p53-Bad* in a Hepatocellular Carcinoma Mouse Model.

Recent advances in liver cancer treatments have not changed the fact that the majority of patients will not survive the disease. In order to advance future liver cancer treatments, this work presents an exploration of various iterations of the liver cancer specific AFP promoter as well as the gene construct p53-Bad*. p53-Bad* is a mitochondrially targeted re-engineered p53 therapy that has shown previous success in a zebrafish HCC model. Both the most promising AFP promoter and p53-Bad* were packaged in an adenoviral delivery system and tested in vitro in liver cancer cell lines. Finally, mixed results for adenoviral p53-Bad* in vivo are presented, and this work suggests future modifications to study parameters in order to further explore the potential of p53-Bad* as a potential liver cancer therapeutic.

The Challenges and Prospects of p53-Based Therapies in Ovarian Cancer.

It has been well established that mutations in the tumor suppressor gene, p53, occur readily in a vast majority of cancer tumors, including ovarian cancer. Typically diagnosed in stages three or four, ovarian cancer is the fifth leading cause of death in women, despite accounting for only 2.5% of all female malignancies. The overall 5-year survival rate for ovarian cancer is around 47%; however, this drops to an abysmal 29% for the most common type of ovarian cancer, high-grade serous ovarian carcinoma (HGSOC). HGSOC has upwards of 96% of cases expressing mutations in p53. Therefore, wild-type (WT) p53 and p53-based therapies have been explored as treatment options via a plethora of drug delivery vehicles including nanoparticles, viruses, polymers, and liposomes. However, previous p53 therapeutics have faced many challenges, which have resulted in their limited translational success to date. This review highlights a selection of these historical p53-targeted therapeutics for ovarian cancer, why they failed, and what the future could hold for a new generation of this class of therapies.

p53-Bad* Fusion Gene Therapy Induces Apoptosis In Vitro and Reduces Zebrafish Tumor Burden in Hepatocellular Carcinoma.

With few curative treatments and a global yearly death rate of over 800,000, hepatocellular carcinoma (HCC) desperately needs new therapies. Although wild-type p53 gene therapy has been shown to be safe in HCC patients, it has not shown enough efficacy to merit approval. This work aims to show how p53 can be re-engineered through fusion to the pro-apoptotic BH3 protein Bcl-2 antagonist of cell death (Bad) to improve anti-HCC activity and potentially lead to a novel HCC therapeutic, p53-Bad*. p53-Bad* is a fusion of p53 and Bad, with two mutations, S112A and S136A. We determined mitochondrial localization of p53-Bad* in liver cancer cell lines with varying p53 mutation statuses via fluorescence microscopy. We defined the apoptotic activity of p53-Bad* in four liver cancer cell lines using flow cytometry. To determine the effects of p53-Bad* in vivo, we generated and analyzed transgenic zebrafish expressing hepatocyte-specific p53-Bad*. p53-Bad* localized to the mitochondria regardless of the p53 mutation status and demonstrated superior apoptotic activity over WT p53 in early, middle, and late apoptosis assays. Tumor burden in zebrafish HCC was reduced by p53-Bad* as measured by the liver-to-body mass ratio and histopathology. p53-Bad* induced significant apoptosis in zebrafish HCC as measured by TUNEL staining but did not induce apoptosis in non-HCC fish. p53-Bad* can induce apoptosis in a panel of liver cancer cell lines with varying p53 mutation statuses and induce apoptosis/reduce HCC tumor burden in vivo in zebrafish. p53-Bad* warrants further investigation as a potential new HCC therapeutic.

Advancing care and research for traumatic brain injury: a roadmap.

Across all segments of society in the United States, millions of adults and children experience a traumatic brain injury (TBI) each year, which may pose lifetime health and financial burdens in the billions of dollars. Efforts have been made to advance research and care with goals to improve awareness of the causes and consequences of TBI, but gaps still remain in understanding TBI and delivering high-quality care to everyone who needs it both in military and civilian life. At the request of the Department of Defense, the National Academies of Sciences, Engineering and Medicine recently convened experts to address existing gaps in brain injury science and systems of care. Although many people who experience a TBI recover fully, others experience long-term physical, emotional, and often financial consequences to the patient and family system, and require ongoing accommodations to support their return to the communities in which they live, learn, and work. A holistic approach within the context of osteopathic medicine may be helpful and enhance contributions within the field. This article will discuss the roadmap to help guide the field, including key conclusions and recommendations for actions to advance progress over the next decade while embracing a comprehensive bio-psycho-socio-ecological model of TBI care bringing in the distinctive osteopathic approach not only to improve care and outcomes, but also to understand patient and family experiences on their TBI journey.

Assessing the Risks and Benefits of Advances in Science and Technology: Exploring the Potential of Qualitative Frameworks.

