US2021380649A1PendingUtilityA1

Wnt/sfrp complexes, wnt-containing compositions, wnt-expressing cells, and methods of making, purifying, and using same

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Assignee: BIO TECHNE CORPPriority: Oct 9, 2017Filed: Jun 18, 2021Published: Dec 9, 2021
Est. expiryOct 9, 2037(~11.2 yrs left)· nominal 20-yr term from priority
Inventors:Anthony PersonLiwen XiongMing BiCamrin Tracy
C07K 1/14C07K 14/475C07K 19/00C07K 14/71C12N 5/0686C07K 14/47C12Q 1/6897
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Claims

Abstract

This disclosure describes isolated protein complexes including a Wnt and a sFRP; compositions including a Wnt; a cell overexpressing a Wnt and a sFRP; compositions including a cell overexpressing a Wnt and a cell overexpressing a sFRP; methods of making the protein complexes, compositions, and cells; and methods of using the isolated protein complexes, compositions, and cells. This disclosure further describes methods of forming a complex including a Wnt and a sFRP and methods for isolating a Wnt. Also described herein are methods that may be used to purify a Wnt without the use of a detergent.

Claims

exact text as granted — not AI-modified
What is claimed is: 
     
         1 . An isolated protein complex comprising a Wnt and a sFRP. 
     
     
         2 . The isolated protein complex of  claim 1 , wherein the Wnt comprises one or more of Wnt1, Wnt2, Wnt2b, Wnt3, Wnt3a, Wnt4, Wnt5a, Wnt5b, Wnt6, Wnt7a, Wnt7b, Wnt8a, Wnt8b, Wnt9a, Wnt9b, Wnt10a, Wnt10b, Wnt11, and Wnt16. 
     
     
         3 . The isolated protein complex of  claim 1 , wherein the sFRP comprises one or more of sFRP1, sFRP2, sFRP3, sFRP4, and sFRP5. 
     
     
         4 . The isolated protein complex of  claim 1 , wherein the Wnt comprises an active Wnt. 
     
     
         5 . The isolated protein complex of  claim 4 , wherein the active Wnt comprises Wnt having Wnt reporter activity as measured using a secreted alkaline phosphatase (SEAP) reporter assay. 
     
     
         6 . The isolated protein complex of  claim 1 , wherein the protein complex is substantially free of a detergent. 
     
     
         7 . The isolated protein complex of  claim 1 , wherein the Wnt comprises a mouse Wnt and the sFRP comprises a mouse sFRP. 
     
     
         8 . The isolated protein complex of  claim 1 , wherein the Wnt comprises a mouse Wnt1 and the sFRP comprises a mouse sFRP1. 
     
     
         9 . The isolated protein complex of  claim 1 , wherein the protein complex exhibits an effective dose of 50 percent (ED 50 ) of less than 500 ng/mL, as measured using a HEK293 TCF9-SEAP hFz4/hLRP5 Wnt Reporter assay. 
     
     
         10 . A composition comprising a Wnt, wherein the composition is substantially free of a detergent. 
     
     
         11 . The composition of  claim 10 , wherein the Wnt comprises one or more of Wnt1, Wnt2, Wnt2b, Wnt3, Wnt3a, Wnt4, Wnt5a, Wnt5b, Wnt6, Wnt7a, Wnt7b, Wnt8a, Wnt8b, Wnt9a, Wnt9b, Wnt10a, Wnt10b, Wnt11, and Wnt16. 
     
     
         12 . The composition of  claim 10 , wherein the Wnt comprises active Wnt. 
     
     
         13 . The composition of  claim 12 , wherein the active Wnt comprises Wnt having Wnt reporter activity as measured using a secreted alkaline phosphatase (SEAP) reporter assay. 
     
     
         14 . The composition of  claim 10 , wherein the composition further comprises a sFRP. 
     
     
         15 . The composition of  claim 14 , wherein the sFRP comprises one or more of sFRP1, sFRP2, sFRP3, sFRP4, and sFRP5. 
     
     
         16 . The composition of  claim 14 , wherein the Wnt comprises a mouse Wnt and the sFRP comprises a mouse sFRP. 
     
     
         17 . The composition of  claim 14 , wherein the Wnt comprises a mouse Wnt1 and the sFRP comprises a mouse sFRP1. 
     
     
         18 . The composition of  claim 10 , wherein the Wnt exhibits an effective dose of 50 percent (ED 50 ) of less than 100 ng/mL, as measured using a HEK293 TCF9-SEAP hFz4/hLRP5 Wnt Reporter assay. 
     
     
         19 . The composition of  claim 10 , the composition further comprising one or more of R-Spondin 1, R-Spondin 2, R-Spondin 3, R-Spondin 4, Lipocalin7, or WIF1. 
     
     
         20 . A method comprising:
 overexpressing a Wnt1 and an sFRP1;   forming a complex comprising the Wnt1 and the sFRP1; and   isolating the Wnt1, wherein isolating the Wnt1 comprises an aqueous purification procedure and wherein isolating the Wnt does not comprise using a detergent.

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