Gene Edited T Cell Therapies for Inborn Errors of Immunity.

Inborn errors of immunity (IEIs) are a heterogeneous group of inherited disorders of the immune system. Many IEIs have a severe clinical phenotype that results in progressive morbidity and premature mortality. Over 450 IEIs have been described and the incidence of all IEIs is 1/1,000-10,000 people. Current treatment options are unsatisfactory for many IEIs. Allogeneic haematopoietic stem cell transplantation (alloHSCT) is curative but requires the availability of a suitable donor and carries a risk of graft failure, graft rejection and graft-versus-host disease (GvHD). Autologous gene therapy (GT) offers a cure whilst abrogating the immunological complications of alloHSCT. Gene editing (GE) technologies allow the precise modification of an organisms' DNA at a base-pair level. In the context of genetic disease, this enables correction of genetic defects whilst preserving the endogenous gene control machinery. Gene editing technologies have the potential to transform the treatment landscape of IEIs. In contrast to gene addition techniques, gene editing using the CRISPR system repairs or replaces the mutation in the DNA. Many IEIs are limited to the lymphoid compartment and may be amenable to T cell correction alone (rather than haematopoietic stem cells). T cell Gene editing has the advantages of higher editing efficiencies, reduced risk of deleterious off-target edits in terminally differentiated cells and less toxic conditioning required for engraftment of lymphocytes. Although most T cells lack the self-renewing property of HSCs, a population of T cells, the T stem cell memory compartment has long-term multipotent and self-renewal capacity. Gene edited T cell therapies for IEIs are currently in development and may offer a less-toxic curative therapy to patients affected by certain IEIs. In this review, we discuss the history of T cell gene therapy, developments in T cell gene editing cellular therapies before detailing exciting pre-clinical studies that demonstrate gene editing T cell therapies as a proof-of-concept for several IEIs.

Earthquakes indicated magma viscosity during Kilauea's 2018 eruption.

Magma viscosity strongly controls the style (for example, explosive versus effusive) of a volcanic eruption and thus its hazard potential, but can only be measured during or after an eruption. The identification of precursors indicative of magma viscosity would enable forecasting of the eruption style and the scale of associated hazards1. The unanticipated May 2018 rift intrusion and eruption of Kilauea Volcano, Hawai'i2 displayed exceptional chemical and thermal variability in erupted lavas, leading to unpredictable effusion rates and explosivity. Here, using an integrated analysis of seismicity and magma rheology, we show that the orientation of fault-plane solutions (which indicate a fault's orientation and sense of movement) for earthquakes preceding and accompanying the 2018 eruption indicate a 90-degree local stress-field rotation from background, a phenomenon previously observed only at high-viscosity eruptions3, and never before at Kilauea4-8. Experimentally obtained viscosities for 2018 products and earlier lavas from the Pu'u 'O'o vents tightly constrain the viscosity threshold required for local stress-field reorientation. We argue that rotated fault-plane solutions in earthquake swarms at Kilauea and other volcanoes worldwide provide an early indication that unrest involves magma of heightened viscosity, and thus real-time monitoring of the orientations of fault-plane solutions could provide critical information about the style of an impending eruption. Furthermore, our results provide insight into the fundamental nature of coupled failure and flow in complex multiphase systems.

The cascading origin of the 2018 Kilauea eruption and implications for future forecasting.

The 2018 summit and flank eruption of Kilauea Volcano was one of the largest volcanic events in Hawai'i in 200 years. Data suggest that a backup in the magma plumbing system at the long-lived Pu'u 'O'o eruption site caused widespread pressurization in the volcano, driving magma into the lower flank. The eruption evolved, and its impact expanded, as a sequence of cascading events, allowing relatively minor changes at Pu'u 'O'o to cause major destruction and historic changes across the volcano. Eruption forecasting is inherently challenging in cascading scenarios where magmatic systems may prime gradually and trigger on small events.

First 3D imaging characterization of Pele's hair from Kilauea volcano (Hawaii).

