Discriminatory ability of next-generation tau PET tracers for Alzheimer's disease.

A next generation of tau PET tracers for the imaging of Alzheimer's disease and other dementias has recently been developed. Whilst the new compounds have now entered clinical studies, there is limited information available to assess their suitability for clinical applications. Head-to-head comparisons are urgently needed to understand differences in the radiotracer binding profiles. We characterized the binding of the tau tracers PI2620, RO948, MK6240 and JNJ067 in human post-mortem brain tissue from a cohort of 25 dementia cases and age-matched controls using quantitative phosphorimaging with tritium-labelled radiotracers in conjunction with phospho-tau specific immunohistochemistry. The four radiotracers depicted tau inclusions composed of paired helical filaments with high specificity, both in cases with Alzheimer's disease and in primary tauopathy cases with concomitant Alzheimer's disease pathology. In contrast, cortical binding to primary tauopathy in cases without paired helical filament tau was found to be within the range of age-matched controls. Off-target binding to monoamine oxidase B has been overcome, as demonstrated by heterologous blocking studies in basal ganglia tissue. The high variability of cortical tracer binding within the Alzheimer's disease group followed the same pattern with each tracer, suggesting that all compounds are suited to differentiate Alzheimer's disease from other dementias.

Evaluation of a Bispidine-Based Chelator for Gallium-68 and of the Porphyrin Conjugate as PET/PDT Theranostic Agent.

In this study a bispidine ligand has been applied to the complexation of gallium(III) and radiolabelled with gallium-68 for the first time. Despite its 5-coordinate nature, the resulting complex is stable in serum for over two hours, demonstrating a ligand system well matched to the imaging window of gallium-68 positron emission tomography (PET). To show the versatility of the bispidine ligand and its potential use in PET, the bifunctional chelator was conjugated to a porphyrin, producing a PET/PDT-theranostic, which showed the same level of stability to serum as the non-conjugated gallium-68 complex. The PET/PDT complex killed >90 % of HT-29 cells upon light irradiation at 50 µm. This study shows bispidines have the versatility to be used as a ligand system for gallium-68 in PET.

Chemo- and regio-selective differential modification of native cysteines on an antibody via the use of dehydroalanine forming reagents.

Protein modification has garnered increasing interest over the past few decades and has become an important tool in many aspects of chemical biology. In recent years, much effort has focused on site-selective modification strategies that generate more homogenous bioconjugates, and this is particularly so in the antibody modification space. Modifying native antibodies by targeting solvent-accessible cysteines liberated by interchain disulfide reduction is, perhaps, the predominant strategy for achieving more site-selectivity on an antibody scaffold. This is evidenced by numerous approved antibody therapeutics that have utilised cysteine-directed conjugation reagents and the plethora of methods/strategies focused on antibody cysteine modification. However, all of these methods have a common feature in that after the reduction of native solvent-accessible cystines, the liberated cysteines are all reacted in the same manner. Herein, we report the discovery and application of dehydroalanine forming reagents (including novel reagents) capable of regio- and chemo-selectively modifying these cysteines (differentially) on a clinically relevant antibody fragment and a full antibody. We discovered that these reagents could enable differential reactivity between light chain C-terminal cysteines, heavy chain hinge region cysteines (cysteines with an adjacent proline residue, Cys-Pro), and other heavy chain internal cysteines. This differential reactivity was also showcased on small molecules and on the peptide somatostatin. The application of these dehydroalanine forming reagents was exemplified in the preparation of a dually modified antibody fragment and full antibody. Additionally, we discovered that readily available amide coupling agents can be repurposed as dehydroalanine forming reagents, which could be of interest to the broader field of chemical biology.

Cytokine elevation and transaminitis after laparoscopic donor nephrectomy.

