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US-12622431-B2 - Insect control nanobodies and uses thereof

US12622431B2US 12622431 B2US12622431 B2US 12622431B2US-12622431-B2

Abstract

Insect control nanobodies are provided. Accordingly there is provided a nanobody which specifically binds to an insect polypeptide selected from the group consisting of: a polypeptide comprising a chitin binding domain (CBD), V-ATPase subunit c, trehalase, cytochrome p450 monooxygenase, chitin deacetylase, chitin synthase and NPC1 sterol transporter, wherein binding of the nanobody to the insect polypeptide confers an insect control activity to the nanobody. Also provided are polynucleotides encoding the nanobody, host cells expressing the nanobody and methods of using it.

Inventors

  • Rony OREN-BENAROYA
  • Amir AYALI
  • Jessica MONSERRATE
  • Jeroen Van Rie

Assignees

  • IBI-AG INNOVATIVE BIO INSECTICIDES LTD.

Dates

Publication Date
20260512
Application Date
20201111

Claims (20)

  1. 1 . A VHH antibody which specifically binds to an insect chitin deacetylase comprising complementarity determining regions (CDRs) as set forth in SEQ ID NOs: 351, 347 and 356 arranged in a sequential order from N to C on said nanobody, wherein binding of said nanobody to said insect chitin deacetylase confers an insect control activity to said VHH antibody.
  2. 2 . A composition comprising a VHH antibody which specifically binds to an insect chitin deacetylase comprising complementarity determining regions (CDRs) as set forth in SEQ ID NOs: 351, 347 and 356 arranged in a sequential order from N to C on said nanobody; and a toxin moiety having an insect control activity.
  3. 3 . The composition of claim 2 , wherein binding of said VHH antibody to said insect chitin deacetylase confers an insect control activity to said VHH antibody.
  4. 4 . The VHH antibody of claim 1 , wherein said VHH antibody downregulates activity of said insect chitin deacetylase.
  5. 5 . The VHH antibody of claim 1 , being formulated for delivery by spraying, irrigation and/or fumigation.
  6. 6 . A polynucleotide encoding the VHH antibody of claim 1 .
  7. 7 . A nucleic acid construct comprising the polynucleotide of claim 6 and a cis-acting regulatory element for directing expression of said polynucleotide.
  8. 8 . A transformed host cell comprising the VHH antibody of claim 1 .
  9. 9 . A method of producing an insect control VHH antibody, the method comprising expressing in a host cell the polynucleotide of claim 6 .
  10. 10 . The method of claim 9 , comprising isolating the VHH antibody.
  11. 11 . A method of insect control, the method comprising contacting the insect with the nanobody of claim 1 , a polynucleotide or a nucleic acid construct encoding same or a host cell expressing same.
  12. 12 . The nanobody of claim 1 , wherein said VHH antibody is formulated as a liquid formulation.
  13. 13 . The nanobody of claim 1 , wherein said VHH antibody is formulated as a dry formulation.
  14. 14 . A plant comprising the VHH antibody of claim 1 or a polynucleotide or a nucleic acid construct encoding same.
  15. 15 . The plant of claim 14 , being a transgenic plant.
  16. 16 . A commodity product comprising the VHH antibody of claim 1 .
  17. 17 . The VHH antibody of claim 1 , wherein said insect is selected from the group consisting of moth, stinkbug, hopper, beetle, aphid and honeybee.
  18. 18 . The VHH antibody of claim 1 , wherein said insect is a moth.
  19. 19 . The VHH antibody of claim 1 , being a synthetic recombinant VHH antibody.
  20. 20 . The VHH antibody of claim 1 , wherein said insect chitin deacetylase is a Helicoverpa armigera chitin deacetylase.

