IP Library Granted Patent US 12,428,649
Granted Patent B2
US 12,428,649 · App. 17/677,648 · Granted Sep 30, 2025

Methods and compositions for making and using compatible insecticidal proteins

Inventors: James A. Baum (Webster Groves, MO); Artem G. Evdokimov (Orchard Park, NY); Agoston Jerga (Chesterfield, MO); Farhad Moshiri (Chesterfield, MO)
Assignee: Monsanto Technology LLC
C12N15/8286G01N33/5085G01N2333/43552
View Patent ↗
Loading inventors, assignments & file history…
Monitor This Case
Get email alerts when status or documents change.
Order Certified Copies
Most orders are placed with the USPTO same day — all within 24 business hours.
Order via The Patent Place →
Pre-filled with this patent's details
Quick Facts
Patent No.
US 12,428,649
App. No.
17/677,648
Granted
Sep 30, 2025
Kind
B2
Abstract

Pesticidal proteins (FFPP's) are used to produce derivatives (DP's) that are ineffective and disabled relative to conferring toxic properties upon a target pest, yet the ability of the DP to bind to the receptor to which said FFPP binds is unaffected. Such DP's are useful in inhibiting the FFPP from which it was derived when both are fed to a target pest and for comparing receptor binding capability and efficiency relative to different FFPP's from which the DP has been derived, providing for an assessment of different FFPP's relative to each other, and providing uniformity and certainty in combinations of such FFPP's for compositions, including transgenic plants, that can be used to control pest populations susceptible to both FFPP's, creating more durable transgenic plant products, inhibiting the development of resistance to such FFPP's when used in plants commercially, and in providing a durable and viable resistance management strategy for crops using such FFPP combinations. Polynucleotide sequences intended for use in expression of the DP's and FFPP's are also provided. Particular embodiments provide methods of designing and preparing DP's, as well as compositions and methods of using DP's and the FFPP's from which the DP's have been derived in more effective pesticidal compositions and products.

Claims (13)

1. A method for selecting a first FFPP (fully functional pesticidal polypeptide) and a second FFPP that are compatible for use together in a composition for controlling a target pest, said method comprising:

a) selecting the first FFPP, which is toxic to a target pest and the second FFPP that is toxic to the target pest and is different from said first FFPP, wherein each of the first FFPP and the second FFP have toxic properties when provided individually in a diet of the target pest;

b) producing a first DT (disabled protein) from said first FFPP that, upon ingestion by said target pest, blocks the toxic properties conferred by said first FFPP;

c) producing a plurality of different mixtures containing a fixed but pesticidally effective amount of the second FFPP and increasing amounts of said first DT;

d) providing a dose of each mixture of step c) in the diet of at least three different individuals of said target pest;

wherein observing toxic properties in any individual in step d) is determinative that said first and second toxic agents are compatible for use together to control said target pest.

2. The method of claim 1 , wherein the composition comprising said first FFPP and said second FFPP is effective in controlling an insect pest infestation wherein said insects are selected from the group consisting of Arachnida, Coleoptera, Ctenocephalides, Diptera, Hemiptera, Heteroptera, Homoptera, Hymenoptera, Lepidoptera and Thysanoptera insects.

3. The method of claim 1 , wherein the first FFPP and the second FFPP bind to different receptors in the target pest.

4. The method of claim 1 , wherein preparing the first DP comprises the step of confirming that said first DP when used alone in a bioassay with said target pest has diminished toxicity against the target pest when compared to the toxicity of the first FFPP.

5. The method of claim 1 , wherein the method identifies a combination of two FFPPs that has a decreased likelihood of development of resistance by said target pest against any one of the FFPPs relative to a combination that has not been selected by the method.

6. The method of claim 1 , wherein the method identifies a combination of two FFPPs that has a delayed onset of resistance by said target pest against any one of the FFPPs relative to a combination that has not been selected by the method.

7. The method of claim 1 , wherein the first DT does not itself confer toxic properties.

8. The method of claim 1 , wherein the plurality of different mixtures comprises a plurality of molar ratios in which the first DP is present in a greater concentration than said second FFPP.

