Rare variants in pharmacogenes influence clozapine metabolism in individuals with schizophrenia.

Clozapine is the only licensed medication for treatment-resistant schizophrenia (TRS). Few predictors for variation in response to clozapine have been identified, but clozapine metabolism is known to influence therapeutic response and adverse side effects. Here, we expand on genome-wide studies of clozapine metabolism, previously focused on common genetic variation, by analysing whole-exome sequencing data from 2062 individuals with schizophrenia taking clozapine in the UK. We investigated whether rare genomic variation in genes and gene sets involved in the clozapine metabolism pathway influences plasma concentrations of clozapine metabolites, assessed through the longitudinal analysis of 6585 pharmacokinetic assays. We observed a statistically significant association between the burden of rare damaging coding variants (MAF ≤ 1 %) in gene sets broadly related to drug pharmacokinetics and lower clozapine (ß = -0.054, SE = 0.019, P-value = 0.005) concentrations in plasma. We estimate that the effects in clozapine plasma concentrations of a single damaging allele in this gene set are akin to reducing the clozapine dose by about 35 mg/day. The gene-based analysis identified rare variants in CYP1A2, which encodes the enzyme responsible for converting clozapine to norclozapine, as having the strongest effects of any gene on clozapine metabolism (ß = 0.324, SE = 0.124, P = 0.009). Our findings support the hypothesis that rare genetic variants in known drug-metabolising enzymes and transporters can markedly influence clozapine plasma concentrations; these results suggest that pharmacogenomic efforts trying to predict clozapine metabolism and personalise drug therapy could benefit from the inclusion of rare damaging variants in pharmacogenes beyond those already identified and catalogued as PGx star alleles.

Mediation and longitudinal analysis to interpret the association between clozapine pharmacokinetics, pharmacogenomics, and absolute neutrophil count.

Clozapine is effective at reducing symptoms of treatment-resistant schizophrenia, but it can also induce several adverse outcomes including neutropenia and agranulocytosis. We used linear mixed-effect models and structural equation modelling to determine whether pharmacokinetic and genetic variables influence absolute neutrophil count in a longitudinal UK-based sample of clozapine users not currently experiencing neutropenia (N = 811). Increased daily clozapine dose was associated with elevated neutrophil count, amounting to a 133 cells/mm3 rise per standard deviation increase in clozapine dose. One-third of the total effect of clozapine dose was mediated by plasma clozapine and norclozapine levels, which themselves demonstrated opposing, independent associations with absolute neutrophil count. Finally, CYP1A2 pharmacogenomic activity score was associated with absolute neutrophil count, supporting lower neutrophil levels in CYP1A2 poor metabolisers during clozapine use. This information may facilitate identifying at-risk patients and then introducing preventative interventions or individualised pharmacovigilance procedures to help mitigate these adverse haematological reactions.

Human dispersals out of Africa via the Levant.

Homo sapiens dispersed from Africa into Eurasia multiple times in the Middle and Late Pleistocene. The route, across northeastern Africa into the Levant, is a viable terrestrial corridor, as the present harsh southern Levant would probably have been savannahs and grasslands during the last interglaciation. Here, we document wetland sediments with luminescence ages falling in the last interglaciation in the southern Levant, showing protracted phases of moisture availability. Wetland sediments in Wadi Gharandal containing Levallois artifacts yielded an age of 84 ka. Our findings support the growing consensus for a well-watered Jordan Rift Valley that funneled migrants into western Asia and northern Arabia.

Lateral Bone Augmentation Using a Three-Dimensional-Printed Polymeric Chamber to Compare Biomaterials.