Acute kidney injury frequently occurs in the critically ill and often progresses into multiorgan dysfunction syndrome, resulting in high mortality. We previously showed that nephrectomized mice had increased interleukin (IL)-6 and tumor necrosis factor (TNF)-α that directly contributed to systemic inflammation and hepatic injury. In this study, we examined whether patients undergoing laparoscopic donor nephrectomy have increased postoperative cytokine levels with injury to the liver and whether the remaining kidney sustains injury. Serial serum and urine samples were collected from 32 patients undergoing laparoscopic donor nephrectomy and 17 patients undergoing nonrenal laparoscopic surgery. Serum IL-6, IL-18, TNF-α and monocyte chemotactic protein-1 (MCP-1) (markers of systemic inflammation) and urinary neutrophil gelatinase-associated lipocalin (NGAL), kidney injury molecule-1 (KIM-1), MCP-1, and IL-18 (markers of acute kidney injury) were quantified by enzyme-linked immunosorbent assay. We also analyzed serum creatinine, aspartate transaminase (AST), and alanine transaminase to assess liver injury. Patients who underwent donor nephrectomy not only demonstrated increased serum creatinine but also had significant increases in serum IL-6, MCP-1, and AST. Serum TNF-α also trended upward in donor nephrectomy patients. Finally, the donor nephrectomy group showed increased urinary NGAL but not KIM-1 at 24 h. Taken together, our findings of increased serum IL-6, MCP-1, and AST after donor nephrectomy suggest that an acute reduction of kidney function induces systemic inflammation and may have distant effects on the liver. Further studies are needed to correlate increased urinary NGAL after donor nephrectomy both as a potential marker for renal tubular stress and/or hypertrophy in the contralateral kidney.

Adenosine and protection from acute kidney injury.

PURPOSE OF REVIEW: Acute kidney injury (AKI) is a major clinical problem without effective therapy. Development of AKI among hospitalized patients drastically increases mortality and morbidity. With increases in complex surgical procedures together with a growing elderly population, the incidence of AKI is rising. Renal adenosine receptor manipulation may have great therapeutic potential in mitigating AKI. In this review, we discuss renal adenosine receptor biology and potential clinical therapies for AKI. RECENT FINDINGS: The four adenosine receptor subtypes (A(1)AR, A(2A)AR, A(2B)AR, and A(3)AR) have diverse effects on the kidney. The pathophysiology of AKI may dictate the specific adenosine receptor subtype activation needed to produce renal protection. The A(1)AR activation in renal tubules and endothelial cells produces beneficial effects against ischemia and reperfusion injury by modulating metabolic demand, decreasing necrosis, apoptosis, and inflammation. The A(2A)AR protects against AKI by modulating leukocyte-mediated renal and systemic inflammation, whereas the A(2B)AR activation protects by direct activation of renal parenchymal adenosine receptors. In contrast, the A(1)AR antagonism may play a protective role in nephrotoxic AKI and radiocontrast induced nephropathy by reversing vascular constriction and inducing naturesis and diuresis. Furthermore, as the A(3)AR activation exacerbates apoptosis and tissue damage due to renal ischemia and reperfusion, selective A(3)AR antagonism may hold promise to attenuate renal ischemia and reperfusion injury. Finally, renal A(1)AR activation also protects against renal endothelial dysfunction caused by hepatic ischemia and reperfusion injury. SUMMARY: Despite the current lack of therapies for the treatment and prevention of AKI, recent research suggests that modulation of renal adenosine receptors holds promise in treating AKI and extrarenal injury.

Acute kidney injury and extrarenal organ dysfunction: new concepts and experimental evidence.

Acute kidney injury (AKI) is a frequent complication in the intensive care unit with limited therapeutic modalities. Although survival from isolated AKI has improved with recent advancements in renal replacement therapy, mortality from AKI complicated by multiorgan dysfunction has remained unchanged and is estimated to be approximately 50%. Hence, defining and better understanding the pathophysiology of distant organ dysfunction associated with AKI is clinically important because it may lead to new treatment strategies. In animal models, it has become increasingly clear that AKI is not an isolated event but results in remote organ dysfunction involving the heart, lungs, liver, intestines, and brain through an inflammatory mechanism that involves neutrophil migration, cytokine expression, and increased oxidative stress. The purpose of this brief review is to summarize the human and basic science evidence for AKI and its detrimental effects on distant organs.

Unearthing the unique stability of thiophosphonium-C-terminal cysteine adducts on peptides and proteins.

Herein we report a fundamental discovery on the use of tris(dialkylamino)phosphine reagents for peptide and protein modification. We discovered that C-terminal thiophosphonium species, which are uniquely stable, could be selectively and rapidly generated from their disulfide counterparts. In sharp and direct contrast, internal thiophosphonium species rapidly degrade to dehydroalanine. We demonstrate this remarkable chemoselectivity on a bis-cysteine model peptide, and the formation of a stable C-terminal-thiophosphonium adduct on an antibody fragment, as well as characterise the species in various small molecule/peptide studies.

