Methods for improving safety of blood-brain barrier transport

Abstract

The present disclosure relates to compositions and methods for improving the safety of blood-brain barrier receptor-mediated blood-brain barrier transport.

What is claimed is: 
     
         1 . A method of transporting a compound across the blood-brain barrier in a subject comprising exposing an antibody which binds with low affinity to a blood-brain barrier receptor (BBB-R) coupled to a compound to the blood-brain barrier such that the antibody transports the compound coupled thereto across the blood-brain barrier, wherein reduction of red blood cell levels in the subject upon antibody administration to the subject is decreased or eliminated. 
     
     
         2 . A method of increasing exposure of the CNS of a subject to a compound, wherein the compound is coupled to an antibody which binds with low affinity to a BBB-R, thereby increasing the exposure of the CNS to the compound, and wherein reduction of red blood cell levels in the subject upon compound-coupled antibody administration to the subject is decreased or eliminated. 
     
     
         3 . A method of decreasing clearance of a compound administered to a subject, wherein the compound is coupled to an antibody which binds with low affinity to a BBB-R, such that the clearance of the compound is decreased, and wherein reduction of red blood cell levels in the subject upon compound-coupled antibody administration to the subject is decreased or eliminated. 
     
     
         4 . A method of increasing retention in the CNS of a compound administered to a subject, wherein the compound is coupled to an antibody which binds with low affinity to a BBB-R, such that the retention in the CNS of the compound is increased, and wherein reduction of red blood cell levels in the subject upon compound-coupled antibody administration to the subject is decreased or eliminated. 
     
     
         5 . A method of optimizing the pharmcokinetics and/or pharmacodynamics of a compound to be efficacious in the CNS in a subject, wherein the compound is coupled to an antibody which binds with low affinity to a BBB-R, and the antibody is selected such that its affinity for the BBB-R after coupling to the compound results in an amount of transport of the antibody conjugated to the compound across the BBB that optimizes the pharmacokinetics and/or pharmacodynamics of the compound in the CNS, wherein reduction of red blood cell levels in the subject upon compound-coupled antibody administration to the subject is decreased or eliminated. 
     
     
         6 . A method of treating a neurological disorder in a mammal comprising treating the mammal with an antibody that binds a BBB-R and is coupled to a compound, wherein the antibody has been selected to have a low affinity for the BBB-R and thereby improves CNS uptake of the antibody and coupled compound, and wherein reduction of red blood cell levels in the subject upon compound-coupled antibody administration to the subject is decreased or eliminated. 
     
     
         7 . The method of any of  claims 1 - 6 , wherein the BBB-R is selected from the group consisting of transferrin receptor (TfR), insulin receptor, insulin-like growth factor receptor (IGF receptor), low density lipoprotein receptor-related protein 8 (LRP8), low density lipoprotein receptor-related protein 1 (LRP1), glucose transporter 1 (Glut1) and heparin-binding epidermal growth factor-like growth factor (HB-EGF). 
     
     
         8 . The method of  claim 7 , wherein the BBB-R is TfR. 
     
     
         9 . The method of  claim 8 , wherein the red blood cells are immature red blood cells. 
     
     
         10 . The method of  claim 9 , wherein the immature red blood cells are reticulocytes. 
     
     
         11 . The method of  claim 10 , wherein reduction of reticulocyte levels is accompanied by acute clinical symptoms. 
     
     
         12 . The method of  claim 11 , wherein one or more properties of the antibody have been modified to reduce the impact of the antibody on reticulocyte levels and/or reduce the severity or presence of acute clinical symptoms in the subject or mammal. 
     
     
         13 . The method of  claim 12 , wherein the one or more properties are selected from the effector function of the antibody Fc region, the complement activation function of the antibody and the affinity of the antibody for the BBB-R. 
     
     
         14 . The method of  claim 13 , wherein the one or more properties are selected from the effector function of the antibody Fc region and the complement activation function of the antibody, and wherein the effector function or complement activation function has been reduced or eliminated relative to a wild-type antibody of the same isotype. 
     
     
         15 . The method of  claim 14 , wherein the effector function is reduced or eliminated by a method selected from reduction of glycosylation of the antibody, modification of the antibody isotype to an isotype that naturally has reduced or eliminated effector function, and modification of the Fc region. 
     
