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Introduction Retinoic acid (RA) is a differentiating agent utilized as maintenance therapy for high-risk neuroblastoma (NB), but associated toxicities limit its use. We have previously shown that a non-toxic, novel rexinoid, 9-cis-UAB30 (UAB30), decreased NB cell proliferation and in vivo tumor growth. A second generation, mono-methylated compound, 6-Methyl-UAB30 (6-Me), has been recently designed having greater potency compared with UAB30. In the current study, we hypothesized that 6-Me would inhibit NB cell proliferation and survival and induce differentiation and cell-cycle arrest. Methods Proliferation and viability were measured in four human NB cell lines following treatment with UAB30 or 6-Me. Cell-cycle was analyzed and tumor cell stemness was evaluated with extreme limiting dilution assays and immunoblotting for expression of stem cell markers. A xenograft murine model was utilized to study the effects of 6-Me in vivo. Results Treatment with 6-Me led to decreased proliferation and viability, induced cell cycle arrest, and increased neurite outgrowth, indicating differentiation of surviving cells. Furthermore, treatment with 6-Me decreased tumorsphere formation and expression of stem cell markers. Finally, inhibition of tumor growth and increased animal survival was observed in vivo following treatment with 6-Me. Conclusion These results indicate a potential therapeutic role for this novel rexinoid in neuroblastoma treatment.

The archaeological rescue excavations occasioned by the construction of the Lot no. 1 of the Highway no. 10, Sebeș-Turda, highlighted an unexpected Bronze Age necropolis belonging to Wietenberg Culture. The Bronze Age necropolis was delimited between 6+600 and 6+650 kilometers, and located on the southwestern part of the Site no. 6, right above the early Vinča settlement defensive trenches (Fig. 1-2). With 74 excavated funerary complexes, the partially excavated Wietenberg cemetery is one of the largest in Transylvania. According the radiocarbon data presented in this study, the necropolis was used in the Middle Bronze Age (1780-1690 cal. BC), associated with the Wietenberg culture (phase II), and is contemporary with the end of the cemetery from Sebeș-Între Răstoace located approximately 7.5 km South - South-West, on the Sebeș River terraces.

Highlights • Rare, high grade neuroendocrine-like pediatric tumor has TYRO3 and ALK mutations. • Crizotinib downregulates STAT3 and ERK1/2 signaling in tumor. • In vivo analysis demonstrated crizotinib resistance. • Crizotinib resistant cells have upregulation of PDGFRß signaling. • PDGFRß inhibition overrides resistance., Patient-derived xenografts provide significant advantages over long-term passage cell lines when investigating efficacy of treatments for solid tumors. Our laboratory encountered a high-grade, metastatic, neuroendocrine-like tumor from a pediatric patient that presented with a unique genetic profile. In particular, mutations in TYRO3 and ALK were identified. We established a human patient-derived xenoline (PDX) of this tumor for use in the current study. We investigated the effect of crizotinib, a chemotherapeutic known to effectively target both TYRO3 and ALK mutations. Crizotinib effectively decreased viability, proliferation, growth, and the metastatic properties of the PDX tumor through downregulation of STAT3 signaling, but expression of PDGFRß was increased. Sunitinib is a small molecule inhibitor of PDGFRß and was studied in this PDX independently and in combination with crizotinib. Sunitinib alone decreased viability, proliferation, and growth in vitro and decreased tumor growth in vivo. In combination, sunitinib was able to overcome potential crizotinib-induced resistance through downregulation of ERK 1/2 activity and PDGFRß receptor expression; consequently, tumor growth was significantly decreased both in vitro and in vivo. Through the use of the PDX, it was possible to identify crizotinib as a less effective therapeutic for this tumor and suggest that targeting PDGFRß would be more effective. These findings may translate to other solid tumors that present with the same genetic mutations.

