The receptor tyrosine kinase ROR1 is a cell-surface onco-fetal protein expressed on a variety of ... more The receptor tyrosine kinase ROR1 is a cell-surface onco-fetal protein expressed on a variety of solid tumours and haematological malignancies, which has limited expression on normal adult tissue. A functional role for ROR1 in tumorigenesis and disease progression has been demonstrated in a number of cancer indications. ROR1 is also involved in mediating drug resistance (for example to chemotherapies as well as to the targeted therapy T-DM1), activation of YAP/TAZ signalling and up-regulation of BMI-1. Consistent with this pleiotropic role, increased ROR1 expression correlates with poor clinical outcome in a number of cancer indications including triple negative breast cancer, ovarian cancer and lung adenocarcinoma. As a result, ROR1 has emerged as a highly attractive target for cancer therapy, and its limited expression on normal tissues make it particularly well suited for antibody and protein-drug conjugate (ADC and PDC) approaches. This has been fuelled by recent Phase I clinica...
An entry from the Cambridge Structural Database, the world's repository for small molecule cr... more An entry from the Cambridge Structural Database, the world's repository for small molecule crystal structures. The entry contains experimental data from a crystal diffraction study. The deposited dataset for this entry is freely available from the CCDC and typically includes 3D coordinates, cell parameters, space group, experimental conditions and quality measures.
An entry from the Cambridge Structural Database, the world's repository for small molecule cr... more An entry from the Cambridge Structural Database, the world's repository for small molecule crystal structures. The entry contains experimental data from a crystal diffraction study. The deposited dataset for this entry is freely available from the CCDC and typically includes 3D coordinates, cell parameters, space group, experimental conditions and quality measures.
An entry from the Cambridge Structural Database, the world's repository for small molecule cr... more An entry from the Cambridge Structural Database, the world's repository for small molecule crystal structures. The entry contains experimental data from a crystal diffraction study. The deposited dataset for this entry is freely available from the CCDC and typically includes 3D coordinates, cell parameters, space group, experimental conditions and quality measures.
An entry from the Cambridge Structural Database, the world's repository for small molecule cr... more An entry from the Cambridge Structural Database, the world's repository for small molecule crystal structures. The entry contains experimental data from a crystal diffraction study. The deposited dataset for this entry is freely available from the CCDC and typically includes 3D coordinates, cell parameters, space group, experimental conditions and quality measures.
The onco-embryonic receptor tyrosine kinase ROR1is a cell-surface protein expressed on a broad ra... more The onco-embryonic receptor tyrosine kinase ROR1is a cell-surface protein expressed on a broad range of solid tumours and haematological malignancies but is largely absent from normal adult tissue. Increased ROR1 expression correlates with poor clinical outcome in a number of cancer indications, consistent with its functional role in tumorigenesis and disease progression. In addition, elevated expression has been associated with increased metastatic potential and drug resistance in breast cancer and non-small cell lung cancer. The oncofetal expression pattern of ROR1 coupled with its role in cancer pathology and therapy resistance makes it a highly attractive target for a protein-drug conjugate approach. Shark-derived Variable New Antigen Receptor domains (VNARs) are the smallest antigen binding domain in the vertebrate kingdom. We have developed a platform to enable VNAR-based drug discovery and used it to identify a series of highly potent protein-drug conjugates that target ROR1....