Continuing rapid advances in science and technology both pose potential risks and offer potential benefits for the effective implementation of the Biological Weapons Convention (BWC). The lack of commonly accepted methods for assessing relevant risks and benefits present significant challenges to building common understandings that could support policy choices. This article argues that qualitative frameworks can provide the basis to structure BWC discussions about potential risks and benefits, reveal areas of agreement and disagreement, and provide a basis for continuing dialogue. It draws on the results of a workshop held in Geneva during the 2019 BWC Meetings of Experts. A diverse group of international experts were given the opportunity to apply 2 qualitative frameworks developed specifically to assess potential biosecurity concerns arising from emerging science and technology to BWC-relevant case examples. Participants discussed how such frameworks might be adapted and put into action to help support the BWC. They also began a discussion of how a comparable framework to assess potential benefits could be developed.

Advances in delivery vectors for gene therapy in liver cancer.

Hepatocellular carcinoma (HCC) is the third most common cause of cancer death globally, mainly due to lack of effective treatments - a problem that gene therapy is poised to solve. Successful gene therapy requires safe and efficient delivery vectors, and recent advances in both viral and nonviral vectors have made an important impact on HCC gene therapy delivery. This review explores how adenoviral, retroviral and adeno-associated viral vectors have been modified to increase safety and delivery capacity, highlighting studies and clinical trials using these vectors for HCC gene therapy. Nanoparticles, liposomes, exosomes and virosomes are also featured in their roles as HCC gene delivery vectors. Finally, new discoveries in gene editing technology and their impacts on HCC gene therapy are discussed.

p53-Bad: A Novel Tumor Suppressor/Proapoptotic Factor Hybrid Directed to the Mitochondria for Ovarian Cancer Gene Therapy.

Clinical trials involving p53 gene therapy for ovarian cancer failed due to the dominant negative inhibition of wild-type p53 and multiple genetic aberrations in ovarian cancer. To overcome this problem, we have designed a more potent chimeric gene fusion, called p53-Bad, that combines p53 with the mitochondrial pro-apoptotic factor Bad. Unlike wild-type p53, which acts as a nuclear transcription factor, this novel p53-Bad construct has multiple unique mechanisms of action including a direct and rapid apoptotic effect at the mitochondria. The mitochondrial localization, transcription activity, and apoptotic activity of the constructs were tested. The results suggest that p53 can be effectively targeted to the mitochondria by controlling the phosphorylation of pro-apoptotic Bad, which can only localize to the mitochondria when Ser-112 and Ser-136 of Bad are unphosphorylated. By introducing S112A and S136A mutations, p53-Bad fusion cannot be phosphorylated at these two sites and always localizes to the mitochondria. p53-Bad constructs also have superior activity over p53 and Bad alone. The apoptotic activity is consistent in many ovarian cancer cell lines regardless of the endogenous p53 status. Both p53 and the BH3 domain of Bad contribute to the superior activity of p53-Bad. Our data suggests that p53-Bad fusions are capable of inducing apoptosis and should be further pursued for gene therapy for ovarian cancer.

Mitochondrially targeted p53 or DBD subdomain is superior to wild type p53 in ovarian cancer cells even with strong dominant negative mutant p53.

BACKGROUND: While tumor suppressor p53 functions primarily as a transcription factor in the nucleus, cellular stress can cause p53 to translocate to the mitochondria and directly trigger a rapid apoptotic response. We have previously shown that fusing p53 (or its DNA binding domain, DBD, alone) to the mitochondrial targeting signal (MTS) from Bak or Bax can target p53 to the mitochondria and induce apoptosis in gynecological cancer cell lines including cervical cancer cells (HeLa; wt p53), ovarian cancer cells (SKOV-3; p53 267del non-expressing), and breast cancer cells (T47D; L194F p53 mutation). However, p53 with Bak or Bax MTSs have not been previously tested in cancers with strong dominant negative (DN) mutant p53 which are capable of inactivating wt p53 by homo-oligomerization. Since p53-Bak or Bax MTS constructs act as monomers, they are not subject to DN inhibition. For this study, the utility of p53-Bak or p53-Bax MTS constructs was tested for ovarian cancers which are known to have varying p53 statuses, including a strong DN contact mutant p53 (Ovcar-3 cells), a p53 DN structural mutant (Kuramochi cells), and a p53 wild type, low expressing cells (ID8). RESULTS: Our mitochondrial p53 constructs were tested for their ability to localize to the mitochondria in both mutant non-expressing p53 (Skov-3) and p53 structural mutant (Kuramochi) cell lines using fluorescence microscopy and a nuclear transcriptional activity assay. The apoptotic activity of these mitochondrial constructs was determined using a mitochondrial outer membrane depolarization assay (TMRE), caspase assay, and a late stage cell death assay (7-AAD). We also tested the possibility of using our constructs with paclitaxel, the current standard of care in ovarian cancer treatment. Our data indicates that our mitochondrial p53 constructs are able to effectively localize to the mitochondria in cancer cells with structural mutant p53 and induce apoptosis in many ovarian cancer cell lines with different p53 statuses. These constructs can also be used in combination with paclitaxel for an increased apoptotic effect. CONCLUSIONS: The results suggest that targeting p53 to mitochondria can be a new strategy for ovarian cancer treatment.