As archaeologists expand the accessibility of legacy data, they have an opportunity to use these datasets to design future research. We argue that legacy data can be a critical resource to help predict characteristics of sites and socioeconomic systems. In this article, we present a combined geographic information system (GIS) and network analysis methodology that turns site location data into testable hypotheses about site characteristics and the organization of regional settlement systems. We demonstrate the utility of this approach with a case study: Bronze Age (2700–1100 BC) settlement patterns in the mining region of Hunedoara in southwest Transylvania, Romania. We leverage unsystematically collected site location information in legacy datasets to develop testable predictions about sites, regional networks, and socioeconomic systems that can be evaluated through future systematic surveys and large-scale excavations. Such testable hypotheses can inform archaeological research design by providing a quantitative basis for determining where to focus research efforts and can also help secure funding and fieldwork permits. The method developed here can be applied in diverse archaeological contexts to reinvigorate legacy data as part of future archaeological research design.

Despite increasing incidence, treatment for hepatoblastoma has not changed significantly over the past 20 years. Chemotherapeutic strategies continue to rely on cisplatin, as it remains the most active single agent against hepatoblastoma. However, chemoresistance remains a significant challenge with 54–80% of patients developing resistance to chemotherapy after 4–5 cycles of treatment. Stem cell-like cancer cells (SCLCCs) are a subset of cells thought to play a role in chemoresistance and disease recurrence. We have previously demonstrated that Proviral Integration site for Moloney murine leukemia virus (PIM) kinases, specifically PIM3, play a role in hepatoblastoma cell proliferation and tumor growth and maintain the SCLCC phenotype. Here, we describe the development of a cisplatin-resistant hepatoblastoma xenograft model of the human HuH6 cell line and a patient-derived xenograft, COA67. We provide evidence that these cisplatin-resistant cells are enriched for SCLCCs and express PIM3 at higher levels than cisplatin-naïve cells. We demonstrate that PIM inhibition with AZD1208 sensitizes cisplatin-resistant hepatoblastoma cells to cisplatin, enhances cisplatin-mediated apoptosis, and decreases the SCLCC phenotype seen with cisplatin resistance. Together, these findings indicate that PIM inhibition may be a promising adjunct in the treatment of hepatoblastoma to effectively target SCLCCs and potentially decrease chemoresistance and subsequent disease relapse.

Simple Summary Children with high-risk neuroblastoma have limited therapeutic options poor survival rates. The neuroblastoma tumor microenvironment contributes the lack of response to many interventions so innovative methods are needed to study the effects of the tumor microenvironment on new therapies. In this manuscript, we review the current literature related to the components of the tumor microenvironment and to the use of three-dimensional printing as modality to study cancer. This review highlights the potential for using three-dimensional printing to create an artificial tumor microenvironment in the presence of neuroblastoma to provide improved preclinical testing of novel therapies. Abstract In the quest to advance neuroblastoma therapeutics, there is a need to have a deeper understanding of the tumor microenvironment (TME). From extracellular matrix proteins to tumor associated macrophages, the TME is a robust and diverse network functioning in symbiosis with the solid tumor. Herein, we review the major components of the TME including the extracellular matrix, cytokines, immune cells, and vasculature that support a more aggressive neuroblastoma phenotype and encumber current therapeutic interventions. Contemporary treatments for neuroblastoma are the result of traditional two-dimensional culture studies and in vivo models that have been translated to clinical trials. These pre-clinical studies are costly, time consuming, and neglect the study of cofounding factors such as the contributions of the TME. Three-dimensional (3D) bioprinting has become a novel approach to studying adult cancers and is just now incorporating portions of the TME and advancing to study pediatric solid. We review the methods of 3D bioprinting, how researchers have included TME pieces into the prints, and highlight present studies using neuroblastoma. Ultimately, incorporating the elements of the TME that affect neuroblastoma responses to therapy will improve the development of innovative and novel treatments. The use of 3D bioprinting to achieve this aim will prove useful in developing optimal therapies for children with neuroblastoma.

Background: Current chemotherapy regimens for hepatoblastoma include platinum-based combinations that have remained virtually unchanged over the last twenty years. The overall survival for children with advanced stage or recurrent disease utilizing these regimens remains poor. We have previously shown that Proviral Integration site for Moloney murine leukemia virus (PIM) kinases act as oncoproteins promoting hepatoblastoma cell survival and the cancer stem cell phenotype, which may contribute to chemoresistance. We hypothesized that PIM inhibition utilizing …

Background: Serine-threonine kinase receptor associated protein (STRAP) is a scaffolding protein that has been implicated in tumor growth and metastasis. The role of STRAP in neuroblastoma (NB) has not been explored. Stem cell-like cancer cells (SCLCCs) are believed to be responsible for tumor cell self-renewal and cancer recurrence. We sought to determine the role of STRAP in neuroblastoma SCLCCs maintenance. …