     
         16 . The method of  claim 15 , wherein the glycosylation of the antibody is reduced by a method selected from: production of the antibody in an environment that does not permit wild-type glycosylation; removal of carbohydrate groups already present on the antibody; and modification of the antibody such that wild-type glycosylation does not occur. 
     
     
         17 . The method of  claim 16 , wherein the antibody is produced in a non-mammalian cell production system, or where the antibody is produced synthetically. 
     
     
         18 . The method of  claim 16 , wherein the Fc region of the antibody comprises a mutation at position 297 such that the wild-type asparagine residue at that position is replaced with another amino acid that interferes with glycosylation at that position. 
     
     
         19 . The method of  claim 15 , wherein the effector function is reduced or eliminated by at least one modification of the Fc region. 
     
     
         20 . The method of  claim 19 , wherein the effector function or complement activation function is reduced or eliminated by deletion of all or a portion of the Fc region, or by engineering the antibody such that it does not include an Fc region or non-Fc region competent for effector function or complement activation function. 
     
     
         21 . The method of  claim 19 , wherein the modification is selected from: a point mutation of the Fc region to impair binding to one or more Fc receptors selected from the following positions: 238, 239, 248, 249, 252, 254, 265, 268, 269, 270, 272, 278, 289, 292, 293, 294, 295, 296, 297, 298, 301, 303, 322, 324, 327, 329, 333, 335, 338, 340, 373, 376, 382, 388, 389, 414, 416, 419, 434, 435, 437, 438, and 439; a point mutation of the Fc region to impair binding to C1q selected from the following positions: 270, 322, 329, and 321; eliminating some or all of the Fc region, and a point mutation at position 132 of the CH1 domain. 
     
     
         22 . The method of  claim 11 , wherein the dose amount and/or frequency of administration is modulated to reduce the concentration of antibody to which the red blood cells are exposed. 
     
     
         23 . The method of  claim 11 , wherein the antibody is modified to comprise pH-sensitive binding to the BBB-R. 
     
     
         24 . The method of  claim 11 , wherein a further compound is administered in addition to the antibody. 
     
     
         25 . The method of  claim 24 , wherein the further compound is responsible for or contributes to the lack of reduction of reticulocyte levels. 
     
     
         26 . The method of  claim 25 , wherein the further compound protects reticulocytes from antibody-related depletion or supports the growth, development, or reestablishment of reticulocytes. 
     
     
         27 . The method of  claim 26 , wherein the further compound is selected from erythropoietin (EPO), an iron supplement, vitamin C, folic acid, and vitamin B12. 
     
     
         28 . The method of  claim 26 , wherein the further compound is red blood cells or reticulocytes from the same or another subject. 
     
     
         29 . The method of  claim 13 , wherein the affinity of the antibody for the BBB-R is further decreased. 
     
     
         30 . The method of any of  claims 1 - 29 , further comprising the step of monitoring the subject for depletion of red blood cells. 
     
     
         31 . The method of any of  claims 1 - 30 , wherein the compound is a neurological disorder drug or an imaging agent. 
     
     
         32 . The method of any of  claims 2 - 30 , wherein the increase or decrease is measured relative to a wild-type antibody of the same isotype not having lowered affinity for the BBB-R. 
     
     
         33 . The method of any of  claims 1 - 30 , wherein the antibody does not impair the binding of the BBB-R to one or more of its native ligands. 
     
     
         34 . The method of any of  claims 1 - 30 , wherein the blood-brain barrier is in a mammal. 
     
     
         35 . The method of  claim 34 , wherein the mammal has a neurological disorder. 
     
     
         36 . The method of  claim 35 , wherein the neurological disorder is selected from the group consisting of Alzheimer's disease (AD), stroke, dementia, muscular dystrophy (MD), multiple sclerosis (MS), amyotrophic lateral sclerosis (ALS), cystic fibrosis, Angelman's syndrome, Liddle syndrome, Parkinson's disease, Pick's disease, Paget's disease, cancer, and traumatic brain injury. 
     
     
         37 . The method of  claim 34  wherein the mammal is a human. 
     
     
         38 . The method of any of  claims 1 - 30 , wherein the antibody has an IC50 for the BBB-R from about 1 nM to about 100 μM. 
     
