Methods of determining susceptibility of olfactory receptor ligands to adaptation

Abstract

The present disclosure generally provides methods for identifying olfactory receptor ligands that are more or less likely to be susceptible to olfactory adaptation, and the assays related to identifying such ligands. In some embodiments, the ligand is an olfactory receptor agonist, and, in other embodiments, the ligand is an olfactory receptor antagonist. In some embodiments, the methods comprise identifying which agonists are more susceptible or less susceptible to olfactory adaptation based on differences in their signaling activity before and after a prolonged exposure of the olfactory receptors to a composition comprising the olfactory receptor ligand.

Description

[0001] METHODS OF DETERMINING SUSCEPTIBILITY OF OLFACTORY RECEPTOR LIGANDS TO ADAPTATION

[0002] TECHNICAL FIELD

[0003] The present disclosure generally provides methods for identifying olfactory receptor ligands that are more or less likely to be susceptible to olfactory adaptation, and the assays related to identifying such ligands. In some embodiments, the ligand is an olfactory receptor agonist, and, in other embodiments, the ligand is an olfactory receptor antagonist. In some embodiments, the methods comprise identifying which agonists are more susceptible or less susceptible to olfactory adaptation based on differences in their signaling activity before and after a prolonged exposure of the olfactory receptors to a composition comprising the olfactory receptor ligand.

[0004] DESCRIPTION OF RELATED ART

[0005] People commonly experience the physiological phenomenon of olfactory adaptation. Certain fragrances or odors seem to linger, and one continues to smell them as long as these fragrances or odors are present. But other fragrances or odors seem not to linger, even though they remain present. Thus, humans have a tendency to adapt to certain fragrances and odors but not to others. Adaptation thus results in a decreased perceived intensity of volatile and mixtures thereof over prolonged exposure.

[0006] There has been no singular explanation for such adaptation, or the lack of it. Some have suggested that it is of psychological nature. After all, some people tend to experience olfactory adaptation in situations where others do not. But it has also been suggested that there may be a biochemical explanation as well. A variety of different biochemical mechanisms for adaptation have been proposed.

[0007] There may be many benefits to identifying a biochemical explanation, as it would permit the rapid and efficient identification of compounds that may be susceptible to adaptation and therefore as less useful as fragrances in situations where a long-term olfactory stimulation is desired or more useful in situations where a long-term olfactory stimulation is not desired. Additionally, it would also permit the rapid and efficient identification of compounds that may be less susceptible to adaptation and therefore more useful as fragrances in situations where a long-term olfactory stimulation is desired or less useful as fragrances in situations where a longterm olfactory stimulation is not desired. Otherwise, one is left to test for olfactory adaptation through human sensory testing, which can be time-intensive, expensive, and inefficient.

[0008] Therefore, there is a continuing need to understand if there is a biochemical explanation for adaptation and, if so, if there may be ways to apply such understandings to identify compounds that may be more susceptible or less susceptible to olfactory adaptation in a manner that is quick and efficient. SUMMARY

[0009] The present disclosure relates to the discovery that measured differences between the intensity of the signaling activity of an olfactory receptor before and after a prolonged exposure to an olfactory receptor ligand, such as an agonist or an antagonist, correlate reasonably well with the observation of olfactory adaptation in human sensory testing.

[0010] In a first aspect, the disclosure provides a method of identifying a substance that is susceptible to olfactory adaptation, the method comprising: (a) introducing an olfactory receptor ligand to a plurality of olfactory receptors, wherein the olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand; (c) reintroducing the olfactory receptor ligand to the plurality of olfactory receptors, such that the olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes; (d) following an optional period during which the olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the olfactory receptor ligand to the plurality of olfactory receptors; and (e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor ligand. In some embodiments, the method further comprises: (f) identifying an adaptation-susceptible olfactory receptor agonist that is susceptible to olfactory adaptation based on a difference between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) deselecting the adaptation-susceptible olfactory receptor agonist as an adaptation-resistant substance.

[0011] In a second aspect, the disclosure provides a method of identifying a substance that is susceptible to olfactory cross-adaptation, the method comprising: (a) introducing a first olfactory receptor ligand to a plurality of olfactory receptors, wherein the first olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand; (c) introducing a second olfactory receptor ligand, which is different from the first olfactory receptor ligand, wherein the second olfactory receptor ligand is an antagonist of the plurality of olfactory receptors, such that the second olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes; (d) following an optional period during which the second olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the first olfactory receptor ligand to the plurality of olfactory receptors; and (e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor agonist. In some embodiments, the method further comprises: (f) identifying an adaptation-susceptible olfactory receptor antagonist that is susceptible to olfactory adaptation based on a difference between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) deselecting the adaptation-susceptible olfactory receptor antagonist as an adaptation-resistant substance.

[0012] In a third aspect, the disclosure provides a method of identifying a substance that is susceptible to olfactory cross-adaptation, the method comprising: (a) introducing a first olfactory receptor ligand to a plurality of olfactory receptors, wherein the first olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand; (c) introducing a second olfactory receptor ligand, which is different from the first olfactory receptor ligand, and wherein the second olfactory receptor ligand is an agonist of the plurality of olfactory receptors, such that the second olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes; (d) following an optional period during which the second olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the first olfactory receptor ligand to the plurality of olfactory receptors; and (e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor agonist. In some embodiments, the method further comprises: (f) identifying an adaptation-susceptible olfactory receptor agonist that is susceptible to olfactory adaptation based on a difference between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) deselecting the adaptation-susceptible olfactory receptor agonist as an adaptation-resistant substance.

[0013] In a fourth aspect, the disclosure provides a method of identifying a substance that is resistant to olfactory adaptation, the method comprising: (a) introducing an olfactory receptor ligand to a plurality of olfactory receptors, wherein the olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand; (c) reintroducing the olfactory receptor ligand to the plurality of olfactory receptors, such that the olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes; (d) following an optional period during which the olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the olfactory receptor ligand to the plurality of olfactory receptors; and (e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor ligand. In some embodiments, the method further comprises: (f) identifying an adaptation-resistant olfactory receptor agonist that is resistant to olfactory adaptation based on a difference between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) selecting the adaptation-resistant olfactory receptor agonist as an adaptation-resistant substance.

[0014] In a fifth aspect, the disclosure provides a method of identifying a substance that is resistant to olfactory adaptation, the method comprising: (a) introducing a first olfactory receptor ligand to a plurality of olfactory receptors, wherein the first olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the first olfactory receptor ligand; (c) introducing a second olfactory receptor ligand, wherein the second olfactory receptor ligand is different from the first olfactory receptor ligand and is an antagonist of the plurality of olfactory receptors, such that the olfactory receptor antagonist remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes; (d) following an optional period during which the second olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the first olfactory receptor ligand to the plurality of olfactory receptors; and (e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the first olfactory receptor ligand. In some embodiments, the method further comprises: (f) identifying an adaptation-resistant olfactory receptor antagonist that is resistant to olfactory adaptation based on a difference (or a lack of a difference) between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) selecting the adaptation-resistant olfactory receptor antagonist as an adaptation-resistant substance.

[0015] In a sixth aspect, the disclosure provides a method of identifying a substance that is resistant to olfactory adaptation, the method comprising: (a) introducing a first olfactory receptor ligand to a plurality of olfactory receptors, wherein the first olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the first olfactory receptor ligand; (c) introducing a second olfactory receptor ligand, wherein the second olfactory receptor ligand is different from the first olfactory receptor ligand and is an agonist of the plurality of olfactory receptors, such that the second olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes; (d) following an optional period during which the second olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the first olfactory receptor ligand to the plurality of olfactory receptors; and (e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the first olfactory receptor ligand. In some embodiments, the method further comprises: (f) identifying an adaptation-resistant olfactory receptor agonist that is resistant to olfactory adaptation based on a difference (or a lack of a difference) between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) selecting the adaptation-resistant olfactory receptor agonist as an adaptation-resistant substance.

[0016] In a seventh aspect, the disclosure provides a method of identifying a substance that is resistant to olfactory adaptation, the method comprising: (a) introducing an olfactory receptor ligand to a plurality of olfactory receptors, wherein the olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand; (c) reintroducing the olfactory receptor ligand to the plurality of olfactory receptors, such that the olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time immediately following the subsequent reintroduction of the olfactory receptor ligand; (d) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor ligand. In some embodiments, the method further comprises: (f) identifying an adaptation-resistant olfactory receptor agonist that is resistant to olfactory adaptation based on a difference between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) selecting the adaptation-resistant olfactory receptor agonist as an adaptation-resistant substance.

[0017] In an eighth aspect, the disclosure provides a method of identifying a substance that is susceptible to olfactory adaptation, the method comprising: (a) introducing an olfactory receptor ligand to a plurality of olfactory receptors, wherein the olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand; (c) reintroducing the olfactory receptor ligand to the plurality of olfactory receptors, such that the olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time immediately following the subsequent reintroduction of the olfactory receptor ligand; (d) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor ligand. In some embodiments, the method further comprises: (f) identifying an adaptation-susceptible olfactory receptor agonist that is resistant to olfactory adaptation based on a difference between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) deselecting the adaptation-susceptible olfactory receptor agonist as an adaptation-resistant substance.

[0018] In a ninth aspect, the disclosure provides the use of an olfactory-resistant substance identified or selected by the method of the previous aspects to provide a long-lasting fragrance to a fragranced article. In some embodiments, the fragranced article is a fiber, such as a natural or synthetic fiber. In some such embodiments, the fiber is comprised by a textile article, such as an article of clothing, a towel, bedding, upholstery, and the like. In some embodiments, the fragranced article is a consumer care product, such as a household cleaning product, an air freshener, a commercial cleaning product, a dish detergent, a laundry detergent, a fabric softener, a scent booster, or a personal care product, such as a shower gel, a shampoo, a hair conditioner, a hair-styling product, a skin care product, a cosmetic product, a deodorant, an antiperspirant, or a self-tanning product. In some embodiments, the fragranced article is a perfuming composition, such as a fine fragrance composition or a fragrance composition for use in various air-care products, such as plug-in diffusers, candles, perfuming reeds, room sprays, and the like.

[0019] Further aspects, and embodiments thereof, are set forth below in the Detailed Description, the Drawings, the Abstract, and the Claims.

[0020] BRIEF DESCRIPTION OF THE DRAWINGS

[0021] The drawings set forth in the attached sheets are provided for purposes of illustrating various embodiments of the compositions and methods disclosed herein. The drawings are provided for illustrative purposes only and are not intended to describe any preferred compositions or preferred methods, or to serve as a source of any limitations on the scope of the claimed inventions.

[0022] FIG. 1 shows a fluorescence-time trace of calcium levels in a cell transfected with the human odorant receptor OR1A1 and Ga15, demonstrating how odor responses after a long exposure to saline are very similar to the responses before the saline soak, indicating that no adaptation has occurred.

[0023] FIG. 2 shows the calculated adaptation index, where “1” is complete adaptation and “0” is no adaptation, for the population of saline-treated cells as in FIG. 1.

[0024] FIG. 3 shows a fluorescence-time trace of calcium levels in a cell transfected with the human odorant receptor OR1A1 and Ga15, demonstrating how odor responses are decreased and only gradually recover after a long exposure to the same odorant, indicating that selfadaptation has occurred.

[0025] FIG. 4 shows the calculated adaptation index for the population of odor-treated cells as in FIG. 3, for one time point. Each tested time point after the long exposure will have its own adaptation index score. FIG. 5 shows a fluorescence-time trace of cAMP levels in a cell transfected with the human odorant receptor OR8D1 and Golf, demonstrating how odor responses after a long exposure to saline are very similar to responses to comparable odor injections before the saline soak, indicating that no adaptation has occurred.

[0026] FIG. 6 shows a fluorescence-time trace of cAMP levels in a cell transfected with the human odorant receptor OR8D1 and Golf, demonstrating how odor responses are decreased after a long exposure to the same odorant, indicating that self-adaptation has occurred.

[0027] FIG. 7 shows a dose-response curve generated by applying pulses of two different odorants to cells expressing 0R1A1 and Ga15 and monitored through fluorescence-based calcium-imaging, with the arrows indicating that concentrations can be found for each odorant that elicit nearly equal activity levels.

[0028] FIG. 8 shows that, on cells expressing OR1A1 and Ga15 and monitored through fluorescence-based calcium-imaging, the adaptation index is not equal for a prolonged exposure of the odorants at the nearly equi-active concentrations determined in FIG. 7.

[0029] FIG. 9 shows the adaptation indices at four time points each for cells expressing various human ORs and Golf and monitored through fluorescence-based cAMP-imaging. Cells were given prolonged exposure of the same odorant that they were initially tested with in pulses, making these examples of self-adaptation.

[0030] FIG. 10 shows the adaptation indices at four time points each for cells expressing various human ORs and Golf and monitored through fluorescence-based cAMP-imaging. Cells were given prolonged exposure of a different activating odorant than they were initially tested with in pulses, making these examples of cross-adaptation.

