The SMC5/6 complex prevents genotoxicity upon APOBEC3A-mediated replication stress.

Mutational patterns caused by APOBEC3 cytidine deaminase activity are evident throughout human cancer genomes. In particular, the APOBEC3A family member is a potent genotoxin that causes substantial DNA damage in experimental systems and human tumors. However, the mechanisms that ensure genome stability in cells with active APOBEC3A are unknown. Through an unbiased genome-wide screen, we define the Structural Maintenance of Chromosomes 5/6 (SMC5/6) complex as essential for cell viability when APOBEC3A is active. We observe an absence of APOBEC3A mutagenesis in human tumors with SMC5/6 dysfunction, consistent with synthetic lethality. Cancer cells depleted of SMC5/6 incur substantial genome damage from APOBEC3A activity during DNA replication. Further, APOBEC3A activity results in replication tract lengthening which is dependent on PrimPol, consistent with re-initiation of DNA synthesis downstream of APOBEC3A-induced lesions. Loss of SMC5/6 abrogates elongated replication tracts and increases DNA breaks upon APOBEC3A activity. Our findings indicate that replication fork lengthening reflects a DNA damage response to APOBEC3A activity that promotes genome stability in an SMC5/6-dependent manner. Therefore, SMC5/6 presents a potential therapeutic vulnerability in tumors with active APOBEC3A.

Enolase Inhibitors as Early Lead Therapeutics against Trypanosoma brucei.

Glucose metabolism is critical for the African trypanosome, Trypanosoma brucei, serving as the lone source of ATP production for the bloodstream form (BSF) parasite in the glucose-rich environment of the host blood. Recently, phosphonate inhibitors of human enolase (ENO), the enzyme responsible for the interconversion of 2-phosphoglycerate (2-PG) to phosphoenolpyruvate (PEP) in glycolysis or PEP to 2-PG in gluconeogenesis, have been developed for the treatment of glioblastoma multiforme (GBM). Here, we have tested these agents against T. brucei ENO (TbENO) and found the compounds to be potent enzyme inhibitors and trypanocides. For example, (1-hydroxy-2-oxopyrrolidin-3-yl) phosphonic acid (deoxy-SF2312) was a potent enzyme inhibitor (IC50 value of 0.60 ± 0.23 µM), while a six-membered ring-bearing phosphonate, (1-hydroxy-2-oxopiperidin-3-yl) phosphonic acid (HEX), was less potent (IC50 value of 2.1 ± 1.1 µM). An analog with a larger seven-membered ring, (1-hydroxy-2-oxoazepan-3-yl) phosphonic acid (HEPTA), was not active. Molecular docking simulations revealed that deoxy-SF2312 and HEX had binding affinities of -6.8 and -7.5 kcal/mol, respectively, while the larger HEPTA did not bind as well, with a binding of affinity of -4.8 kcal/mol. None of these compounds were toxic to BSF parasites; however, modification of enzyme-active phosphonates through the addition of pivaloyloxymethyl (POM) groups improved activity against T. brucei, with POM-modified (1,5-dihydroxy-2-oxopyrrolidin-3-yl) phosphonic acid (POMSF) and POMHEX having EC50 values of 0.45 ± 0.10 and 0.61 ± 0.08 µM, respectively. These findings suggest that HEX is a promising lead against T. brucei and that further development of prodrug HEX analogs is warranted.

Rising Mortality in Pediatric Self-Inflicted Firearm Trauma Associated With Distinct Anatomic Injury.