In this work the morphologic features of Pele's hair formed during three different eruptions of Kilauea volcano have been investigated: fountaining from Kilauea Iki's 1959 Episode 1, weak explosive activity from Halemaumau lava lake and littoral explosions at Waikupanaha (2009). Morphological studies were performed by optical, stereo- and scanning electron microscopy. For the first time 3D image analysis was carried out by synchrotron radiation X-ray computed microtomography, which allowed a high-resolution 3D reconstruction of the internal structure of each Pele's hair, highlighting several differences in terms of number density, elongation and shape of the vesicles between the samples from the three eruptions. We identified three main parameters determining these differences: initial size of the magma droplet, ejection velocity and magma viscosity. Pele's hair erupted during the Kilauea Iki's fountaining shows the highest thickness and the least elongated shape of the vesicles, though it is related to fast ejection of a low viscosity magma. We therefore suggest that the size of magma droplets is the main parameter influencing the morphology and inner textures of the Pele's hair. The comparison with Pele's hair of similar eruptions elsewhere demonstrates that there is no univocal correspondence between eruptive style and Pele's hair texture.

The 2018 rift eruption and summit collapse of Kilauea Volcano.

In 2018, Kilauea Volcano experienced its largest lower East Rift Zone (LERZ) eruption and caldera collapse in at least 200 years. After collapse of the Pu'u 'O'o vent on 30 April, magma propagated downrift. Eruptive fissures opened in the LERZ on 3 May, eventually extending ~6.8 kilometers. A 4 May earthquake [moment magnitude (M w) 6.9] produced ~5 meters of fault slip. Lava erupted at rates exceeding 100 cubic meters per second, eventually covering 35.5 square kilometers. The summit magma system partially drained, producing minor explosions and near-daily collapses releasing energy equivalent to M w 4.7 to 5.4 earthquakes. Activity declined rapidly on 4 August. Summit collapse and lava flow volume estimates are roughly equivalent-about 0.8 cubic kilometers. Careful historical observation and monitoring of Kilauea enabled successful forecasting of hazardous events.

Spatter matters - distinguishing primary (eruptive) and secondary (non-eruptive) spatter deposits.

Spatter is a common pyroclastic product of hawaiian fountaining, which typically forms vent-proximal ramparts or cones. Based on textural characteristics and field relations of spatter from the 1969 Mauna Ulu eruption of Kilauea, Hawai'i, three spatter types were identified: (1) Primary spatter deposited as spatter ramparts and isolated cones during the peak of episode 1; (2) Late-stage spatter comprising dense, small volume, vent proximal deposits, formed at the end of episode 1; (3) Secondary spatter preserved in isolated mounds around tectonic ground cracks that we interpret to have formed by the disruption of overlying lava. We propose that not all spatter deposits are evidence of primary magmatic fountaining. Rather, deposits can be "secondary" in nature and associated with lava drain-back, disruption, and subsequent ejection from tectonic cracks. Importantly, these secondary pyroclastic deposits are difficult to distinguish from primary eruptive features based on field relations and bulk clast vesicularity alone, allowing for the potential misinterpretation of eruption vents, on Earth and in remotely sensed planetary data, thereby misinforming hazard maps and probabilistic assessments. Here, we show that vesicle number density provides a statistically-robust metric by which to discriminate primary and secondary spatter, supporting accurate identification of eruptive vents.

The effect of pulmonary function testing on bleomycin dosing in germ cell tumours.

BACKGROUND/AIM: The utility of pulmonary function testing (PFT) to detect bleomycin-induced pneumonitis is controversial. We describe its impact on bleomycin dosing in a phase 2 trial of accelerated BEP (bleomycin, etoposide, cisplatin) for advanced germ cell tumours. METHODS: There were 12 planned weekly bleomycin doses for intermediate-risk and poor-risk disease and nine for good-risk disease. Clinical assessments, chest X-ray, diffusing capacity of lung for carbon monoxide (DLCO) and forced vital capacity (FVC) were performed bi-weekly. Bleomycin was ceased for predefined clinical/radiological evidence of pulmonary toxicity and a >25% reduction in DLCO or FVC. We determined doses planned, received and omitted and patients receiving all, ≥two-thirds, two-thirds of planned bleomycin doses. RESULTS: Of 43 eligible patients, 30% had lung metastases. Of 471, 375 (80%) of planned bleomycin doses were received, and 30% received 25% reduction in DLCO (35 vs 24%, P = 0.4) and 1.5 times as likely to receive

Accelerated BEP for metastatic germ cell tumours: a multicenter phase II trial by the Australian and New Zealand Urogenital and Prostate Cancer Trials Group (ANZUP).