[0031] FIG. 11 shows a fluorescence-time trace of cAMP levels in a cell transfected with the human odorant receptor OR8D1 and Golf, demonstrating how odor responses are decreased after a long exposure to a non-activating antagonist, making this an unexpected case of crossadaptation.

[0032] FIG. 12 shows the adaptation indices at four time points each for cells expressing various human ORs and Golf and monitored through fluorescence-based cAMP-imaging. Cells were given prolonged exposure of a different antagonist odorants than the activating odorants they were initially tested with in pulses, making these examples of unexpected crossadaptation.

[0033] FIG. 13 shows a concordance between human sensory self-adaptation and the selfadaptation indices for cells expressing the human odorant receptor OR10K1 and Golf and monitored through fluorescence-based cAMP-imaging. In both systems, odor C adapted more than odor D.

[0034] FIG. 14 shows a concordance between human sensory self-adaptation and the selfadaptation indices for cells expressing the human odorant receptor OR5K1 and Golf and monitored through fluorescence-based cAMP-imaging. In both systems, the accord (odorant mixture) A adapted more than accord B.

[0035] FIG. 15 shows that in cells expressing OR1A1 and Ga15 and monitored through fluorescence-based calcium-imaging, that increasing the time of the prolonged exposure to odor increases the extent of adaptation; in this system, it plateaus after ~10min.

[0036] FIG. 16 shows that in cells expressing 0R11A1 and Golf and monitored through fluorescence-based cAMP-imaging, the increasing the concentration of the prolonged exposure to odor increases the extent of adaptation.

[0037] FIG. 17 shows that in cells expressing 0R1A1 and Ga15 and monitored through fluorescence-based calcium-imaging, the time of the prolonged exposure and concentration of that odor exposure can be varied to elicit similar levels of adaptation.

[0038] DETAILED DESCRIPTION

[0039] The following Detailed Description sets forth various aspects and embodiments provided herein. The description is to be read from the perspective of the person of ordinary skill in the relevant art. Therefore, information that is well known to such ordinarily skilled artisans is not necessarily included.

[0040] Definitions

[0041] The following terms and phrases have the meanings indicated below, unless otherwise provided herein. This disclosure may employ other terms and phrases not expressly defined herein. Such other terms and phrases have the meanings that they would possess within the context of this disclosure to those of ordinary skill in the art. In some instances, a term or phrase may be defined in the singular or plural. In such instances, it is understood that any term in the singular may include its plural counterpart and vice versa, unless expressly indicated to the contrary.

[0042] As used herein, the term “polypeptide” refers to a molecule composed of amino acid monomers linearly linked by amide bonds (also known as peptide bonds). The term “polypeptide” refers to any chain of two or more amino acids and does not refer to a specific length of the chain. Thus, peptides, dipeptides, tripeptides, oligopeptides, “protein,” “amino acid chain,” or any other term used herein to refer to a chain of two or more amino acids, are included within the definition of “polypeptide,” and the term “polypeptide” may be used instead of, or interchangeably with, any of these terms.

[0043] As used herein, the term “polynucleotide” refers to a compound that includes a polymer of nucleotides. Each nucleotide is composed of a base, specifically a purine or pyrimidine base (namely, cytosine (C), guanine (G), adenine (A), thymine (T) or uracil (U)), a sugar (namely, deoxyribose or ribose), and a phosphate group. In general, the polynucleotide is described by a sequence of bases, whereby the bases represent the primary structure (linear structure) of the polynucleotide. The sequence of bases is typically represented from 5’ to 3’. Herein, the term polynucleotide encompasses deoxyribonucleic acid (DNA), including, for example, complementary DNA (cDNA) and genomic DNA, ribonucleic acid (RNA), in particular, messenger RNA (mRNA), synthetic forms of DNA or RNA, and mixed polymers including two or more of these molecules. The polynucleotide may be linear or circular. In addition, the term polynucleotide includes both sense and antisense strands, as well as single stranded and double stranded forms. Moreover, the polynucleotides described herein can contain naturally occurring or non-naturally occurring nucleotides.

[0044] As used herein, the term “functional fragment” refers to a portion of a polypeptide sequence to which the ligand binds. Polypeptide sequences often contain certain amino acids that do not actively participate in binding, but which may serve other purposes. In some instances, these non-functioning parts of the polypeptide sequence can be removed or partially replaced, while leaving the functional portion of the sequence intact. These modified proteins are said to comprise a functional fragment of the original polypeptide sequence.

[0045] As used herein, the term “adaptation” refers to a reduction in perceived intensity to a compound or a substance after exposure to a compound or a substance for prolonged periods of time such as minutes, hours or days.

[0046] As used herein, the term “ligand” refers to a compound or substance that binds orthosterically or allosterically as an agonist or an antagonist to an olfactory receptor, when the olfactory receptor expressed in a eukaryotic cell in a cell-based assay.

[0047] As used herein, the term “olfactory receptor agonist” refers to a compound or substance that binds as an agonist to an olfactory receptor, when the olfactory receptor expressed in a eukaryotic cell in a cell-based assay, with an EC50 of no more than 600 pM.

[0048] As used herein, the term “olfactory receptor antagonist” refers to a compound or substance that binds as an antagonist to an olfactory receptor, when the olfactory receptor expressed in a eukaryotic cell in a cell-based assay, with an IC50 of no more than 600 pM.

[0049] As used herein, the term “allele” refers to one of several alternative forms of a gene or polynucleotide sequence at a specific chromosomal location (locus).

[0050] As used herein, the term “human OR1A1 olfactory receptor” refers to the polypeptide sequence that in humans is encoded by the OR1A1 alleles, whose sequence is well known in the art.

[0051] As used herein, the term “human OR8D1 olfactory receptor” refers to the polypeptide sequence that in humans is encoded by the OR8D1 alleles, whose sequence is well known in the art.

[0052] As used herein, the term “human OR51 I2 olfactory receptor” refers to the polypeptide sequence that in humans is encoded by the OR51I2 alleles, whose sequence is well known in the art. As used herein, the term “human 0R51V1 olfactory receptor” refers to the polypeptide sequence that in humans is encoded by the 0R51V1 alleles, whose sequence is well known in the art.

[0053] As used herein, the term “human OR10J5 olfactory receptor” refers to the polypeptide sequence that in humans is encoded by the OR10J5 alleles, whose sequence is well known in the art.

[0054] As used herein, the term “human OR10G4 olfactory receptor” refers to the polypeptide sequence that in humans is encoded by the OR10G4 alleles, whose sequence is well known in the art.

[0055] As used herein, the term “human OR5K1 olfactory receptor” refers to the polypeptide sequence that in humans is encoded by the OR5K1 alleles, whose sequence is well known in the art.

[0056] As used herein, the term “human OR10K1 olfactory receptor” refers to the polypeptide sequence that in humans is encoded by the OR10K1 alleles, whose sequence is well known in the art.

[0057] As used herein, the term “human 0R11A1 olfactory receptor” refers to the polypeptide sequence that in humans is encoded by the 0R11A1 alleles, whose sequence is well known in the art.

[0058] As used herein, the singular forms “a,” “an,” and “the” include plural referents unless the context clearly dictates otherwise. For example, reference to “a substituent” encompasses a single substituent as well as two or more substituents, and the like.

[0059] As used herein, “for example,” “for instance,” “such as,” or “including” are meant to introduce examples that further clarify more general subject matter. Unless otherwise expressly indicated, such examples are provided only as an aid for understanding embodiments illustrated in the present disclosure and are not meant to be limiting in any fashion. Nor do these phrases indicate any kind of preference for the disclosed embodiment.

[0060] As used herein, “comprise” or “comprises” or “comprising” or “comprised of” refer to groups that are open, meaning that the group can include additional members in addition to those expressly recited. For example, the phrase, “comprises A” means that A must be present, but that other members can be present too. The terms “include,” “have,” and “composed of” and their grammatical variants have the same meaning. In contrast, “consist of” or “consists of” or “consisting of” refer to groups that are closed. For example, the phrase “consists of A” means that A and only A is present.

[0061] As used herein, “optionally” means that the subsequently described event(s) may or may not occur. In some embodiments, the optional event does not occur. In some other embodiments, the optional event does occur one or more times. As used herein, “or” is to be given its broadest reasonable interpretation and is not to be limited to an either / or construction. Thus, the phrase “comprising A or B” means that A can be present and not B, or that B is present and not A, or that A and B are both present. Further, if A, for example, defines a class that can have multiple members, e.g., Ai and A2, then one or more members of the class can be present concurrently.

[0062] Other terms are defined in other portions of this description, even though not included in this subsection.

[0063] As used herein, “indole” refers to 1H-indole.

[0064] As used herein, “isobutylquinoline” refers to 2-isobutylquinoline., ,

[0065] As used herein, “lyral” refers to (+-)-3-(4-hydroxy-4-methylpentyl)-3-cyclohexene-1- carbaldehyde + (+-)-4-(4-hydroxy-4-methylpentyl)-3-cyclohexene-1-carbaldehyde.

[0066] As used herein, “beta demascenone” refers to (2E)-1-(2,6,6-trimethyl-1 ,3- cyclohexadien-1-yl)-2-buten-1-one.

[0067] As used herein, “nerol” refers to (Z)-3,7-dimethyl-2,6-octadien-1-ol.

[0068] As used herein, “delta-decalactone” refers to (+-)-6-pentyltetrahydro-2H-pyran-2-one.

[0069] Screening Methods

[0070] In certain aspects, the disclosure provides a method of identifying a substance that is susceptible to olfactory adaptation, the method comprising: (a) introducing an olfactory receptor ligand to a plurality of olfactory receptors, wherein the olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand; (c) reintroducing the olfactory receptor ligand to the plurality of olfactory receptors, such that the olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes; (d) following an optional period during which the olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the olfactory receptor ligand to the plurality of olfactory receptors; and (e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor ligand. In some embodiments, the method further comprises: (f) identifying an adaptation-susceptible olfactory receptor agonist that is susceptible to olfactory adaptation based on a difference between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) deselecting the adaptation-susceptible olfactory receptor agonist as an adaptation-resistant substance.

[0071] In certain aspects, the disclosure provides a method of identifying a substance that is susceptible to olfactory cross-adaptation, the method comprising: (a) introducing a first olfactory receptor ligand to a plurality of olfactory receptors, wherein the first olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand; (c) introducing a second olfactory receptor ligand, which is different from the first olfactory receptor ligand, wherein the second olfactory receptor ligand is an antagonist of the plurality of olfactory receptors, such that the second olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes; (d) following an optional period during which the second olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the first olfactory receptor ligand to the plurality of olfactory receptors; and (e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor agonist. In some embodiments, the method further comprises: (f) identifying an adaptation-susceptible olfactory receptor antagonist that is susceptible to olfactory adaptation based on a difference between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) deselecting the adaptation-susceptible olfactory receptor antagonist as an adaptation-resistant substance.

[0072] In certain aspects, the disclosure provides a method of identifying a substance that is susceptible to olfactory cross-adaptation, the method comprising: (a) introducing a first olfactory receptor ligand to a plurality of olfactory receptors, wherein the first olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand; (c) introducing a second olfactory receptor ligand, which is different from the first olfactory receptor ligand, and wherein the second olfactory receptor ligand is an agonist of the plurality of olfactory receptors, such that the second olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes; (d) following an optional period during which the second olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the first olfactory receptor ligand to the plurality of olfactory receptors; and (e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor agonist. In some embodiments, the method further comprises: (f) identifying an adaptation-susceptible olfactory receptor agonist that is susceptible to olfactory adaptation based on a difference between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) deselecting the adaptation-susceptible olfactory receptor agonist as an adaptation-resistant substance.

[0073] In certain aspects, the disclosure provides a method of identifying a substance that is resistant to olfactory adaptation, the method comprising: (a) introducing an olfactory receptor ligand to a plurality of olfactory receptors, wherein the olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand; (c) reintroducing the olfactory receptor ligand to the plurality of olfactory receptors, such that the olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes; (d) following an optional period during which the olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the olfactory receptor ligand to the plurality of olfactory receptors; and (e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor ligand. In some embodiments, the method further comprises: (f) identifying an adaptation-resistant olfactory receptor agonist that is resistant to olfactory adaptation based on a difference between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) selecting the adaptation-resistant olfactory receptor agonist as an adaptation-resistant substance.

[0074] In certain aspects, the disclosure provides a method of identifying a substance that is resistant to olfactory adaptation, the method comprising: (a) introducing a first olfactory receptor ligand to a plurality of olfactory receptors, wherein the first olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the first olfactory receptor ligand; (c) introducing a second olfactory receptor ligand, wherein the second olfactory receptor ligand is different from the first olfactory receptor ligand and is an antagonist of the plurality of olfactory receptors, such that the olfactory receptor antagonist remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes; (d) following an optional period during which the second olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the first olfactory receptor ligand to the plurality of olfactory receptors; and (e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the first olfactory receptor ligand. In some embodiments, the method further comprises: (f) identifying an adaptation-resistant olfactory receptor antagonist that is resistant to olfactory adaptation based on a difference (or a lack of a difference) between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) selecting the adaptation-resistant olfactory receptor antagonist as an adaptation-resistant substance.