The aim of this study was to test the suitability of calcium phosphate cement mixed with poly(lactic-co-glycolic acid) (CPC-PLGA) microparticles into a ring-shaped polymeric space-maintaining device as bone graft material for lateral bone augmentation. Therefore, the bone chambers were installed on the lateral portion of the anterior region of the mandibular body of mini-pigs. Chambers were filled with either CPC-PLGA or BioOss® particles for comparison and left for 4 and 12 weeks. Histology and histomorphometry were used to obtain temporal insight in material degradation and bone formation. Results indicated that between 4 and 12 weeks of implantation, a significant degradation of the CPC-PLGA (from 75.1% to 23.1%), as well as BioOss material, occurred (from 40.6% to 14.4%). Degradation of both materials was associated with the presence of macrophage-like and osteoclast-like cells. Furthermore, a significant increase in bone formation occurred between 4 and 12 weeks for the CPC-PLGA (from 0.1% to 7.2%), as well as BioOss material (from 8.3% to 23.3%). Statistical analysis showed that bone formation had progressed significantly better using BioOss compared to CPC-PLGA (p < 0.05). In conclusion, this mini-pig study showed that CPC-PLGA does not stimulate lateral bone augmentation using a bone chamber device. Both treatments failed to achieve "clinically" meaningful alveolar ridge augmentation.

Influence of bisphosphonate treatment on bone substitute performance in osteoporotic conditions.

OBJECTIVE: Considering the elevated number of osteoporotic patients in need of bone graft procedures, we here evaluated the effect of alendronate (ALN) treatment on the regeneration of bone defects in osteoporotic rats. Bone formation was histologically and histomorphometrically assessed in rat femoral condyle bone defects filled with bone graft (Bio-Oss®) or left empty. METHODS: Male Wistar rats were induced osteoporotic through orchidectomy (ORX) and SHAM-operated. The animals were divided into three groups: osteoporotic (ORX), osteoporotic treated with ALN (ORX + ALN) and healthy (SHAM). Six weeks after ORX or SHAM surgeries, bone defects were created bilaterally in femoral condyles; one defect was filled with Bio-Oss® and the other one left empty. Bone regeneration within the defects was analyzed by histology and histomorphometry after 4 and 12 weeks. RESULTS: Histological samples showed new bone surrounding Bio-Oss® particles from week 4 onward in all three groups. At week 12, the data further showed that ALN treatment of osteoporotic animals enhanced bone formation to a 10-fold increase compared to non-treated osteoporotic control. Bio-Oss® filling of the defects promoted bone formation at both implantation periods compared to empty controls. CONCLUSION: Our histological and histomorphometric results demonstrate that the enteral administration of alendronate under osteoporotic bone conditions leverages bone defect regeneration to a level comparable to that in healthy bone. Additionally, Bio-Oss® is an effective bone substitute, increasing bone formation, and acting as an osteoconductive scaffold guiding bone growth in both healthy and osteoporotic bone conditions. SIGNIFICANCE: Based on the results of this study, enteral use of ALN mitigates adverse effects of an osteoporotic condition on bone defect regeneration.

Calcium Phosphate Ceramics and Synergistic Bioactive Agents for Osteogenesis in Implant Dentistry.

Implant-supported dental prosthetics are widely used in dental practice. Sufficient peri-implant bone tissue is a crucial prerequisite for the long-term success of this treatment, as insufficient peri-implant bone volume hampers dental implant installation and negatively influences dental implant stability. However, due to tooth extraction, bone metabolism diseases, and trauma, bone defects in the jaw are common in patients, particularly in the elderly and those suffering from underlying conditions. If this is the case, the alveolar ridge has to be augmented for reliable implant placement. Various biomaterials, growth factors (GFs) or GF-based products, and trace elements have been tested and used for alveolar ridge augmentation. Among those biomaterials, calcium phosphates (CaPs) are the most popular due to their promising biocompatibility, great osteoconductivity, and distinguishing osteogenesis. Combining CaPs with GFs or trace elements can further favor bone defect repair. This review mainly focuses on applying artificial CaP biomaterials and their combination with bioactive agents to repair bone defects in implant dentistry.

Repeated Application and Removal of Polyisocyanopeptide Hydrogel Wound Dressings in a Splinted Full-Thickness Wound Model.

