Oral cancer markers and their detection
AssigneeZila Biotechnology, Inc.
InventorsDouglas D. Burkett, David Sidransky, Antonette C.P. Allen, Francis A. Chiafari, Mark Bride, Yu Ping Maguire
Life of this patent
Estimating the term…
Abstract
Methods of detecting progression from precancer to cancer are provided utilizing toluidine blue staining as well as detecting allelic variation at microsatellite loci. An allelic variation in one or more locus is indicative of a progression from precancer to cancer.
Claims22
A method for detecting cancer or precancer in a subject, the method comprising:
- (a) determining a first ratio of a level of microsatellite DNA present at a first allele to a level of microsatellite DNA present at a second allele in an oral epithelial cell of the subject;
- determining a second ratio of a level of microsatellite DNA present at a first allele to a level of microsatellite DNA present at a second allele in a nonepithelial cell of the subject;
- and
- (b) comparing the first ratio to the second ratio;
- wherein the subject is heterozygous for the genetic locus;
- wherein the first and second alleles of the oral epithelial cell and the nonepithelial cell are at the genetic locus, and the genetic locus comprises microsatellite DNA;
- wherein the genetic locus is at least one of D3S3597, D3S1067, D3S1300, D3S4103, D9S171, IFN-A, D9S1748, D17S695, or tp53;
- and wherein a difference between the first and second ratios is indicative of cancer or precancer.
A method of analyzing a microsatellite locus, the method comprising:
- a) extracting DNA from paraffin embedded samples of an oral epithelial cell from a subject and of a nonepithelial cell of the subject;
- b) providing primers for amplifying a first and a second allele in the oral epithelial cell at the microsatellite locus and a first and second allele in the nonepithelial cell at the microsatellite locus;
- c) amplifying the microsatellite locus, wherein the microsatellite locus comprises at least one of D3S1067, D3S1300, DS4103, D3S3597, D9S171, IFN-A, D9S1748, D17S695, and tp53;
- and
- d) determining a first ratio of a level of microsatellite DNA present at the first allele to a level of microsatellite DNA present at the second allele in the oral epithelial cell, and determining a second ratio of a level of microsatellite DNA present at the first allele and a level of microsatellite DNA present at the second allele in the nonepithelial cell;
- and
- e) comparing the first and second ratios.
A method of analyzing microsatellite loci, the method comprising detecting allelic variation at a genetic locus comprising D3S1067, D3S3597, D3S1300, D3S4103, D9S171, IFN-A, D9S1748, D17S695, and tp53.
A method of detecting cancer or precancer in a subject, the method comprising:
- a) determining a size of a DNA fragment amplified at a minimum of one locus selected from the group consisting of D3S3597, D3S1067, D3S1300, D3S4103, D9S171, IFN-A, D9S1748, D17S695, or tp53 using SEQ. ID NO. 1, SEQ. ID NO. 2, SEQ. ID NO. 3, SEQ. ID NO. 4, SEQ. ID NO. 5, SEQ. ID NO. 6, SEQ. ID NO. 7, SEQ. ID NO. 8, SEQ. ID NO. 9, SEQ. ID NO. 10, SEQ. ID NO. 11, SEQ. ID NO. 12, SEQ. ID NO. 13, SEQ. ID NO. 14, SEQ. ID NO. 15, SEQ. ID NO. 16, SEQ. ID NO. 17, or SEQ. ID NO. 18 as primers in a non-cancerous biological sample and comparing said size to a size of a DNA fragment at the same locus in a cancerous biological sample;
- wherein a difference in size is indicative of microsatellite instability.
A method of analyzing microsatellite loci, the method comprising:
- (a) providing primers for amplifying a set of at least two microsatellite loci of human DNA, wherein the set of at least two microsatellite loci are selected from the group consisting of D3S1067, D3S3597, D3S1300, D3S4103, D9S171, IFN-A, D9S1748, D17S695, and tp53;
- (b) amplifying the set of at least two microsatellite loci from a sample of genomic DNA in a multiplex amplification reaction, using the primers, thereby producing amplified DNA fragments;
- and
- c) determining the size of amplified DNA fragments.
A method of detecting cancer or precancer in a subject, the method comprising:
- (a) administering a toluidine blue O stain;
- (b) providing a control sample DNA and a test sample DNA;
- (c) amplifying at least one microsatellite locus selected from the group consisting of D3S3597, D3S1067, D3S1300, D3S4103, D9S171, IFN-A, D9S1748, D17S695, and tp53 in the control sample DNA and the test sample DNA;
- and
- (d) determining a microsatellite allele ratio of the control sample DNA and a microsatellite allele ratio of the test sample DNA at the microsatellite locus;
- (e) detecting a difference between the microsatellite allele ratio of the test sample DNA and the microsatellite allele ratio of the control sample DNA.
A kit for detecting a DNA mutation, the kit comprising oligonucleotide primers that are complementary to a nucleotide sequence that flanks nucleotide repeats of microsatellite DNA, wherein the nucleotide repeats of microsatellite DNA comprise at least one of D3S3597, D3S1067, D3S1300, D3S4103, D9S171, IFN-A, D9S1748, D17S695, and tp53.
A method of detecting cancer or precancer in a subject, the method comprising:
- (a) administering a toluidine blue O stain and (b) detecting allelic variation in at least one locus selected from the group comprising D3S3597, D3S1067, D3S1300, D3S4103, D9S171, IFN-A, D9S1748, D17S695, and tp53.
Dependencies are inferred from the claim text.