[0075] In certain aspects, the disclosure provides a method of identifying a substance that is resistant to olfactory adaptation, the method comprising: (a) introducing a first olfactory receptor ligand to a plurality of olfactory receptors, wherein the first olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the first olfactory receptor ligand; (c) introducing a second olfactory receptor ligand, wherein the second olfactory receptor ligand is different from the first olfactory receptor ligand and is an agonist of the plurality of olfactory receptors, such that the second olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes; (d) following an optional period during which the second olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the first olfactory receptor ligand to the plurality of olfactory receptors; and (e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the first olfactory receptor ligand. In some embodiments, the method further comprises: (f) identifying an adaptation-resistant olfactory receptor agonist that is resistant to olfactory adaptation based on a difference (or a lack of a difference) between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) selecting the adaptation-resistant olfactory receptor agonist as an adaptation-resistant substance.

[0076] In certain aspects, the disclosure provides a method of identifying a substance that is resistant to olfactory adaptation, the method comprising: (a) introducing an olfactory receptor ligand to a plurality of olfactory receptors, wherein the olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand; (c) reintroducing the olfactory receptor ligand to the plurality of olfactory receptors, such that the olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time immediately following the subsequent reintroduction of the olfactory receptor ligand; (d) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor ligand. In some embodiments, the method further comprises: (f) identifying an adaptation-resistant olfactory receptor agonist that is resistant to olfactory adaptation based on a difference between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) selecting the adaptation-resistant olfactory receptor agonist as an adaptation-resistant substance.

[0077] In certain aspects, the disclosure provides a method of identifying a substance that is susceptible to olfactory adaptation, the method comprising: (a) introducing an olfactory receptor ligand to a plurality of olfactory receptors, wherein the olfactory receptor ligand is an agonist of the plurality of olfactory receptors; (b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand; (c) reintroducing the olfactory receptor ligand to the plurality of olfactory receptors, such that the olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time immediately following the subsequent reintroduction of the olfactory receptor ligand; (d) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor ligand. In some embodiments, the method further comprises: (f) identifying an adaptation-susceptible olfactory receptor agonist that is resistant to olfactory adaptation based on a difference between the first measurement and the second measurement. In some further embodiments, the method further comprises: (g) deselecting the adaptation-susceptible olfactory receptor agonist as an adaptation-resistant substance.

[0078] The foregoing screening methods are not limited to any particular olfactory receptor or its ligands. It is believed that the methods are generalizable to a wide range of olfactory receptors in addition to those for which working Examples are provided. For example, in some embodiments, the plurality of olfactory receptors are a plurality of human olfactory receptors, such as a plurality of human 0R1A1, 0R8D1, OR51I2, 0R51V1, OR10J5, OR10G4, 0R5K1 , OR10K1, or OR11A1 olfactory receptors, or a polypeptide having an amino acid sequence that is at least 70%, or at least 75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97%, or at least 99%, equivalent thereto. In some other embodiments, the plurality of olfactory receptors are a plurality of non-human mammalian olfactory receptors, such as mouse or rat olfactory receptors.

[0079] Any suitable olfactory receptor ligand can be used in the screening method, so long as the compound or substance is a ligand of at least one of the plurality of olfactory receptors employed in the method. For example, in embodiments where the plurality of olfactory receptors are a plurality of human OR1A1 olfactory receptors, the olfactory receptor ligand is a human OR1A1 olfactory receptor ligand. In general, the particular ligands of various human olfactory receptors are well known in the art. Moreover, such ligands can be identified without undue experimentation by a person of skill in the art using standard cellular screening assays, where the olfactory receptor is expressed in a eukaryotic cell and test compounds or substances are introduced to the assay and an activation or inhibition dose-response curve is generated. The skilled artisan can readily calculate an EC50 from the resulting activation curve or an IC50 for the inhibition curve. In general, compounds or substances having an activation curve with respect to an olfactory receptor with an EC50 of no more than 600 pM are classified as agonists of the olfactory receptor in question. In general, compounds or substances having an inhibition curve with respect to an olfactory receptor with an IC50 of no more than 600 pM are classified as antagonists of the olfactory receptor in question. Examples of suitable olfactory receptor ligands for use in the methods described herein include, not are not limited to, nerol, indole, beta-damascenone, lyral, isobutylquinoline, delta- decalactone, and the like.

[0080] The plurality of olfactory receptors can be provided in any suitable way. For example, in some embodiments, the olfactory receptors are comprised by olfactory sensory neurons (OSNs). Such OSNs can be obtained in any suitable way. For example, in some embodiments, such OSNs are separated from the tissue of a mammal, such as the tissue of a human, mouse, rat, cat, dog, and the like. In some embodiments, the OSNs are mouse OSNs. In some other embodiments, the OSNs are human OSNs. In general, each OSN contains a single olfactory receptor. Thus, by observing the signaling activity of an OSN, one can indirectly measure the signaling activity of the olfactory receptor comprised by the OSN. Methods of separating OSNs from mammalian tissue are known to the skilled artisan.

[0081] In some other embodiments, the plurality of olfactory receptors are expressed on the surface of a plurality of cells. In general, the eukaryotic cells are engineered to overexpress the polypeptides corresponding to the olfactory receptor agonist, a functional fragment thereof, or a polypeptide having at least 70% equivalence to the foregoing. The eukaryotic cells can be engineered to overexpress any of the plurality of olfactory receptors, such as the human OR1A1 olfactory receptor. In some embodiments, the eukaryotic cells are engineered to overexpress one or more of the polypeptides corresponding to different olfactory receptors.

[0082] In some embodiments, the eukaryotic cell further comprises an exogenous nucleic acid sequence that encodes for a G protein. In some further embodiments, the G-protein is the a- subunit Golf or any chimera thereof. In some further embodiments, the G protein is gustducin, such as an a-gustducin. In some other embodiments, the G protein is a promiscuous G protein. Non-limiting examples of promiscuous G proteins include Ga16, Ga15, Ga16gust25, Ga15gust25, Ga16gust44, Ga15gust44, and Ga15-i / 3-5, which are described in detail in U.S. Patent No. 10,107,794. In some other embodiments, the G protein is a G(i / o) protein.

[0083] In some embodiments, the eukaryotic cell further comprises a G protein coupled to the olfactory receptors. In some further embodiments, the G-protein is the a-subunit Golf or any chimera thereof. In some further embodiments, the G protein is gustducin, such as an a- gustducin. In some other embodiments, the G protein is a promiscuous G protein. Non-limiting examples of promiscuous G proteins include Ga16, Ga15, Ga16gust25, Ga15gust25, Ga16gust44, Ga15gust44, and Ga15-i / 3-5, which are described in U.S. Patent No. 10,107,794. In some other embodiments, the G protein is a G(i / o) protein.

[0084] Any suitable cell can be used. For example, human H EK-293 and human U2OS cells are well known in the art and are generally available commercially. In some embodiments, the eukaryotic cells are isolated eukaryotic cells. In general, the eukaryotic cells are contained in an assay. In some embodiments, the eukaryotic cells are adhered to a substrate. Various functional assays can be used, including assays that measure of changes in intracellular calcium cation concentration (for example, Fluorometric Imaging Plate Reader-based Ca2+mobilization assay, FLIPR, or a genetically encoded calcium indicator such as GCaMP), intracellular cyclic adenosine monophosphate (cAMP) (for example Homogenous Time Resolved Fluorescence, Pherastar, or a genetically encoded cAMP indicator such as GcAMP or cADDis), pERK1 / 2 activation (for example, high content imaging, HCI), and receptor internalization (for example, TRANSFLUOR). A change in olfactory receptor activity by a test compound or by an olfactory receptor ligand indicates modulation of the olfactory receptor by the test compound or the agonist.