Polyisocyanopeptide (PIC) hydrogels are proposed as promising wound dressings. These gels are thermo-sensitive, allow application as a cold liquid, and rely on gelation through body heat. It is supposed that the gel can be easily removed by reversing the gelation and washing it away with a cold irrigation solution. The impact on wound healing of the regular application and removal of PIC dressings is compared to a single application of PIC and the clinically used Tegaderm™ in murine splinted full-thickness wounds for up to 14 days. SPECT/CT analysis of 111In-labelled PIC gels showed that, on average, 58% of the PIC gel could be washed out of the wounds with the employed method, which is, however, heavily influenced by personal technique. Evaluation with photography and (immuno-)histology showed that wounds in which PIC dressings were regularly removed and replaced were smaller at 14 days post-injury but performed on par with the control treatment. Moreover, the encapsulation of PIC in wound tissue was less severe and occurred less often when PIC was regularly refreshed. In addition, no morphological damage related to the removal procedure was observed. Thus, PIC gels are atraumatic and perform similarly to currently employed wound dressing materials, offering possible future benefits for both clinicians and patients.

Repair of complex ovine segmental mandibulectomy utilizing customized tissue engineered bony flaps.

Craniofacial defects require a treatment approach that provides both robust tissues to withstand the forces of mastication and high geometric fidelity that allows restoration of facial architecture. When the surrounding soft tissue is compromised either through lack of quantity (insufficient soft tissue to enclose a graft) or quality (insufficient vascularity or inducible cells), a vascularized construct is needed for reconstruction. Tissue engineering using customized 3D printed bioreactors enables the generation of mechanically robust, vascularized bony tissues of the desired geometry. While this approach has been shown to be effective when utilized for reconstruction of non-load bearing ovine angular defects and partial segmental defects, the two-stage approach to mandibular reconstruction requires testing in a large, load-bearing defect. In this study, 5 sheep underwent bioreactor implantation and the creation of a load-bearing mandibular defect. Two bioreactor geometries were tested: a larger complex bioreactor with a central groove, and a smaller rectangular bioreactor that were filled with a mix of xenograft and autograft (initial bone volume/total volume BV/TV of 31.8 ± 1.6%). At transfer, the tissues generated within large and small bioreactors were composed of a mix of lamellar and woven bone and had BV/TV of 55.3 ± 2.6% and 59.2 ± 6.3%, respectively. After transfer of the large bioreactors to the mandibular defect, the bioreactor tissues continued to remodel, reaching a final BV/TV of 64.5 ± 6.2%. Despite recalcitrant infections, viable osteoblasts were seen within the transferred tissues to the mandibular site at the end of the study, suggesting that a vascularized customized bony flap is a potentially effective reconstructive strategy when combined with an optimal stabilization strategy and local antibiotic delivery prior to development of a deep-seated infection.

Pharmacokinetics and pharmacogenomics of clozapine in an ancestrally diverse sample: a longitudinal analysis and genome-wide association study using UK clinical monitoring data.