     
         39 . The method of  claim 38 , wherein the IC50 is from about 5 nM to about 100 μM. 
     
     
         40 . The method of  claim 38 , wherein the IC50 is from about 50 nM to about 100 μM. 
     
     
         41 . The method of  claim 38 , wherein the IC50 is from about 100 nM to about 100 μM. 
     
     
         42 . The method of any of  claims 1 - 30 , wherein the antibody has an affinity for the BBB-R from about 5 nM to about 50 μM. 
     
     
         43 . The method of any of  claims 1 - 30 , wherein the antibody coupled to the compound has an affinity for the BBB-R from about 30 nM to about 30 μM. 
     
     
         44 . The method of any of  claims 1 - 30 , wherein the antibody coupled to the compound has an affinity for the BBB-R from about 30 nM to about 1 μM. 
     
     
         45 . The method of any of  claims 1 - 30 , wherein the antibody coupled to the compound has a dissociation half-life for the BBB-R from about 30 seconds to about 5 minutes, or from about 30 seconds to about 2 minutes. 
     
     
         46 . The method of  claim 8 , wherein the antibody does not inhibit the binding of TfR to transferrin. 
     
     
         47 . The method of any of  claims 1 - 30 , wherein the antibody coupled to the compound is administered at a therapeutic dose. 
     
     
         48 . The method of  claim 47 , wherein the therapeutic dose is BBB-R-saturating. 
     
     
         49 . The method of any of  claims 1 - 30 , wherein the antibody is a multispecific antibody and the compound optionally forms one portion of the multispecific antibody. 
     
     
         50 . The method of  claim 49  wherein the multispecific antibody comprises a first antigen binding site which binds the BBB-R and a second antigen binding site which binds a brain antigen. 
     
     
         51 . The method of  claim 50 , wherein the brain antigen is selected from the group consisting of: beta-secretase 1 (BACE1), Abeta, epidermal growth factor receptor (EGFR), human epidermal growth factor receptor 2 (HER2), tau, apolipoprotein E4 (ApoE4), alpha-synuclein, CD20, huntingtin, prion protein (PrP), leucine rich repeat kinase 2 (LRRK2), parkin, presenilin 1, presenilin 2, gamma secretase, death receptor 6 (DR6), amyloid precursor protein (APP), p75 neurotrophin receptor (p75NTR), and caspase 6. 
     
     
         52 . The method of  claim 51 , wherein the multispecific antibody binds both TfR and BACE1. 
     
     
         53 . The method of  claim 51 , wherein the multispecific antibody binds both TfR and Abeta. 
     
     
         54 . A method of improving the safety in a subject of an antibody that binds a BBB-R comprising modifying one or more properties of the antibody such that administration of the antibody decreases or eliminates reduction of red blood cell levels in the subject observed upon administration of the unmodified antibody. 
     
     
         55 . A method of making an antibody useful for transporting a compound across the BBB with improved safety comprising selecting an antibody specific for a blood-brain barrier receptor (BBB-R) that has a desirably low affinity for the BBB-R, and modifying one or more properties of the antibody such that administration of the antibody decreases or eliminates reduction of red blood cell levels in the subject observed upon administration of the unmodified antibody. 
     
     
         56 . The method of  claim 54  or  55  wherein the BBB-R is selected from the group consisting of transferrin receptor (TfR), insulin receptor, insulin-like growth factor receptor (IGF receptor), low density lipoprotein receptor-related protein 8 (LRP8), low density lipoprotein receptor-related protein 1 (LRP1), glucose transporter 1 (Glut1) and heparin-binding epidermal growth factor-like growth factor (HB-EGF). 
     
     
         57 . The method of  claim 56  wherein the BBB-R is transferrin receptor (TfR). 
     
     
         58 . The method of  claim 57 , wherein the red blood cells are reticulocytes. 
     
     
         59 . The method of  claim 58 , wherein reduction of reticulocyte levels is accompanied by acute clinical symptoms. 
     
     
         60 . The method of  claim 59 , wherein the one or more properties is selected from the effector function of the antibody Fc region, the complement activation function of the antibody and the affinity of the antibody for the BBB-R. 
     