Description

RELATED APPLICATIONS This application is a National Phase of PCT Patent Application No. PCT/IL2020/051170 having the International filing date of Nov. 11, 2020, which claims the benefit of priority under 35 USC § 119(e) of U.S. Provisional Patent Application No. 62/933,533 filed on Nov. 11, 2019. The contents of the above applications are all incorporated by reference as if fully set forth herein in their entirety. SEQUENCE LISTING STATEMENT The ASCII file, entitled 92382SequenceListing.txt, created on 2022, 10 May, comprising 387,592 bytes, submitted concurrently with the filing of this application is incorporated herein by reference. The sequence listing submitted herewith is identical to the sequence listing forming part of the international application. FIELD AND BACKGROUND OF THE INVENTION The present invention, in some embodiments thereof, relates to insect control nanobodies and uses thereof. Management of insect pests in the past 70 years has been achieved mainly through application of synthetic pesticides. Since the discovery of the insecticidal properties of DDT by Paul Müller in 1939, hundreds of insecticidal compounds have been developed, accompanied by a relatively steady increase in insecticide use. Most of the currently used chemical insecticides have a high potential for damaging the ecosystem, they are toxic to humans directly or through the food chain and their use is also impeded by development of genetic insect resistance. The most prominent biological solutions currently in use include beneficial organisms or natural enemies that reduce harm caused by insects, pheromones that act as bait or harm reproduction, release of sterile males, spraying with Bacillus thuringiensis toxin or genetically modified crops containing a gene for Bacillus thuringiensis toxin that is lethal to the insects but not to humans. However, these biological solutions also have disadvantage, including low efficiency, danger of violating the natural ecological balance and development genetic insect resistance (e.g. in the case of the use of Bacillus toxin). Nanobodies, also known as VHH antibodies, are single domain antibodies which practically contain the heavy chain of an antibody (HCAb) and completely lack the light chain. They were discovered in the blood of camels by Raymond Hamers who was credited with this discovery in 1989 at Vrije Universiteit Brussel. Nanobodies are the smallest available intact antigen binding fragment (Cortez-Retamozo et al., 2004; Revets et al., 2005) with a size of approximately 15 kDa. The nanobodies have significant advantages including high production yield in a broad variety of expression systems, their minimal size allows high accessibility to their epitopes, high physical-chemical stability, reversible refolding and high solubility in aqueous solutions, highly homogenous showing no signs of spontaneous dimerization and ability to specifically recognize unique epitopes with sub-nanomolar affinities. The use of nanobodies as insecticides has been previously suggested (see e.g. EP Patent Application Publication Nos: EP3415010 and EP2609116; US Patent Publication No: U.S. Pat. No. 9,516,879; US Patent Application Publication No: U.S. Pat. No. 9,803,003B2; and International Patent Application Publication No. WO2014191146). SUMMARY OF THE INVENTION According to an aspect of some embodiments of the present invention there is provided a nanobody which specifically binds to an insect polypeptide selected from the group consisting of: (i) a polypeptide comprising a chitin binding domain (CBD), wherein the nanobody binds the CBD;(ii) V-ATPase subunit c, wherein the nanobody comprises complementarity determining regions (CDRs) as set forth in SEQ ID NOs: 167-169; 171 and 168-169; 174 and 168-169; 167, 178 and 169; or 180-182 arranged in a sequential order from N to C on the nanobody;(iii) trehalase;(iv) cytochrome p450 monooxygenase;(v) chitin deacetylase;(vi) chitin synthase; and(vii) NPC1 sterol transporter,wherein binding of the nanobody to the insect polypeptide confers an insect control activity to the nanobody. According to an aspect of some embodiments of the present invention there is provided a composition comprising a nanobody which specifically binds to an insect polypeptide selected from the group consisting of: (i) a polypeptide comprising a chitin binding domain (CBD), wherein the nanobody binds the CBD;(ii) V-ATPase subunit c, wherein the nanobody comprises complementarity determining regions (CDRs) as set forth in SEQ ID NOs: 167-169; 171 and 168-169; 174 and 168-169; 167, 178 and 169; 180-182; 187-189; or 191-193 arranged in a sequential order from N to C on the nanobody;(iii) trehalase;(iv) cytochrome p450 monooxygenase;(v) chitin deacetylase;(vi) chitin synthase; and(vii) NPC1 sterol transporter, and a toxin moiety having an insect control activity. According to some embodiments of the invention, the binding of the nanobody to the insect polypeptide confers an insect cont