Introduction The poor therapeutic efficacy seen with current treatments for neuroblastoma may be attributed to stem cell-like cancer cells (SCLCCs), a subpopulation of cancer cells associated with poor prognosis and disease recurrence. Retinoic acid (RA) is a differentiating agent used as maintenance therapy for high-risk neuroblastoma but nearly half of children treated with RA relapse. We hypothesized that 6-Methyl-UAB30 (6-Me), a second-generation rexinoid recently developed with a favorable toxicity profile compared to RA, would reduce cancer cell stemness in human neuroblastoma patient-derived xenografts (PDXs). Methods Cells from three neuroblastoma PDXs were treated with 6-Me and proliferation, viability, motility, and cell-cycle progression were assessed. CD133 expression, sphere formation, and mRNA abundance of stemness and differentiation markers were evaluated using flow cytometry, in vitro extreme limiting dilution analysis, and real-time PCR, respectively. Results Treatment with 6-Me decreased proliferation, viability, and motility, and induced cell-cycle arrest and differentiation in all three neuroblastoma PDXs. In addition, 6-Me treatment led to decreased CD133 expression, decreased sphere-forming ability, and decreased mRNA abundance of Oct4, Nanog, and Sox2, indicating decreased cancer cell stemness. Conclusions 6-Me decreased oncogenicity and reduced cancer cell stemness of neuroblastoma PDXs, warranting further exploration of 6-Me as potential novel therapy for neuroblastoma.

Background Novel therapies are needed for patients with hepatoblastoma because of an increasing incidence of disease and poor prognosis for advanced, refractory, and recurrent disease. PIM kinases promote tumorigenesis in hepatoblastoma. A novel PIM inhibitor, PIM447, has shown promise in inhibiting oncogenesis in hematologic and lymphoid malignancies. We hypothesized that PIM inhibition with PIM447 would result in decreased tumorigenesis in hepatoblastoma. Methods The effects of PIM447 on hepatoblastoma viability, proliferation, motility, apoptosis, and tumor cell stemness were assessed in HuH6, a human hepatoblastoma cell line, and COA67, a human hepatoblastoma patient-derived xenograft. Results PIM447 significantly decreased the viability, proliferation, and motility of HuH6 and COA67 cells. Apoptosis significantly increased following PIM447 treatment. PIM447 had a significant impact on tumor cell stemness as evidenced by decreased expression of CD133 and reduced ability of HuH6 and COA67 cells to form tumorspheres. Furthermore, combining PIM447 with cisplatin resulted in a significant decrease in cell viability compared to either treatment alone. Conclusion We showed that PIM447 inhibits oncogenesis and potentiates the effects of cisplatin in hepatoblastoma and, therefore, warrants further investigation as a potential therapeutic agent for hepatoblastoma.

The Bronze Age was a period of significant socio-economic transformation that gave rise to the first complex regional polities with institutionalized inequality in Europe. Communities in southwest Transylvania, a major source of gold, copper, and salt, played a critical role in this transformation. This article examines how socio-economic changes affected how people situated settlements in resource procurement zones during the Early and Middle Bronze Age (2700–1500 cal. BCE). Taking advantage of the heterogeneous distribution of natural resources across the landscape, a GIS catchment analysis of the orientation of settlements toward particular constellations of resources is presented. Our results show increased preference for access to high quality agricultural land and access to interregional trade through the Mureș River corridor over the course of the Bronze Age. Despite the increased importance of metal within Bronze Age economies, there is no evidence that Transylvanian communities placed their settlements to maximize their ability to contest or secure access to the metal ore sources in the Apuseni Mountains. The organization of settlement systems in the Bronze Age demonstrates that Transylvanian communities prioritized socio-economic institutions beyond metal procurement. This study demonstrates that tracing how humans situate themselves in variable landscapes can provide new insights into the conditions and mechanisms of social change.

