Raymond Tong | Genentech - Academia.edu (original) (raw)

Papers by Raymond Tong

Research paper thumbnail of Addressing Safety Liabilities of TfR Bispecific Antibodies That Cross the Blood-Brain Barrier

Science Translational Medicine, 2013

Bispecific antibodies using the transferrin receptor (TfR) have shown promise for boosting antibo... more Bispecific antibodies using the transferrin receptor (TfR) have shown promise for boosting antibody uptake in brain. Nevertheless, there are limited data on the therapeutic properties including safety liabilities that will enable successful development of TfR-based therapeutics. We evaluate TfR/BACE1 bispecific antibody variants in mouse and show that reducing TfR binding affinity improves not only brain uptake but also peripheral exposure and the safety profile of these antibodies. We identify and seek to address liabilities of targeting TfR with antibodies, namely, acute clinical signs and decreased circulating reticulocytes observed after dosing. By eliminating Fc effector function, we ameliorated the acute clinical signs and partially rescued a reduction in reticulocytes. Furthermore, we show that complement mediates a residual decrease in reticulocytes observed after Fc effector function is eliminated. These data raise important safety concerns and potential mitigation strategies for the development of TfR-based therapies that are designed to cross the blood-brain barrier.

Research paper thumbnail of MR angiogenesis imaging with Robo4- vs.  V 3-targeted nanoparticles in a B16/F10 mouse melanoma model

Faseb Journal, 2010

The primary objective of this study was to utilize MR molecular imaging to compare the 3-dimensio... more The primary objective of this study was to utilize MR molecular imaging to compare the 3-dimensional spatial distribution of Robo4 and ␣ V ␤ 3integrin as biosignatures of angiogenesis, in a rapidly growing, syngeneic tumor. B16-F10 melanoma-bearing mice were imaged with magnetic resonance (MR; 3.0 T) 11 d postimplantation before and after intravenous administration of either Robo4-or ␣ V ␤ 3 -targeted paramagnetic nanoparticles. The percentage of MR signalenhanced voxels throughout the tumor volume was low and increased in animals receiving ␣ V ␤ 3 -and Robo4targeted nanoparticles. Neovascular signal enhancement was predominantly associated with the tumor periphery (i.e., outer 50% of volume). Microscopic examination of tumors coexposed to the Robo4-and ␣ V ␤ 3 -targeted nanoparticles corroborated the MR angiogenesis mapping results and further revealed that Robo4 expression generally colocalized with ␣ V ␤ 3integrin. Robo4-and ␣ V ␤ 3 -targeted nanoparticles were compared to irrelevant or nontargeted control groups in all modalities. These results suggest that ␣ V ␤ 3integrin and Robo4 are useful biomarkers for noninvasive MR molecular imaging in syngeneic mouse tumors, but ␣ V ␤ 3 -integrin expression was more detectable by MR at 3.0 T than Robo4. Noninvasive, neovascular assessments of the MR signal of Robo4, particularly combined with ␣ V ␤ 3 -integrin expression, may help define tumor character prior to and following cancer therapy. MR angiogenesis imaging with Robo4-vs. ␣ V ␤ 3 -targeted nanoparticles in a B16/F10 mouse melanoma model. FASEB J. 24, 4262-4270 (2010). www.fasebj.org

Research paper thumbnail of Identification of circulating neuropilin-1 and dose-dependent elevation following anti-neuropilin-1 antibody administration