BACKGROUND: This Australian single-arm, multicenter, phase II trial evaluated feasibility, tolerability and activity of accelerated bleomycin, etoposide and cisplatin (BEP) as first-line chemotherapy for metastatic germ cell tumours. PATIENTS AND METHODS: Patients were planned to receive cisplatin 20 mg/m(2) and etoposide 100 mg/m(2) days 1-5, and pegfilgrastim 6 mg day 6, all repeated every 2 weeks for four cycles (three cycles for good prognosis). Bleomycin was given at 30 000 IU weekly to a total of 12 doses (9 doses for good prognosis). Primary end point was feasibility, defined as the proportion of patients able to complete the etoposide and cisplatin components of BEP and be eligible to receive a fourth cycle of BEP by day 50. RESULTS: Twelve poor, 16 intermediate and 15 good prognosis (n = 43) eligible patients were enrolled. Two patients aged >40 years were ineligible and excluded from analyses. The regimen was feasible in 86%, not feasible in 7% and not assessable in 7% of patients. Most common grade 3/4 adverse events were non-neutropenic infection (16%) and febrile neutropenia (12%). Complete response (CR) to chemotherapy and surgery was achieved in 33% poor-prognosis, 81% intermediate-prognosis and 100% good-prognosis patients. At median follow-up of 27 months (range 6-42), the 2-year progression-free survival was 50% for poor-prognosis, 94% for intermediate-prognosis and 92% for good-prognosis patients. CONCLUSION: Accelerated BEP is feasible and tolerable. Efficacy data appear to be promising. This trial and a similar UK study provide the rationale for a randomised trial comparing accelerated versus standard BEP. Australian New Zealand Clinical Trials Registry Registration number. ACTRN 12607000294459.

Epididymal specific, selenium-independent GPX5 protects cells from oxidative stress-induced lipid peroxidation and DNA mutation.