10040 Background: Osteosarcoma is the most common primary bone tumor in children. For those with relapsed or metastatic disease, the five-year survival rate is approximately 20%, and survivors often suffer from long-term disability from current therapies. The high morbidity and mortality for these patients highlight a great need for improved therapies. One such novel therapeutic approach is oncolytic herpes simplex virus (oHSV) immunovirotherapy. We previously demonstrated that M002, an engineered oHSV that contains deletions of the neurovirulence gene preventing infection of normal cells, effectively infects and kills neuroblastoma and rhabdomyosarcoma. Currently, similar oHSVs are being evaluated in early phase clinical trials for children and adults with relapsed or refractory brain tumors. To date, there has been limited investigation of oncolytic virotherapy in osteosarcoma. Thus, we sought to examine the ability of oHSV, M002, to infect and kill osteosarcoma cells in vitro. Methods: We evaluated two long-term passaged human osteosarcoma cell lines, U2-OS and MG-63. Flow cytometry was used to assess baseline expression of oHSV viral entry-mediated receptors (CD111, CD112, syndecan, HVEM). Single and multi-step viral recovery experiments measured virus infectivity and replication. Cells were infected with increasing multiplicity of infection (MOI) of M002, and cell viability was measured 72 hours post-infection via alamarBlue assay. Results: Both MG-63 and U2-OS cells expressed HSV entry molecules (Table) including high levels of the primary HSV entry molecule CD111. Single step virus recovery experiments in MG-63 cells infected at a MOI of 10 plaque-forming units (PFU)/cell demonstrated a 3 log-fold increase in virus titer from 12 to 24 hours post-infection. For multi-step experiments, MG-63 cells were infected with a MOI of 0.1 PFU/cell; viral replication significantly increased from 1.1x103 PFU at 6 hours post-infection to 3.8x1010 PFU at 72 hours post-infection. M002 successfully decreased osteosarcoma viability with a lethal dose in 50% of cells (LD50)of 2.82 and0.67 PFU/cell for MG-63 and U2-OS cells, respectively. Notably, at a virus MOI of 5 PFU/cell, viability was decreased by 64% ± 0.1% (p

The use of prior exercise training has shown promise in minimizing doxorubicin (DOX)-induced physical impairments. The purpose of this study was to compare changes in thymus mass, thymocyte (T-cell) number, and tissue peroxidation following chronic endurance exercise and DOX treatment in the rat. The thymus mass, number of viable T-cells, and levels of malondialdehyde and 4-hydroxyalkenals (MDA+4-HAE) were compared 3 days post-injection between rats assigned to the following treatment conditions: ( a) 10 weeks of endurance training, followed by a saline injection 24 hours after the last training session (TM+SAL); ( b) treadmill training as above, followed by a single, bolus 10-mg/kg injection of DOX (TM+10); ( c) treadmill training with 12.5 mg/kg of DOX (TM+12.5); ( d) sedentary (without exercise) and a saline injection (SED+SAL); ( e) sedentary with 10 mg/kg of DOX (SED+10); and ( f) sedentary with 12.5 mg/kg (SED+12.5). Thymic mass and T-cell numbers significantly decreased following DOX injections. TM rats exhibited significantly less lipid peroxidation compared with paired-dose SED groups. TM+10 did not significantly differ from SED+SAL in thymic levels of lipid peroxidation. We conclude that chronic endurance exercise decreases levels of lipid peroxidation in the thymus seen with acute DOX treatment.

Diarrheal diseases are a leading cause of morbidity and mortality in Africa. Although research documents the magnitude and pattern of diarrheal diseases are associated with weather in particular locations, there is limited quantification of this association in sub-Saharan Africa and no studies conducted in Mozambique. Our study aimed to determine whether variation in diarrheal disease was associated with precipitation in Mozambique. In secondary analyses we investigated the associations between temperature and diarrheal disease. We obtained weekly time series data for weather and diarrheal disease aggregated at the administrative district level for 1997–2014. Weather data include modeled estimates of precipitation and temperature. Diarrheal disease counts are confirmed clinical episodes reported to the Mozambique Ministry of Health (n = 7,315,738). We estimated the association between disease counts and precipitation, defined as the number of wet days (precipitation > 1 mm) per week, for the entire country and for Mozambique’s four regions. We conducted time series regression analyses using an unconstrained distributed lag Poisson model adjusted for time, maximum temperature, and district. Temperature was similarly estimated with adjusted covariates. Using a four-week lag, chosen a priori, precipitation was associated with diarrheal disease. One additional wet day per week was associated with a 1.86% (95% CI: 1.05–2.67%), 1.37% (95% CI: 0.70–2.04%), 2.09% (95% CI: 1.01–3.18%), and 0.63% (95% CI: 0.11–1.14%) increase in diarrheal disease in Mozambique’s northern, central, southern, and coastal regions, respectively. Our study indicates a strong association between diarrheal disease and precipitation. Diarrheal disease prevention efforts should target areas forecast to experience increased rainfall. The burden of diarrheal disease may increase with increased precipitation associated with climate change, unless additional health system interventions are undertaken.