     
         61 . The method of  claim 60 , wherein the one or more properties is selected from the effector function of the antibody Fc region and the complement activation function of the antibody, and wherein the effector function or the complement activation function has been reduced or eliminated relative to a wild-type antibody of the same isotype. 
     
     
         62 . The method of  claim 61 , wherein the effector function is reduced or eliminated by a method selected from reduction of glycosylation of the antibody, modification of the antibody isotype to an isotype that naturally has reduced or eliminated effector function, and modification of the Fc region. 
     
     
         63 . The method of  claim 62 , wherein the glycosylation of the antibody is reduced by a method selected from: production of the antibody in an environment that does not permit wild-type glycosylation; removal of carbohydrate groups already present on the antibody; and modification of the antibody such that wild-type glycosylation does not occur. 
     
     
         64 . The method of  claim 63 , wherein the antibody is produced in a non-mammalian cell production system, or where the antibody is produced synthetically. 
     
     
         65 . The method of  claim 63 , wherein the Fc region of the antibody comprises a mutation at position 297 such that the wild-type asparagine residue at that position is replaced with another amino acid that interferes with glycosylation at that position 
     
     
         66 . The method of  claim 62 , wherein the effector function or the complement activation function is reduced or eliminated by at least one modification of the Fc region or the non-Fc region of the antibody. 
     
     
         67 . The method of  claim 66 , wherein the effector function or complement activation function is reduced or eliminated by deletion of all or a portion of the Fc region, or by engineering the antibody such that it does not include an Fc region competent for effector function or complement activation function. 
     
     
         68 . The method of  claim 66 , wherein the modification is selected from: a point mutation of the Fc region to impair binding to one or more Fc receptors selected from the following positions: 238, 239, 248, 249, 252, 254, 265, 268, 269, 270, 272, 278, 289, 292, 293, 294, 295, 296, 297, 298, 301, 303, 322, 324, 327, 329, 333, 338, 340, 373, 376, 382, 388, 389, 414, 416, 419, 434, 435, 437, 438, and 439; a point mutation of the Fc region to impair binding to C1q selected from the following positions: 270, 322, 329, and 321, eliminating some or all of the Fc region, and a point mutation at position 132 of the CH1 domain. 
     
     
         69 . The method of  claim 59 , wherein the dose amount and/or frequency of administration is modulated to reduce the concentration of antibody to which the red blood cells are exposed. 
     
     
         70 . The method of  claim 59 , wherein the antibody is modified to comprise pH-sensitive binding to the BBB-R. 
     
     
         71 . The method of  claim 60 , wherein the affinity of the antibody for the BBB-R is further decreased. 
     
     
         72 . The method of  claim 71 , wherein the decrease is measured relative to a wild-type antibody of the same isotype not having lowered affinity for the BBB-R. 
     
     
         73 . The method of  claim 54  or  55 , wherein the affinity for the BBB-R is from about 1 nM to about 100 μM. 
     
     
         74 . The method of  claim 73 , wherein the antibody has an IC50 of from about 1 nM to about 100 μM. 
     
     
         75 . The method of  claim 73 , wherein the IC50 is from about 5 nM to about 100 μM. 
     
     
         76 . The method of  claim 73 , wherein the IC50 is from about 50 nM to about 100 μM. 
     
     
         77 . The method of  claim 73 , wherein the IC50 is from about 100 nM to about 100 μM. 
     
     
         78 . The method of any of  claims 54 - 71 , wherein the antibody has an affinity for the BBB-R from about 5 nM to about 50 μM. 
     
     
         79 . The method of any of  claims 54 - 71 , wherein the antibody has a dissociation half-life for the BBB-R from about 30 seconds to about 5 minutes, or from about 30 seconds to about 2 minutes. 
     
     
         80 . The method of  claim 54  or  55  wherein the antibody is selected from a panel of antibodies based upon the affinity of the selected antibody. 
     
     
         81 . The method of  claim 54  or  55 , wherein the antibody is engineered to have the affinity. 
     
     
         82 . The method of  claim 54  or  55  comprising coupling the antibody with a therapeutic compound. 
     
     
         83 . The method of  claim 82  wherein the therapeutic compound is a neurological disorder drug. 
     
     
         84 . The method of  claim 82 , wherein the antibody coupled to the compound has an affinity for the BBB-R from about 30 nM to about 30 μM. 
     