Whilst antibody drug conjugates (ADCs) have been successfully used to target highly potent cytoto... more Whilst antibody drug conjugates (ADCs) have been successfully used to target highly potent cytotoxic agents to tumours, there is a continued emphasis on developing technologies for improving the therapeutic effectiveness of this class of agents. The vast majority of ADCs that have entered the clinic have been based on large intact immunoglobulins, however, there is significant interest in exploiting small protein domains as the payload delivery vehicle. These offer a number of potential benefits over full length antibodies including increased tumour penetration, amenability to protein engineering and site-specific conjugation, and improved tolerability. To this end, we have developed a therapeutics platform based on shark Variable New Antigen Receptor (VNAR) proteins. Shark VNAR domains are the smallest naturally occurring antigen binding domain (~11 kDa), with a binding mechanism distinct from traditional antibodies. Here we report the application of this platform to the development of homogenous VNAR-drug conjugates targeting the onco-embryonic receptor tyrosine kinase ROR1. ROR1 expression has been observed across a broad range of solid tumours and haematological malignancies, and is reported to correlate with poor clinical outcome for a number of cancer indications, but is largely absent from normal adult tissue. As such, the expression pattern of ROR1, coupled with its functional role in tumourogenesis and disease progression, make it an attractive PDC target. Using a combination of direct immunisation and synthetic VNAR library screening, we have generated high affinity binders to the extracellular domain of ROR1 which show species cross-reactivity, do not bind the closely related family member ROR2, and can target distinct non-overlapping regions of the protein. By exploiting the ability of VNAR domains to be flexibly reformatted, the lead sequences have been configured as a series of bivalent Fc fusions, VNAR multimers and novel bi-paratopic binders, and constructs identified that show further enhanced binding to ROR1 and which can induce internalisation and lysosomal trafficking in ROR1 expressing cancer cell-lines. Subsequent attachment of highly potent DNA-damaging cytotoxic payloads yielded homogenous VNAR-drug conjugates which demonstrated extremely potent (sub-nanomolar) in vitro cytotoxicity against selected cancer cell-lines in a receptor mediated fashion. Studies to assess the in vivo efficacy of these conjugates are in progress. These ROR1 VNAR constructs have also been successfully fused to sdAbs and scFv directed to other cell-surface protein targets, demonstrating the power of the platform and opening the way for novel bi-specific approaches for targeting both solid and haematological tumours. Citation Format: Graham Cotton, Jennifer Thom, Paul Trumper, Stacey Bell, Andrei Kamenski, Mark Wappett, Caroline Barelle, Marina Kovaleva, John Steven, Andy Porter, Estelle McLean, Chiara Saladino, Aidan McCann, Aaron Cranston, Tim Harrison. Novel protein drug conjugates targeting ROR1 through the development and exploitation of a drug discovery platform based on small, engineered VNAR domains [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 222.
This work demonstrates the feasibility of making sensitive nanometer distance measurements betwee... more This work demonstrates the feasibility of making sensitive nanometer distance measurements between Fe(III) heme centers and nitroxide spin labels in proteins using the double electron-electron resonance (DEER) pulsed EPR technique at 94 GHz. Techniques to measure accurately long distances in many classes of heme proteins using DEER are currently strongly limited by sensitivity. In this paper we demonstrate sensitivity gains of more than 30 times compared with previous lower frequency (X-band) DEER measurements on both human neuroglobin and sperm whale myoglobin. This is achieved by taking advantage of recent instrumental advances, employing wideband excitation techniques based on composite pulses and exploiting more favorable relaxation properties of low-spin Fe(III) in high magnetic fields. This gain in sensitivity potentially allows the DEER technique to be routinely used as a sensitive probe of structure and conformation in the large number of heme and many other metalloproteins.
The receptor tyrosine kinase ROR1 is a cell-surface onco-fetal protein expressed on a variety of ... more The receptor tyrosine kinase ROR1 is a cell-surface onco-fetal protein expressed on a variety of solid tumours and haematological malignancies, which has limited expression on normal adult tissue. A functional role for ROR1 in tumorigenesis and disease progression has been demonstrated in a number of cancer indications. ROR1 is also involved in mediating drug resistance (for example to chemotherapies as well as to the targeted therapy T-DM1), activation of YAP/TAZ signalling and up-regulation of BMI-1. Consistent with this pleiotropic role, increased ROR1 expression correlates with poor clinical outcome in a number of cancer indications including triple negative breast cancer, ovarian cancer and lung adenocarcinoma. As a result, ROR1 has emerged as a highly attractive target for cancer therapy, and its limited expression on normal tissues make it particularly well suited for antibody and protein-drug conjugate (ADC and PDC) approaches. This has been fuelled by recent Phase I clinica...
An entry from the Cambridge Structural Database, the world's repository for small molecule cr... more An entry from the Cambridge Structural Database, the world's repository for small molecule crystal structures. The entry contains experimental data from a crystal diffraction study. The deposited dataset for this entry is freely available from the CCDC and typically includes 3D coordinates, cell parameters, space group, experimental conditions and quality measures.
An entry from the Cambridge Structural Database, the world's repository for small molecule cr... more An entry from the Cambridge Structural Database, the world's repository for small molecule crystal structures. The entry contains experimental data from a crystal diffraction study. The deposited dataset for this entry is freely available from the CCDC and typically includes 3D coordinates, cell parameters, space group, experimental conditions and quality measures.
An entry from the Cambridge Structural Database, the world's repository for small molecule cr... more An entry from the Cambridge Structural Database, the world's repository for small molecule crystal structures. The entry contains experimental data from a crystal diffraction study. The deposited dataset for this entry is freely available from the CCDC and typically includes 3D coordinates, cell parameters, space group, experimental conditions and quality measures.
An entry from the Cambridge Structural Database, the world's repository for small molecule cr... more An entry from the Cambridge Structural Database, the world's repository for small molecule crystal structures. The entry contains experimental data from a crystal diffraction study. The deposited dataset for this entry is freely available from the CCDC and typically includes 3D coordinates, cell parameters, space group, experimental conditions and quality measures.
