Compositions and Methods for Detecting Oral Neoplasms

Abstract

Methods and kits for assessing the presence of, and for detecting cancer, notably oral cancer, are disclosed. Such methods and kits use one or more lectins operably linked to a fluorophore, wherein the lectin binds differentially to cancerous and non-cancerous tissues, and wherein the fluorophore facilitates visualizing the differential binding. The lectins are applied to the oral mucosa of a subject, the fluorophore is exposed to light, and cancerous regions of the mucosa are visualized. In certain embodiments, the lectin specifically binds to one or more of a β-galactoside, an α- or β-N-acetylglucosamine, or a sialic acid moiety.

The invention claimed is: 
     
         1 . A method of in vivo imaging an oral mucosal surface of a subject to assess aberrant glycosylation patterns; the method comprising the steps of:
 (a) topically applying one or more lectin-fluorophore conjugates consisting of a lectin covalently bonded to a fluorescent label directly to the oral mucosal surface of the subject in vivo, wherein the one or more lectins target saccharides of β-galatoside, α- or β-N-acetylglucosamine, and/or sialic acid;   (b) exciting the one or more lectin-fluorophore conjugates in vivo; and   (c) observing the emitted fluorescent signal in vivo, wherein an observable fluorescence difference between adjacent regions is indicative of aberrant glycosylation;   
       wherein the fluorescence difference is observed without extracting tissue from the subject. 
     
     
         2 . The method of  claim 1 , wherein the step of topically applying the one or more lectin-fluorophore conjugates directly to the oral mucosal surface of the subject in vivo is performed by one or more of:
 using a cotton swab or similar device, applying a solution comprising the one or more lectin-fluorophore conjugates as an oral rinse, applying a solution comprising the one or more lectin-fluorophore conjugates as a lotion, injecting a solution comprising the one or more lectin-fluorophore conjugates onto the oral mucosal surface, mechanically agitating a solution comprising the one or more lectin-fluorophore conjugates, or ultrasonically agitating a solution comprising the one or more lectin-fluorophore conjugates.   
     
     
         3 . The method of  claim 1 , further comprising the step of directly applying in vivo to at least a portion of the oral mucosal surface a pretreatment solution that facilitates lectin binding, wherein the step of directly applying the pretreatment solution is performed prior to topically applying the one or more lectin-fluorophore conjugates to the oral mucosal surface. 
     
     
         4 . The method of  claim 1 , wherein the one or more lectin-fluorophore conjugates are topically applied using a solution that further comprises a buffering agent. 
     
     
         5 . The method of  claim 1 , wherein the one or more lectin-fluorophore conjugates are topically applied using a solution that further comprises one or more permeation enhancers. 
     
     
         6 . The method of  claim 1 , wherein the step of exciting the one or more lectin-fluorophore conjugates comprises one or more of:
 waiting to perform the observing step to allow time for the lectin-fluorophore conjugates to penetrate the oral mucosal surface, rinsing the oral mucosal surface with a solution to clear any unbound lectin-fluorophore conjugate present, serially rinsing the oral mucosal surface to clear any unbound lectin-fluorophore conjugate present, mechanically agitating the oral mucosal surface to clear any unbound lectin-fluorophore conjugate present, ultrasonically agitating the oral mucosal surface to clear any unbound lectin-fluorophore conjugate present, swabbing or rubbing the oral mucosal surface to clear any unbound lectin-fluorophore conjugate present, or illuminating the oral mucosal surface with visible, ultraviolet, or near infrared light.   
     
     
         7 . The method of  claim 1 , wherein the step of observing the emitted fluorescent signal comprises one or more of:
 visualizing the oral mucosal surface by the naked eye, visualizing the oral mucosal surface through a handheld filter or filter glasses, using a camera, using an image processing device and software, or using an optical element or elements selected from a dichroic mirror, a neutral density filter, a polarizer, a microscope, or a combination thereof.   
     
     
         8 . A method of in vivo imaging a mucosal epithelium surface of a subject to assess aberrant glycosylation patterns; the method comprising the steps of:
 (a) topically applying one or more lectin-fluorophore conjugates consisting of a lectin covalently bonded to a fluorescent label directly to the mucosal epithelium surface of the subject in vivo, wherein the lectin is selected from the group consisting of wheat germ agglutinin lectin (WGA), peanut agglutinin lectin (PNA), and  griffonia simplicifolia  lectin (GS-II);   (b) exciting the one or more lectin-fluorophore conjugates in vivo; and   (c) observing the emitted fluorescent signal in vivo, wherein an observable fluorescence difference between adjacent regions is indicative of aberrant glycosylation;   
       wherein the fluorescence difference is observed without extracting tissue from the subject. 
     
     
         9 . The method of  claim 8 , wherein the mucosal epithelium is a keratinized or non-keratinized stratified squamous epithelium. 
     
     
         10 . The method of  claim 8 , further comprising the step of directly applying in vivo to at least a portion of the mucosal epithelium surface a pretreatment solution that facilitates lectin binding, wherein the step of directly applying the pretreatment solution is performed prior to topically applying the one or more lectin-fluorophore conjugates to the mucosal epithelium surface. 
     