     
         85 . The method of  claim 83 , wherein the antibody is a multispecific antibody and the compound optionally forms one portion of the multispecific antibody. 
     
     
         86 . The method of  claim 54  or  55  wherein the antibody is a multispecific antibody which comprises a first antigen binding site which binds the BBB-R and a second antigen binding site which binds a brain antigen. 
     
     
         87 . The method of  claim 86 , wherein the brain antigen is selected from the group consisting of: beta-secretase 1 (BACE1), Abeta, epidermal growth factor receptor (EGFR), human epidermal growth factor receptor 2 (HER2), tau, apolipoprotein E4 (ApoE4), alpha-synuclein, CD20, huntingtin, prion protein (PrP), leucine rich repeat kinase 2 (LRRK2), parkin, presenilin 1, presenilin 2, gamma secretase, death receptor 6 (DR6), amyloid precursor protein (APP), p75 neurotrophin receptor (p75NTR), and caspase 6. 
     
     
         88 . The method of  claim 87 , wherein the multispecific antibody binds both TfR and BACE1. 
     
     
         89 . The method of  claim 87 , wherein the multispecific antibody binds both TfR and Abeta. 
     
     
         90 . The method of any of  claims 54 - 88 , wherein the antibody does not impair the binding of the BBB-R to one or more of its native ligands. 
     
     
         91 . The method of  claim 90 , wherein the antibody does not inhibit the binding of TfR to transferrin. 
     
     
         92 . An antibody which binds to a BBB-R, wherein the affinity of the antibody for the BBB-R is from about 5 nM to about 50 μM or the dissociation half-life of the antibody for the BBB-R is from about 30 seconds to about 2 minutes, and wherein one or more properties of the antibody have been modified to reduce at least one undesired side effect on red blood cells. 
     
     
         93 . The antibody of  claim 92  wherein the BBB-R is selected from the group consisting of transferrin receptor (TfR), insulin receptor, insulin-like growth factor receptor (IGF receptor), low density lipoprotein receptor-related protein 8 (LRP8), low density lipoprotein receptor-related protein 1 (LRP1), glucose transporter 1 (Glut1) and heparin-binding epidermal growth factor-like growth factor (HB-EGF). 
     
     
         94 . The antibody of  claim 93  wherein the BBB-R is transferrin receptor (TfR). 
     
     
         95 . The antibody of  claim 94 , wherein the red blood cells are reticulocytes. 
     
     
         96 . The antibody of  claim 95 , wherein the at least one undesired side effect on red blood cells is selected from reduction of reticulocyte levels and acute clinical symptoms. 
     
     
         97 . The antibody of  claim 96 , wherein the one or more properties is selected from the effector function of the antibody Fc region, the complement activation function of the antibody and the affinity of the antibody for the BBB-R. 
     
     
         98 . The antibody of  claim 97 , wherein the one or more properties is selected from the effector function of the antibody Fc region and the complement activation function of the antibody, and wherein the effector function or complement activation function has been reduced or eliminated relative to a wild-type antibody of the same isotype. 
     
     
         99 . The antibody of  claim 98 , wherein the effector function is reduced or eliminated by a method selected from reduction of glycosylation of the antibody, modification of the antibody isotype to an isotype that naturally has reduced or eliminated effector function, and modification of the Fc region. 
     
     
         100 . The antibody of  claim 99 , wherein the glycosylation of the antibody is reduced by a method selected from: production of the antibody in an environment that does not permit wild-type glycosylation; removal of carbohydrate groups already present on the antibody; and modification of the antibody such that wild-type glycosylation does not occur. 
     
     
         101 . The antibody of  claim 100 , wherein the antibody is produced in a non-mammalian cell production system, or where the antibody is produced synthetically. 
     
     
         102 . The antibody of  claim 100 , wherein the Fc region of the antibody comprises a mutation at position 297 such that the wild-type asparagine residue at that position is replaced with another amino acid that interferes with glycosylation at that position. 
     
     
         103 . The antibody of  claim 99 , wherein the effector function or complement activation function is reduced or eliminated by at least one modification of the Fc region or the non-Fc region. 
     