mAbs, 2009

Neuropilin-1 (NRP1) acts as a co-receptor for class 3 semaphorins and vascular endothelial growth... more Neuropilin-1 (NRP1) acts as a co-receptor for class 3 semaphorins and vascular endothelial growth factor and is an attractive angiogenesis target for cancer therapy. In addition to the transmembrane form, naturally occurring soluble NRP1 proteins containing part of the extracellular domain have been identified in tissues and a cell line. We developed ELISAs to study the existence of circulating NRP1 and to quantify it in serum. As measured by ELISAs, circulating NRP1 levels in mice, rats, monkeys and humans were 427 ± 77, 20 ± 3, 288 ± 86 and 322 ± 82 ng/ml (mean ± standard deviation; n ≥ 10), respectively. Anti-NRP1 B , a human monoclonal antibody, has been selected from a synthetic phage library. A 4-fold increase in circulating NRP1 was observed in mice receiving a single dose of 10 mg/kg anti-NRP1 B antibody. In rats and monkeys receiving single injections of anti-NRP1 B at different dose levels, higher doses of antibody resulted in greater and more prolonged increases in circulating NRP1. Maximum increases were 56-and 7-fold for rats and monkeys receiving 50 mg/kg anti-NRP1 B , respectively. In addition to the soluble NRP1 isoforms, for the first time, a ~120 kDa circulating NRP1 protein containing the complete extracellular domain was detected in serum by western blot and mass spectrometry analysis. This protein increased more than the putative soluble NRP1 bands in anti-NRP1 B treated mouse, rat and monkey sera compared with untreated controls, suggesting that anti-NRP1 B induced membrane NRP1 shedding.

Research paper thumbnail of Supplemental Data Blocking Neuropilin-2 Function Inhibits Tumor Cell Metastasis

Research paper thumbnail of Addressing Safety Liabilities of TfR Bispecific Antibodies That Cross the Blood-Brain Barrier

Science Translational Medicine, 2013

Bispecific antibodies using the transferrin receptor (TfR) have shown promise for boosting antibo... more Bispecific antibodies using the transferrin receptor (TfR) have shown promise for boosting antibody uptake in brain. Nevertheless, there are limited data on the therapeutic properties including safety liabilities that will enable successful development of TfR-based therapeutics. We evaluate TfR/BACE1 bispecific antibody variants in mouse and show that reducing TfR binding affinity improves not only brain uptake but also peripheral exposure and the safety profile of these antibodies. We identify and seek to address liabilities of targeting TfR with antibodies, namely, acute clinical signs and decreased circulating reticulocytes observed after dosing. By eliminating Fc effector function, we ameliorated the acute clinical signs and partially rescued a reduction in reticulocytes. Furthermore, we show that complement mediates a residual decrease in reticulocytes observed after Fc effector function is eliminated. These data raise important safety concerns and potential mitigation strategies for the development of TfR-based therapies that are designed to cross the blood-brain barrier.

Research paper thumbnail of Antiparallel Conformation of Knob and Hole Aglycosylated Half-Antibody Homodimers Is Mediated by a CH2–CH3 Hydrophobic Interaction

Journal of Molecular Biology, 2014

Bispecific antibody and antibody-like molecules are of wide interest as potential therapeutics th... more Bispecific antibody and antibody-like molecules are of wide interest as potential therapeutics that can recognize two distinct targets. Among the variety of ways such molecules have been engineered is by creating "knob" and "hole" heterodimerization sites in the CH3 domains of two antibody heavy chains. The molecules produced in this manner maintain their biological activities while differing very little from the native human IgG sequence. To better understand the knob-into-hole interface, the molecular mechanism of heterodimerization, and to engineer Fc domains that could improve the assembly and purity of heterodimeric reaction products, we sought crystal structures of aglycosylated heterodimeric and homodimeric "knob" and "hole" Fc fragments derived from bacterial expression. The structure of the knob-into-hole Fc was determined at 2.64 Å. Except for the sites of mutation, the structure is very similar to that of the native human IgG1 Fc, consistent with a heterodimer interaction kinetic K(D) of…

Research paper thumbnail of Robo4 Maintains Vessel Integrity and Inhibits Angiogenesis by Interacting with UNC5B