Alcohol intake is associated with a decreased risk of developing primary biliary cholangitis.

BACKGROUND: Primary biliary cholangitis (PBC) is a chronic progressive liver disease of unknown aetiology characterised by immune-mediated destruction of small and medium-sized intrahepatic bile ducts. There are few well-established risk factors and epidemiological studies are needed to further evaluate the pathogenesis of the disease. AIM: To evaluate the relationship between alcohol intake, smoking and marijuana use with PBC development. METHODS: We conducted a prevalent case control study of 200 cases and 200 age (within a five year age band) and sex-matched controls, identified from the Victorian PBC prevalence study. We assessed lifetime alcohol intake and smoking behaviour (both tobacco and marijuana) prior to PBC onset and used conditional logistic regression for analyses. RESULTS: Alcohol intake consistently showed a dose-dependent inverse association with case status, and this was most substantial for 21-30 years and 31-40 years (P trend < 0.001). Smoking was associated with PBC, with a stronger association with a longer duration of smoking [e.g., adjusted OR 2.27 (95%CI: 1.12- 4.62) for those who had smoked for 20-35 years]. There was no association between marijuana use and PBC. CONCLUSION: Alcohol appears to have an inverse relationship with PBC. Smoking has been confirmed as an environmental risk factor for PBC. There was no association between marijuana use and PBC.

Synthesis of a porphyrin with histidine-like chelate: an efficient path towards molecular PDT/SPECT theranostics.

The goal of "personalised" medicine has seen a growing interest in the development of theranostic agents. Bifunctional, and targeted-trifunctional, theranostic water-soluble porphyrins with a histidine-like chelating group have been synthesised via copper-catalysed azide-alkyne cycloaddition (CuAAC) "click" chemistry in high yield and purity. They are capable of photodynamic treatment and [99mTc(CO)3]+ complexation for single-photon emission computed tomography (SPECT) imaging, with a radiochemical yield of >95%. The toxicity and phototoxicity were evaluated on HT-29 cells, DU145, and DU145-PSMA cell lines, with the targeted theranostic showing more potent phototoxicity towards DU145-PSMA expressing cells.

Selective radiolabelling with 68Ga under mild conditions: a route towards a porphyrin PET/PDT theranostic agent.

A theranostic conjugate for use as a positron emission tomography (PET) radiotracer and as a photosensitiser for photodynamic therapy (PDT) has been synthesised. A water-soluble porphyrin was coupled with the bifunctional chelate, H4Dpaa.ga. This conjugate is capable of rapid 68Ga complexation under physiological conditions; with 93% and 80% radiochemical yields achieved, at pH 4.5 and pH 7.4 respectively, in 15 min at 25 °C. Photocytotoxicity was evaluated on HT-29 cells and showed the conjugate was capable of >50% cell death at 50 µM upon irradiation with light, while causing minimal toxicity in the absence of light (>95% cell survival).

In vivo liver electroporation: optimization and demonstration of therapeutic efficacy.

Adverse effects (death and leukemogenesis) from viral vector-mediated gene therapy have renewed interest in plasmids as safer, more scalable, simple, and cost-effective vectors. Electroporation and hydrodynamic delivery are two techniques that improve the efficiency of plasmid-mediated gene transfer. The liver is a good tissue platform for targeted transfer of therapeutically relevant genes for correction of metabolic disorders, for example, hemophilia A. However, in vivo electroporation of liver has not yet been shown to achieve therapeutic efficacy of systemically active, secreted transgenic proteins. We have investigated the effect of field strength, pulse duration, pulse number, electrical waveforms, electrode contact area, plasmid administration routes, and injection technique on the efficiency of in vivo electrotransfer of naked plasmid to liver. Plasmid injection into a systemic vein was superior to intrahepatic injection. Unlike in vivo muscle electroporation, high-voltage pulses and microsecond pulses offered no advantage. Optimal electroporation conditions were 8-10 uni- or bipolar pulses of 20 msec, each at 250 V/cm. Using a nonhydrodynamic technique that greatly enhanced electrotransfer efficiency with minimal tissue injury, we demonstrate for the first time that liverdirected in vivo electroporation of factor VIII cDNA achieved significant phenotypic correction in hemophilic mice.