[0085] In some instances, cells besides human HEK-293 or human U2OS cells can also be used. Non-limiting examples of such cells include the following cell types: 1A2, ARH-77, RWPE-1, WI-38, EJM, NCI-H1155, L-1236, NCI-H526, JM1, SHP-77, SNU-878, NCI-H2196, C3A, CA46, SNU-466, KS-1, SNU-738, MOLP-2, HDLM-2, Pfeiffer, HCC-15, Alexander cells, L-540, KMS-12-BM, JK-1, NCI-H1092, SW 1990, NCI-H1184, SU-DHL-1, Hep 3B2.1-7, P3HR- 1 , NCI-H2029, SU-DHL-5, SNU-1, MOLP-8, SUP-M2, MONO-MAC-1 , SNU-1040, KYM-1, HEC-59, HCC1569, OCI-LY3, Hs 819.T, DU4475, CI-1, S-117, OVCAR-8, SNU-626, HL-60, SUIT-2, T3M-4, RKO, MOR / CPR, DK-MG, GA-10, OCUM-1, HCT-15, HT, MONO-MAC-6, G- 402, Toledo, COV362, SU-DHL-8, Daoy, NCI-H1435, LS513, Hs 839.T, Hs 172.T, BT-483, KMS-21 BM, AGS, NCI-H2172, LC-1 / sq-SF, SNU-201 , NUGC-4, SK-HEP-1 , SUP-B15, SNU-5, HT-1197, SUP-T1, AMO-1 , KU812, AN3 CA, AML-193, VMRC-RCW, HLE, HuH28, Hs 751.T, NCI-H2110, MEG-01 , MV-4-11, Hep G2, KYSE-30, KALS-1, BICR 6, RMUG-S, JHH-6, Ki-JK, IST-MES1 , HCC-95, HPB-ALL, HSC-3, 697, LOU-NH91, KARPAS-299, GI-1, COLO 792, SK- N-FI, D341 Med, HGC-27, SR-786, COLO-818, MHH-CALL-2, SF126, NCI-H322, A-253, NCI- H1623, MCF7, HCC-44, FU97, OCI-LY-19, Hs 766T, NCI-H522, RL, HCC1428, RPMI 6666, U- 937, NCI-H460, SW 1088, NCI-H1792, NCI-H1693, UACC-257, JHUEM-2, HuT 78, UACC-893, NCI-H929, A-704, OV56, LN-229, OE19, SK-MEL-24, RD-ES, NCI-H211 , KCI-MOH1, NCI- H1963, Hs 706.T, ChaGo-K-1, EPLC-272H, OPM-2, KHM-1 B, A549, HuG1-N, NCI-H508, MHH-CALL-3, SNU-1076, A3 / KAW, MEL-HO, TO 175.T, Caki-1 , Hs 936.T, SK-LU-1, WM- 983B, K-562, EFE-184, SNU-520, NCI-H2291, HCC-1195, ABC-1, KE-39, NH-6, HCC2218, CMK, RS4;11 , KYSE-450, OV7, KYSE-510, SK-UT-1, SNU-C1, OE33, P12-ICHIKAWA, DLD-1, COV434, HuNS1 , SNU-899, SW480, COLO-678, LU99, KOPN-8, NCI-H2227, SW1463, Hs 675.T, JHH-4, NCI-H1703, HEC-1-A, BDCM, MIA PaCa-2, PC-3, TE-15, PK-45H, MKN-45, HCC-366, CAL-29, HEC-50B, CPC-N, KMRC-20, SW1116, EOL-1, COLO 205, EHEB, YD-38, MC116, SK-N-BE(2), BV-173, NCI-H2347, LU65, RT4, U-87 MG, LK-2, KP-N-YN, HEC-251, NCI-H1651 , GP2d, RERF-LC-MS, NB-4, NCI-H2286, SNU-61, T-47D, huH-1, KYSE-180, ST486, SW 1353, M-07e, KASUMI-1 , YH-13, NCI-H28, GAMG, JeKo-1 , GOS-3, SNU-324, PA- TU-8902, MFE-280, SNU-245, NALM-1, RERF-LC-Sq1, BICR 22, ZR-75-1, COR-L23, SW579, COR-L88, KM12, Hs 611.T, OUMS-23, RERF-LC-Ad1, NCI-H1385, SK-LMS-1 , COLO-320, BL- 70, GRANTA-519, MCAS, Pane 08.13, AM-38, KMS-11, SIG-M5, SNU-407, JHOS-2, OVCAR- 4, Set-2, OV-90, MeWo, HEL, HT-29, MDA-MB-231, TOV-21G, NCI-H1355, KMS-27, NALM-6, KMS-26, Caov-4, KASUMI-2, UACC-62, U266B1 , Hs 695T, HT55, BICR 31 , TCC-PAN2, KMS- 20, Hs 578T, RI-1, Hs 606.T, NCI-H1341, THP-1, BCP-1 , Hs 737.T, SW1417, MOLT-4, Raji, ESS-1 , MEL-JUSO, SH-10-TC, Hs 683, ME-1, EB2, PLC / PRF / 5, NCI-H1339, A4 / Fuk, SEM, HEC-265, IST-MES2, KE-97, NCI-H1437, COLO-704, NCI-H1915, TE-5, NCI-H2023, NCI-H82, T1-73, SNU-840, HuT 102, NCI-H1944, KYSE-520, Kasumi-6, 1321N1 , Hs 742.T, IM95, PL45, CL-40, WM1799, KMM-1 , SNU-449, JHUEM-1, KARPAS-620, Loucy, SNU-1079, Daudi, HCC-56, HSC-2, COR-L47, PA-TU-89885, OAW28, COR-L311, L-363, Malme-3M, NOMO-1 , Hs 870.T, SU-DHL-10, Hs 229.T, NCI-H810, KYSE-410, RPMI-8402, SNU-175, EBC-1, RVH- 421 , K029AX, PA-TU-8988T, LXF-289, OVSAHO, CAL-12T, Hs 940.T, MM1-S, SUP-HD1 , LNCaP clone FGC, HSC-4, NU-DHL-1 , NCI-H2228, BEN, CAL-78, Sq-1, NCI-H1793, SNU- C2A, M DA- MB- 134- VI, COV318, KE-37, TYK-nu, MOTN-1 , T98G, SW837, EB1 , Becker, PE / CA-PJ34 (clone C12), Hs 616.T, NCI-H446, WM-88, CHP-126, Calu-1, SNU-283, NCI- H1573, SW 1271, SNU-16, JHOS-4, ACHN, Calu-3, KMRC-1 , SW 1783, TE-11, TE-9, HuH-6, P31 / FUJ, HT-1376, NCI-H520, 786-0, KNS-60, Caki-2, OVK18, PL-21, NCI-H2452, JURL- MK1, TEN, JHH-7, MDA-MB-157, Calu-6, RKN, NUGC-2, ONS-76, J82, OUMS-27, SNU-1196, Hs 739.T, RPMI-7951 , NCI-H854, JHH-5, JVM-2, Hey-A8, 5637, KYSE-140, Capan-2, KYSE- 150, HEC-1-B, BICR 16, HEL 92.1.7, MHH-NB-11, SNU-387, SK-OV-3, SK-MEL-28, IGROV1, ML-1 , HLF-a, CHL-1 , YKG1 , A-204, OCI-M1 , 8505C, JVM-3, NCI-H647, DB, COLO-800, PK- 59, FaDu, HLF, OVMANA, EFO-27, PF-382, NCI-H747, LS123, SU-DHL-6, SJRH30, PANC-1, NCI-H2342, KM-H2, DND-41 , HH, HuCCTI, F-36P, DMS 454, Hs 274.T, AU565, NCI-H1666, EN, RH-41, NCI-H1373, NCI-H838, SK-MEL-30, MOLM-6, DEL, NCI-H226, NCI-H1648, NCI- H661 , 143B, Mino, C32, KMS-34, NCI-H1694, SK-ES-1 , UACC-812, GDM-1 , NCI-H23, Pane 02.03, CCF-STTG1 , LOX IMVI, SJSA-1, MDST8, PK-1, NCI-H716, SU-DHL-4, MPP 89, MJ, COLO 829, PE / CA-PJ15, HD-MY-Z, BxPC-3, WM-793, COLO 668, T84, JHOM-1, PEER, LS411 N, GMS-10, KMBC-2, RMG-I, KELLY, SNU-761, NALM-19, HEC-151, G-361, OVTOKO, A-498, SW 900, LCLC-103H, FTC-133, QGP-1, Reh, CMK-11-5, NU-DUL-1, BT-20, Hs 600.T, Hs 604.T, KATO III, SNU-410, NCI-H2126, SK-MEL-5, MDA-MB-468, AsPC-1 , HUP-T3, KP-N- SI9s, L-428, SNU-1105, HUP-T4, 769-P, LMSU, NCI-H1869, NCO2, MOLM-16, CAL 27, HCC70, NCI-H1930, COV644, Hs 863.T, HCC-2279, D283 Med, Hs 944.T, HCC1599, MDA- MB-415, HCC2157, NCI-H1618, SNU-308, HCC1954, DMS 153, HPAF-II, T24, CJM, VM- CUB1, UM-UC-3, LAMA-84, NCI-H1734, JHH-2, VMRC-RCZ, MFE-319, MDA-MB-453, SNU- 503, TOV-112D, B-CPAP, GSU, HCC-78, NCI-H2171 , CAMA-1, HEC-108, HCC4006, CAL-85- 1 , NCI-H2122, COLO-699, NCI-H196, LUDLU-1 , SW 780, RPMI 8226, LP-1, PC-14, HuTu 80, T.T, SW948, 22Rv1 , HARA, NCI-H596, IPC-298, SCaBER, NCI-H1838, NB-1, Hs 934.T, Hs 895. T, DMS 114, KYSE-70, KP-3, KP4, DAN-G, NCI-H2009, OC 316, SCC-25, U-138 MG, RCC10RGB, MFE-296, NCI-H1755, RERF-LC-KJ, 8305C, WSU-DLCL2, ES-2, MSTO-211 H, SCC-15, ZR-75-30, PSN1 , SNU-423, NCI-H2106, TE-1 , UT-7, KMS-28BM, NCI-H2081, SK- MM-2, COLO 741 , OC 314, HCC1395, MOLT-13, LN-18, Pane 10.05, PE / CA-PJ41 (clone D2), Hs 746T, CW-2, SKM-1 , NUGC-3, TE-10, NCI-H358, NCI-H69, BFTC-909, HOS, BICR 18, NCI-H1395, OVKATE, Hs 698.T, EFM-19, COLO-783, MHH-CALL-4, ACC-MESO-1 , NCI- H1436, KP-N-RT-BM-1, SK-MEL-31, NCI-H1105, CAL-51 , YD-15, NCI-H2085, NCI-H2444, HCC1187, Hs 939.T, CAL-120, SCC-9, TUHR14TKB, KMRC- 2, KG-1-C, ECC10, CGTH-W-1, NCI-H841, C2BBe1 , SUP-T11, RCH-ACV, CADO-ES1, JURKAT, 647-V, SK-MEL-2, MDA-MB-175-VII, MKN74, SNU-C4, LCLC-97TM1, SCC-4, BHY, IGR-37, KYO-1, HS 281.T, TT, TUHR4TKB, HT-1080, NCI-H660, TE 441.T, LS1034, KNS-42, Pane 04.03, HCC1419, AZ-521 , SNG-M, NCI-N87, G-292, clone A141 B1 , KPL-1 , MDA-MB- 361 , CL-14, NCI-H2170, HuH-7, RD, NCI-H2066, IGR-1, TE-14, VCaP, BL-41 , SNU-620, SK- MES-1, MEC-2, NCI-H1299, IGR-39, RT112 / 84, SF-295, DV-90, A2780, BICR 56, NCI-H510, NCI-H2141 , YD-8, NCI-H2405, TF-1, MEC-1 , CCK-81, NCI-H1048, Hs 822.T, NCI-H2052, K052, CAL- 54, Hs 840.T, SW620, SK-CO-1, BT-474, CL-11, KNS-62, NCI-H1650, G-401, MOLT-16, SNU-398, COLO-680N, EM-2, Hs 294T, CAL-62, KMRC-3, A101D, KG-1, BT-549, HT115, A-375, SW-1710, WM-115, KLE, JHUEM-3, MKN7, CHP-212, HCC202, BC-3C, NCI- H1568, KMS-18, PE / CA-PJ49, COLO-849, SIMA, OCI-AML3, GSS, EC-GI-10, EFO-21 , RCM- 1 , DMS 273, KU-19-19, RERF-GC-1B, SH-4, SK-MEL-3, RERF-LC-Ad2, M059K, JHOM-2B, MDA PCa 2b, Hs 852.T, RL95-2, Pane 03.27, SNU-216, Pane 02.13, CFPAC-1 , SK-N-SH, OCI-AML2, LoVo, SBC-5, NCI-H1876, NCI-H441, SK-N-AS, COR-L24, HCC38, NCI-H1781, DOHH-2, NCI-H1563, U-251 MG, HPAC, JIMT-1, U-2 OS, A-673, TC-71, NCI-H650, NIH:OVCAR-3, CAS-1, JL-1, SK-MEL-1 , MDA-MB-4355, Ishikawa (Heraklio) 02 ER-, TE 617.T, SU.86.86, RERF-LC-AI, TT2609-C02, LS 180, YAPC, HDQ-P1, KNS-81, FU-OV-1, KP-2, DMS 53, SNU-1272, Detroit 562, 42-MG-BA, L3.3, COLO-679, NCI-H2087, NCI-H2030, GCT, NCI- H889, Caov-3, MDA-MB-436, NCI-H524, MKN1 , KCL-22, Capan-1 , CML-T1, H4, NCI-H727, Hs 343.T, MHH-ES-1, NMC-G1 , HCC-1171, REC-1, Hs 618.T, A172, YD-10B, SW48, MUTZ-5, TE-6, JHH-1, HCT 116, TE-4, IA-LM, MG-63, NCI-H1975, TALL-1, HCC1806, HMCB, SCLC- 21H, HCC1500, CL-34, Pane 05.04, SW403, TM-31 , HCC1937, JMSU-1, DMS 79, SNB-19, NCI-H1836, Li-7, HCC827, 639-V, MOLM-13, SK-BR-3, IMR-32, TUHR10TKB, OAW42, SK-N- MC, TGBC11TKB, NCI-H1581, EFM-192A, YMB-1 , HCC2935, ECC12, HCC-33, DU 145, NCI- H146, SNU-1214, SNU-1077, 23132 / 87, HT-144, SNU-182, Hs 888.T, SNU-475, GCIY, Hs 729, JHOC-5, SW 1573, HEC-6, OCI-AMLS, Hs 688(A).T, Hs 821. T, PCM6, RT-112, SK-N-DZ, SNU-478, SNU-119, HCC1143, NCI-H209, 8-MG-BA, COR-L105, COR-L95, SNU-46, COV504, CAL-148, SNU-05, DBTRG-05MG, BHT-101, WM-266-4, BFTC-905, KYSE-270, TE- 8, SNU-213, and SH-SYSY. The methods disclosed herein include two separate steps involving measuring of a parameter indicative of signaling activity of the plurality of olfactory receptors following the initial introduction and subsequent reintroduction of the olfactory receptor ligand. In the methods disclosed herein, the measuring generally continues until the indicative parameter reaches a reasonable plateau. For example, in some embodiments, the measuring continues for 20 minutes, or 40 minutes, or 60 minutes, or 80 minutes, or 100 minutes, or 120 minutes following introduction or re introduction of the olfactory receptor ligand to the olfactory receptor. The measuring can occur at any suitable intervals throughout the duration of the measuring, such as at intervals of 10 seconds, 20 seconds, 30 seconds, 45 seconds, 1 minute, 2 minutes, 5 minutes, 10 minutes, and the like.

[0086] Any suitable parameter can be measured, so long as the parameter is indicative of the signaling activity of an olfactory receptor in response to the introduction or reintroduction of an agonist or antagonist of that olfactory receptor.

[0087] For example, one of the downstream signaling events from receptor activation is an influx of calcium ions into the cell expressing the olfactory receptor. Therefore, in some embodiments, the measuring comprises measuring the calcium ion concentration in the cell following the introduction or reintroduction of an agonist or antagonist of that olfactory receptor. This can be measured in various ways. For example, in some embodiments, a fluorescent calcium ion indicator is introduced to the cells and the calcium concentration is measured indirectly by measuring or observing the intensity of the fluorescence of the indicator, such as fluorescent imaging, and the like. Such techniques are known to the skilled artisan.

[0088] Another downstream signaling event involves from receptor activation involves the increase in the concentration of cyclic adenosine monophosphate (cAMP). Techniques for measuring downstream cAMP concentration in a cell are known to the skilled artisan. For example, indicators having fluorescent tags can be used to measure the cAMP concentration through fluorescent imaging or other techniques for assessing fluorescent intensity.