Developmental Cell, 2011

Robo4 is an endothelial cell-specific member of the Roundabout axon guidance receptor family. To ... more Robo4 is an endothelial cell-specific member of the Roundabout axon guidance receptor family. To identify Robo4 binding partners, we performed a proteinprotein interaction screen with the Robo4 extracellular domain. We find that Robo4 specifically binds to UNC5B, a vascular Netrin receptor, revealing unexpected interactions between two endothelial guidance receptors. We show that Robo4 maintains vessel integrity by activating UNC5B, which inhibits signaling downstream of vascular endothelial growth factor (VEGF). Function-blocking monoclonal antibodies against Robo4 and UNC5B increase angiogenesis and disrupt vessel integrity. Soluble Robo4 protein inhibits VEGF-induced vessel permeability and rescues barrier defects in Robo4 À/À mice, but not in mice treated with anti-UNC5B. Thus, Robo4-UNC5B signaling maintains vascular integrity by counteracting VEGF signaling in endothelial cells, identifying a novel function of guidance receptor interactions in the vasculature.

Research paper thumbnail of CCBE1 Is Essential for Mammalian Lymphatic Vascular Development and Enhances the Lymphangiogenic Effect of Vascular Endothelial Growth Factor-C In Vivo

Circulation Research, 2011

Collagen- and calcium-binding EGF domains 1 (CCBE1) has been associated with Hennekam syndrome, i... more Collagen- and calcium-binding EGF domains 1 (CCBE1) has been associated with Hennekam syndrome, in which patients have lymphedema, lymphangiectasias, and other cardiovascular anomalies. Insight into the molecular role of CCBE1 is completely lacking, and mouse models for the disease do not exist. CCBE1 deficient mice were generated to understand the function of CCBE1 in cardiovascular development, and CCBE1 recombinant protein was used in both in vivo and in vitro settings to gain insight into the molecular function of CCBE1. Phenotypic analysis of murine Ccbe1 mutant embryos showed a complete lack of definitive lymphatic structures, even though Prox1(+) lymphatic endothelial cells get specified within the cardinal vein. Mutant mice die prenatally. Proximity ligation assays indicate that vascular endothelial growth factor receptor 3 activation appears unaltered in mutants. Human CCBE1 protein binds to components of the extracellular matrix in vitro, and CCBE1 protein strongly enhances vascular endothelial growth factor-C-mediated lymphangiogenesis in a corneal micropocket assay. Our data identify CCBE1 as a factor critically required for budding and migration of Prox-1(+) lymphatic endothelial cells from the cardinal vein. CCBE1 probably exerts these effects through binding to components of the extracellular matrix. CCBE1 has little lymphangiogenic effect on its own but dramatically enhances the lymphangiogenic effect of vascular endothelial growth factor-C in vivo. Thus, our data suggest CCBE1 to be essential but not sufficient for lymphangiogenesis.

Research paper thumbnail of Blocking Neuropilin-1 Function Has an Additive Effect with Anti-VEGF to Inhibit Tumor Growth

Cancer Cell, 2007

Neuropilin-1 (NRP1) guides the development of the nervous and vascular systems. Binding to either... more Neuropilin-1 (NRP1) guides the development of the nervous and vascular systems. Binding to either semaphorins or VEGF, NRP1 acts with plexins to regulate neuronal guidance, or with VEGFR2 to mediate vascular development. We have generated two monoclonal antibodies that bind to the Sema-and VEGF-binding domains of NRP1, respectively. Both antibodies reduce angiogenesis and vascular remodeling, while having little effect on other VEGFR2-mediated events. Importantly, anti-NRP1 antibodies have an additive effect with anti-VEGF therapy in reducing tumor growth. Vessels from tumors treated with anti-VEGF show a close association with pericytes, while tumors treated with both anti-NRP1 and anti-VEGF lack this organization. We propose that blocking NRP1 function inhibits vascular remodeling, rendering vessels more susceptible to anti-VEGF therapy.

