Diagnosis of melanoma by nucleic acid analysis
The present invention provides methods for diagnosing melanoma and/or solar lentigo in a subject by analyzing nucleic acid molecules obtained from the subject. The present invention also provides methods for distinguishing melanoma from solar lentigo and/or 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.
1. A method for detecting an expression level of preferentially expressed antigen in melanoma (PRAME) in a subject, the method comprising:
(a) obtaining a biological sample of a skin lesion suspected of comprising melanoma from the subject with an adhesive tape, wherein the biological sample comprises or is suspected of comprising a nucleic acid molecule expressed from preferentially expressed antigen in melanoma (PRAME), and wherein the biological sample comprises cells from the stratum corneum; and
(b) detecting whether the expression level of preferentially expressed antigen in melanoma (PRAME) in the biological sample is at a level that is above an expression level of PRAME in a non-melanoma sample by application of a detectably labeled probe that hybridizes to a nucleic acid molecule expressed from PRAME, whereby the presence of a nucleic acid molecule expressed from PRAME in the biological sample in an amount that is greater than the presence of PRAME in a non-melanoma sample is indicative of melanoma in the biological sample.
2. The method of claim 1 , wherein the adhesive tape is a pliable tape comprising rubber adhesive.
3. The method of claim 2 , provided that the adhesive tape further comprises a polyurethane film.
4. The method of claim 3 , provided that about one to about ten adhesive tapes or one to ten applications of the tape are applied to and removed from the skin lesion.
5. The method of claim 1 , provided that the nucleic acid molecule comprises an RNA molecule.
6. The method of claim 5 , wherein about 200-500 picograms of the RNA molecule are analyzed.
7. The method of claim 5 , provided that determining an expression level of the nucleic acid molecule comprises generating a cDNA product from the RNA molecule, and amplifying the cDNA product by real-time quantitative polymerase chain reaction (PCR).
8. The method of claim 1 , provided that determining an expression level of the nucleic acid molecule comprises microarray analysis or a direct sequencing method.
9. The method of claim 1 , provided that the expression level of the corresponding nucleic acid molecule in the non-melanoma sample is contained within a database.
10. The method of claim 9 , provided that the expression level of the nucleic acid molecule in the biological sample is compared to the expression level of the corresponding nucleic acid molecule in the non-melanoma sample using a computer.
11. A method for detecting melanoma in a biological sample, comprising:
(a) obtaining a biological sample of a skin lesion suspected of comprising melanoma from a subject with an adhesive tape; wherein the biological sample comprises or is suspected of comprising a nucleic acid molecule expressed from preferentially expressed antigen in melanoma (PRAME), wherein the nucleic acid molecule comprises an RNA molecule, and
wherein the biological sample comprises cells from the stratum corneum;
(b) generating a complementary DNA (cDNA) product from the RNA molecule;
(c) detecting an expression level of the nucleic acid molecule expressed from preferentially expressed antigen in melanoma (PRAME) in the biological sample by performing real-time quantitative PCR to amplify and quantify the cDNA product; and
(d) comparing the expression level of the nucleic acid molecule expressed from PRAME based on quantification of the cDNA product in the biological sample to the expression level of a corresponding nucleic acid molecule expressed from PRAME in a non-melanoma sample, whereby an increased expression level of the nucleic acid molecule expressed from PRAME in the biological sample as compared to the non-melanoma sample is indicative of melanoma, thereby detecting melanoma in the biological sample of a skin lesion.
12. The method of claim 11 , provided that about one to about ten adhesive tapes or one to ten applications of the tape are applied to and removed from the skin lesion.
13. The method of claim 11 , wherein the adhesive tape is a pliable tape comprising rubber adhesive.
14. The method of claim 13 , provided that the adhesive tape further comprises a polyurethane film.