STUDY QUESTION: Can selenium (Se) independent, epididymal-specific glutathione peroxidase 5 (GPX5) protect CHO-K1 cells from oxidative damage and, more specifically, from lipid peroxidation and DNA mutation? SUMMARY ANSWER: CHO-K1 cells expressing GPX5 have increased resistance to oxidative challenge and, more specifically, decreased levels of lipid peroxidation and decreased levels of the downstream DNA lesion 8-oxo-7,8-dihydroguanine (8-oxodG) compared with control cells. WHAT IS KNOWN ALREADY: GPX5 associates with sperm during transit of the epididymis, and has been postulated to protect sperm from peroxide-mediated attack. However, its function as an active glutathione peroxidase has been questioned due to substitution of the classical selenocysteine residue at its active site. Indirect evidence for a functional role for GPX5 has been provided by in vivo studies, in particular from the GPX5 knockout mouse whereby offspring sired by GPX5(-/-) males have a higher rate of spontaneous abortion and developmental defects, attributed to increased oxidative injury (8-oxodG) to sperm DNA, but only when the GPX5(-/-) males are over 1 year of age. Interestingly, we have previously shown severely reduced levels of GPX5 in humans. STUDY DESIGN, SIZE, DURATION: To look more directly at its role in protection against oxidative damage, we have used an in vitro system, generating a CHO-K1 mammalian cell line expressing recombinant rat GPX5. PARTICIPANTS/MATERIALS, SETTING, METHODS: We have used the recombinant CHO-K1 cells to determine whether GPX5 is able to protect these cells from an administered oxidative challenge, using a range of approaches. We compared the viability of GPX5-expressing cells with control cells by both MTT and trypan blue exclusion assays. We next investigated whether GPX5 protects the cells specifically from lipid peroxidation, by using the fluorescent reporter molecule C11-BODIPY(581/591), and thus from downstream DNA mutation, by comparing levels of the DNA lesion 8-oxodG. We also investigated whether GPX5 can be transferred to rat sperm via epididymosomes. MAIN RESULTS AND THE ROLE OF CHANCE: GPX5-expressing CHO-K1 cells had increased viability compared with control cells following oxidative challenge (P < 0.005). We also found that GPX5-expressing CHO-K1 cells had significantly lower levels of C11-BODIPY(581/591) oxidation, and hence lipid peroxidation, compared with control cells. Levels of 8-oxodG DNA damage were also markedly lower in the nuclei of GPX5-expressing cells than in control cells. Finally, we showed that GPX5 can be transferred to rat sperm via epididymosomes. LIMITATIONS, REASONS FOR CAUTION: GPX5 is not active in glutathione peroxidase assays using H2O2 as the substrate. However, the related non-mammalian Se-independent GPXs show preference for electron donors other than glutathione, with a number utilizing thioredoxin as a reducing equivalent. Hence, the in vitro activity of GPX5 needs to be assessed using a range of alternative substrates and electron donors. GPX5 is secreted by the epididymis and associates with the sperm plasma membrane. We showed that this transfer can occur via epididymosomes; however, the mechanism for transfer and the identity of a potential binding partner in the sperm membrane needs to be determined. Finally, our study utilized an in vitro system that needs to be translated to human sperm. WIDER IMPLICATIONS OF THE FINDINGS: Our study supports an important role for GPX5 as an antioxidant, possibly acting as a phospholipid hydroperoxidase and participating in the maintenance of cell and DNA integrity.

The 'sweet killer'. Can you recognize the symptoms of ethylene glycol poisoning?

Ingested ethylene glycol is readily absorbed and metabolized into toxic metabolites that can cause CNS depression, cardiopulmonary failure, and renal failure. Thorough history taking, physical examination, and laboratory testing are essential for diagnosis. Careful differential diagnosis is important because symptoms of ethylene glycol poisoning are similar to those of other intoxicants. Early, aggressive treatment with appropriate therapies, such as ethanol therapy, hemodialysis, vitamin cofactors, and antidotal agents, is necessary to prevent permanent disability or death.

Discharge planners and cost containment.

Discharge planners play an important role in managing healthcare costs. Until recent years, the actual dollar costs of healthcare were not a concern of either healthcare providers or patients. This author studied the level of knowledge discharge planners have of actual dollar costs of healthcare, their perception of the patient's level of knowledge, and the impact of the knowledge on healthcare decisions.

The source and significance of raised serum enzymes in rheumatoid arthritis.

Hepatobiliary dysfunction in rheumatoid arthritis has been suggested on the basis of raised serum activity of alkaline phosphatase, 5-nucleotidase, lactic dehydrogenase and gamma-glutamyl transferase, but a specific pathological lesion has not been demonstrated and serum transaminases and bilirubin are almost invariably normal. This paper reports a series of studies designed to determine the tissues of origin of the enzymes and offers an alternative interpretation of the enzymological findings. The results suggest that only alkaline phosphatase originates from the liver, while lactic dehydrogenase and 5-nucleotidase originate from synovial fluid polymorphs and synovial lining cells, respectively. Serum alkaline phosphatase may be induced by inflammatory mediators such as interleukin-1 because it correlates with the acute phase response. Serum lactic dehydrogenase is an integrated measure of polymorph lysis in all joints and offers a marker of joint inflammation more specific than measures such as the ESR. Levels of serum 5-nucleotidase provide information about the activity of the synovium. Finally, because hepatic necrosis does not normally occur, the transaminases may be used to monitor drug toxicity.

Corticosteroid replacement therapy: twice or thrice daily?

