Choice of fusion proteins, expression host, and analytics solves difficult-to-produce protein challenges in discovery research.

High quality biological reagents are a prerequisite for pharmacological research. Herein a protein production screening approach, including quality assessment methods, for protein-based discovery research is presented. Trends from 2895 expression constructs representing 253 proteins screened in mammalian and bacterial hosts-91% of which are successfully expressed and purified-are discussed. Mammalian expression combined with the use of solubility-promoting fusion proteins is deemed suitable for most targets. Furthermore, cases utilizing stable cell line generation and choice of fusion protein for higher yield and quality of difficult-to-produce proteins (Leucine-rich repeat-containing G-protein coupled receptor 4 (LGR4) and Neurturin) are presented and discussed. In the case of Neurturin, choice of fusion protein impacted the target binding 80-fold. These results highlight the need for exploration of construct designs and careful Quality Control (QC) of difficult-to-produce protein reagents.

A Verbal Card Sorting Task to Measure Executive Functions.

PURPOSE: The Wisconsin Card Sorting Test (WCST) is commonly used to measure nonverbal executive functions (EFs) in a variety of clinical populations. However, in some clinical populations (e.g., people with aphasia), deficits may be present in more linguistic (or verbal) domains and less pronounced in nonverbal domains. Thus, when determining possible deficits in these individuals, it is critical to assess both verbal and nonverbal cognitive abilities. The purpose of this study was to create a verbal card sorting task (VCST) to complement the WCST. METHOD: We created the VCST by modifying a computerized version of the WCST, the Berg Card Sorting Task (BCST). We then compared 35 individuals with mild traumatic brain injury (mTBI) and 33 matched controls' performance on each task. We tested the VCST in individuals with mTBI first because they demonstrate impaired EFs but unimpaired language. We therefore expected the mTBI group to perform similarly on the VCST and BCST, suggesting that the two tasks measure EFs similarly. RESULTS: In line with our hypothesis, the mTBI group had unimpaired inhibition and sustained attention but impaired shifting on each task. Component loadings for both tasks were also similar, and participants' inhibition and shifting scores positively correlated across the two tasks. CONCLUSIONS: Together, these findings suggest that the VCST is a potentially useful tool for measuring verbal EF deficits. Our results also provide important insights into the EF impairments experienced by individuals with mTBI. SUPPLEMENTAL MATERIAL: https://doi.org/10.23641/asha.23230475.

Adverse events in the placebo arm of maintenance therapy trials in advanced ovarian cancer: A systematic review and meta-analysis.

BACKGROUND: Maintenance treatment is standard of care for front-line (FL) and platinum-sensitive recurrent ovarian cancer (PSROC) following response to chemotherapy. Adverse events (AEs) on maintenance therapies are common and usually attributable to investigational treatments but could also be unrelated. Randomised controlled trial (RCT) with blinded placebo design is the gold standard for determining the relative differences in efficacy and AEs between treatment arms. We performed a meta-analysis to quantify AE rates in placebo arms of RCTs to determine AEs not due to investigational agents. METHODS: We performed an electronic search to identify eligible RCTs in FL and PSROC settings. Data from placebo arms were extracted and pooled using the inverse variance method to determine the risk of any AE, overall and specific grade 3 or higher (G ≥ 3) AEs, and AE-related treatment delay, reduction and discontinuation. RESULTS: We identified 13 eligible RCTs (FL, N = 8; PSROC, N = 5) with 2224 patients who received placebo (FL, N = 1541; PSROC, N = 683). The majority experienced an AE of any grade (FL, 93.0%; PSROC, 95.2%). Substantial proportions experienced G ≥ 3 AEs (FL, 14.6%; PSROC, 18.2%). In the FL setting, AEs led to treatment delay in 14.4%, dose reduction in 4.1% and discontinuation in 2.6%. Findings were similar for PSROC: 8.4%, 5.5% and 2.1%, respectively. CONCLUSIONS: AEs not due to investigational agents are common in ovarian cancer patients in maintenance therapy RCTs. Potential explanations include the nocebo effect, residual toxicities from previous treatment or underlying disease. Further research is required to identify better approaches to assessing AEs in this population.

Dual targeting of the epigenome via FACT complex and histone deacetylase is a potent treatment strategy for DIPG.

Diffuse intrinsic pontine glioma (DIPG) is an aggressive and incurable childhood brain tumor for which new treatments are needed. CBL0137 is an anti-cancer compound developed from quinacrine that targets facilitates chromatin transcription (FACT), a chromatin remodeling complex involved in transcription, replication, and DNA repair. We show that CBL0137 displays profound cytotoxic activity against a panel of patient-derived DIPG cultures by restoring tumor suppressor TP53 and Rb activity. Moreover, in an orthotopic model of DIPG, treatment with CBL0137 significantly extends animal survival. The FACT subunit SPT16 is found to directly interact with H3.3K27M, and treatment with CBL0137 restores both histone H3 acetylation and trimethylation. Combined treatment of CBL0137 with the histone deacetylase inhibitor panobinostat leads to inhibition of the Rb/E2F1 pathway and induction of apoptosis. The combination of CBL0137 and panobinostat significantly prolongs the survival of mice bearing DIPG orthografts, suggesting a potential treatment strategy for DIPG.