BACKGROUND: The antipsychotic, clozapine, is the only licensed drug against the treatment-resistant symptoms that affect 20-30% of people with schizophrenia. Clozapine is markedly underprescribed, partly because of concerns about its narrow therapeutic range and adverse drug reaction profile. Both concerns are linked to drug metabolism, which varies across populations globally and is partly genetically determined. Our study aimed to use a cross-ancestry genome-wide association study (GWAS) design to investigate variations in clozapine metabolism within and between genetically inferred ancestral backgrounds, to discover genomic associations to clozapine plasma concentrations, and to assess the effects of pharmacogenomic predictors across different ancestries. METHODS: In this GWAS, we analysed data from the UK Zaponex Treatment Access System clozapine monitoring service as part of the CLOZUK study. We included all available individuals with clozapine pharmacokinetic assays requested by their clinicians. We excluded people younger than 18 years, or whose records contained clerical errors, or with blood drawn 6-24 h after dose, a clozapine or norclozapine concentration less than 50 ng/mL, a clozapine concentration of more than 2000 ng/mL, a clozapine-to-norclozapine ratio outside of the 0·5-3·0 interval, or a clozapine dose of more than 900 mg/day. Using genomic information, we identified five biogeographical ancestries: European, sub-Saharan African, north African, southwest Asian, and east Asian. We did pharmacokinetic modelling, a GWAS, and a polygenic risk score association analysis using longitudinal regression analysis with three primary outcome variables: two metabolite plasma concentrations (clozapine and norclozapine) and the clozapine-to-norclozapine ratio. FINDINGS: 19 096 pharmacokinetic assays were available for 4760 individuals in the CLOZUK study. After data quality control, 4495 individuals (3268 [72·7%] male and 1227 [27·3%] female; mean age 42·19 years [range 18-85]) linked to 16 068 assays were included in this study. We found a faster average clozapine metabolism in people of sub-Saharan African ancestry than in those of European ancestry. By contrast, individuals with east Asian or southwest Asian ancestry were more likely to be slow clozapine metabolisers than those with European ancestry. Eight pharmacogenomic loci were identified in the GWAS, seven with significant effects in non-European groups. Polygenic scores generated from these loci were associated with clozapine outcome variables in the whole sample and within individual ancestries; the maximum variance explained was 7·26% for the metabolic ratio. INTERPRETATION: Longitudinal cross-ancestry GWAS can discover pharmacogenomic markers of clozapine metabolism that, individually or as polygenic scores, have consistent effects across ancestries. Our findings suggest that ancestral differences in clozapine metabolism could be considered for optimising clozapine prescription protocols for diverse populations. FUNDING: UK Academy of Medical Sciences, UK Medical Research Council, and European Commission.

Fast Degradable Calcium Phosphate Cement for Maxillofacial Bone Regeneration.

The aim of this preclinical study was to test the applicability of calcium phosphate cement (CPC)-poly(lactic-co-glycolic acid) (PLGA)-carboxymethylcellulose (CMC) as a bone substitute material for guided bone regeneration (GBR) procedures in a clinically relevant mandibular defect model in minipigs. In the study, a predicate device (i.e., BioOss®) was included for comparison. Critical-sized circular mandibular bone defects were created and filled with either CPC-PLGA-CMC without coverage with a GBR membrane or BioOss covered with a GBR membrane and left to heal for 4 and 12 weeks to obtain temporal insight in material degradation and bone formation. Bone formation increased significantly for both CPC-PLGA-CMC and BioOss with increasing implantation time. Further, no significant differences were found for bone formation at either 4 or 12 weeks between CPC-PLGA-CMC and BioOss. Finally, bone substitute material degradation increased significantly for both CPC-PLGA-CMC and BioOss from 4 to 12 weeks of implantation, showing the highest degradation for CPC-PLGA-CMC (∼85%) compared to BioOss (∼12%). In conclusion, this minipig study showed that CPC-PLGA-CMC can be used as a bone-grafting material and stimulates bone regeneration to a comparable extent as with BioOss particles. Importantly, CPC-PLGA-CMC degrades faster compared to BioOss, is easier to apply into a bone defect, and does not need the use of an additional GBR membrane. Consequently, the data support the further investigation of CPC-PLGA-CMC in human clinical trials. Impact statement Guided bone regeneration (GBR) is a frequently used dental surgical technique to regenerate the alveolar ridge to allow stable implant installation. However, stabilization of the GBR membrane and avoidance of bone graft movement remain a challenge. Consequently, there is need for the development of alternative materials to be used in GBR procedures that are easier to apply and induce predictable bone regeneration. In this minipig study, we focused on the applicability of calcium phosphate cement-poly(lactic-co-glycolic acid)-carboxymethylcellulose as an alternative bone substitute material for GBR procedures without the need of an additional GBR membrane.

Genomic Stratification of Clozapine Prescription Patterns Using Schizophrenia Polygenic Scores.