Although glucocorticoid replacement is conventionally administered twice daily, the pharmacokinetics of hydrocortisone would predict very low levels of plasma cortisol by mid-afternoon. This study compared plasma cortisol day profiles in 7 hypoadrenal patients while on twice daily and thrice daily hydrocortisone replacement. The twice daily regimen was associated with very low levels of cortisol at 16.00 and 18.00 h. This was eliminated by administering the same total dose in a thrice daily regimen. Furthermore, estimates of 'well-being' by visual analogue scale correlated significantly with simultaneous plasma cortisol levels and 5 of the patients expressed a preference for the thrice daily regimen. The findings suggest that thrice daily glucocorticoid replacement therapy should be adopted routinely.

Plasma arginine in cancer of the gastrointestinal tract: effect of surgical treatment.

The concentration of 21 amino acids was measured in the venous plasma of 41 patients with cancer of the gastrointestinal tract who had lost weight, 12 patients who had lost a similar amount of weight from non-malignant, non-septic conditions (benign weight loss), 12 patients with cancer of the gastrointestinal tract who had not lost weight and 21 control patients. Sixteen patients with localised tumours were restudied eight and 12 weeks after successful removal of the primary growth. Six patients with pyloric stenosis (benign weight loss) were similarly studied for comparison after corrective surgery. The concentration of the amino acid arginine was significantly greater in patients with cancer both with weight loss (71.2 +/- 4.1 mumol/l mean +/- SEM) and without weight loss (66.8 +/- 4.1 mumol/l) when compared both with patients with benign weight loss (34.6 +/- 3.2 mumol/l) and with control patients (48.2 +/- 3.5 mumol/l) (p less than 0.05). In patients with cancer subjected to surgery the concentration of arginine (76.4 +/- 7.5 mumol/l) fell to normal levels eight weeks after operation and remained normal 12 weeks after surgery (48.6 +/- 4.4 mumol/l) (p less than 0.05). This was in contrast with the rise in plasma arginine in patients with pyloric stenosis after surgery, suggesting that the raised level of arginine was due to the presence of the primary tumour.

Alteration of pituitary-thyroid function in patients with chronic renal failure treated by haemodialysis or continuous ambulatory peritoneal dialysis.

Pituitary-thyroid function was investigated in 60 patients with chronic renal failure and 18 normal subjects. Serum triiodothyronine and free thyroxine levels were lower in patients treated by either haemodialysis or continuous ambulatory peritoneal dialysis compared with the normal subjects. Serum thyroxine and free thyroxine index were significantly lower in the haemodialysis-treated patients than in the normal subjects. There were no differences in serum thyroxine and free thyroxine index between the continuous ambulatory peritoneal dialysis-treated and normal groups. Serum thyrotrophin was not raised in any of the groups but the serum thyrotrophin response to thyrotrophin-releasing hormone was blunted in both groups of patients. Basal serum prolactin and growth hormone were raised in both groups of patients but there was no significant difference between them. This study confirmed that pituitary-thyroid function is abnormal in patients receiving haemodialysis and established that a similar pattern of abnormalities occurs in patients undergoing continuous ambulatory peritoneal dialysis.

Muscle glycogen and diet in elite soccer players.

The top players in an elite soccer team volunteered as subjects in a study to determine intramuscular glycogen concentrations after a regular season match, and whether optimal glycogen levels could be re-attained prior to the team's next match. Fifteen players were biopsied in the M. vastus lateralis following a regular season game (Day I). Of these, eight were biopsied 24 h later in the evening of a free day during which no training session was held (Day II), and again 24 h later after a very light training session (Day III). Muscle glycogen concentrations for the eight players averaged (+/- SD) 45.9 +/- 7.9, 68.9 +/- 2.7, and 72.8 +/- 8.3 mmol glucose units x kg-1 wet muscle weight on Day I, Day II, and Day III respectively. Dietary records were analysed during a week of peak, season competition and training. The average daily total energy consumption and total quantity of carbohydrates consumed were less than what is normally consumed by athletes in similar sports. The inability of the players to maintain even normal, resting levels of muscle glycogen is probably related to their dietary practices.