Research paper thumbnail of Blocking Neuropilin-2 Function Inhibits Tumor Cell Metastasis

Cancer Cell, 2008

Metastasis, which commonly uses lymphatics, accounts for much of the mortality associated with ca... more Metastasis, which commonly uses lymphatics, accounts for much of the mortality associated with cancer. The vascular endothelial growth factor (VEGF)-C coreceptor, neuropilin-2 (Nrp2), modulates but is not necessary for developmental lymphangiogenesis, and its significance for metastasis is unknown. An antibody to Nrp2 that blocks VEGFC binding disrupts VEGFC-induced lymphatic endothelial cell migration, but not proliferation, in part independently of VEGF receptor activation. It does not affect established lymphatics in normal adult mice but reduces tumoral lymphangiogenesis and, importantly, functional lymphatics associated with tumors. It also reduces metastasis to sentinel lymph nodes and distant organs, apparently by delaying the departure of tumor cells from the primary tumor. Our results demonstrate that Nrp2, which was originally identified as an axon-guidance receptor, is an attractive target for modulating metastasis.

Research paper thumbnail of Therapeutic bispecific antibodies cross the blood-brain barrier in nonhuman primates

Science translational medicine, Jan 5, 2014

Using therapeutic antibodies that need to cross the blood-brain barrier (BBB) to treat neurologic... more Using therapeutic antibodies that need to cross the blood-brain barrier (BBB) to treat neurological disease is a difficult challenge. We have shown that bispecific antibodies with optimized binding to the transferrin receptor (TfR) that target β-secretase (BACE1) can cross the BBB and reduce brain amyloid-β (Aβ) in mice. Can TfR enhance antibody uptake in the primate brain? We describe two humanized TfR/BACE1 bispecific antibody variants. Using a human TfR knock-in mouse, we observed that anti-TfR/BACE1 antibodies could cross the BBB and reduce brain Aβ in a TfR affinity-dependent fashion. Intravenous dosing of monkeys with anti-TfR/BACE1 antibodies also reduced Aβ both in cerebral spinal fluid and in brain tissue, and the degree of reduction correlated with the brain concentration of anti-TfR/BACE1 antibody. These results demonstrate that the TfR bispecific antibody platform can robustly and safely deliver therapeutic antibody across the BBB in the primate brain.

Research paper thumbnail of Addressing Safety Liabilities of TfR Bispecific Antibodies That Cross the Blood-Brain Barrier

Science Translational Medicine, 2013

Bispecific antibodies using the transferrin receptor (TfR) have shown promise for boosting antibo... more Bispecific antibodies using the transferrin receptor (TfR) have shown promise for boosting antibody uptake in brain. Nevertheless, there are limited data on the therapeutic properties including safety liabilities that will enable successful development of TfR-based therapeutics. We evaluate TfR/BACE1 bispecific antibody variants in mouse and show that reducing TfR binding affinity improves not only brain uptake but also peripheral exposure and the safety profile of these antibodies. We identify and seek to address liabilities of targeting TfR with antibodies, namely, acute clinical signs and decreased circulating reticulocytes observed after dosing. By eliminating Fc effector function, we ameliorated the acute clinical signs and partially rescued a reduction in reticulocytes. Furthermore, we show that complement mediates a residual decrease in reticulocytes observed after Fc effector function is eliminated. These data raise important safety concerns and potential mitigation strategies for the development of TfR-based therapies that are designed to cross the blood-brain barrier.

Research paper thumbnail of MR angiogenesis imaging with Robo4- vs.  V 3-targeted nanoparticles in a B16/F10 mouse melanoma model