BACKGROUND: Treatment-resistant schizophrenia affects approximately 30% of individuals with the disorder. Clozapine is the medication of choice in treatment-resistant schizophrenia, but optimizing administration and dose titration is complex. The identification of factors influencing clozapine prescription and response, including genetics, is of interest in a precision psychiatry framework. METHODS: We used linear regression models accounting for demographic, pharmacological, and clinical covariates to determine whether a polygenic risk score (PRS) for schizophrenia would be associated with the highest dose recorded during clozapine treatment. Analyses were performed across 2 independent multiancestry samples of individuals from a UK patient monitoring system, CLOZUK2 (n = 3133) and CLOZUK3 (n = 909), and a European sample from a Norwegian therapeutic drug monitoring service (n = 417). In a secondary analysis merging both UK cohorts, logistic regression models were used to estimate the relationship between schizophrenia PRSs and clozapine doses classified as low, standard, or high. RESULTS: After controlling for relevant covariates, the schizophrenia PRS was correlated with the highest clozapine dose on record for each individual across all samples: CLOZUK2 (ß = 12.22, SE = 3.78, p = .001), CLOZUK3 (ß = 12.73, SE = 5.99, p = .034), and the Norwegian cohort (ß = 46.45, SE = 18.83, p = .014). In a secondary analysis, the schizophrenia PRS was associated with taking clozapine doses >600 mg/day (odds ratio = 1.279, p = .006). CONCLUSIONS: The schizophrenia PRS was associated with the highest clozapine dose prescribed for an individual in records from 3 independent samples, suggesting that the genetic liability for schizophrenia might index factors associated with therapeutic decisions in cohorts of patients with treatment-resistant schizophrenia.

Applied Geophysics for Managed Aquifer Recharge.

Increasing water stress and decreasing supplies caused by growth and climate variability have expanded demand for managed aquifer recharge (MAR) projects to provide water supply resilience. Some of the most important factors in determining the performance of a MAR project include site selection, subsurface hydrogeologic characteristics and associated properties of the storage zone. Costs for invasive subsurface investigations to address these factors have slowly increased over the past two decades, with drilling costs increasing dramatically by as much as 30% or more since COVID-19 hit, a result of supply chain issues, steel prices, and manpower challenges. This paper provides a high-level review of major geophysical methods that have become more mainstream over the past decade or two to supplement invasive subsurface investigations and are very cost effective when compared to drilling boreholes and installing wells, which provide only point data. The more commonly used surface geophysical methods include ground-based and airborne time-domain electromagnetic methods (TEM), electrical resistivity, and seismic reflection. Airborne TEM methods (AEM) collect data very quickly, avoiding ground-based access constraints, and land-based methods are especially efficient using towed arrays. Electrical resistivity measurements provide resolution comparable to TEM but require more time than towed methods. Seismic reflection surveys are more expensive than other methods but typically have a much greater depth of penetration and can provide high resolution information on aquifer geometry, geology, and faults. Borehole geophysics is one of the more common methods used in MAR, providing near hole formation data and ground truths surface geophysics.

A Mini-Pig Mandibular Defect Model for Evaluation of Craniomaxillofacial Bone Regeneration.

Craniomaxillofacial bone defects represent a clinical challenge in the fields of maxillofacial surgery and (implant) dentistry. Regeneration of these bone defects requires the application of bone graft materials that facilitate new bone formation in a safe, reliable, and predictive manner. In addition to autologous bone graft, several types of (synthetic) bone substitute materials have become clinically available, and still major efforts are focused on improving such bone substitute materials by optimizing their properties. Given the regulatory necessity to evaluate the performance of new bone substitute materials for craniomaxillofacial bone regeneration in a large animal model with similarity to human bone before clinical application, we here describe a mini-pig mandibular bone defect model that allows for the creation of multiple (critical-size) bone defects within the mandibular body of a single animal. As examples of bone substitute materials, we utilize both the clinically used BioOss granules and an experimental calcium phosphate cement for filling the created defects. Regarding the latter, its advantages are the injectable application within the defect site, in which the material rapidly sets, and the tailorable degradation properties via the inclusion of hydrolytically degrading polymeric particles. For both bone substitute materials, we show the suitability of the bone defect model to assess bone regeneration via histology and micro-computed tomography. Impact statement Given the regulatory necessity to evaluate the performance of new bone substitute materials for craniomaxillofacial bone regeneration in a large animal model with similarity to the human bone before clinical application, we here describe a mini-pig mandibular bone defect model that allows for the creation of multiple (critical-size) bone defects within the mandibular body of a single animal that can be used for the evaluation of the bone regenerative capacity of new bone grafting materials as well as tissue-engineered products for alveolar bone regeneration.

