Further delineation of short-chain enoyl-CoA hydratase deficiency in the Pacific population.

Short-chain enoyl-coA hydratase (SCEH) deficiency due to biallelic pathogenic ECHS1 variants was first reported in 2014 in association with Leigh syndrome (LS) and increased S-(2-carboxypropyl)cysteine excretion. It is potentially treatable with a valine-restricted, high-energy diet and emergency regimen. Recently, Simon et al. described four Samoan children harbouring a hypomorphic allele (c.489G > A, p.Pro163=) associated with reduced levels of normally-spliced mRNA. This synonymous variant, missed on standard genomic testing, is prevalent in the Samoan population (allele frequency 0.17). Patients with LS and one ECHS1 variant were identified in NZ and Australian genomic and clinical databases. ECHS1 sequence data were interrogated for the c.489G > A variant and clinical data were reviewed. Thirteen patients from 10 families were identified; all had Pacific ancestry including Samoan, Maori, Cook Island Maori, and Tokelauan. All developed bilateral globus pallidi lesions, excluding one pre-symptomatic infant. Symptom onset was in early childhood, and was triggered by illness or starvation in 9/13. Four of 13 had exercise-induced dyskinesia, 9/13 optic atrophy and 6/13 nystagmus. Urine S-(2-carboxypropyl)cysteine-carnitine and other SCEH-related metabolites were normal or mildly increased. Functional studies demonstrated skipping of exon four and markedly reduced ECHS1 protein. These data provide further support for the pathogenicity of this ECHS1 variant which is also prevalent in Maori, Cook Island Maori, and Tongan populations (allele frequency 0.14-0.24). It highlights the need to search for a second variant in apparent heterozygotes with an appropriate phenotype, and has implications for genetic counselling in family members who are heterozygous for the more severe ECHS1 alleles. SYNOPSIS: Short-chain enoyl-CoA hydratase deficiency is a frequent cause of Leigh-like disease in Maori and wider-Pacific populations, due to the high carrier frequency of a hypomorphic ECHS1 variant c.489G > A, p.[Pro163=, Phe139Valfs*65] that may be overlooked by standard genomic testing.

Uncovering social structures and informational prejudices to reduce inequity in delivery and uptake of new molecular technologies.

Advances in molecular technologies have the potential to help remedy health inequities through earlier detection and prevention; if, however, their delivery and uptake (and therefore any benefits associated with such testing) are not more carefully considered, there is a very real risk that existing inequities in access and use will be further exacerbated. We argue this risk relates to the way that information and knowledge about the technology is both acquired and shared, or not, between health practitioners and their patients.A healthcare system can be viewed as a complex social network comprising individuals with different worldviews, hierarchies, professional cultures and subcultures and personal beliefs, both for those giving and receiving care. When healthcare practitioners are not perceived as knowledge equals, they would experience informational prejudices, and the result is that knowledge dissemination across and between them would be impeded. The uptake and delivery of a new technology may be inequitable as a result. Patients would also experience informational prejudice when they are viewed as not being able to understand the information that is presented to them, and information may be withheld.Informational prejudices driven by social relations and structures have thus far been underexplored in considering (in)equitable implementation and uptake of new molecular technologies. Every healthcare interaction represents an opportunity for experiencing informational prejudice, and with it the risk of being inappropriately informed for undertaking (or offering) such screening or testing. Making knowledge acquisition and information dissemination, and experiences of informational prejudice, explicit through sociologically framed investigations would extend our understandings of (in)equity, and offer ways to affect network relationships and structures that support equity in delivery and uptake.

Synthesis and biological evaluation of solubilized sulfonamide analogues of the phosphatidylinositol 3-kinase inhibitor ZSTK474.

Replacing one of the morpholine groups of the phosphatidylinositol 3-kinase (PI3K) inhibitor ZSTK474 with a variety of sulfonamide-linked solubilizing substituents produced a new class of active and potent PI3Kα inhibitors, with several derivatives demonstrating high PI3Kα enzyme potency and good cellular potency in two human derived cell lines. The overall results suggest a preference for linear and somewhat flexible solubilizing functions. From this series, compound 16, also known as SN32976, was selected for advanced preclinical evaluation.