Faseb Journal, 2010

The primary objective of this study was to utilize MR molecular imaging to compare the 3-dimensio... more The primary objective of this study was to utilize MR molecular imaging to compare the 3-dimensional spatial distribution of Robo4 and ␣ V ␤ 3integrin as biosignatures of angiogenesis, in a rapidly growing, syngeneic tumor. B16-F10 melanoma-bearing mice were imaged with magnetic resonance (MR; 3.0 T) 11 d postimplantation before and after intravenous administration of either Robo4-or ␣ V ␤ 3 -targeted paramagnetic nanoparticles. The percentage of MR signalenhanced voxels throughout the tumor volume was low and increased in animals receiving ␣ V ␤ 3 -and Robo4targeted nanoparticles. Neovascular signal enhancement was predominantly associated with the tumor periphery (i.e., outer 50% of volume). Microscopic examination of tumors coexposed to the Robo4-and ␣ V ␤ 3 -targeted nanoparticles corroborated the MR angiogenesis mapping results and further revealed that Robo4 expression generally colocalized with ␣ V ␤ 3integrin. Robo4-and ␣ V ␤ 3 -targeted nanoparticles were compared to irrelevant or nontargeted control groups in all modalities. These results suggest that ␣ V ␤ 3integrin and Robo4 are useful biomarkers for noninvasive MR molecular imaging in syngeneic mouse tumors, but ␣ V ␤ 3 -integrin expression was more detectable by MR at 3.0 T than Robo4. Noninvasive, neovascular assessments of the MR signal of Robo4, particularly combined with ␣ V ␤ 3 -integrin expression, may help define tumor character prior to and following cancer therapy. MR angiogenesis imaging with Robo4-vs. ␣ V ␤ 3 -targeted nanoparticles in a B16/F10 mouse melanoma model. FASEB J. 24, 4262-4270 (2010). www.fasebj.org

Research paper thumbnail of Identification of circulating neuropilin-1 and dose-dependent elevation following anti-neuropilin-1 antibody administration

mAbs, 2009

Neuropilin-1 (NRP1) acts as a co-receptor for class 3 semaphorins and vascular endothelial growth... more Neuropilin-1 (NRP1) acts as a co-receptor for class 3 semaphorins and vascular endothelial growth factor and is an attractive angiogenesis target for cancer therapy. In addition to the transmembrane form, naturally occurring soluble NRP1 proteins containing part of the extracellular domain have been identified in tissues and a cell line. We developed ELISAs to study the existence of circulating NRP1 and to quantify it in serum. As measured by ELISAs, circulating NRP1 levels in mice, rats, monkeys and humans were 427 ± 77, 20 ± 3, 288 ± 86 and 322 ± 82 ng/ml (mean ± standard deviation; n ≥ 10), respectively. Anti-NRP1 B , a human monoclonal antibody, has been selected from a synthetic phage library. A 4-fold increase in circulating NRP1 was observed in mice receiving a single dose of 10 mg/kg anti-NRP1 B antibody. In rats and monkeys receiving single injections of anti-NRP1 B at different dose levels, higher doses of antibody resulted in greater and more prolonged increases in circulating NRP1. Maximum increases were 56-and 7-fold for rats and monkeys receiving 50 mg/kg anti-NRP1 B , respectively. In addition to the soluble NRP1 isoforms, for the first time, a ~120 kDa circulating NRP1 protein containing the complete extracellular domain was detected in serum by western blot and mass spectrometry analysis. This protein increased more than the putative soluble NRP1 bands in anti-NRP1 B treated mouse, rat and monkey sera compared with untreated controls, suggesting that anti-NRP1 B induced membrane NRP1 shedding.

Research paper thumbnail of Supplemental Data Blocking Neuropilin-2 Function Inhibits Tumor Cell Metastasis

Research paper thumbnail of Addressing Safety Liabilities of TfR Bispecific Antibodies That Cross the Blood-Brain Barrier

Science Translational Medicine, 2013

Bispecific antibodies using the transferrin receptor (TfR) have shown promise for boosting antibo... more Bispecific antibodies using the transferrin receptor (TfR) have shown promise for boosting antibody uptake in brain. Nevertheless, there are limited data on the therapeutic properties including safety liabilities that will enable successful development of TfR-based therapeutics. We evaluate TfR/BACE1 bispecific antibody variants in mouse and show that reducing TfR binding affinity improves not only brain uptake but also peripheral exposure and the safety profile of these antibodies. We identify and seek to address liabilities of targeting TfR with antibodies, namely, acute clinical signs and decreased circulating reticulocytes observed after dosing. By eliminating Fc effector function, we ameliorated the acute clinical signs and partially rescued a reduction in reticulocytes. Furthermore, we show that complement mediates a residual decrease in reticulocytes observed after Fc effector function is eliminated. These data raise important safety concerns and potential mitigation strategies for the development of TfR-based therapies that are designed to cross the blood-brain barrier.