Background The skeletal muscle toxicity that accompanies the chemotherapy drug doxorubicin (DOX) may lead to cancer patient weakness and fatigue. This myotoxicity involves myogenic regulatory factor (MRF) disruption which alters muscle integrity and regeneration. Endurance exercise enhances MRF expression and thereby may mitigate DOX-induced MRF disruptions. This study examined the effects of endurance training and DOX treatment on myogenic regulatory factor (MRF) expression. Methods Male rats were exercise trained (EXER) or remained sedentary (SED) for two weeks. EXER and SED then received either DOX (15 mg/kg) or saline (SAL). Soleus, extensor digitorum longus (EDL), and diaphragm were excised 24 hours post injection, and MRF expression was analyzed. Results Significant Myf5 drug and activity effects were observed in the soleus with EXER+DOX expressing higher Myf5 than SED+DOX. A significant drug effect was detected in soleus MyoD, and a significant activity effect was detected in soleus Mrf4. No main effects or interactions were observed in the EDL, but in the diaphragm, a significant activity effect was observed for Myf5 with EXER+DOX expressing higher levels than SED+DOX. Conclusion Doxorubicin treatment increased soleus MRFs and exercise boosted MRF response in soleus and diaphragm suggesting that exercise may enhance regenerative signaling with DOX treatment. Level of evidence I b, individual randomized controlled trial.

Lisano, JK, Smith, JD, Mathias, AB, Christensen, M, Smoak, P, Phillips, KT, Quinn, CJ, and Stewart, LK. Performance and health-related characteristics of physically active men using marijuana. J Strength Cond Res 33(6): 1659-1669, 2019-The influence of chronic marijuana use on the performance and health of physically active individuals has yet to be fully elucidated. The purpose of this study was to explore pulmonary function, aerobic and anaerobic fitness, strength, serum testosterone, cortisol, C-reactive protein (CRP), Δ-9-tetrahydrocannabinol (THC), 11-nor-9-carboxy-Δ-9-tetrahydrocannabinol (THC-COOH), and 11-hydroxy-Δ-9-tetrahydrocannabinol (THC-OH) concentrations in a physically active population either using or not using marijuana. Healthy, physically active males (N = 24) were compared based on their marijuana-use status: marijuana users (MU; n = 12) and nonusers (NU; n = 12). Statistical analysis (p = 0.05) revealed no difference between groups for age, body mass, body mass index, body fat, forced expiratory volume in 1 second percentage, VO2max, anaerobic power output, strength measures, testosterone, or cortisol concentrations. Although not statistically significant, MU showed a trend to fatigue to a greater percentage of absolute power output than NU from the beginning to the end of the Wingate Anaerobic Power Assessment (p = 0.08, effect size = 0.75). C-reactive protein in MU (1.76 ± 2.81 mg·L) and NU (0.86 ± 1.49 mg·L) was not significantly different (p = 0.60) but placed MU at moderate risk and NU at low risk for cardiovascular disease. Anaerobic fatigue was the only performance variable to show a trend for difference between groups. These results suggest that marijuana use in physically active males may not have significant effects on performance; however, it may be linked to elevated concentrations of CRP which place users at a higher risk for cardiovascular disease.