Identification, structure modification, and characterization of potential small-molecule SGK3 inhibitors with novel scaffolds.

The serum and glucocorticoid-regulated kinase (SGK) family has been implicated in the regulation of many cellular processes downstream of the PI3K pathway. It plays a crucial role in PI3K-mediated tumorigenesis, making it a potential therapeutic target for cancer. SGK family consists of three isoforms (SGK1, SGK2, and SGK3), which have high sequence homology in the kinase domain and similar substrate specificity with the AKT family. In order to identify novel compounds capable of inhibiting SGK3 activity, a high-throughput screening campaign against 50,400 small molecules was conducted using a fluorescence-based kinase assay that has a Z' factor above 0.5. It identified 15 hits (including nitrogen-containing aromatic, flavone, hydrazone, and naphthalene derivatives) with IC50 values in the low micromolar to sub-micromolar range. Four compounds with a similar scaffold (i.e., a hydrazone core) were selected for structural modification and 18 derivatives were synthesized. Molecular modeling was then used to investigate the structure-activity relationship (SAR) and potential protein-ligand interactions. As a result, a series of SGK inhibitors that are active against both SGK1 and SGK3 were developed and important functional groups that control their inhibitory activity identified.

High-throughput screening campaigns against a PI3Kα isoform bearing the H1047R mutation identified potential inhibitors with novel scaffolds.

The phosphatidylinositol 3-kinase (PI3K) pathway is involved in many cellular functions including cell growth, metabolism, and transformation. Hyperactivation of this pathway contributes to tumorigenesis, therefore, PI3K is a major target for anticancer drug discovery. Since the PI3Kα isoform is implicated mostly in cancer, we conducted a high-throughput screening (HTS) campaign using a 3-step PI3K homogenous time-resolved fluorescence assay against this isoform bearing the H1047R mutation. A total of 288,000 synthetic and natural product-derived compounds were screened and of which, we identified 124 initial hits that were further selected by considering the predicted binding mode, relationship to known pan-assay interference compounds and previous descriptions as a lipid kinase inhibitor. A total of 24 compounds were then tested for concentration-dependent responses. These hit compounds provide novel scaffolds that can potentially be optimized to create novel PI3K inhibitors.

Combining properties of different classes of PI3Kα inhibitors to understand the molecular features that confer selectivity.

Phosphoinositide 3-kinases (PI3Ks) are major regulators of many cellular functions, and hyperactivation of PI3K cell signalling pathways is a major target for anticancer drug discovery. PI3Kα is the isoform most implicated in cancer, and our aim is to selectively inhibit this isoform, which may be more beneficial than concurrent inhibition of all Class I PI3Ks. We have used structure-guided design to merge high-selectivity and high-affinity characteristics found in existing compounds. Molecular docking, including the prediction of water-mediated interactions, was used to model interactions between the ligands and the PI3Kα affinity pocket. Inhibition was tested using lipid kinase assays, and active compounds were tested for effects on PI3K cell signalling. The first-generation compounds synthesized had IC50 (half maximal inhibitory concentration) values >4 µM for PI3Kα yet were selective for PI3Kα over the other Class I isoforms (ß, δ and γ). The second-generation compounds explored were predicted to better engage the affinity pocket through direct and water-mediated interactions with the enzyme, and the IC50 values decreased by ∼30-fold. Cell signalling analysis showed that some of the new PI3Kα inhibitors were more active in the H1047R mutant bearing cell lines SK-OV-3 and T47D, compared with the E545K mutant harbouring MCF-7 cell line. In conclusion, we have used a structure-based design approach to combine features from two different compound classes to create new PI3Kα-selective inhibitors. This provides new insights into the contribution of different chemical units and interactions with different parts of the active site to the selectivity and potency of PI3Kα inhibitors.

Novel pyrazolo[1,5-a]pyridines with improved aqueous solubility as p110α-selective PI3 kinase inhibitors.

As part of our investigation into pyrazolo[1,5-a]pyridines as novel p110α selective PI3 kinase inhibitors, we report a range of analogues with improved aqueous solubility by the addition of a basic amine. The compounds demonstrated comparable p110α potency and selectivity to earlier compounds but with up to 1000× greater aqueous solubility, as the hydrochloride salts. The compounds also displayed good activity in a cellular assay of PI3 kinase activity.