[0089] The methods disclosed herein further comprise a step of reintroducing the olfactory receptor ligand to the plurality of olfactory receptors, such that the olfactory receptor ligand remains in contact with the plurality of olfactory receptors for an extended period of time, such as a period ranging from 1 minute to 120 minutes. This soaking step occurs after the initial introduction and before a subsequent reintroduction of the olfactory receptor ligand. This soaking can occur for any suitable period of time, such as a time period ranging from 2 minutes to 90 minutes, or from 5 minutes to 60 minutes, or from 10 minutes to 30 minutes. Following this soaking step and before reintroduction of the olfactory receptor ligand, there is a period of time during which the olfactory receptor is not contacted with the olfactory receptor ligand. This period of time can last for any suitable duration, for example, from 1 minute to 60 minutes, or from 1 minute to 30 minutes, or from 1 minute to 20 minutes. In some embodiments, the methods disclosed herein include calculating a difference between the first and second measurement of the parameter indicative of signaling activity. In general, the measuring steps result in one or more curves showing an increase in signaling activity followed by a drop off in signaling activity. Various parameters can be extracted from these curves, such as peak intensity (amplitude), area under the curve, the initial rate, the onset slope, the onset half-life (ti / 2), the peak response, the peak time, the plateau, the offset slope, the offset half-life (ti / 2), and the like. A small difference in a relevant parameter between the first and second measurement indicates that a particular odorant is resistant to adaptation, meaning that one will generally continue to experience a sensory response to the presence of the odorant over time. For example, one does not adapt to the odorant over time. By contrast, a large difference in a relevant parameter between the first and second measurement indicates that a particular odorant is susceptible to adaptation, meaning that one will discontinue to experience a sensory response to the presence of the odorant over time. For example, one adapts to the odorant over time.

[0090] In some further embodiments of any of the foregoing embodiments, the methods further comprise identifying an adaptation-susceptible olfactory receptor ligand that is susceptible to olfactory adaptation based on the determined degree of cellular olfactory receptor internalization or an adaptation-resistant olfactory receptor ligand that is resistant to olfactory adaptation, based on the determined degree of cellular internalization from step (b). The ligands shown to promote internalization of the olfactory receptor are classified as adaptation- susceptible agonists, and those ligands shown not to promote significant internalization of the olfactory receptor are classified as adaptation-resistant agonists. The skilled artisan is capable of making such distinctions based on differences in the determined degree of internalization for a series of ligands of a particular olfactory receptor.

[0091] In some further embodiments, the methods further comprise selecting adaptationresistant ligands or deselecting adaptation-susceptible ligands for use in various applications where olfactory adaptation may be undesirable.

[0092] The olfactory receptor ligand can be any suitable compound or substance that is amenable for use in cellular screening assays. For example, in some embodiments, the olfactory receptor ligand is an organic compound. In some embodiments, the olfactory receptor ligand is a naturally occurring compound. In some other embodiments, the olfactory receptor ligand is a non-naturally occurring compound. In some embodiments, the olfactory receptor ligand is a peptide, such as a polypeptide or an oligopeptide, an amino acid, an amino acid derivative, an amide, a nucleotide, an oligonucleotide. In some embodiments, the olfactory receptor ligand is a plant or food extract, such as a plant or food extract obtained by bioassay- guided fractionation. In some embodiments, the olfactory receptor ligand is a terpene, a diterpene or a triterpene. In some embodiments, the olfactory receptor ligands is a ribonucleotide or derivative thereof. In some embodiments, the olfactory receptor ligand is an organic molecule having a molecular weight of no more than 1000 amu.

[0093] Uses and Methods

[0094] In other aspects, the disclosure provides use of an olfactory-resistant substance identified or selected by the method of the foregoing aspects to provide a long-lasting fragrance to a fragranced article. The disclosure also provides methods that correspond to each of the foregoing uses. Thus, in certain related aspects, the disclosure provides methods of providing a long-lasting fragrance to a fragranced article, the method comprising introducing to an article an adaptation-resistant substance identified or selected by the method of the foregoing aspects. In certain related aspects, the disclosure provides methods of providing a long-lasting fragrance to a fragranced article, the method comprising removing from an article an adaptation-susceptible substance identified or deselected by the method of the foregoing aspects.

[0095] Any suitable fragranced articles are contemplated. In some embodiments, the fragranced article is a fiber, such as a natural or synthetic fiber. In some such embodiments, the fiber is comprised by a textile article, such as an article of clothing, a towel, bedding, upholstery, and the like. In some embodiments, the fragranced article is a consumer care product, such as a household cleaning product, an air freshener, a commercial cleaning product, a dish detergent, a laundry detergent, a fabric softener, a scent booster, or a personal care product, such as a shower gel, a shampoo, a hair conditioner, a hair-styling product, a skin care product, a cosmetic product, a deodorant, an antiperspirant, or a self-tanning product. In some embodiments, the fragranced article is a perfuming composition, such as a fine fragrance composition or a fragrance composition for use in various air-care products, such as plug-in diffusers, candles, perfuming reeds, room sprays, and the like.

[0096] Fiber and Textiles

[0097] In certain related aspects, the disclosure provides uses to impart a long-lasting fragrance to a fiber, the method comprising introducing one or more selected or identified substances to the fiber. In general, the fiber is a textile fiber, such as a natural or synthetic fiber comprised by a textile article, such as an article of clothing, a towel, bedding, upholstery, and the like. In some embodiments, the identified or selected substance is introduced to the fiber as part of a perfuming composition included within a personal care composition, such as those described in further detail below.

[0098] Consumer Care Compositions and Related Uses and Products

[0099] In certain related aspects, the disclosure provides uses to impart a long-lasting fragrance to a consumer care composition, the method comprising introducing one or more selected or identified substances to the consumer care composition. Further embodiments of the consumer care compositions referenced in connection with the foregoing methods and uses are set forth in more detail below.

[0100] In certain aspects, the disclosure provides a consumer care composition comprising a plurality of microparticles of the first aspect or any embodiments thereof. In some embodiments, the consumer care composition is in the form of a household cleaning product, a commercial cleaning product, a dish detergent, a laundry detergent, a fabric softener, a scent booster, a shower gel, a shampoo, a hair conditioner, a hair-styling product, a skin care product, a cosmetic product, a deodorant, an antiperspirant, or a self-tanning product.

[0101] In some embodiments, the consumer care composition comprises a plurality of microparticles and at least one active ingredient, such as an active ingredient selected from the group consisting of a cosmetic ingredient, skin caring ingredient, perfume ingredient, flavor ingredient, malodor counteracting ingredient, bactericide ingredient, fungicide ingredient, pharmaceutical or agrochemical ingredient, a sanitizing ingredient, an insect repellent or attractant, and mixtures thereof.

[0102] In some embodiments, the consumer care composition is a perfuming composition. In some such embodiments, the consumer care composition comprises a plurality of microparticles, as defined above where the microparticle is a coacervate core-shell microcapsule whose core comprises at least one perfume compound, and, optionally, at least one perfumery adjuvant or liquid perfumery carrier.

[0103] Liquid perfumery carriers are well known in the art. Some non-limiting examples include an emulsifying system, such as a solvent and a surfactant system, or a solvent commonly used in perfumery. Some non-limiting examples of suitable such solvents include dipropyleneglycol, diethyl phthalate, isopropyl myristate, benzyl benzoate, 2-(2-ethoxyethoxy)-1 -ethanol or ethyl citrate, which are the most commonly used. For the compositions which comprise both a perfumery carrier and a perfumery co-ingredient, other suitable perfumery carriers than those previously specified, can be also ethanol, water / ethanol mixtures, limonene or other terpenes, isoparaffins such as those known under the trademark ISOPAR (Exxon Chemical, Houston, Tex., US) or glycol ethers and glycol ether esters such as those known under the trademark DOWANOL (Dow Chemical Company, Midland, Mich., US). By “perfumery co-ingredient” it is meant here a compound, which is used in a perfuming preparation or a composition to impart a hedonic effect and which is not a microcapsule as defined above. In other words such a co- ingredient, to be considered as being a perfuming one, must be recognized by a person skilled in the art as being able to at least impart or modify in a positive or pleasant way the odor of a composition, and not just as having an odor.

[0104] Perfuming co-ingredients are well known in the art. In general terms, these perfuming co-ingredients belong to chemical classes as varied as alcohols, lactones, aldehydes, ketones, esters, ethers, acetates, nitriles, terpenoids, nitrogenous or sulfurous heterocyclic compounds and essential oils, and said perfuming co-ingredients can be of natural or synthetic origin. Many of these co-ingredients are in any case listed in reference texts such as the book by Arctander, PERFUME AND FLAVOR CHEMICALS (1969) or its more recent versions, or in other works of a similar nature, as well as in the abundant patent literature in the field of perfumery. It is also understood that said co-ingredients may also be compounds known to release in a controlled manner various types of perfuming compounds. Non-limiting examples of co-ingredients include 4-(dodecylthio)-4-(2,6,6-trimethyl-2-cyclohexen-1-yl)-2-butanone, 4-(dodecylthio)-4- (2,6,6-trimethyl-1-cyclohexen-1-yl)-2-butanone, trans-3-(dodecylthio)-1-(2,6,6-trimethyl-3- cyclohexen-1-yl)-1-butanone, 2-(dodecylthio)octan-4-one, 2-phenylethyl oxo(phenyl)acetate, 3,7-dimethylocta-2,6-dien-1-yl oxo(phenyl)acetate, (Z)-hex-3-en-1-yl oxo(phenyl)acetate, 3,7- dimethyl-2,6-octadien-1-yl hexadecanoate, bis(3,7-dimethylocta-2,6-dien-1-yl) succinate, (2-((2- methylundec-1-en-1-yl)oxy)ethyl)-benzene, 1-methoxy-4-(3-methyl-4-phenethoxybut-3-en-1- yl)benzene, (3-methyl-4-phenethoxybut-3-en-1-yl)benzene, 1-(((Z)-hex-3-en-1-yl)oxy)-2- methylundec-1-ene, (2-((2-methylundec-1-en-1-yl)oxy)ethoxy)benzene, 2-methyl-1-(octan-3- yloxy)undec-1 -ene, 1 -methoxy-4-(1 -phenethoxyprop-1 -en-2-yl)benzene, 1 -methyl-4-(1 - phenethoxyprop- 1-en-2-yl)benzene, 2-(1-phenethoxyprop-1-en-2-yl)naphthalene, (2- phenethoxyvinyl)benzene, 2-(1-((3,7-dimethyloct-6-en-1-yl)oxy)prop-1-en-2-yl)naphthalene, (2- ((2-pentylcyclopentylidene)-methoxy)ethyl)-benzene, 4-allyl-2-methoxy-1-((2-methoxy-2- phenylvinyl)oxy)benzene, (2-((2-heptyl-cyclopentylidene)methoxy)ethyl)benzene, 1-isopropyl-4- methyl-2-((2-pentyl-cyclopentylidene)methoxy)benzene, 2-methoxy-1-((2- pentylcyclopentylidene)methoxy)-4-propylbenzene, 3-methoxy-4-((2-methoxy-2- phenylvinyl)oxy)benzaldehyde, 4-((2-(hexyloxy)-2-phenylvinyl)oxy)-3-methoxybenzaldehyde or a mixture thereof or a mixture thereof.

[0105] The term “perfumery adjuvant” refers to an ingredient capable of imparting additional added benefit such as a color, a particular light resistance, chemical stability, etc. Such compounds are well known in the art.

[0106] The personal care composition can contain any suitable amount of the microparticles of the present disclosure. In some embodiments, the personal care composition comprises from 0.01% by weight to 30% by weight of microparticles, based on the total weight of the consumer care composition.

[0107] The consumer care composition can be in any suitable physical state, such as a solid (such as a powder), liquid, or a gas. In some embodiments, the consumer care composition is a liquid. In some such embodiments, the liquid consumer care composition has one or more of the following characteristics: a) comprises from 2% by weight to 65% by weight of a surfactant, based on the total weight of the consumer care composition; b) comprises water or a water-miscible hydrophilic organic solvent; c) is in the form of a microparticle slurry; and d) comprises a non-encapsulated perfume.

[0108] In some embodiments, the consumer care composition is in the form of a powder. In some such embodiments, the powder consumer care composition has one or more of the following characteristics: a) comprises from 2% by weight to 65% by weight of a surfactant, based on the total weight of the consumer care composition; b) is in the form of a microcapsule powder; and c) comprises a perfume powder that is different from any perfume comprised by the microparticles.

[0109] In embodiments where the plurality of microparticles comprise encapsulated cores containing perfume compounds, the consumer care composition comprising these microparticles can be of used in various perfumed consumer products, such as products belonging to fine fragrance or “functional” perfumery. Functional perfumery includes personal care products, including hair care products, body cleansing products, skin care products, hygiene products, as well as home care products, including laundry care products, surface care products, and air care products. The term “perfumed consumer product” broadly refers to any consumer product that is expected to deliver, among different benefits, a perfuming effect to the surface to which it is applied, including, but not limited to, skin, hair, textiles, paper, countertops, sinks, toilets, floors, furniture, or other household surfaces, or in the air, for example, in the case of an air freshener, a room deodorizer, a candle, a reed diffuser, and the like.

[0110] Such perfumed consumer products can contain any other ingredients that are commonly used in the industry. Methods of formulating perfume-containing microcapsules are such products is also well known and can be used to develop formulations of such products containing microparticles of the present disclosure.

