more of nucleic acid molecules in any of Tables 1-6, or to a nucleic acid or protein expression product of a nucleic acid molecule in any of Tables 1-6.

59. The kit of claim 58, wherein the probes are detectably labeled.