Aptamers and uses thereof
Methods and compositions are provided for specific aptamers and aptamer pools that bind biomarkers of interest such as microvesicle surface antigens or functional fragments of microvesicle surface antigens. In various embodiments, aptamers of the invention are used in diagnostic, prognostic, or theranostic processes to screen a biological sample for the presence or levels of biomarkers such as microvesicles that are determined to provide a diagnostic, prognostic, or theranostic readout. The diagnosis, prognosis, or theranosis may be related to cancer or other diseases and disorders. The invention also provides methods and composition to facilitate aptamer library screening and aptamer detection methods.
1. A method comprising;
(a) contacting a biological sample comprising microvesicles with a pool of different aptamers enriched to target one or more microvesicles;
(b) removing unbound aptamers;
(c) eluting bound aptamers;
(d) identifying the aptamers that are eluted by sequencing, and
(e) determining the presence and copy number of the identified aptamers; wherein it is not necessary to know the precise targets of the different aptamers.
2. The method of claim 1 , wherein the pool of different aptamers are pre-selected through at least one step of positive and/or negative selection, wherein positive selection comprises selection of aptamers against a sample having substantially similar characteristics compared to the biological sample, and wherein negative selection comprises selection of aptamers against a sample having substantially different characteristics compared to the biological sample.
3. The method of claim 1 , wherein the sequencing comprises performing high-throughput sequencing.
4. The method of claim 1 , wherein the biological sample is from a subject suspected of having or being predisposed to having a cancer.
5. The method of claim 4 , wherein the cancer comprises an acute lymphoblastic leukemia; acute myeloid leukemia; adrenocortical carcinoma; AIDS-related cancers; AIDS-related lymphoma; anal cancer; appendix cancer; astrocytomas; atypical teratoid/rhabdoid tumor; basal cell carcinoma; bladder cancer; brain stem glioma; brain tumor (including brain stem glioma, central nervous system atypical teratoid/rhabdoid tumor, central nervous system embryonal tumors, astrocytomas, craniopharyngioma, ependymoblastoma, ependymoma, medulloblastoma, medulloepithelioma, pineal parenchymal tumors of intermediate differentiation, supratentorial primitive neuroectodermal tumors and pineoblastoma); breast cancer; bronchial tumors; Burkitt lymphoma; cancer of unknown primary site; carcinoid tumor; carcinoma of unknown primary site; central nervous system atypical teratoid/rhabdoid tumor; central nervous system embryonal tumors; cervical cancer; childhood cancers; chordoma; chronic lymphocytic leukemia; chronic myelogenous leukemia; chronic myeloproliferative disorders; colon cancer; colorectal cancer; craniopharyngioma; cutaneous T-cell lymphoma; endocrine pancreas islet cell tumors; endometrial cancer; ependymoblastoma; ependymoma; esophageal cancer; esthesioneuroblastoma; Ewing sarcoma; extracranial germ cell tumor; extragonadal germ cell tumor; extrahepatic bile duct cancer; gallbladder cancer; gastric (stomach) cancer; gastrointestinal carcinoid tumor; gastrointestinal stromal cell tumor; gastrointestinal stromal tumor (GIST); gestational trophoblastic tumor; glioma; hairy cell leukemia; head and neck cancer; heart cancer; Hodgkin lymphoma; hypopharyngeal cancer; intraocular melanoma; islet cell tumors; Kaposi sarcoma; kidney cancer; Langerhans cell histiocytosis; laryngeal cancer; lip cancer; liver cancer; lung cancer; malignant fibrous histiocytoma bone cancer; medulloblastoma; medulloepithelioma; melanoma; Merkel cell carcinoma; Merkel cell skin carcinoma; mesothelioma; metastatic squamous neck cancer with occult primary; mouth