Histological evaluation of titanium fiber mesh-coated implants in a rabbit femoral condyle model.

OBJECTIVES: This study was aimed to comparatively evaluate new bone formation into the pores of a flexible titanium fiber mesh (TFM) applied on the surface of implant. METHODS: Twenty-eight custom made cylindrical titanium implants (4 ×10 mm) with and without a layer of two different types of TFM (fiber diameter of 22 µm and 50 µm, volumetric porosity ~70%) were manufactured and installed bilaterally in the femoral condyles of 14 rabbits. The elastic modulus for these two TFM types was ~20 GPa and ~5 GPa respectively, whereas the solid titanium was ~110 GPa. The implants (Control, TFM-22, TFM-50) were retrieved after 14 weeks of healing and prepared for histological assessment. The percentage of the bone area (BA%), the bone-to-implant contact (BIC%) and amount were determined. RESULTS: Newly formed bone into mesh porosity was observed for all three types of implants. Histomorphometric analyses revealed significantly higher (~2.5 fold) BA% values for TFM-22 implants (30.9 ± 9.5%) compared to Control implants (12.7 ± 6.0%), whereas BA% for TMF-50 did not significantly differ compared with Control implants. Furthermore, both TFM-22 and TFM-50 implants showed significantly higher BIC% values (64.9 ± 14.0%, ~2.5 fold; 47.1 ± 14.1%, ~2 fold) compared to Control (23.6 ± 17.4%). Finally, TFM-22 implants showed more and thicker trabeculae in the peri-implant region. SIGNIFICANCE: This in vivo study demonstrated that implants with a flexible coating of TFM improve bone formation within the inter-fiber space and the peri-implant region.

Desertification of Iran in the early twenty-first century: assessment using climate and vegetation indices.

Remote sensing of specific climatic and biogeographical parameters is an effective means of evaluating the large-scale desertification status of drylands affected by negative human impacts. Here, we identify and analyze desertification trends in Iran for the period 2001-2015 via a combination of three indices for vegetation (NPP-net primary production, NDVI-normalized difference vegetation index, LAI-leaf area index) and two climate indices (LST-land surface temperature, P-precipitation). We combine these indices to identify and map areas of Iran that are susceptible to land degradation. We then apply a simple linear regression method, the Mann-Kendall non-parametric test, and the Theil-Sen estimator to identify long-term temporal and spatial trends within the data. Based on desertification map, we find that 68% of Iran shows a high to very high susceptibility to desertification, representing an area of 1.1 million km2 (excluding 0.42 million km2 classified as unvegetated). Our results highlight the importance of scale in assessments of desertification, and the value of high-resolution data, in particular. Annually, no significant change is evident within any of the five indices, but significant changes (some positive, some negative) become apparent on a seasonal basis. Some observations follow expectations; for instance, NDVI is strongly associated with cooler, wet spring and summer seasons, and milder winters. Others require more explanation; for instance, vegetation appears decoupled from climatic forcing during autumn. Spatially, too, there is much local and regional variation, which is lost when the data are considered only at the largest nationwide scale. We identify a northwest-southeast belt spanning central Iran, which has experienced significant vegetation decline (2001-2015). We tentatively link this belt of land degradation with intensified agriculture in the hinterlands of Iran's major cities. The spatial and temporal trends identified with the three vegetation and two climate indices afford a cost-effective framework for the prediction and management of future environmental trends in developing regions at risk of desertification.