     
         11 . The method of  claim 8 , wherein the one or more lectin-fluorophore conjugates are topically applied using a solution that further comprises a buffering agent. 
     
     
         12 . The method of  claim 8 , wherein the one or more lectin-fluorophore conjugates are topically applied using a solution that further comprises one or more permeation enhancers. 
     
     
         13 . The method of  claim 8 , wherein the step of exciting the one or more lectin-fluorophore conjugates comprises one or more of:
 waiting to perform the observing step to allow time for the lectin-fluorophore conjugates to penetrate the mucosal epithelium surface, rinsing the mucosal epithelium surface with a solution to clear any unbound lectin-fluorophore conjugate present, serially rinsing the mucosal epithelium surface to clear any unbound lectin-fluorophore conjugate present, mechanically agitating the mucosal epithelium surface to clear any unbound lectin-fluorophore conjugate present, ultrasonically agitating the mucosal epithelium surface to clear any unbound lectin-fluorophore conjugate present, swabbing or rubbing the mucosal epithelium surface to clear any unbound lectin-fluorophore conjugate present, or illuminating the mucosal epithelium surface with visible, ultraviolet, or near infrared light.   
     
     
         14 . The method of  claim 8 , wherein the step of observing the emitted fluorescent signal comprises one or more of;
 visualizing the mucosal epithelium surface by the naked eye, visualizing the mucosal epithelium surface through a handheld filter or filter glasses, using a camera, using an image processing device and software, or using an optical element or elements selected from a dichroic mirror, a neutral density filter, a polarizer, a microscope, or a combination thereof.   
     
     
         15 . A method of in vivo imaging a mucosal epithelium surface of a subject to assess aberrant glycosylation patterns; the method comprising the steps of:
 (a) topically applying one or more lectin-fluorophore conjugates consisting of a lectin covalently bonded to a fluorescent label directly to the mucosal epithelium surface of the subject in vivo, wherein the lectin(s) target(s) saccharides of β-galatoside, α- or β-N-acetylglucosamine, and/or sialic acid;   (b) exciting the one or more lectin-fluorophore conjugates in vivo; and   (c) observing the emitted fluorescent signal in vivo, wherein an observable fluorescence difference between adjacent regions is indicative of aberrant glycosylation;   
       wherein the fluorescence difference is observed without extracting tissue from the subject. 
     
     
         16 . The method of  claim 15 , wherein the mucosal epithelium is a keratinized or non-keratinized stratified squamous epithelium. 
     
     
         17 . The method of  claim 15 , further comprising the step of directly applying in vivo to at least a portion of the mucosal epithelium surface a pretreatment solution that facilitates lectin binding, wherein the step of directly applying the pretreatment solution is performed prior to topically applying the one or more lectin-fluorophore conjugates to the mucosal surface. 
     
     
         18 . The method of  claim 15 , wherein the one or more lectin-fluorophore conjugates are topically applied using a solution that further comprises a buffering agent. 
     
     
         19 . The method of  claim 15 , wherein the one or more lectin-fluorophore conjugates are topically applied using a solution that further comprises one or more permeation enhancers. 
     
     
         20 . The method of  claim 15 , wherein the step of exciting the one or more lectin-fluorophore conjugates comprises one or more of:
 waiting to perform the observing step to allow time for the lectin-fluorophore conjugates to penetrate the mucosal epithelium surface, rinsing the mucosal epithelium surface with a solution to clear any unbound lectin-fluorophore conjugate present, serially rinsing the mucosal epithelium surface to clear any unbound lectin-fluorophore conjugate present, mechanically agitating the mucosal epithelium surface to clear any unbound lectin-fluorophore conjugate present, ultrasonically agitating the mucosal epithelium surface to clear any unbound lectin-fluorophore conjugate present, swabbing or rubbing the mucosal epithelium surface to clear any unbound lectin-fluorophore conjugate present, or illuminating the mucosal epithelium surface with visible, ultraviolet, or near infrared light.   
     
     
         21 . The method of  claim 15 , wherein the step of observing the emitted fluorescent signal comprises one or more of;
 visualizing the mucosal epithelium surface by the naked eye, visualizing the mucosal epithelium surface through a handheld filter or filter glasses, using a camera, using an image processing device and software, or using an optical element or elements selected from a dichroic mirror, a neutral density filter, a polarizer, a microscope, or a combination thereof.   
     
     
         22 . The method of  claim 15 , wherein the step of topically applying the one or more lectin-fluorophore conjugates directly to the mucosal epithelium surface of the subject in vivo is performed by one or more of:
 using a cotton swab or similar device, applying a solution comprising the one or more lectin-fluorophore conjugates as an rinse, applying a solution comprising the one or more lectin-fluorophore conjugates as a lotion, injecting a solution comprising the one or more lectin-fluorophore conjugates onto the mucosal epithelium surface, mechanically agitating a solution comprising the one or more lectin-fluorophore conjugates, or ultrasonically agitating a solution comprising the one or more lectin-fluorophore conjugates.