     
         104 . The antibody of  claim 103 , wherein the effector function or complement activation function is reduced or eliminated by deletion of all or a portion of the Fc region, or by engineering the antibody such that it does not include an Fc region competent for effector function. 
     
     
         105 . The antibody of  claim 103 , wherein the modification is selected from: a point mutation of the Fc region to impair binding to one or more Fc receptors selected from the following positions: 238, 239, 248, 249, 252, 254, 265, 268, 269, 270, 272, 278, 289, 292, 293, 294, 295, 296, 297, 298, 301, 303, 322, 324, 327, 329, 333, 338, 340, 373, 376, 382, 388, 389, 414, 416, 419, 434, 435, 437, 438, and 439; a point mutation of the Fc region to impair binding to C1q selected from the following positions: 270, 322, 329, and 321; eliminating some or all of the Fc region, and a point mutation at position 132 of the CH1 domain. 
     
     
         106 . The antibody of  claim 97 , wherein the antibody is modified to comprise pH-sensitive binding to the BBB-R. 
     
     
         107 . The antibody of  claim 97 , wherein the affinity of the antibody for the BBB-R is further decreased. 
     
     
         108 . The antibody of  claim 107 , wherein the decrease is measured relative to a wild-type antibody of the same isotype not having lowered affinity for the BBB-R. 
     
     
         109 . The antibody of  claim 92 , wherein the affinity is measured as an IC50. 
     
     
         110 . The antibody of  claim 92  wherein the antibody is selected from a panel of antibodies based upon the affinity of the selected antibody. 
     
     
         111 . The antibody of  claim 92  wherein the antibody is engineered to have the affinity. 
     
     
         112 . The antibody of  claim 92 , further coupled with a therapeutic compound. 
     
     
         113 . The antibody of  claim 112  wherein the therapeutic compound is a neurological disorder drug. 
     
     
         114 . The antibody of  claim 112 , wherein the antibody coupled to the compound has an affinity for the BBB-R from about 30 nM to about 30 μM. 
     
     
         115 . The antibody of  claim 112 , wherein the antibody is a multispecific antibody and the compound optionally forms one portion of the multispecific antibody. 
     
     
         116 . The antibody of  claim 92  wherein the antibody is a multispecific antibody which comprises a first antigen binding site which binds the BBB-R and a second antigen binding site which binds a brain antigen. 
     
     
         117 . The antibody of  claim 116 , wherein the brain antigen is selected from the group consisting of: beta-secretase 1 (BACE1), Abeta, epidermal growth factor receptor (EGFR), human epidermal growth factor receptor 2 (HER2), tau, apolipoprotein E4 (ApoE4), alpha-synuclein, CD20, huntingtin, prion protein (PrP), leucine rich repeat kinase 2 (LRRK2), parkin, presenilin 1, presenilin 2, gamma secretase, death receptor 6 (DR6), amyloid precursor protein (APP), p75 neurotrophin receptor (p75NTR), and caspase 6. 
     
     
         118 . The antibody of  claim 117 , wherein the multispecific antibody binds both TfR and BACE1. 
     
     
         119 . The antibody of  claim 117 , wherein the multispecific antibody binds both TfR and Abeta. 
     
     
         120 . The antibody of any of  claims 92 - 119 , wherein the antibody does not impair the binding of the BBB-R to one or more of its native ligands. 
     
     
         121 . The antibody of  claim 120 , wherein the antibody does not inhibit the binding of TfR to transferrin. 
     
     
         122 . Use of an antibody that binds with low affinity to a BBB-R and that does not impact red blood cell levels for the manufacture of a medicament for treating a neurological disorder. 
     
     
         123 . Use of the antibody of any of  claims 92 - 121  for the manufacture of a medicament for treating a neurological disorder. 
     
     
         124 . An antibody that binds with low affinity to a BBB-R and which does not impact red blood cell levels for use in treating a neurological disorder. 
     
     
         125 . An antibody of any of  claims 92 - 121  for use in treating a neurological disorder. 
     
     
         126 . A method of treating a disease or disorder associated with or caused by elevated red blood cell levels in a subject comprising administering an anti-TfR antibody comprising at least partial effector function to the subject. 
     
     
         127 . The method of  claim 126 , wherein the administering step is at a dose and/or dose frequency calibrated to minimize acute clinical symptoms of the antibody administration.