Continuity (3)
Division 16296886 · Mar 8, 2019
Provisional Application 62640927 · Mar 9, 2018
Related Publication 20220243221A1 · Aug 4, 2022
References Cited (38)
US 5500365A · Fischhoff et al. · 1996 [cited by applicant]
US 5866784A · Van Mellaert et al. · 1999 [cited by applicant]
US 8344207B2 · Bogdanova et al. · 2013 [cited by applicant]
US 9121035B2 · Baum et al. · 2015 [cited by applicant]
US 9322033B2 · Baum et al. · 2016 [cited by applicant]
US 10188115B2 · Baum et al. · 2019 [cited by applicant]
US 10897910B2 · Baum et al. · 2021 [cited by applicant]
US 20100180351A1 · Gossele et al. · 2010 [cited by applicant]
US 20110318272A1 · Street · 2011 [cited by examiner]
US 20130269060A1 · Baum et al. · 2013 [cited by applicant]
US 20150047076A1 · Anderson · 2015 [cited by examiner]
US 20150274786A1 · Bowen · 2015 [cited by examiner]
Badran et al., “Continuous evolution of Bacillus thuringiensis toxins overcomes insect resistance,” Nature 533 (7601):58-63, 2016. [cited by applicant]
Bates et al., “Insect resistance management in GM crops: past, present and future,” Nat. Biotechnol. 1:57-62, 2005. [cited by applicant]
Carrière et al., “Optimizing pyramided transgenic Bt crops for sustainable pest management,” Nat. Biotechnol. 33:161-168, 2015. [cited by applicant]
De et al., “Crystal structure of the Vibrio cholerae cytolysin heptamer reveals common features among disparate pore-forming toxins,” Proc. Natl. Acad. Sci. 108:7385-7390, 2011. [cited by applicant]
De Maagd et al, “Structure, diversity, and evolution of protein toxins from spore-forming entomopathogenic bacteria,” Annu. Rev. Genet. 37:409-433, 2003. [cited by applicant]
Deitloff et al., “Effects of refuges on the evolution of resistance to transgenic corn by western corn rootworm, [cited by applicant]
Devos et al. “Resistance evolution to plant-produced Bt-toxins of the first generation of genetically engineered Diabrotica-active Bt-maize events by western corn rootworm: management and monitoring considerations,” ISB… [cited by applicant]
Estela et al., “Interaction of Bacillus thuringiensis toxins with larval midgut binding sites of Helicoverpa armigera (Lepidoptera: Noctuidae),” Appl. Environ. Microbiol. 70(3):1378-1384, 2004. [cited by applicant]
Girard et al., “Cysteine scanning mutagenesis of 4, a putative pore-lining helix of the Bacillus thuringiensis insecticidal toxin Cry1Aa,” Applied and Environmental Microbiology 74(9):2565-2572, 2008. [cited by applicant]
Girard et al., “Helix α4 of the Bacillus thuringiensis Cry1aa toxin plays a critical role in the postbinding steps of pore formation,” Applied and Environmental Microbiology 75(2):359-365, 2009. [cited by applicant]
González-Cabrera et al., “Binding of Bacillus thuringiensis toxins in resistant and susceptible strains of pink bollworm ( [cited by applicant]
Gowda et al., “A transgenic approach for controlling Lygus in cotton,” Nature Communications 12213, 2016. [cited by applicant]
Granero et al., “Bacillus thuringiensis crystal proteins Cry1Ab and Cry1Fa share a high affinity binding site in [cited by applicant]
Jimenez-Juarez et al., “Bacillus thuringiensis Cry1Ab mutants affecting oligomer formation are non-toxic to Manduca sexta larvae,” J Biol. Chem. 282(29): 21222-9, 2007. [cited by applicant]
Jurat-Fuentes et al., “Specificity determinants for Cry insecticidal proteins: Insights from their mode of action,” J. Invertebr. Pathol. 142:5-10, 2017. [cited by applicant]
Melo et al., “Bacillus thuringiensis: mechanism of action, resistance, and new applications: a review,” Crit Rev Biotechnol. 36(2):317-26, 2014. [cited by applicant]
Pardo-Lopez et al., “Bacillus thuringiensis insecticidal three-domain Cry toxins: mode of action, insect resistance and consequences for crop protection,” FEMS Microbiology Reviews 37(1): 3-22, 2013. [cited by applicant]
Rodríguez-Almazán et al., “Dominant negative mutants of Bacillus thuringiensis Cry1Ab toxin function as anti-toxins: demonstration of the role of oligomerization in toxicity,” PloS One 4(5):e5545, 2009. [cited by applicant]
Siebert et al., Evaluation of corn hybrids expressing Cry1F, Cry1A.105, Cry2Ab2, Cry34Ab1/Cry35Ab1, and Cry3Bb1 against southern United States insect pests, Journal of Economic Entomology 105(5):1825-1834, 2012. [cited by applicant]
Schwartz et al., “Restriction of intramolecular movements within the Cry1Aa toxin molecule of Bacillus thuringiensis through disulfide bond engineering,” FEBS Letters 410:397-402, 1997. [cited by applicant]
Tabashnik, “Pest adaptation,” Nature 389: 778, 1997. [cited by applicant]
Tabashnik et al., “Suppressing resistance to Bt cotton with sterile insect releases,” Nat. Biotechnol. 28(12):1304-7, 2010. [cited by applicant]
Tabashnik et al.,“Insect resistance to Bt crops: lessons from the first billion acres,” Nat. Biotechnol. 31(6):510-21, 2013. [cited by applicant]
Tanaka et al.,“2-Methyl-2,4-pentanediol induces spontaneous assembly of [cited by applicant]
Vachon et al., “Helix 4 mutants of the Bacillus thuringiensis insecticidal toxin Cry1Aa display altered pore-forming abilities,” Applied and Environmental Microbiology 70(10):6123-6130, 2004. [cited by applicant]
Zhao et al., “Transgenic plants expressing two Bacillus thuringiensis toxins delay insect resistance evolution,” Nat. Biotechnol. 21:1493-1497, 2003. [cited by applicant]