Research paper thumbnail of Antiparallel Conformation of Knob and Hole Aglycosylated Half-Antibody Homodimers Is Mediated by a CH2–CH3 Hydrophobic Interaction

Journal of Molecular Biology, 2014

Bispecific antibody and antibody-like molecules are of wide interest as potential therapeutics th... more Bispecific antibody and antibody-like molecules are of wide interest as potential therapeutics that can recognize two distinct targets. Among the variety of ways such molecules have been engineered is by creating "knob" and "hole" heterodimerization sites in the CH3 domains of two antibody heavy chains. The molecules produced in this manner maintain their biological activities while differing very little from the native human IgG sequence. To better understand the knob-into-hole interface, the molecular mechanism of heterodimerization, and to engineer Fc domains that could improve the assembly and purity of heterodimeric reaction products, we sought crystal structures of aglycosylated heterodimeric and homodimeric "knob" and "hole" Fc fragments derived from bacterial expression. The structure of the knob-into-hole Fc was determined at 2.64 Å. Except for the sites of mutation, the structure is very similar to that of the native human IgG1 Fc, consistent with a heterodimer interaction kinetic K(D) of…

Research paper thumbnail of Robo4 Maintains Vessel Integrity and Inhibits Angiogenesis by Interacting with UNC5B

Developmental Cell, 2011

Robo4 is an endothelial cell-specific member of the Roundabout axon guidance receptor family. To ... more Robo4 is an endothelial cell-specific member of the Roundabout axon guidance receptor family. To identify Robo4 binding partners, we performed a proteinprotein interaction screen with the Robo4 extracellular domain. We find that Robo4 specifically binds to UNC5B, a vascular Netrin receptor, revealing unexpected interactions between two endothelial guidance receptors. We show that Robo4 maintains vessel integrity by activating UNC5B, which inhibits signaling downstream of vascular endothelial growth factor (VEGF). Function-blocking monoclonal antibodies against Robo4 and UNC5B increase angiogenesis and disrupt vessel integrity. Soluble Robo4 protein inhibits VEGF-induced vessel permeability and rescues barrier defects in Robo4 À/À mice, but not in mice treated with anti-UNC5B. Thus, Robo4-UNC5B signaling maintains vascular integrity by counteracting VEGF signaling in endothelial cells, identifying a novel function of guidance receptor interactions in the vasculature.

Research paper thumbnail of CCBE1 Is Essential for Mammalian Lymphatic Vascular Development and Enhances the Lymphangiogenic Effect of Vascular Endothelial Growth Factor-C In Vivo

Circulation Research, 2011

Collagen- and calcium-binding EGF domains 1 (CCBE1) has been associated with Hennekam syndrome, i... more Collagen- and calcium-binding EGF domains 1 (CCBE1) has been associated with Hennekam syndrome, in which patients have lymphedema, lymphangiectasias, and other cardiovascular anomalies. Insight into the molecular role of CCBE1 is completely lacking, and mouse models for the disease do not exist. CCBE1 deficient mice were generated to understand the function of CCBE1 in cardiovascular development, and CCBE1 recombinant protein was used in both in vivo and in vitro settings to gain insight into the molecular function of CCBE1. Phenotypic analysis of murine Ccbe1 mutant embryos showed a complete lack of definitive lymphatic structures, even though Prox1(+) lymphatic endothelial cells get specified within the cardinal vein. Mutant mice die prenatally. Proximity ligation assays indicate that vascular endothelial growth factor receptor 3 activation appears unaltered in mutants. Human CCBE1 protein binds to components of the extracellular matrix in vitro, and CCBE1 protein strongly enhances vascular endothelial growth factor-C-mediated lymphangiogenesis in a corneal micropocket assay. Our data identify CCBE1 as a factor critically required for budding and migration of Prox-1(+) lymphatic endothelial cells from the cardinal vein. CCBE1 probably exerts these effects through binding to components of the extracellular matrix. CCBE1 has little lymphangiogenic effect on its own but dramatically enhances the lymphangiogenic effect of vascular endothelial growth factor-C in vivo. Thus, our data suggest CCBE1 to be essential but not sufficient for lymphangiogenesis.

