Methods for sequencing samples

Abstract

Personalized medicine involves the use of a patient's molecular markers to guide treatment regimens for the patient. The scientific literature provides multiple examples of correlations between drug treatment efficacy and the presence or absence of molecular markers in a patient sample. Methods are provided herein that permit efficient dissemination of scientific findings regarding treatment efficacy and molecular markers found in patient tumors to health care providers.

What is claimed is: 
     
         1 . A method of determining a status of more than one gene from a formalin-fixed paraffin embedded biopsy from a subject, said method comprising:
 (a) obtaining a formalin-fixed paraffin embedded (FFPE) biopsy, wherein said FFPE biopsy comprises nucleic acid; and   (b) determining a status of more than one gene in said nucleic acid, wherein said determining comprises de novo DNA sequencing of said nucleic acid.   
     
     
         2 . The method of  claim 1 , wherein said FFPE biopsy comprises a tumor cell. 
     
     
         3 . The method of  claim 1 , wherein said nucleic acid comprises DNA. 
     
     
         4 . The method of  claim 1 , wherein said nucleic acid comprises genomic DNA. 
     
     
         5 . The method of  claim 1 , further comprising amplifying said nucleic acid from said FFPE biopsy. 
     
     
         6 . The method of  claim 1 , wherein said determining said status of said more than one gene comprises determining a presence or absence of a point mutation, a single nucleotide polymorphism, copy number variation, substitution, insertion, deletion, rearrangement, de novo mutation, nonsense mutation, missense mutation, silent mutation, frameshift mutation, amplification, chromosomal translocation, interstitial deletion, chromosomal inversion, loss of heterozygosity, loss of function mutation, gain of function mutation, dominant negative mutation, or lethal mutation. 
     
     
         7 . The method of  claim 1 , wherein said determining said status of said more than one gene comprises determining a presence or absence of a substitution, insertion, deletion, rearrangement, or amplification. 
     
     
         8 . The method of  claim 1 , wherein said determining said status of said more than one gene comprises determining a presence or absence of a copy number variation. 
     
     
         9 . The method of  claim 1 , wherein said determining said status of said more than one gene comprises determining a presence or absence of a point mutation. 
     
     
         10 . The method of  claim 1 , wherein said determining said status of said more than one gene comprises determining a presence or absence of a single nucleotide polymorphism. 
     
     
         11 . The method of  claim 1 , wherein said determining said status of said more than one gene comprises determining a presence or absence of an amplification. 
     
     
         12 . The method of  claim 1 , wherein said de novo DNA sequencing comprises attaching said nucleic acid to a solid surface. 
     
     
         13 . The method of  claim 1 , wherein said de novo DNA sequencing comprises bridge amplification. 
     
     
         14 . The method of  claim 1 , wherein said de novo DNA sequencing comprises use of reversibly terminating nucleotides. 
     
     
         15 . The method of  claim 1 , further comprising performing digital polymerase chain reaction (PCR) on said FFPE biopsy. 
     
     
         16 . The method of  claim 1 , wherein said more than one gene comprises KIT, BRAF, Hsp90AA1, EGFR, KRAS, ESR1, TOPOI, PTEN, PIK3CA, NFE2L2, and ERBB2. 
     
     
         17 . The method of  claim 1 , wherein said more than one gene comprises KIT, BRAF, EGFR, ERBB2, ESR1, KRAS, and PIK3CA. 
     
     
         18 . The method of  claim 1 , wherein said more than one gene comprises KRAS, EGFR, BRAF, ERBB2, and KIT. 
     
     
         19 . The method of  claim 1 , wherein said more than one gene comprises KRAS, EGFR, BRAF, and KIT. 
     
     
         20 . The method of  claim 1 , wherein said more than one gene comprises KRAS. 
     
     
         21 . The method of  claim 1 , wherein said more than one gene comprises PIK3CA. 
     
     
         22 . The method of  claim 1 , wherein said FFPE biopsy comprises a tumor cell from a colon cancer, a bone cancer, a breast cancer, a central nervous system cancer, a gastric cancer, a cervical cancer, a blood cancer, an esophageal cancer, a head and neck cancer, a kidney cancer, a skin cancer, a lung cancer, or a carcinoma. 
     
     
         23 . The method of  claim 1 , wherein said FFPE biopsy comprises a tumor cell from a colon cancer. 
     
     
         24 . The method of  claim 1 , wherein said FFPE biopsy comprises a tumor cell from blood. 
     
     
         25 . The method of  claim 1 , wherein said determining said status of more than one gene comprises hybridizing a probe nucleic acid to nucleic acids corresponding to said more than one gene from a tumor cell. 
     
     
         26 . The method of  claim 1 , wherein said de novo DNA sequencing comprises use of a chemical-sensitive field effect transistor (chemFET) array. 
     
     
         27 . The method of  claim 1 , wherein said de novo DNA sequencing comprises using a laser to excite a fluorophore. 
     
     
         28 . The method of  claim 1 , further comprising ligating an adaptor to said nucleic acid. 
     
     
         29 . The method of  claim 1 , wherein said subject is a human. 
     
     
         30 . The method of  claim 1 , wherein said obtaining comprises sending said sample through a mail or delivery service. 
     
     
         31 . The method of  claim 1 , further comprising obtaining additional samples after obtaining an initial sample from said subject, wherein said additional samples comprise FFPE biopsy comprising DNA from a tumor cell. 
     
     
         32 . The method of  claim 1 , wherein said de novo DNA sequencing occurs in a Clinical Laboratory Improvement Amendment (CLIA) certified laboratory.