Synthesis and biological evaluation of sulfonamide analogues of the phosphatidylinositol 3-kinase inhibitor ZSTK474.

Replacement of one of the morpholine groups of the phosphatidylinositol 3-kinase (PI3K) inhibitor ZSTK474 (1) with sulfonamide containing substituents produced a new class of active and potent PI3Kα inhibitors. Solubility issues prevented all but the 6-amino derivative 17 from being evaluated in vivo, but the clear activity of this compound demonstrated that this class of PI3K inhibitor shows great promise.

Effects of a Custom Bite-Aligning Mouthguard on Performance in College Football Players.

Besides injury prevention, mouthguards can also be employed to improve physical performance. The effects of personalization of mouthguards have rarely been investigated. This 3-armed, randomized, controlled crossover trial investigated the difference of wearing (a) personalized or custom-made (CM, e.g., bite-aligned), (b) standard (BB, boil and bite), and (c) no (CON) mouthguards on general fitness parameters in experienced collegiate football players. A group of 10 upperclassmen (age, 19-22 years; mean ± SD: age = 20.7 ± 0.8 years; body mass = 83 ± 7.4 kg; height = 179.1 ± 5.2 cm; body mass index = 25.9 ± 2.2 kg·cm), National Collegiate Athletic Association Division II football players with at least 2 years of playing experience, were randomly assigned to the 3 mouthguard conditions: a randomized, within-subjects repeated-measures design was applied. All participants were randomly tested on strength and endurance performance V[Combining Dot Above]O2max testing, with Bruce treadmill protocol including (a) time to fatigue, (b) blood lactate concentration in millimoles per liter at stage 2 and (c) at peak fatigue, (d) flexibility, (e) reaction time, (f) squat vertical jump, (g) countermovement vertical jump, and (h) 1 repetition maximum bench press. Repeated-measures analysis of variance showed no significant differences between the 3 conditions for each outcome variable (0.23 < p < 0.94; 0.007 < (Equation is included in full-text article.)< 0.15). These data indicate that CM mouthguards did not superiorly affect general fitness parameters compared with BB and CON. In turn, protective BB or CM mouthpieces did not appear to impair general fitness performance vs. CON. The recommendation of a custom bite-aligning mouthguards for performance enhancement in young Division II football players is questioned. Further studies with larger sample sizes, gender comparison, and (sport) discipline-specific performance testing are needed.

Preptin Deficiency Does Not Protect against High-Fat Diet-Induced Metabolic Dysfunction or Bone Loss in Mice.

Preptin is derived from the cleavage of the E-peptide of pro-insulin-like growth factor (IGF)-II and is an insulin secretagogue. Observational studies have linked elevated circulating preptin to metabolic dysfunction in humans; however, a causal role for preptin in metabolic dysfunction has not been established. Additionally, preptin can promote osteoblast proliferation and differentiation, suggesting a link with skeletal health. We previously described a global preptin knockout (KO) model. In this study, we sought to uncover the impact of preptin KO in mice on the response to a moderately high-fat diet (HFD) and low-fat diet (LFD). HFD groups had higher weight and fat mass gain, lower trabecular and cortical bone volume and fracture load, and higher liver triglycerides. In males, preptin deficiency led to lower blood glucose than wild-type (WT) mice under LFD conditions. This was accompanied by differences in bone microarchitecture, including lower trabecular bone volume fraction, trabecular number, and lower cortical thickness. These differences were absent in female mice, although KO females had a HFD-driven increase in fat mass and liver triglycerides that was absent in WT mice. Female WT mice had increased glucose-stimulated insulin secretion under HFD conditions that was absent in female KO mice. Overall, preptin may have a detrimental impact on metabolism and a positive impact on bone health in male mice and may protect against liver fat storage in females while enabling islet compensation under HFD conditions. When we consider that serum preptin levels are elevated in humans of both sexes in pathological states in which insulin levels are elevated, the impact of preptin on comorbidity risk needs to be better understood. © 2023 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

DMXAA (Vadimezan, ASA404) is a multi-kinase inhibitor targeting VEGFR2 in particular.