1. Doxorubicin (DOX) is a highly effective and commonly used anthracycline antibiotic used to treat cancer patients. The side effects of DOX are manifested in a more delayed manner in children and multidrug resistant proteins (MRPs) may factor into this phenomenon. MRPs are known to extrude DOX and may factor into the degree of cardiac DOX accumulation. 2. The purpose of this study was to examine age-related differences in muscle MRP expression and DOX accumulation. 3. Female Sprague-Dawley rats were randomly selected to receive a 15-mg DOX/kg body weight bolus injection (i.p.) at various ages. 4. Cardiac and extensor digitorum longus DOX accumulation was markedly increased as animals aged from 4 to 24 weeks. In contrast, no differences in soleus accumulation were observed. A significant age-related reduction in MRP-2 and MRP-7 expression was detected in cardiac and extensor digitorum longus tissues with no age differences in MRP-1 expression in any tissues analyzed. MRP-6 was not detected in any tissues. 5. These data suggest that aging is associated with increased DOX accumulation and an age-related decrease in MRP expression may be a factor.

Chemotherapy treatment with doxorubicin (DOX) can have a negative effect on normal skeletal muscle function. Recent research demonstrates the potential value of exercise in alleviating DOX-induced cardiotoxicity. Yet up to now, little research has been done to examine whether exercise might also be effective in addressing DOX's skeletal muscle adverse effects, especially because posttreatment skeletal muscle dysfunction may cause patient difficulties with completing activities of daily living. The main aim of this study was to examine how resistance training (RT) and treadmill (TM) training play a role in preventing DOX-induced skeletal muscle dysfunction.Male Sprague-Dawley rats were randomly placed into an RT, TM, or sedentary (SED) group for 10 wk and then received either a bolus injection of DOX (15 mg·kg) or saline as a control. Skeletal muscle function was then assessed ex vivo 5 d after injection.SED animals treated with DOX showed significantly lower maximal twitch force, maximal rate of force production, and maximal rate of force decline versus SED + saline in the soleus (SOL) (Type I muscle). In the extensor digitorum longus (Type II muscle), treatment with DOX resulted in a significantly lower maximal rate of force production and maximal rate of force decline. RT preserved maximal twitch force and maximal rate of force decline in the SOL. TM attenuated DOX-induced fatigue in the SOL but not in the extensor digitorum longus.These findings suggest that RT and TM before DOX could be useful in preserving skeletal muscle function and minimizing fatigue after chemotherapy, but this protection may be dependent on the skeletal muscle type.

Doxorubicin (DOX), an anthracycline antiobiotic, has been shown to decrease skeletal muscle function via underlying mechanisms associated with the generation of reactive oxygen species. Prior studi...

This study analyzed the effects of prior endurance exercise or resistance training exercise on skeletal muscle dysfunction induced by the chemotherapy drug doxorubicin (DOX). Furthermore, this stud...

The chemotherapy drug doxorubicin (DOX) has been shown to cause skeletal muscle dysfunction when measured ex vivo, but little is known about the effects of DOX on in vivo skeletal muscle function. ...

Introduction The element selenium (Se), named after the moon (Greek selene), has long been known for its toxicity. In 1295, Marco Polo was the 1st to describe the typical symptoms of selenosis in his pack animals. Centuries later, selenosis may have played a role in the demise of General Custer in the battle of Little Big Horn. Still today, chronic or acute Se poisoning is responsible for the loss of thousands of head of livestock every year in the western United States, where Se is prevalent in soils. Due to its chemical similarity to sulfur, Se in excess will nonspecifi cally replace sulfur in proteins, leading to the observed toxicity. At low levels, however, Se is also an essential trace element for many organisms, including humans, as a component of specifi c selenoproteins that often play a role in cancer prevention. Selenosis in animals mostly is due to ingestion of Se hyperaccumulator plants, commonly termed “locoweeds” by ranchers. These plants can accumulate Se to around 1% of their dry weight, 100-fold higher levels than those found in surrounding vegetation, and 1,000-fold higher than soil Se. Selenium hyperaccumulator species mainly occur in the genera Astragalus (Fabaceae) and Stanleya (Brassicaceae), and predominantly are found on Se-rich soils as seen in the western United States. Several hypotheses exist for the functional signifi cance of the intriguing phenomenon of hyperaccumulation. We currently study the elemental defense hypothesis, which proposes that the accumulated Se protects the plants from herbivory or microbial infection. We also are interested in how accumulated Se in plants affects local ecological interactions (Pilon-Smits and Freeman 2006; Fig. 1A). In this paper, we give an overview of our fi ndings so far, followed by an overall discussion of their implications with respect to the evolution of hyperaccumulator plants and the management of Se-rich plants in natural and agricultural settings.