[0111] Non-limiting examples of suitable perfumed consumer products include a perfume, such as a fine perfume, a cologne, an after-shave lotion, a body splash, a fabric care product, such as a liquid or solid detergent, tablets and unit dose (single or multi-chambers), a fabric softener, a dryer sheet, a fabric refresher, an ironing water, a bleach, a hair care product, such as a shampoo, a hair conditioner, a coloring preparation, or a hair spray, a cosmetic preparation, such as a vanishing cream, a body lotion, or a deodorant or antiperspirant, or a skin-care product, such as a perfumed soap, a shower or bath mousse, a body wash, an oil or gel, bath salts, or a hygiene product, an air care product, such as an air freshener or a “ready to use” powdered air freshener, or a home care product, such all-purpose cleaners, liquid or power or tablet dishwashing products, toilet cleaners, or products for cleaning various surfaces, for example sprays and wipes intended for the treatment or refreshment of textiles or hard surfaces like floors, tiles, and stone, or a hygiene product such as sanitary napkins, diapers, or toilet paper.

[0112] In some embodiments, the consumer care composition comprises: a personal care active base, and a plurality of microparticles according to any of the embodiments set forth herein, wherein the consumer care composition is in the form of a personal care product.

[0113] Any suitable personal care active base or combination of such materials can be used. Such materials are well known to the skilled artisan and are widely discussed in the relevant patent literature. Non-limiting examples of a consumer care active base include surfactants, oils, hydrophobic solvents, hydrophilic solvents, water, and the like, as well as auxiliary agents, such as bleaching agents, buffering agent, builders, soil release or soil suspension polymers, granulated enzyme particles, corrosion inhibitors, antifoaming, sud suppressing agents, dyes, fillers, and mixtures thereof.

[0114] In some embodiments, the personal care product is a hair-care product, such as a shampoo, a hair conditioner, a coloring preparation, or a hair spray, a cosmetic preparation such as a vanishing cream, a body lotion, or a deodorant or antiperspirant, or a skin care product, such as a perfumed soap, a shower, or a bath mousse, a body wash, an oil or gel, bath salts, or a hygiene product.

[0115] In some embodiments, the consumer care composition comprises: a home care active base, and a plurality of microparticles according to any of the embodiments set forth herein, wherein the consumer care composition is in the form of a home care product.

[0116] Any suitable home care active base or combination of such materials can be used. Such materials are well known to the skilled artisan and are widely discussed in the relevant patent literature. Non-limiting examples of a consumer care active base include surfactants, oils, hydrophobic solvents, hydrophilic solvents, water, and the like, as well as auxiliary agents, such as bleaching agents, buffering agent, builders, soil release or soil suspension polymers, granulated enzyme particles, corrosion inhibitors, antifoaming, sud suppressing agents, dyes, fillers, and mixtures thereof.

[0117] In some embodiments, the home care product is an air care product, such as an air freshener or a “ready to use” powdered air freshener, or a home care product, such all-purpose cleaners, liquid or power or tablet dishwashing products, toilet cleaners, or products for cleaning various surfaces, for example sprays and wipes intended for the treatment or refreshment of textiles or hard surfaces like floors, tiles, and stone, or a hygiene product such as sanitary napkins, diapers, or toilet paper.

[0118] In some embodiments, the consumer care composition comprises the plurality of microparticles in an amount ranging from 0.1% by weight to 15 % by weight, or from 0.2% by weight to 5% by weight, of the plurality of microparticles, based on the total weight of the consumer care composition. The consumer care composition can have any suitable pH. For example, in some embodiments, the consumer care composition has a pH of less than 7. In some other embodiments, the consumer care product has a pH of at least 7.

[0119] Fabric Softener

[0120] In some embodiments, the consumer care composition is in the form of a fabric softener composition. In some such embodiments, the consumer care composition comprises a fabric softener active base and a plurality of microparticles according to any of the embodiments set forth above.

[0121] Any suitable fabric softener active base can be used. For example, in some embodiments, the fabric softener active base includes dialkyl quaternary ammonium salts, dialkyl ester quaternary ammonium salts, 1,2-dioleoyl-3-trimethylammonium propane, triethanolamine quaternary salts, silicones, and mixtures thereof, as well as various surfactants, hydrophilic organic solvents, and water. In some embodiments, the consumer care composition comprises the fabric softener active base in an amount ranging from 85% by weight and 99.95% by weight, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises the plurality of microparticles in an amount ranging from 0.1% by weight to 15 % by weight, or from 0.2% by weight to 5% by weight, of the plurality of microparticles, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises nonencapsulated perfume compounds.

[0122] Liquid Detergent

[0123] In some embodiments, the consumer care composition is in the form of a liquid detergent composition. In some such embodiments, the consumer care composition comprises a liquid detergent active base and a plurality of microparticles according to any of the embodiments set forth above.

[0124] Any suitable liquid detergent active base can be used. For example, in some embodiments, the liquid detergent active base includes anionic surfactant such as alkylbenzenesulfonate (ABS), secondary alkyl sulfonate (SAS), primary alcohol sulfate (PAS), lauryl ether sulfate (LES), methyl ester sulfonate (MES) and nonionic surfactant such as alkyl amines, alkanolamide, fatty alcohol poly(ethylene glycol) ether, fatty alcohol ethoxylate (FAE), ethylene oxide (EO) and propylene oxide (PO) copolymers, amine oxydes, alkyl polyglucosides, alkyl polyglucosamides, as well as various surfactants, hydrophilic organic solvents, and water. In some embodiments, the consumer care composition comprises the liquid detergent active base in an amount ranging from 85% by weight and 99.95% by weight, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises the plurality of microparticles in an amount ranging from 0.1% by weight to 15 % by weight, or from 0.2% by weight to 5% by weight, of the plurality of microparticles, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises non-encapsulated perfume compounds.

[0125] Solid Detergent

[0126] In some embodiments, the consumer care composition is in the form of a solid detergent composition. In some such embodiments, the consumer care composition comprises a solid detergent active base and a plurality of microparticles according to any of the embodiments set forth above.

[0127] Any suitable solid detergent active base can be used. For example, in some embodiments, the solid detergent active base includes anionic surfactant such as alkylbenzenesulfonate (ABS), secondary alkyl sulfonate (SAS), primary alcohol sulfate (PAS), lauryl ether sulfate (LES), methyl ester sulfonate (MES) and nonionic surfactant such as alkyl amines, alkanolamide, fatty alcohol poly(ethylene glycol) ether, fatty alcohol ethoxylate (FAE), ethylene oxide (EO) and propylene oxide (PO) copolymers, amine oxydes, alkyl polyglucosides, alkyl polyglucosamides, as well as various surfactants, hydrophilic organic solvents, water, and fatty acid carboxylates. In some embodiments, the consumer care composition comprises the solid detergent active base in an amount ranging from 85% by weight and 99.95% by weight, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises the plurality of microparticles in an amount ranging from 0.1 % by weight to 15 % by weight, or from 0.2% by weight to 5% by weight, of the plurality of microparticles, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises non-encapsulated perfume compounds.

[0128] Shampoo or Shower Gel

[0129] In some embodiments, the consumer care composition is in the form of a shampoo or shower gel composition. In some such embodiments, the consumer care composition comprises a shampoo or shower gel active base and a plurality of microparticles according to any of the embodiments set forth above.

[0130] Any suitable shampoo or shower gel active base can be used. For example, in some embodiments, the shampoo or shower gel active base includes anionic surfactant such as sodium alkylether sulfate, ammonium alkylether sulfates, alkylamphoacetate, cocamidopropyl betaine, cocamide MEA, alkylglucosides and aminoacid based surfactants and mixtures thereof, as well as various surfactants, hydrophilic organic solvents, and water. In some embodiments, the consumer care composition comprises the shampoo or shower gel active base in an amount ranging from 85% by weight and 99.95% by weight, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises the plurality of microparticles in an amount ranging from 0.1% by weight to 15 % by weight, or from 0.2% by weight to 5% by weight, of the plurality of microparticles, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises non-encapsulated perfume compounds.

[0131] Rinse-Off Conditioner

[0132] In some embodiments, the consumer care composition is in the form of a rinse-off conditioner composition. In some such embodiments, the consumer care composition comprises a rinse-off conditioner active base and a plurality of microparticles according to any of the embodiments set forth above.

[0133] Any suitable rinse-off conditioner active base can be used. For example, in some embodiments, the rinse-off conditioner active base includes cetyltrimonium chloride, stearyl trimonium chloride, benzalkonium chloride, behentrimonium chloride and mixture thereof, as well as various surfactants, hydrophilic organic solvents, and water. In some embodiments, the consumer care composition comprises the rinse-off conditioner active base in an amount ranging from 85% by weight and 99.95% by weight, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises the plurality of microparticles in an amount ranging from 0.1% by weight to 15 % by weight, or from 0.2% by weight to 5% by weight, of the plurality of microparticles, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises non-encapsulated perfume compounds.

[0134] Solid Scent Booster

[0135] In some embodiments, the consumer care composition is in the form of a solid scent booster composition. In some such embodiments, the consumer care composition comprises a solid scent booster active base and a plurality of microparticles according to any of the embodiments set forth above.

[0136] Any suitable solid scent booster active base can be used. For example, in some embodiments, the solid scent booster active base includes urea, sodium chloride, sodium sulphate, sodium acetate, zeolite, sodium carbonate, sodium bicarbonate, clay, talc, calcium carbonate, magnesium sulfate, gypsum, calcium sulfate, magnesium oxide, zinc oxide, titanium dioxide, calcium chloride, potassium chloride, magnesium chloride, zinc chloride, saccharides such as sucrose, mono-, di-, and polysaccharides and derivatives such as starch, cellulose, methyl cellulose, ethyl cellulose, propyl cellulose, polyols / sugar alcohols such as sorbitol, maltitol, xylitol, erythritol, and isomalt, PEG, PVP, citric acid or any water soluble solid acid, fatty alcohols or fatty acids and mixtures thereof, as well as various surfactants, hydrophilic organic solvents, and water. In some embodiments, the consumer care composition comprises the solid scent booster active base in an amount ranging from 85% by weight and 99.95% by weight, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises the plurality of microparticles in an amount ranging from 0.1 % by weight to 15 % by weight, or from 0.2% by weight to 5% by weight, of the plurality of microparticles, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises non-encapsulated perfume compounds.

[0137] Liquid Scent Booster

[0138] In some embodiments, the consumer care composition is in the form of a liquid scent booster composition. In some such embodiments, the consumer care composition comprises a liquid scent booster active base and a plurality of microparticles according to any of the embodiments set forth above.

[0139] Any suitable liquid scent booster active base can be used. For example, in some embodiments, the liquid scent booster active base includes ethoxylated aliphatic alcohols, POE / PPG (polyoxyethylene and polyoxypropylene) ethers, mono and polyglyceryl esters, sucrose ester compounds, polyoxyethylene hydroxylesters, alkyl polyglucosides, amine oxides, alcohols, salts and esters of carboxylic acids, salts and esters of hydroxyl carboxylic acids, fatty acids, fatty acid salts, glycerol fatty acids, and mixtures thereof, as well as various surfactants, hydrophilic organic solvents, and water. In some embodiments, the consumer care composition comprises the liquid scent booster active base in an amount ranging from 85% by weight and 99.95% by weight, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises the plurality of microparticles in an amount ranging from 0.1% by weight to 15 % by weight, or from 0.2% by weight to 5% by weight, of the plurality of microparticles, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises nonencapsulated perfume compounds.

[0140] Hair Colorant

[0141] In some embodiments, the consumer care composition is in the form of a hair colorant composition. In some such embodiments, the consumer care composition comprises a hair colorant active base and a plurality of microparticles according to any of the embodiments set forth above.

[0142] Any suitable hair colorant active base can be used. For example, in some embodiments, the hair colorant active base includes oxidizing agents, an alkakine agent, dye precursors, coupling agents, as well as various surfactants, hydrophilic organic solvents, and water. In some embodiments, the consumer care composition comprises the hair colorant active base in an amount ranging from 85% by weight and 99.95% by weight, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises the plurality of microparticles in an amount ranging from 0.1% by weight to 15 % by weight, or from 0.2% by weight to 5% by weight, of the plurality of microparticles, based on the total weight of the consumer care composition. In some embodiments, the consumer care composition comprises non-encapsulated perfume compounds.

[0143] Perfuming Composition

[0144] In some embodiments, the consumer care composition is in the form of a perfuming composition. In some such embodiments, the consumer care composition comprises perfume compounds, ethanol, and a plurality of microparticles according to any of the embodiments set forth above. In some embodiments, the consumer care composition comprises the plurality of microparticles in an amount ranging from 0.1% by weight to 30 % by weight, or from 0.2% by weight to 20% by weight, of the plurality of microparticles, based on the total weight of the consumer care composition. In some embodiments, the perfume compounds are present in an amount ranging from 0% by weight to 40% by weight, or from 3% by weight to 40% by weight, based on the total weight of the consumer care composition. In some embodiments, the ethanol is present in an amount ranging from 20% by weight to 90% by weight, or from 40% by weight to 90% by weight, based on the total weight of the consumer care composition.