The flavone acetic acid derivative DMXAA [5,6-dimethylXAA (xanthenone-4-acetic acid), Vadimezan, ASA404] is a drug that displayed vascular-disrupting activity and induced haemorrhagic necrosis and tumour regression in pre-clinical animal models. Both immune-mediated and non-immune-mediated effects contributed to the tumour regression. The vascular disruption was less in human tumours, with immune-mediated effects being less prominent, but nonetheless DMXAA showed promising effects in Phase II clinical trials in non-small-cell lung cancer. However, these effects were not replicated in Phase III clinical trials. It has been difficult to understand the differences between the pre-clinical findings and the later clinical trials as the molecular targets for the agent have never been clearly established. To investigate the mechanism of action, we sought to determine whether DMXAA might target protein kinases. We found that, at concentrations achieved in blood during clinical trials, DMXAA has inhibitory effects against several kinases, with most potent effects being on members of the VEGFR (vascular endothelial growth factor receptor) tyrosine kinase family. Some analogues of DMXAA were even more effective inhibitors of these kinases, in particular 2-MeXAA (2-methylXAA) and 6-MeXAA (6-methylXAA). The inhibitory effects were greatest against VEGFR2 and, consistent with this, we found that DMXAA, 2-MeXAA and 6-MeXAA were able to block angiogenesis in zebrafish embryos and also inhibit VEGFR2 signalling in HUVECs (human umbilical vein endothelial cells). Taken together, these results indicate that at least part of the effects of DMXAA are due to it acting as a multi-kinase inhibitor and that the anti-VEGFR activity in particular may contribute to the non-immune-mediated effects of DMXAA on the vasculature.

A brief relaxation intervention reduces stress and improves surgical wound healing response: a randomised trial.

Psychological stress has been shown to impair wound healing, but experimental research in surgical patients is lacking. This study investigated whether a brief psychological intervention could reduce stress and improve wound healing in surgical patients. This randomised controlled trial was conducted at a surgical centre. Inclusion criteria were English-speaking patients over 18 years booked to undergo elective laparoscopic cholecystectomy; exclusion criteria were cancellation of surgery, medical complications, and refusal of consent. Seventy five patients were randomised and 15 patients were excluded; 60 patients completed the study (15 male, 45 female). Participants were randomised to receive standard care or standard care plus a 45-min psychological intervention that included relaxation and guided imagery with take-home relaxation CDs for listening to for 3 days before and 7 days after surgery. In both groups ePTFE tubes were inserted during surgery and removed at 7 days after surgery and analysed for hydroxyproline as a measure of collagen deposition and wound healing. Change in perceived stress from before surgery to 7-day follow-up was assessed using questionnaires. Intervention group patients showed a reduction in perceived stress compared with the control group, controlling for age. Patients in the intervention group had higher hydroxyproline deposition in the wound than did control group patients (difference in means 0.35, 95% CI 0.66-0.03; t(43)=2.23, p=0.03). Changes in perceived stress were not associated with hydroxyproline deposition. A brief relaxation intervention prior to surgery can reduce stress and improve the wound healing response in surgical patients. The intervention may have particular clinical application for those at risk of poor healing following surgery.

Discovery of pyrazolo[1,5-a]pyridines as p110α-selective PI3 kinase inhibitors.

We have made a novel series of pyrazolo[1,5-a]pyridines as PI3 kinase inhibitors, and demonstrated their selectivity for the p110α isoform over the other Class Ia PI3 kinases. We investigated the SAR around the pyrazolo[1,5-a]pyridine ring system, and found compound 5x to be a particularly potent example (p110α IC(50) 0.9nM). This compound inhibits cell proliferation and phosphorylation of Akt/PKB, a downstream marker of PI3 kinase activity, and showed in vivo activity in an HCT-116 human xenograft model.

Novel pyrazolo[1,5-a]pyridines as p110α-selective PI3 kinase inhibitors: Exploring the benzenesulfonohydrazide SAR.

