title: The effects of allelic hinge variation in IgG3 antibodies on protein dynamics and effector functions

authors: Kuplich Barcellos, Marina

abstract: Antibodies play a crucial role in eliminating infections by recognising foreign antigens with high specificity and affinity. If a target is not directly neutralised, antibodies can mediate potent effector function responses by coordinating the recruitment of immune cells through their constant regions. These powerful immune functions are being widely explored in the form of IgG-based monoclonal antibody therapies for the treatment of infections, cancer, and autoimmune diseases. However, the antibody constant region exhibits significant genetic polymorphism, and new variants are rapidly being discovered by advances in genomic sequencing techniques. The documented effects of this diversity include differences in structural dynamics, thermal stability, and Fc-mediated effector functions among a number of antibody subclasses, including IgG1, IgG2, and IgG3. Some variants are reported to be a contributing factor to variability in the outcome of disease and responsiveness to vaccines in humans. The IgG3 subclass, in particular, displays striking differences in the antibody hinge region – responsible for connecting target binding with constant-region-mediated immune function. Here, exon duplications mean the hinge region can vary by as many as 45 amino acids in length. This work seeks to better understand some of the functional consequences of this unusual IgG3 variation. 

To assess the effects of allelic hinge variation on protein dynamics and effector functions, we have expressed a panel of seven anti-CD20 antibodies with unique constant region allele sequences from subclasses IgG1, IgG2, and IgG3. Sequence verified plasmids were used to express full-length antibodies in HEK293 cells at high purity. We first used small angle X-ray scattering (SAXS) to assess the variability in size and structural flexibility between the allelic variants and compared our data with previous research on the same variant panel with antiHER2 variable domain sequences. Following this structural analysis, we next undertook ADCC (antibody-dependent cellular cytotoxicity) and ADCP (antibody dependent cellular phagocytosis) assays to determine if differences in structural dynamics between the polymorphic variants could be linked to differences in effector function. These assays specifically used genome-engineered reporter cell lines to evaluate the ability of allelic variants to engage constant region binding FcyRIIIa and FcyRIIa Fc receptors, typically found on peripheral blood mononuclear cells. Activation was measured by quantifying the levels of luciferase expressed as a reporter.  

The findings provided in this study demonstrate that allelic variants with longer hinges have larger and more flexible structures and tend to present weaker ADCC and ADCP responses. The data presented here is the basis for future work on the influence of IgG constant region diversity on antibody structural dynamics and effector functions, which could contribute to improvements in the design of therapeutic antibodies.

text: The effects of allelic hinge variation in IgG3 antibodies on protein dynamics and effector functions Antibodies play a crucial role in eliminating infections by recognising foreign antigens with high specificity and affinity. If a target is not directly neutralised, antibodies can mediate potent effector function responses by coordinating the recruitment of immune cells through their constant regions. These powerful immune functions are being widely explored in the form of IgG-based monoclonal antibody therapies for the treatment of infections, cancer, and autoimmune diseases. However, the antibody constant region exhibits significant genetic polymorphism, and new variants are rapidly being discovered by advances in genomic sequencing techniques. The documented effects of this diversity include differences in structural dynamics, thermal stability, and Fc-mediated effector functions among a number of antibody subclasses, including IgG1, IgG2, and IgG3. Some variants are reported to be a contributing factor to variability in the outcome of disease and responsiveness to vaccines in humans. The IgG3 subclass, in particular, displays striking differences in the antibody hinge region – responsible for connecting target binding with constant-region-mediated immune function. Here, exon duplications mean the hinge region can vary by as many as 45 amino acids in length. This work seeks to better understand some of the functional consequences of this unusual IgG3 variation. 

To assess the effects of allelic hinge variation on protein dynamics and effector functions, we have expressed a panel of seven anti-CD20 antibodies with unique constant region allele sequences from subclasses IgG1, IgG2, and IgG3. Sequence verified plasmids were used to express full-length antibodies in HEK293 cells at high purity. We first used small angle X-ray scattering (SAXS) to assess the variability in size and structural flexibility between the allelic variants and compared our data with previous research on the same variant panel with antiHER2 variable domain sequences. Following this structural analysis, we next undertook ADCC (antibody-dependent cellular cytotoxicity) and ADCP (antibody dependent cellular phagocytosis) assays to determine if differences in structural dynamics between the polymorphic variants could be linked to differences in effector function. These assays specifically used genome-engineered reporter cell lines to evaluate the ability of allelic variants to engage constant region binding FcyRIIIa and FcyRIIa Fc receptors, typically found on peripheral blood mononuclear cells. Activation was measured by quantifying the levels of luciferase expressed as a reporter.  