Research paper thumbnail of Blocking Neuropilin-1 Function Has an Additive Effect with Anti-VEGF to Inhibit Tumor Growth

Cancer Cell, 2007

Neuropilin-1 (NRP1) guides the development of the nervous and vascular systems. Binding to either... more Neuropilin-1 (NRP1) guides the development of the nervous and vascular systems. Binding to either semaphorins or VEGF, NRP1 acts with plexins to regulate neuronal guidance, or with VEGFR2 to mediate vascular development. We have generated two monoclonal antibodies that bind to the Sema-and VEGF-binding domains of NRP1, respectively. Both antibodies reduce angiogenesis and vascular remodeling, while having little effect on other VEGFR2-mediated events. Importantly, anti-NRP1 antibodies have an additive effect with anti-VEGF therapy in reducing tumor growth. Vessels from tumors treated with anti-VEGF show a close association with pericytes, while tumors treated with both anti-NRP1 and anti-VEGF lack this organization. We propose that blocking NRP1 function inhibits vascular remodeling, rendering vessels more susceptible to anti-VEGF therapy.

Research paper thumbnail of Blocking Neuropilin-2 Function Inhibits Tumor Cell Metastasis

Cancer Cell, 2008

Metastasis, which commonly uses lymphatics, accounts for much of the mortality associated with ca... more Metastasis, which commonly uses lymphatics, accounts for much of the mortality associated with cancer. The vascular endothelial growth factor (VEGF)-C coreceptor, neuropilin-2 (Nrp2), modulates but is not necessary for developmental lymphangiogenesis, and its significance for metastasis is unknown. An antibody to Nrp2 that blocks VEGFC binding disrupts VEGFC-induced lymphatic endothelial cell migration, but not proliferation, in part independently of VEGF receptor activation. It does not affect established lymphatics in normal adult mice but reduces tumoral lymphangiogenesis and, importantly, functional lymphatics associated with tumors. It also reduces metastasis to sentinel lymph nodes and distant organs, apparently by delaying the departure of tumor cells from the primary tumor. Our results demonstrate that Nrp2, which was originally identified as an axon-guidance receptor, is an attractive target for modulating metastasis.

Research paper thumbnail of Therapeutic bispecific antibodies cross the blood-brain barrier in nonhuman primates

Science translational medicine, Jan 5, 2014

Using therapeutic antibodies that need to cross the blood-brain barrier (BBB) to treat neurologic... more Using therapeutic antibodies that need to cross the blood-brain barrier (BBB) to treat neurological disease is a difficult challenge. We have shown that bispecific antibodies with optimized binding to the transferrin receptor (TfR) that target β-secretase (BACE1) can cross the BBB and reduce brain amyloid-β (Aβ) in mice. Can TfR enhance antibody uptake in the primate brain? We describe two humanized TfR/BACE1 bispecific antibody variants. Using a human TfR knock-in mouse, we observed that anti-TfR/BACE1 antibodies could cross the BBB and reduce brain Aβ in a TfR affinity-dependent fashion. Intravenous dosing of monkeys with anti-TfR/BACE1 antibodies also reduced Aβ both in cerebral spinal fluid and in brain tissue, and the degree of reduction correlated with the brain concentration of anti-TfR/BACE1 antibody. These results demonstrate that the TfR bispecific antibody platform can robustly and safely deliver therapeutic antibody across the BBB in the primate brain.