Structure-activity relationship studies of the pyrazolo[1,5-a]pyridine class of PI3 kinase inhibitors show that substitution off the hydrazone nitrogen and replacement of the sulfonyl both gave a loss of p110α selectivity, with the exception of an N-hydroxyethyl analogue. Limited substitutions were tolerated around the phenyl ring; in particular the 2,5-substitution pattern was important for PI3 kinase activity. The N-hydroxyethyl compound also showed good inhibition of cell proliferation and inhibition of phosphorylation of Akt/PKB, a downstream marker of PI3 kinase activity. It had suitable pharmacokinetics for evaluation in vivo, and showed tumour growth inhibition in two human tumour cell lines in xenograft studies. This work has provided suggestions for the design of more soluble analogues.

A drug targeting only p110α can block phosphoinositide 3-kinase signalling and tumour growth in certain cell types.

Genetic alterations in PI3K (phosphoinositide 3-kinase) signalling are common in cancer and include deletions in PTEN (phosphatase and tensin homologue deleted on chromosome 10), amplifications of PIK3CA and mutations in two distinct regions of the PIK3CA gene. This suggests drugs targeting PI3K, and p110α in particular, might be useful in treating cancers. Broad-spectrum inhibition of PI3K is effective in preventing growth factor signalling and tumour growth, but suitable inhibitors of p110α have not been available to study the effects of inhibiting this isoform alone. In the present study we characterize a novel small molecule, A66, showing the S-enantiomer to be a highly specific and selective p110α inhibitor. Using molecular modelling and biochemical studies, we explain the basis of this selectivity. Using a panel of isoform-selective inhibitors, we show that insulin signalling to Akt/PKB (protein kinase B) is attenuated by the additive effects of inhibiting p110α/p110ß/p110δ in all cell lines tested. However, inhibition of p110α alone was sufficient to block insulin signalling to Akt/PKB in certain cell lines. The responsive cell lines all harboured H1047R mutations in PIK3CA and have high levels of p110α and class-Ia PI3K activity. This may explain the increased sensitivity of these cells to p110α inhibitors. We assessed the activation of Akt/PKB and tumour growth in xenograft models and found that tumours derived from two of the responsive cell lines were also responsive to A66 in vivo. These results show that inhibition of p110α alone has the potential to block growth factor signalling and reduce growth in a subset of tumours.

Neonatal Hypoxic Ischemic Encephalopathy and Planned Home Birth.

INTRODUCTION: As planned home births increase, emerging evidence on the perinatal outcomes of newborns who were planned hospital births versus planned home births has been inconsistent, and a growing number of states have attempted to legislate community births. We sought to determine whether an association exists between neonatal hypoxic ischemic encephalopathy (HIE), a complication of ischemic birth injury, and planned location of birth. METHODS: A case-control study design was used to compare data from neonates with HIE obtained from electronic health records at Kapiolani Medical Center for Women and Children in Honolulu, Hawaii, with data from neonates without HIE obtained from Hawaii state birth certificate data. A penalized backward stepwise logistic regression was performed to control for confounders. RESULTS: We included 164 neonates with HIE and 656 neonates in the control group. The odds of having been a planned home birth were 2.77 times higher in neonates with HIE compared with those without HIE (95% CI, 1.05-6.87). After adjusting for insurance, mode of birth, meconium fluid, maternal hypertension, and chorioamnionitis, neonates with HIE were still more likely to have been a planned home birth compared with those without HIE (odds ratio, 11.56; 95% CI, 1.37-118.77). DISCUSSION: Neonates with HIE were more likely to have been a planned home birth compared with neonates without HIE.

Response to BRAF-targeted Therapy Is Enhanced by Cotargeting VEGFRs or WNT/ß-Catenin Signaling in BRAF-mutant Colorectal Cancer Models.