The onco-embryonic receptor tyrosine kinase ROR1is a cell-surface protein expressed on a broad ra... more The onco-embryonic receptor tyrosine kinase ROR1is a cell-surface protein expressed on a broad range of solid tumours and haematological malignancies but is largely absent from normal adult tissue. Increased ROR1 expression correlates with poor clinical outcome in a number of cancer indications, consistent with its functional role in tumorigenesis and disease progression. In addition, elevated expression has been associated with increased metastatic potential and drug resistance in breast cancer and non-small cell lung cancer. The oncofetal expression pattern of ROR1 coupled with its role in cancer pathology and therapy resistance makes it a highly attractive target for a protein-drug conjugate approach. Shark-derived Variable New Antigen Receptor domains (VNARs) are the smallest antigen binding domain in the vertebrate kingdom. We have developed a platform to enable VNAR-based drug discovery and used it to identify a series of highly potent protein-drug conjugates that target ROR1....
Whilst antibody drug conjugates (ADCs) have been successfully used to target highly potent cytoto... more Whilst antibody drug conjugates (ADCs) have been successfully used to target highly potent cytotoxic agents to tumours, there is a continued emphasis on developing technologies for improving the therapeutic effectiveness of this class of agents. The vast majority of ADCs that have entered the clinic have been based on large intact immunoglobulins, however, there is significant interest in exploiting small protein domains as the payload delivery vehicle. These offer a number of potential benefits over full length antibodies including increased tumour penetration, amenability to protein engineering and site-specific conjugation, and improved tolerability. To this end, we have developed a therapeutics platform based on shark Variable New Antigen Receptor (VNAR) proteins. Shark VNAR domains are the smallest naturally occurring antigen binding domain (~11 kDa), with a binding mechanism distinct from traditional antibodies. Here we report the application of this platform to the development of homogenous VNAR-drug conjugates targeting the onco-embryonic receptor tyrosine kinase ROR1. ROR1 expression has been observed across a broad range of solid tumours and haematological malignancies, and is reported to correlate with poor clinical outcome for a number of cancer indications, but is largely absent from normal adult tissue. As such, the expression pattern of ROR1, coupled with its functional role in tumourogenesis and disease progression, make it an attractive PDC target. Using a combination of direct immunisation and synthetic VNAR library screening, we have generated high affinity binders to the extracellular domain of ROR1 which show species cross-reactivity, do not bind the closely related family member ROR2, and can target distinct non-overlapping regions of the protein. By exploiting the ability of VNAR domains to be flexibly reformatted, the lead sequences have been configured as a series of bivalent Fc fusions, VNAR multimers and novel bi-paratopic binders, and constructs identified that show further enhanced binding to ROR1 and which can induce internalisation and lysosomal trafficking in ROR1 expressing cancer cell-lines. Subsequent attachment of highly potent DNA-damaging cytotoxic payloads yielded homogenous VNAR-drug conjugates which demonstrated extremely potent (sub-nanomolar) in vitro cytotoxicity against selected cancer cell-lines in a receptor mediated fashion. Studies to assess the in vivo efficacy of these conjugates are in progress. These ROR1 VNAR constructs have also been successfully fused to sdAbs and scFv directed to other cell-surface protein targets, demonstrating the power of the platform and opening the way for novel bi-specific approaches for targeting both solid and haematological tumours. Citation Format: Graham Cotton, Jennifer Thom, Paul Trumper, Stacey Bell, Andrei Kamenski, Mark Wappett, Caroline Barelle, Marina Kovaleva, John Steven, Andy Porter, Estelle McLean, Chiara Saladino, Aidan McCann, Aaron Cranston, Tim Harrison. Novel protein drug conjugates targeting ROR1 through the development and exploitation of a drug discovery platform based on small, engineered VNAR domains [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 222.
This work demonstrates the feasibility of making sensitive nanometer distance measurements betwee... more This work demonstrates the feasibility of making sensitive nanometer distance measurements between Fe(III) heme centers and nitroxide spin labels in proteins using the double electron-electron resonance (DEER) pulsed EPR technique at 94 GHz. Techniques to measure accurately long distances in many classes of heme proteins using DEER are currently strongly limited by sensitivity. In this paper we demonstrate sensitivity gains of more than 30 times compared with previous lower frequency (X-band) DEER measurements on both human neuroglobin and sperm whale myoglobin. This is achieved by taking advantage of recent instrumental advances, employing wideband excitation techniques based on composite pulses and exploiting more favorable relaxation properties of low-spin Fe(III) in high magnetic fields. This gain in sensitivity potentially allows the DEER technique to be routinely used as a sensitive probe of structure and conformation in the large number of heme and many other metalloproteins.
Uploads
Papers by Stacey Bell