Narrowing the field: cancer-specific promoters for mitochondrially-targeted p53-BH3 fusion gene therapy in ovarian cancer.

BACKGROUND: Despite years of research, the treatment options and mortality rate for ovarian cancer remain relatively stagnant. Resistance to chemotherapy and high heterogeneity in mutations contribute to ovarian cancer's lethality, including many mutations in tumor suppressor p53. Though wild type p53 gene therapy clinical trials failed in ovarian cancer, mitochondrially-targeted p53 fusion constructs, including a fusion with pro-apoptotic protein Bad, have shown much higher apoptotic potential than wild type p53 in vitro. Due to the inherent toxicities of mitochondrial apoptosis, cancer-specificity for the p53 fusion constructs must be developed. Cancer-specific promoters such as hTERT, hTC, Brms1, and Ran have shown promise in ovarian cancer. RESULTS: Of five different lengths of hTERT promoter, the - 279/+ 5 length relative to the transcription start site showed the highest activity across a panel of ovarian cancer cells. In addition to - 279/+ 5, promoters hTC (an hTERT/CMV promoter hybrid), Brms1, and Ran were tested as drivers of mitochondrially-targeted p53-Bad and p53-Bad* fusion gene therapy constructs. p53-Bad* displayed cancer-specific killing in all ovarian cancer cell lines when driven by hTC, - 279/+ 5, or Brms1. CONCLUSIONS: Cancer-specific promoters hTC, - 279/+ 5, and Brms1 all display promise in driving p53-Bad* gene therapy for treatment of ovarian cancer and should be moved forward into in vivo studies. -279/+ 5 displays lower expression levels in fewer cells, but greater cancer specificity, rendering it most useful for gene therapeutics with high toxicity to normal cells. hTC and Brms1 show higher transfection and expression levels with some cancer specificity, making them ideal for lowering toxicity in order to increase dose without as much of a reduction in the number of cancer cells expressing the gene construct. Having a variety of promoters available means that patient genetic testing can aid in choosing a promoter, thereby increasing cancer-specificity and giving patients with ovarian cancer a greater chance at survival.

Personal Information Management for Nurses Returning to School.

Registered nurses with a diploma or an associate's degree are encouraged to return to school to earn a Bachelor of Science in Nursing degree. Until they return to school, many RNs have little need to regularly write, store, and retrieve work-related papers, but they are expected to complete the majority of assignments using a computer when in the student role. Personal information management (PIM) is a system of organizing and managing electronic information that will reduce computer clutter, while enhancing time use, task management, and productivity. This article introduces three PIM strategies for managing school work. Nesting is the creation of a system of folders to form a hierarchy for storing and retrieving electronic documents. Each folder, subfolder, and document must be given a meaningful unique name. Numbering is used to create different versions of the same paper, while preserving the original document.

Can I cut it? Medical students' perceptions of surgeons and surgical careers.

BACKGROUND: Recent years have seen a significant drop in applications to surgical residencies. Existing research has yet to explain how medical students make career decisions. This qualitative study explores students' perceptions of surgery and surgeons, and the influence of stereotypes on career decisions. METHODS: Exploratory questionnaires captured students' perceptions of surgeons and surgery. Questionnaire data informed individual interviews, exploring students' perceptions in depth. Rigorous qualitative interrogation of interviews identified emergent themes from which a cohesive analysis was synthesized. RESULTS: Respondents held uniform stereotypes of surgeons as self-confident and intimidating; surgery was competitive, masculine, and required sacrifice. To succeed in surgery, students felt they must fit these stereotypes, excluding those unwilling, or who felt unable, to conform. Deviating from the stereotypes required displaying such characteristics to a level exceptional even for surgery; consequently, surgery was neither an attractive nor realistic career option. CONCLUSIONS: Strong stereotypes of surgery deterred students from a surgical career. As a field, surgery must actively engage medical students to encourage participation and dispel negative stereotypes that are damaging recruitment into surgery.

You've got to know the rules to play the game: how medical students negotiate the hidden curriculum of surgical careers.