cancer; multiple endocrine neoplasia syndromes; multiple myeloma; multiple myeloma/plasma cell neoplasm; mycosis fungoides; myelodysplastic syndromes; myeloproliferative neoplasms; nasal cavity cancer; nasopharyngeal cancer; neuroblastoma; Non-Hodgkin lymphoma; nonmelanoma skin cancer; non-small cell lung cancer; oral cancer; oral cavity cancer; oropharyngeal cancer; osteosarcoma; other brain and spinal cord tumors; ovarian cancer; ovarian epithelial cancer; ovarian germ cell tumor; ovarian low malignant potential tumor; pancreatic cancer; papillomatosis; paranasal sinus cancer; parathyroid cancer; pelvic cancer; penile cancer; pharyngeal cancer; pineal parenchymal tumors of intermediate differentiation; pineoblastoma; pituitary tumor; plasma cell neoplasm/multiple myeloma; pleuropulmonary blastoma; primary central nervous system (CNS) lymphoma; primary hepatocellular liver cancer; prostate cancer; rectal cancer; renal cancer; renal cell (kidney) cancer; renal cell cancer; respiratory tract cancer; retinoblastoma; rhabdomyosarcoma; salivary gland cancer; Sezary syndrome; small cell lung cancer; small intestine cancer; soft tissue sarcoma; squamous cell carcinoma; squamous neck cancer; stomach (gastric) cancer; supratentorial primitive neuroectodermal tumors; T-cell lymphoma; testicular cancer; throat cancer; thymic carcinoma; thymoma; thyroid cancer; transitional cell cancer; transitional cell cancer of the renal pelvis and ureter; trophoblastic tumor; ureter cancer; urethral cancer; uterine cancer; uterine sarcoma; vaginal cancer; vulvar cancer; Waldenstrm macroglobulinemia; or Wilm's tumor.
6. The method of claim 1 , wherein the biological sample comprises a cell culture or a biological fluid.
7. The method of claim 6 , wherein the biological fluid comprises a bodily fluid.
8. The method of claim 6 , wherein the biological fluid comprises peripheral blood, sera, plasma, ascites, urine, cerebrospinal fluid (CSF), sputum, saliva, bone marrow, synovial fluid, aqueous humor, amniotic fluid, cerumen, breast milk, broncheoalveolar lavage fluid, semen, prostatic fluid, cowper's fluid or pre-ejaculatory fluid, female ejaculate, sweat, fecal matter, tears, cyst fluid, pleural and peritoneal fluid, pericardial fluid, lymph, chyme, chyle, bile, interstitial fluid, menses, pus, sebum, vomit, vaginal secretions, mucosal secretion, stool water, pancreatic juice, lavage fluids from sinus cavities, bronchopulmonary aspirates, blastocyl cavity fluid, or umbilical cord blood.
9. The method of claim 1 , wherein the biological sample comprises isolated microvesicles.
10. The method of claim 9 , wherein the biological sample is contacted with the plurality of aptamers before isolating microvesicles.
11. The method of claim 10 , wherein the microvesicles are isolated using chromatography, filtration, ultrafiltration, centrifugation, ultracentrifugation, flow cytometry, affinity capture, and/or microfluidics.
12. The method of claim 1 , wherein at least one member of the pool of aptamers binds a polypeptide or fragment thereof.
13. The method of claim 1 , wherein at least one member of the pool of aptamers binds a polypeptide or fragment thereof and the polypeptide or fragment thereof is soluble or membrane bound.
14. The method of claim 1 , wherein at least one member of the pool of aptamers binds a microvesicle surface antigen.
15. The method of claim 1 , wherein the pool of aptamers comprises at least 20 different aptamer members.
16. The method of claim 1 , wherein the pool of aptamers comprises at least 50 different aptamer members.
17. The method of claim 1 , wherein an altered presence or copy number of the pool of aptamers that formed a complex with the microvesicles as compared to a reference sample indicates that the sample is cancerous or predisposed to be cancerous.
18. The method of claim 1 , wherein each of the pool of aptamers binds to a different target.