The fact that 10% of colorectal cancer tumors harbor BRAF V600E mutations suggested targeting BRAF as a potential therapy. However, BRAF inhibitors have only limited single-agent efficacy in this context. The potential for combination therapy has been shown by the BEACON trial where targeting the EGF receptor with cetuximab greatly increased efficacy of BRAF inhibitors in BRAF-mutant colorectal cancer. Therefore, we explored whether efficacy of the mutant BRAF inhibitor vemurafenib could be enhanced by cotargeting of either oncogenic WNT/ß-catenin signaling or VEGFR signaling. We find the WNT/ß-catenin inhibitors pyrvinium, ICG-001 and PKF118-310 attenuate growth of colorectal cancer cell lines in vitro with BRAF-mutant lines being relatively more sensitive. Pyrvinium combined with vemurafenib additively or synergistically attenuated growth of colorectal cancer cell lines in vitro. The selective and potent VEGFR inhibitor axitinib was most effective against BRAF-mutant colorectal cancer cell lines in vitro, but the addition of vemurafenib did not significantly increase these effects. When tested in vivo in animal tumor models, both pyrvinium and axitinib were able to significantly increase the ability of vemurafenib to attenuate tumor growth in xenografts of BRAF-mutant colorectal cancer cells. The magnitude of these effects was comparable with that induced by a combination of vemurafenib and cetuximab. This was associated with additive effects on release from tumor cells and tumor microenvironment cell types of substances that would normally aid tumor progression. Taken together, these preclinical data indicate that the efficacy of BRAF inhibitor therapy in colorectal cancer could be increased by cotargeting either WNT/ß-catenin or VEGFRs with small-molecule inhibitors.

The chaperone proteins HSP70, HSP40/DnaJ and GRP78/BiP suppress misfolding and formation of ß-sheet-containing aggregates by human amylin: a potential role for defective chaperone biology in Type 2 diabetes.

Misfolding of the islet ß-cell peptide hA (human amylin) into ß-sheet-containing oligomers is linked to ß-cell apoptosis and the pathogenesis of T2DM (Type 2 diabetes mellitus). In the present study, we have investigated the possible effects on hA misfolding of the chaperones HSP (heat-shock protein) 70, GRP78/BiP (glucose-regulated protein of 78 kDa/immunoglobulin heavy-chain-binding protein) and HSP40/DnaJ. We demonstrate that hA underwent spontaneous time-dependent ß-sheet formation and aggregation by thioflavin-T fluorescence in solution, whereas rA (rat amylin) did not. HSP70, GRP78/BiP and HSP40/DnaJ each independently suppressed hA misfolding. Maximal molar protein/hA ratios at which chaperone activity was detected were 1:200 (HSP70, HSP40/DnaJ and GRP78/BiP). By contrast, none of the chaperones modified the secondary structure of rA. hA, but not rA, was co-precipitated independently with HSP70 and GRP78/BiP by anti-amylin antibodies. As these effects occur at molar ratios consistent with chaperone binding to relatively rare misfolded hA species, we conclude that HSP70 and GRP78/BiP can detect and bind misfolded hA oligomers, thereby effectively protecting hA against bulk misfolding and irreversible aggregation. Defective ß-cell chaperone biology could contribute to hA misfolding and initiation of apoptosis in T2DM.

Evolution of Molecular Targets in Melanoma Treatment.

Melanoma is the deadliest type of skin cancers, accounting for more than 80% of skin cancer mortality. Although melanoma was known very early in the history of medicine, treatment for this disease had remained largely the same until very recently. Previous treatment options, including removal surgery and systemic chemotherapy, offered little benefit in extending the survival of melanoma patients. However, the last decade has seen breakthroughs in melanoma treatment, which all emerged following new insight into the oncogenic signaling of melanoma. This paper reviewed the evolution of drug targets for melanoma treatment based on the emergence of novel findings in the molecular signaling of melanoma. One of the findings that are most influential in melanoma treatment is that more than 50% of melanoma tumors contain BRAF mutations. This is fundamental for the development of BRAF inhibitors, which is the first group of drugs that significantly improves the overall survival of melanoma patients compared to the traditional chemotherapeutic dacarbazine. More recently, findings of the role of immune checkpoint molecules such as CTLA-4 and PD1/PD-L1 in melanoma biology have led to the development of a new therapeutic category: immune checkpoint inhibitors, which, for the first time in the history of cancer treatment, produced a durable response in a subset of melanoma patients. However, as this paper discussed next, there is still an unmet need for melanoma treatment. A significant population of patients did not respond to either BRAF inhibitors or immune checkpoint inhibitors. Of those patients who gained an initial response from those therapies, a remarkable percentage would develop drug resistance even when MEK inhibitors were added to the treatment. Finally, this paper discusses some possible targets for melanoma treatment.