INTRODUCTION: Self-inflicted injuries are the second leading cause of pediatric (10-18 y old) mortality. Self-inflicted firearm trauma (SIFT) was responsible for up to half of these deaths in certain age groups. We hypothesized that SIFT prevalence has increased and is associated with specific demographics, injury patterns, and outcomes. MATERIALS AND METHODS: Data were abstracted from the 2007-2018 American College of Surgeons (ACS) Trauma Quality Programs Participant Use Files (TQP-PUF). Pediatric (1-17 yold) victims of firearm violence were eligible. Age, race, gender, anatomic region, and intent were abstracted. Variables were analyzed using chi-squared tests, t-tests, and single-variate linear regression models. Temporal trends were analyzed using ANCOVA tests. Multivariate logistic regressions were conducted to identify factors influencing mortality. Significance was P < 0.05. RESULTS: There were 41,239 pediatric firearm trauma patients (SIFT: 5.5% [n = 2272]). SIFT incidence increased over the 12-y period (2007 (n = 67) versus 2018 (n = 232), P < 0.05). SIFT was significantly associated with Caucasian race, 67% (n = 1537), teenagers, 90% (n = 2056), male gender, 87% (n = 1978), and a higher median injury severity score (ISS) than other intents of injury (SIFT: 20.0 (IQR: 9.0, 25.0) versus other: 9.0 (IQR: 1.0-13.0), P < 0.001). The SIFT mortality rate was 44% (n = 1005). On multivariate regression head gunshot wounds (OR: 21.1, 95% C.I.: 9.9-45.2, P = 0.001), and ISS (OR:1.1, 95% C.I.: 1.1-1.1, P = 0.001) were significantly associated with mortality. Compared to other intents, SIFT mortality rates increased at a higher annual rate (P < 0.001). CONCLUSIONS: Comprehensive local and federal policy changes to reduce firearms access and increase pediatric mental health support may mitigate these injuries.

Current Advances Toward the Encapsulation of Cas9.

Genetic diseases present formidable hurdles in maintaining a good quality of life for those suffering from these ailments. Often, patients look to inadequate treatments to manage symptoms, which can result in harmful effects on the body. Through genetic engineering, scientists utilize the clustered regularly short palindromic repeat (CRISPR)-associated protein, known as Cas9, to treat the root of the problem. The Cas9 protein is often codelivered with guide RNAs or in ribonucleoprotein complexes (RNP) to ensure targeted delivery of the genetic tool as well as to limit off-target effects. This paper provides an overview of the current advances made toward the encapsulation and delivery of Cas9 to desired locations in the body through encapsulating nanoparticles. Several factors must be considered when employing the Cas9 system to allow gene editing to occur. Material selection is crucial to protect the payload of the delivery vector. Current literature indicates that lipid- and polymer-based nanoparticles show the most potential as delivery vessels for Cas9. Lipid nanoparticles greatly outpace polymer-based nanoparticles in the clinic, despite the benefits that polymers may introduce. When developing translatable systems, there are factors that have not yet been considered that are relevant to Cas9 delivery that are highlighted in this Viewpoint. The proper functioning of Cas9 is dependent on maintaining a proper internal environment; however, there are gaps in the literature regarding these optimal conditions. Interactions between charges of the Cas9 protein, codelivered molecules, and delivery vehicles could impact the effectiveness of the gene editing taking place. While the internal charges of nanoparticles and their effects on Cas9 are presently undetermined, nanoparticles currently offer the ideal delivery method for the Cas9 protein due to their adequate size, modifiable external charge, and ability to be modified. Overall, a cationic lipid-/polymer-based nanoparticle system was found to have the most prospects in Cas9 delivery thus far. By understanding the successes of other systems, translatable, polymer-based delivery vehicles may be developed.

Influence of exercise order on maximum strength and muscle volume in nonlinear periodized resistance training.

The purpose of this study was to examine the influence of exercise order on strength and muscle volume (MV) after 12 weeks of nonlinear periodized resistance training. The participants were randomly assigned into 3 groups. One group began performing large muscle group exercises and progressed to small muscle group exercises (LG-SM), whereas another group started with small muscle group exercises and advanced to large muscle group exercises (SM-LG). The exercise order for LG-SM was bench press (BP), machine lat pull-down (LPD), triceps extension (TE), and biceps curl (BC). The order for the SM-LG was BC, TE, LPD, and BP. The third group did not exercise and served as a control group (CG). Training frequency was 2 sessions per week with at least 72 hours of rest between sessions. Muscle volume was assessed at baseline and after 6 weeks and 12 weeks of training by ultrasound techniques. One repetition maximum strength for all exercises was assessed at baseline and after 12 weeks of training. Effect size data demonstrated that differences in strength and MV were exhibited based on exercise order. Both training groups demonstrated greater strength improvements than the CG, but only BP strength increased to a greater magnitude in the LG-SM group as compared with the SM-LG. In all other strength measures (LPD, TE, and BC), the SM-LG group showed significantly greater strength increases. Triceps MV increased in the SM-LG group; however, biceps MV did not differ significantly between the training groups. In conclusion, if an exercise is important for the training goals of a program, then it should be placed at the beginning of the training session, regardless of whether or not it is a large muscle group exercise or a small muscle group exercise.