[0145] EXAMPLES

[0146] To further illustrate this invention, examples are set forth in the attached appendix sheets. The examples should not, of course, be construed as specifically limiting the invention. Variations of these examples within the scope of the claims are within the purview of one skilled in the art and are considered to fall within the scope of the invention as described and claimed herein. The reader will recognize that the skilled artisan, armed with the present disclosure, and skill in the art is able to prepare and use the invention without exhaustive examples.

[0147] Example 1 - Calcium imaging protocol to assess cellular adaptation properties of volatile chemicals

[0148] Eukaryotic cells were transfected with the odorant receptor (OR) of interest and the G- protein Gais. Upon OR activation, Gais-mediated pathways lead to an increase in calcium that can be monitored using a calcium indicator, such as fura-2 AM. Activation is seen as an upwards deflection in the fluorescence-time trace. Cells are mounted on a chamber in a microscope and subjected to constant saline flow. At the start of the protocol, there is a negative control injection of dimethylsulfoxide (dmso), the solvent used to dissolve the odorants. Any cells that respond to the negative control injection are discarded from the analysis. Following, brief odorant pulse(s) (i.e. the “pre-probe(s)”) are given to assess baseline odor responsiveness. Following is a “soak” period where the cells are continuously bathed in a substance of interest. After return to saline perfusion, brief pulse(s) (i.e. the “post-probe(s)”) are given. The post-probe(s) and pre-probe(s) should be of the same odorant identity and concentration to evaluate the change in odor responsiveness from baseline. Finally, a positive control injection is used to distinguish between cells that fail to recover from adaptation and those that are no longer viable at the end of the recording. Cells that fail to respond to the positive control are not considered for further analyses. One measure of adaptation is to take the ratio of a post-probe’s height relative to a pre-probe (or average of the pre-probes if more than one was given) height to return the percent activity remaining for that timepoint. This percent activity remaining is then subtracted from 1 to get an “Adaptation Index”. In the Adaptation Index, “1” denotes complete adaptation and “0” denotes no adaptation. Thus, a high adaptation index denotes a high amount of adaptation.

[0149] FIG. 1 shows a fluorescence-time trace of calcium levels in heterologous cells transfected with the odorant receptor OR1A1 and Gais, and visualized using fura-2 AM. The odorant probes are 100pM 2-(2-methylpropyl)quinoline, the soak duration is 5 minutes with a 5 min washout in saline before the first post-probe, and the positive control is 10uM ATP. Cell 1 received a saline soak. It shows little decrement in the height of the post-probe peaks, indicating no adaptation.

[0150] FIG. 2 shows the Adaptation Index for the population of cells treated with a saline soak as in panel FIG. 1. There was a negative average Adaptation Index, indicating that for the first post-probe there was not only no adaptation, but rather a small increase in height above the average baseline fluorescence signal.

[0151] FIG. 3 shows the odorant receptor is OR1A1, the odorant probes are 100 pM 2-(2-methylpropyl)quinoline, the odorant soak duration is 5 minutes in 100 pM 2-(2-methylpropyl)quinoline with a 5 min washout in saline before the first post-probe, and the positive control is 10 pM ATP. After receiving an odorant soak, Cell 2 shows very reduced height of the post-probe peaks, indicating strong adaptation at the first two post-probe timepoints. Later post-probe responses recover in a graded manner but plateau to a level of incomplete recovery to the pre-probe baseline levels, indicating residual adaptation.

[0152] FIG. 4 shows the Adaptation Index for the population of cells treated with the odor soak as in FIG. 3, there is a large positive average Adaptation Index, indicating a lot of adaptation of the response to the odorant probe at the first measured post-probe timepoint.

[0153] Example 2 - Cyclic Adenosine Monophosphate imaging protocol to assess cellular adaptation properties of volatile chemicals

[0154] Eukaryotic cells were transfected with the odorant receptor (OR) of interest and the G-protein Goif- Upon OR activation, Goif-mediated pathways lead to an increase in cAMP that can be monitored using a cAMP indicator, such as the Green Upward cADDis indicator. Activation is seen as an upwards deflection in the fluorescence-time trace. The general protocol is as described in Example 1 and applied here to another human receptor. FIG. 5 shows a fluorescence-time trace of cAMP levels in a heterologous cell transfected with the odorant receptor OR8D1 and Goif, and visualized using Green Upward cADDis. In this example, the odorant receptor is OR8D1. The pre-probe is 3pM 4-hydroxy- 2,3-dimethyl-2H-furan-5-one (grey arrowheads). Although 30pM 4-hydroxy-2,3-dimethyl- 2H-furan-5-one pulses (black arrowheads) were also given before the soak, 3pM 4-hydroxy- 2,3-dimethyl-2H-furan-5-one is considered the pre-probe as it is the same odor identity and concentration as used for the post-probes. The soak was 10 minutes with 8 min washout in saline before the first post-probe application. The positive control is 10pM forskolin. In FIG. 5, cell 3, soaked in saline, shows little decrement in the height of the post-probe peaks, indicating no adaptation.

[0155] FIG. 6 shows a fluorescence-time trace of cAMP levels in heterologous cell transfected with the odorant receptor OR8D1 and Goif, and visualized using Green Upward cADDis. Cell 4, soaked in 30pM 4-hydroxy-2,3-dimethyl-2H-furan-5-one, has reduced post-probe peaks, especially at early timepoints. Some residual adaptation is also noted for this cell. The average Adaptation Index for the population at each time point is summarized in FIG. 9.

[0156] Example 3 - Adaptation is not readily predicted from cell sensitivity to the odorant

[0157] Dose response curves can be generated by brief injections of an odor A or an odor B on eukaryotic cells transfected with OR1A1 and Gais, and imaged using the calcium-indicator fura- 2 AM. As highlighted by the arrows in FIG. 7, 10pM of odor A (2-(2-methylpropyl)quinoline, black triangles) and 300pM of odor B ((+-)-6-pentyltetrahydro-2H-pyran-2-one, white circles) elicit very similar levels of activity when applied acutely. FIG. 8 shows a comparison between the adaptation elicited by the two compounds in a calcium-imaging assay. Heterologous cells were transfected with the odorant receptor OR1A1 and Gais. 100pM 2-(2-methylpropyl)- quinoline was used for the probes. The soaks were for 15 minutes of the odorant and concentration indicated. Bar graphs are the average Adaptation Index over the cell population, where 1 indicates complete adaptation. Although the odorants and concentrations were selected to be equi-active, the extent of adaptation is not identical. This unexpected finding underscores that cellular adaptation needs to be measured empirically and independently of activity measures.

[0158] Example 4 - Self-adaptation at various odorant receptors

[0159] Examples of monitoring adaptation and recovery over time in a cAMP-imaging assay. Heterologous cells were transfected with the indicated odorant receptor and Goif. The same odorants were tested as both the probe and soak, making these examples of self-adaptation. All soaks were for 10 minutes. FIG. 9 shows the average Adaptation Index over the cell population as bar graphs, where 1 indicates complete adaptation. In all of the tested conditions, there is a gradual decrease in adaptation over time. Many conditions exhibit residual adaptation at the latest time point tested (t = 26 min post end of soak). Each test condition is shown separately (panels A to I): A: OR51 I2: (E)-3-methylhex-2-enoic acid , 300pM probe, 300pM soak; B: 0R51V1: ((Z)-4-methylhex-3-enoic acid + (E)-4-methylhex-3-enoic acid), 10 pM probe, 10 pM soak; C: OR10J5: 3-(4-hydroxy-4-methylpentyl)cyclohex-3-ene-1- carbaldehyde, 30pM probe, 300pM soak; D: OR1A1: 2-(2-methylpropyl)quinoline, 3pM probe, 3pM soak; E: OR10G4: 3-ethoxy-4-hydroxybenzaldehyde, 100pM probe, 100pM soak; F: OR5K1 : 2-methoxy-3-propan-2-ylpyrazine, 100pM probe, 100pM soak; G: OR10K1: nonanal, 100pM probe, 100pM soak; H: OR8D1 : 4-hydroxy-2,3-dimethyl-2H-furan-5-one, 3pM probe, 30pM soak; I: OR11A1: 4,8a-dimethyl-1,2,3,4,5,6,7,8-octahydronaphthalen-4a-ol, 30pM probe, 10pM soak. Panels A and B are Class I ORs; the remaining panels are Class II ORs.

[0160] Example 5: Cross-adaptation at various odorant receptors

[0161] Examples of monitoring adaptation and recovery over time in a cAMP-imaging assay. Heterologous cells were transfected with the odorant receptor indicated and Goif- Different odorants were tested as the probe and soak, making these examples of cross-adaptation. Both odorants are individually activators of the receptor, as confirmed when given as pulses on other cells (data not shown). All soaks were for 10 minutes. FIG. 10 shows the average Adaptation Index over the cell population as bar graphs, where 1 indicates complete adaptation. Crossadaptation with distinct agonists show a gradual decrease in adaptation over time. Each test condition is shown separately (panels A to D): A: OR10G4, probe = 30 M (2-methoxy- 4-methylphenol + 3-methylphenol + 2,3 / 2,4 / 2,5-dimethylphenol + 4-ethyl-2-methoxyphenol + 2-hydroxy-3-methyl-2-cyclopenten-1-one); soak = 100pM 3-ethoxy-4-hydroxybenzaldehyde; B: OR8D1, probe = 10pM 4-hydroxy-2,3-dimethyl-2H-furan-5-one; soak = 30pM ((3Z)- 3-propylidene-2-benzofuran-1(3H)-one + (3E)-3-propylidene-2-benzofuran-1(3H)-one); C: OR1A1, probe = 100pm (+-)-(e)-8-decen-5-olide + (+-)-(z)-8-decen-5-olide; soak = 3 pM 2-(2-methylpropyl)quinoline; D: OR10K1 , probe = 3pM (Z)-dodec-4-enal; soak = 100 pM nonanal.

[0162] Example 6: Cross-adaptation at various odorant receptors by antagonists

[0163] The general stimulation protocol is as indicated in Example 1. FIG. 11 shows OR8D1 activated by its agonist odor probe 4-hydroxy-2,3-dimethyl-2H-furan-5-one at 10pM. The soak odorant ((1-[(1 RS,2RS)-1 ,2,8,8-tetramethyl-1 ,2,3,4,5,6,7,8-octahydro-2-naphthalenyl]ethanone + 1-((2RS,3RS)-2,3,8,8-tetramethyl-1 ,2,3,4,5,6,7,8-octahydronaphthalen-2-yl)ethanone + 1-[(2RS,3RS,8aRS)-2,3,8,8-tetramethyl-1 ,2,3,5,6,7,8,8a-octahydro-2-naphthalenyl]ethanone + 1-[(1 RS,2RS,8aSR)-1 ,2,8,8-tetramethyl-1 ,2,3,5,6,7,8,8a-octahydro-2-naphthalenyl]ethanone + 1-[(2RS,3RS,8aRS)-2,3,8,8-tetramethyl-1 ,2,3,4,6,7,8,8a-octahydro-2-naphthalenyl]ethenone), 300pM) is an antagonist of OR8D1 , as previously demonstrated by inhibition dose-response experiments (data not shown). Because different odorants were tested as the probe and soak, this is an example of cross-adaptation. Unexpectedly, despite a lack of activation by (1-[(1 RS,2RS)-1 ,2,8,8-tetramethyl-1,2,3,4,5,6,7,8-octahydro-2-naphthalenyl]ethanone + 1-((2RS,3RS)-2,3,8,8-tetramethyl-1,2,3,4,5,6,7,8-octahydronaphthalen-2-yl)ethanone + 1-[(2RS,3RS,8aRS)-2,3,8,8-tetramethyl-1 ,2,3,5,6,7,8,8a-octahydro-2-naphthalenyl]ethanone + 1-[(1 RS,2RS,8aSR)-1 ,2,8,8-tetramethyl-1 ,2,3,5,6,7,8,8a-octahydro-2-naphthalenyl]ethanone + 1-[(2RS,3RS,8aRS)-2,3,8,8-tetramethyl-1 ,2,3,4,6,7,8,8a-octahydro-2-naphthalenyl]ethenone), during the soak period, the post-probe responses are markedly smaller in height while the positive control remains large, indicating adaptation and not general cell rundown. Unlike what was observed for 0R8D1 agonist self-adaptation (FIG. 9 panel H), or agonist cross-adaptation (FIG. 10 panel B) the adaptation induced by the antagonist is relatively stable throughout the imaging period.