The findings provided in this study demonstrate that allelic variants with longer hinges have larger and more flexible structures and tend to present weaker ADCC and ADCP responses. The data presented here is the basis for future work on the influence of IgG constant region diversity on antibody structural dynamics and effector functions, which could contribute to improvements in the design of therapeutic antibodies.

year: 2024

title: Monitoring the landslide at Bramley Drive, Tauranga, NZ

authors: Moon, Vicki G.

subjects: landslide monitoring

abstract: Omokoroa Peninsula, Tauranga Harbour, is prone to landslides in sensitive pyroclastic soils, especially in coastal bluffs. The largest is the landslide at Bramley Drive that first occurred in 1979, and was reactivated in 2011 and 2012. Since 2012 the landslide has been monitored with laser scans, vibro-and static-CPT, pore water logging at 3 depths, and a borehole inclinometer. Laser scan results track degradation of the scarp and allow development of a preliminary magnitude-frequency curve for failure events on the scarp. To date the borehole inclinometer has shown no obvious shear surface development. However, deformations in phase with the solid earth tides are evident in the cumulative displacement plots. Layers of weak soils separated by sharp boundaries are believed to exaggerate the deformations of the solid earth tides to the extent that they are measurable with a simple inclinometer. Residual deformation after subtraction of the earth tide effects indicate some movement over winter of 2014 associated with sensitive soils at or near the failure surface. The depth of this movement corresponds with a zone of high induced pore water pressures under vibratory CPTu. Pore water pressures indicate two discrete aquifers: an upper aquifer in tephra layers high in the upper part of the sequence that responds to atmospheric pressures; and a second aquifer in the underlying ignimbrites. Pressures in the bottom aquifer are lower than in the overlying aquifer in summer and higher in winter. Large spikes in pore water pressure have been observed during winter of 2014; these coincide with the time of deformation noted in the inclinometer traces.

text: Monitoring the landslide at Bramley Drive, Tauranga, NZ Omokoroa Peninsula, Tauranga Harbour, is prone to landslides in sensitive pyroclastic soils, especially in coastal bluffs. The largest is the landslide at Bramley Drive that first occurred in 1979, and was reactivated in 2011 and 2012. Since 2012 the landslide has been monitored with laser scans, vibro-and static-CPT, pore water logging at 3 depths, and a borehole inclinometer. Laser scan results track degradation of the scarp and allow development of a preliminary magnitude-frequency curve for failure events on the scarp. To date the borehole inclinometer has shown no obvious shear surface development. However, deformations in phase with the solid earth tides are evident in the cumulative displacement plots. Layers of weak soils separated by sharp boundaries are believed to exaggerate the deformations of the solid earth tides to the extent that they are measurable with a simple inclinometer. Residual deformation after subtraction of the earth tide effects indicate some movement over winter of 2014 associated with sensitive soils at or near the failure surface. The depth of this movement corresponds with a zone of high induced pore water pressures under vibratory CPTu. Pore water pressures indicate two discrete aquifers: an upper aquifer in tephra layers high in the upper part of the sequence that responds to atmospheric pressures; and a second aquifer in the underlying ignimbrites. Pressures in the bottom aquifer are lower than in the overlying aquifer in summer and higher in winter. Large spikes in pore water pressure have been observed during winter of 2014; these coincide with the time of deformation noted in the inclinometer traces.

year: 2015

title: Māori men, relationships, and everyday practices: towards broadening domestic violence research

authors: King, Pita

subjects: Māori

abstract: Relationships are central to the health and wellbeing of Māori (indigenous people of New Zealand). Through processes of colonisation, cultural ways of relatedness embedded within Māori social structures experienced disruption and were reshaped over decades of assimilation. Māori knowledge and everyday practices that assisted in protecting Māori from societal problems, such as domestic violence, began to dwindle. In contemporary New Zealand, Māori are over-represented in domestic violence statistics. Utilising an auto-ethnographic approach and case studies, our research focuses on five Māori men’s experiences within intimate relationships and whānau (extended family) life. A significant feature of this research is that it provides insights into the ways Māori men draw on their cultural ways-of-being to enhance intimate relationships and maintain bonds within whānau and community life to forge new ways-of-being. Such insights have the potential to inform preventative measures against domestic violence within Māori communities.

text: Māori men, relationships, and everyday practices: towards broadening domestic violence research Relationships are central to the health and wellbeing of Māori (indigenous people of New Zealand). Through processes of colonisation, cultural ways of relatedness embedded within Māori social structures experienced disruption and were reshaped over decades of assimilation. Māori knowledge and everyday practices that assisted in protecting Māori from societal problems, such as domestic violence, began to dwindle. In contemporary New Zealand, Māori are over-represented in domestic violence statistics. Utilising an auto-ethnographic approach and case studies, our research focuses on five Māori men’s experiences within intimate relationships and whānau (extended family) life. A significant feature of this research is that it provides insights into the ways Māori men draw on their cultural ways-of-being to enhance intimate relationships and maintain bonds within whānau and community life to forge new ways-of-being. Such insights have the potential to inform preventative measures against domestic violence within Māori communities.

year: 2017