[0164] FIG. 12 shows additional examples of monitoring adaptation and recovery over time in a cAMP-imaging assay. Heterologous cells were transfected with the odorant receptor (OR) indicated and Goif- Two different odorants were tested as the probe and soak, making these examples of cross-adaptation. In all examples, the soak was comprised of an antagonist previously characterized by inhibition dose-response experiments (data not shown). All soaks were for 10 minutes. Bar graphs are the average Adaptation Index over the cell population, where 1 indicates complete adaptation. Each test condition is shown separately (panels A to E): A: OR8D1, probe = 10pM 4-hydroxy-2,3-dimethyl-2H-furan-5-one; soak = 300pM (1-[(1 RS,2RS)-1 ,2,8,8-tetramethyl-1,2,3,4,5,6,7,8-octahydro-2-naphthalenyl]ethanone + 1-((2RS,3RS)-2,3,8,8-tetramethyl-1,2,3,4,5,6,7,8-octahydronaphthalen-2-yl)ethanone + 1-[(2RS,3RS,8aRS)-2,3,8,8-tetramethyl-1 ,2,3,5,6,7,8,8a-octahydro-2-naphthalenyl]ethanone + 1-[(1 RS,2RS,8aSR)-1 ,2,8,8-tetramethyl-1 ,2,3,5,6,7,8,8a-octahydro-2-naphthalenyl]ethanone + 1-[(2RS,3RS,8aRS)-2,3,8,8-tetramethyl-1 ,2,3,4,6,7,8,8a-octahydro-2-naphthalenyl]ethenone),; B: OR10J5, probe = 30pM 3-(4-hydroxy-4-methylpentyl)cyclohex-3-ene-1-carbaldehyde; soak = 300pM (+-)-(4Z,8E)-1 ,5,8-trimethyl-13-oxabicyclo[10.1.0]trideca-4,8-diene + (+-)-(4Z,8E)-1,4,8- trimethyl-13-oxabicyclo[10.1.0]trideca-4,8-diene; C: OR11A1 , probe = 30pM 4,8a-dimethyl-

[0165] 1.2.3.4.5.6.7.8-octahydronaphthalen-4a-ol; soak = 300pM (2E,6Z)-nona-2,6-dienal; D: OR51 I2, probe = 300pM (E)-3-methylhex-2-enoic acid; soak = 300pM 2,6,6,8-tetramethyl-9- oxatetracyclo[5.4.1.01,5.08,10]dodecane; E: OR51V1, probe = 10pM ((Z)-4-methylhex-3-enoic acid + (E)-4-methylhex-3-enoic acid), antagonist soak = 300pM ((+-)-(4Z,8E)-1 ,5,8-trimethyl-13- oxabicyclo[10.1 ,0]trideca-4,8-diene + (+-)-(4Z,8E)-1 ,4,8-trimethyl-13-oxabicyclo[10.1.0]trideca-

[0166] 4.8-diene). Panels D and E are Class I ORs; remaining panels are Class II ORs.

[0167] Example 7 - Cellular adaptation correlates with sensory adaptation to odorants or to mixtures thereof

[0168] A comparison between experimental cellular adaptation data and experimental sensory adaptation shows strong correlation between the two approaches and assess the predictive nature of the cellular approach for both individual compounds or mixtures such as perfumery accords. In FIG. 13, heterologous cells were transfected with the odorant receptor OR10K1 and Goif. The same odorants were tested as the probe and soak, making these examples of selfadaptation. The soaks were for 10 minutes. Bar graphs are the average Adaptation Index over the cell population, where 1 indicates complete adaptation. More adaptation was elicited by odor C (black bars) than odor D (white bars) at all time points. The corresponding sensory adaptation measurements are also shown. Human subjects rated the perceived intensity of each odorant before and after a 3-minute continuous exposure to that odorant on an olfactometer. Ratings were on a 4-point scale. The bar graph plots the reported intensity loss such that a larger bar indicates a greater loss, indicating more adaptation. The odorant C, which elicited more adaptation in the cellular assay, also elicited more adaptation in the sensory assay. The assay conditions were the following: odor C, black bars: (Z)-dodec-4-enal, probe = 10pM; soak = 100pM; odor D, white bars: nonanal, probe = 100pM; soak = 100pM.

[0169] In FIG. 14, heterologous cells were transfected with the odorant receptor OR5K1 and Goif. The same accords (small mixtures of odorants) were tested as the probe and soak, making these examples of self-adaptation. The soaks are of equal concentration and were for 10 minutes. Bar graphs are the average Adaptation Index over the cell population, where 1 indicates complete adaptation. Adaptation was stronger for accord A (black bars) than accord B (white bars) at all but the last tested time point. The corresponding sensory adaptation measurements are also shown. Human subjects rated the perceived intensity of each accord before and after a 3-minute continuous exposure to that accord on an olfactometer. Ratings were on a 4-point scale. The bar graph plots the reported intensity loss such that a larger bar indicates a greater loss, indicating more adaptation. Accord A, which elicited more adaptation in the cellular assay, also elicited more adaptation in the sensory assay. The assay conditions were the following: accord A, black bars: probe and soak = 1 :3000 vokvol ; accord B, white bars: probe and soak = 1 :3000 vokvol.

[0170] Example 8 - Adaptation levels can be adjusted by soak duration or concentration

[0171] This Example delineates how to monitor the impact of soak as a function of time and concentration in a calcium-imaging assay. FIG. 15 shows heterologous cells transfected with the odorant receptor OR1A1 and Gais. The odorant 2-(2-methylpropyl)quinoline at 100pM was used for both the probe and soak, making this an example of self-adaptation. Bar graphs are the average Adaptation Index over the cell population, where 1 indicates complete adaptation. Increasing the odorant soak time leads to increasing adaptation. FIG. 16 shows heterologous cells transfected with the odorant receptor OR11A1 and Goif. All were probed and soaked in the same odorant, making this an example of self-adaptation. All probes were with 30pM 4,8a-dimethyl-1 ,2,3,4,5,6,7,8-octahydronaphthalen-4a-ol, and soaks were of various concentrations of 4,8a-dimethyl-1 ,2,3,4,5,6,7,8-octahydronaphthalen-4a-ol for 10 minutes. The Y-axis shows the average Adaptation Index over the cell population, where 1 indicates complete adaptation. Higher concentrations of the soak odorant lead to greater adaptation at any matched time point during the imaging period. By reading across the graph horizontally, the curves provide a way of estimating when comparable recovery from adaptation can be achieved. For example, to recover to the same level of adaptation as 8 minutes after a moderate 4,8a-dimethyl-1,2,3,4,5,6,7,8-octahydronaphthalen-4a-ol soak (medium grey triangles), it would take about an additional 12 minutes (to 20 minutes post soak total) had a high 4,8a-dimethyl-1 ,2,3,4,5,6,7,8-octahydronaphthalen-4a-ol soak (black squares) been given. As a result, similar adaptation levels can be obtained from distinct conditions providing a quantitative approach to adaptation modulation. FIG. 17 shows the monitoring the interplay of soak concentration and duration in a calcium-imaging assay. Heterologous cells were transfected with the odorant receptor OR1A1 and Gais. Both probes and soaks were with 2-(2-methylpropyl)quinoline, making this an example of self-adaptation. Bar graphs are the average Adaptation Index over the cell population, where 1 indicates complete adaptation. Adaptation was measured at 5 minutes post end of soak. Decreasing the soak odorant concentration allows for a longer soak time before a similar level of adaptation is reached.

Claims

CLAIMS1. A method of identifying a substance that is susceptible to olfactory adaptation, the method comprising:(a) introducing an olfactory receptor ligand to a plurality of olfactory receptors, wherein the olfactory receptor ligand is an olfactory receptor agonist;(b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand;(c) reintroducing the olfactory receptor ligand to the plurality of olfactory receptors, such that the olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes;(d) following an optional period during which the olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the olfactory receptor ligand to the plurality of olfactory receptors; and(e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor ligand.

2. The method of any one of claim 1, further comprising: (f) identifying an adaptation- susceptible olfactory receptor agonist that is susceptible to olfactory adaptation based on a difference between the first measurement and the second measurement.

3. The method of claim 2, further comprising: (g) deselecting the adaptation-susceptible olfactory receptor agonist as an adaptation-resistant substance.

4. A method of identifying a substance that is resistant to olfactory adaptation, the method comprising:(a) introducing an olfactory receptor ligand to a plurality of olfactory receptors, wherein the olfactory receptor ligand is an olfactory receptor agonist;(b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand;(c) reintroducing the olfactory receptor ligand to the plurality of olfactory receptors, such that the olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes;(d) following an optional period during which the olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the olfactory receptor ligand to the plurality of olfactory receptors; and(e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor ligand.

5. The method of any one of claim 4, further comprising: (f) identifying an adaptation-resistant olfactory receptor agonist that is resistant to olfactory adaptation based on a difference between the first measurement and the second measurement.

6. The method of claim 5, further comprising: (g) selecting the adaptation-resistant olfactory receptor agonist as an adaptation-resistant substance.

7. A method of identifying a substance that is susceptible to olfactory adaptation, the method comprising:(a) introducing an olfactory receptor ligand to a plurality of olfactory receptors, wherein the olfactory receptor ligand is an agonist of the plurality of olfactory receptors;(b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand;(c) introducing a second olfactory receptor ligand to the plurality of olfactory receptors, such that the second olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes, wherein the second olfactory receptor ligand is an antagonist of the plurality of olfactory receptors;(d) following an optional period during which the olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the olfactory receptor ligand to the plurality of olfactory receptors; and(e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor ligand.

8. The method of any one of claim 7, further comprising: (f) identifying an adaptation- susceptible olfactory receptor antagonist that is susceptible to olfactory adaptation based on a difference between the first measurement and the second measurement.

9. The method of claim 8, further comprising: (g) deselecting the adaptation-susceptible olfactory receptor antagonist as an adaptation-resistant substance.

10. A method of identifying a substance that is resistant to olfactory adaptation, the method comprising:(a) introducing an olfactory receptor ligand to a plurality of olfactory receptors, wherein the olfactory receptor ligand is an agonist of the plurality of olfactory receptors;(b) conducting a first measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following introducing the olfactory receptor ligand;(c) introducing a second olfactory receptor ligand to the plurality of olfactory receptors, such that the second olfactory receptor ligand remains in contact with the plurality of olfactory receptors for a period of time ranging from 1 minute to 120 minutes, wherein the second olfactory receptor ligand is an antagonist of the plurality of olfactory receptors;(d) following an optional period during which the olfactory receptor ligand is not in contact with the plurality of olfactory receptors, reintroducing the olfactory receptor ligand to the plurality of olfactory receptors; and(e) conducting a second measurement of a parameter indicative of signaling activity of the plurality of olfactory receptors over a period of time immediately following reintroducing the olfactory receptor ligand.

11. The method of any one of claim 10, further comprising: (f) identifying an adaptation- susceptible olfactory receptor antagonist that is resistant to olfactory adaptation based on a difference between the first measurement and the second measurement.

12. The method of claim 11, further comprising: (g) deselecting the adaptation-resistant olfactory receptor antagonist as an adaptation-resistant substance.

13. The method of any one of claims 1 to 12, wherein the olfactory receptor is selected from the group consisting of: the human OR1A1, OR8D1, OR51 I2, OR51V1 , OR10J5, OR10G4, OR5K1 , OR10K1, and OR11A1 olfactory receptors, and a polypeptide having an amino acid sequence that is at least 70%, or at least 75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97%, or at least 99%, equivalent thereto.

14. Use of an adaptation-resistant substance identified or selected by the method of claim 5, 6, 11, or 12 to provide a long-lasting fragrance to a fragranced article.

15. The use of claim 14, wherein the fragranced article is a fiber, such as a synthetic or natural fiber.

16. The use of claim 14, wherein the fragranced article is a consumer care product, such as a household cleaning product, an air freshener, a commercial cleaning product, a dish detergent, a laundry detergent, a fabric softener, a scent booster, or a personal care product such as a shower gel, a shampoo, a hair conditioner, a hair-styling product, a skin care product, a cosmetic product, a deodorant, an antiperspirant, or a self-tanning product.

17. The use of claim 14, wherein the fragranced article is a perfuming composition, such as a fine fragrance composition.

18. A method of providing a long-lasting fragrance to a fragranced article, the method comprising introducing an adaptation-resistant substance identified or selected by the method of claim 5, 6, 11 , or 12 to the fragranced article.

19. A method of providing a long-lasting fragrance to a fragranced article, the method comprising removing an adaptation-susceptible substance identified or selected by the method of claim 2, 3, 8, or 9 to the fragranced article.

20. The method of claim 18 or 19, wherein the fragranced article is a fiber, such as a synthetic or natural fiber.

21. The method of claim 18 or 19, wherein the fragranced article is a consumer care product, such as a household cleaning product, a commercial cleaning product, a dish detergent, a laundry detergent, a fabric softener, a scent booster, a shower gel, a shampoo, a hair conditioner, a hair-styling product, a skin care product, a cosmetic product, a deodorant, an antiperspirant, or a self-tanning product.

22. The method of claim 18 or 19, wherein the fragranced article is a perfuming composition, such as a fine fragrance composition.

23. A perfuming composition for providing long-lasting fragrance, which comprises an adaptation-resistant substance identified or selected by the method of claim 5, 6, 11, or 12.

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