OBJECTIVES: The hidden curriculum may be framed as the culture, beliefs and behaviours of a community that are passed to students outside formal course offerings. Medical careers involve diverse specialties, each with a different culture, yet how medical students negotiate these cultures has not been fully explored. Using surgery as a case study, we aimed to establish, first, whether a specialty-specific hidden curriculum existed for students, and second, how students encountered and negotiated surgical career options. METHODS: Using a constructivist grounded theory approach, we explored students' thoughts, beliefs and experiences regarding career decisions and surgery. An exploratory questionnaire informed the discussion schedule for semi-structured individual interviews. Medical students were purposively sampled by year group, gender and career intentions in surgery. Data collection and analysis were iterative: analysis followed each interview and guided the adaptation of our discussion schedule to further our evolving model. RESULTS: Students held a clear sense of a hidden curriculum in surgery. To successfully negotiate a surgical career, students perceived that they must first build networks because careers information flows through relationships. They subsequently enacted what they learned by accruing the accolades ('ticking the boxes') and appropriating the dispositions ('walking the talk') of 'future surgeons'. This allowed them to identify themselves and to be identified by others as 'future surgeons' and to gain access to participation in the surgical world. Participation then enabled further network building and access to careers information in a positive feedback loop. For some, negotiating the hidden curriculum was more difficult, which, for them, rendered a surgical career unattractive or unattainable. CONCLUSIONS: Students perceive a clear surgery-specific hidden curriculum. Using a constructivist grounded theory approach, we have developed a model of how students encounter, uncover and enact this hidden curriculum to succeed. Drawing on concepts of Bourdieu, we discuss unequal access to the hidden curriculum, which was found to exclude many from the possibility of a surgical career.

Research recognition, the first step to research literacy.

Nurses are expected to use research as a form of evidence to support clinical practice. When selecting articles, some nurses may be unable to recognize the articles that report research. Research recognition is essential before study results may be used as evidence and translated into practice. The purpose of this article is to describe a typical research article, outline the major types of research, and identify the unique characteristics of research that may be used to recognize those articles that report research.

Childhood maltreatment among community dwelling adult Latinas.

There is a lack of research about childhood maltreatment among adult Latinas in the United States. The aim of this secondary analysis was to explore the occurrence and co-occurrence of maltreatment during childhood in a community-dwelling sample of adult Latinas. Participants were 62 women ages 18-45 who read and spoke English, attended work or school on a regular basis, and had no health or functional interferences with daily living. On average, participants were 29 years of age (M = 29.08, SD = 9.30), high school graduates (n = 36, 58.1%), married (n = 36, 58.1%), and employed (n = 27, 43.5%). Over three quarters of participants (n = 49, 79.0%) reported at least one form of childhood maltreatment: emotional abuse (n = 24, 38.7%), physical abuse (n = 29, 46.8%), sexual abuse (n = 29, 46.8%), emotional neglect (n = 27, 43.5%), and physical neglect (n = 23, 37.1%). Maltreatment severity increased with the number of maltreatment forms. Women sexually abused during childhood were more likely than women without this history to be employed and high school graduates. These Latinas may have developed more adaptive coping mechanisms than other Latinas.

Examining the relationship between a childhood history of sexual abuse and later dissociation, breast-feeding practices, and parenting anxiety.

The purpose of this study is to compare Mexican American adolescent mothers with and without childhood sexual abuse (CSA) histories to examine the influence of CSA on dissociation, selection of infant feeding method, and intimate parenting anxiety. Participants are 78 English-speaking adolescents between 15 and 19 years of age and recruited from the southwestern United States. Nearly one third of the sample ( n = 24, 30.77%) reports CSA histories. There is no correlation between CSA history and intimate parenting anxiety, no difference between breast-feeding and formula-feeding mothers in CSA severity, and intimate parenting anxiety does not predict dissociation. These findings are inconsistent with previous research. Supportive resources may explain the inconsistency and play a role in adolescent mothers' responses to CSA. Further research is necessary to explore these possibilities.

A neural code for three-dimensional object shape in macaque inferotemporal cortex.

Previous investigations of the neural code for complex object shape have focused on two-dimensional pattern representation. This may be the primary mode for object vision given its simplicity and direct relation to the retinal image. In contrast, three-dimensional shape representation requires higher-dimensional coding derived from extensive computation. We found evidence for an explicit neural code for complex three-dimensional object shape. We used an evolutionary stimulus strategy and linear/nonlinear response models to characterize three-dimensional shape responses in macaque monkey inferotemporal cortex (IT). We found widespread tuning for three-dimensional spatial configurations of surface fragments characterized by their three-dimensional orientations and joint principal curvatures. Configural representation of three-dimensional shape could provide specific knowledge of object structure to support guidance of complex physical interactions and evaluation of object functionality and utility.

Gene transfer to hemophilia A mice via oral delivery of FVIII-chitosan nanoparticles.

Effective oral delivery of a non-viral gene carrier would represent a novel and attractive strategy for therapeutic gene transfer. To evaluate the potential of this approach, we studied the oral gene delivery efficacy of DNA polyplexes composed of chitosan and Factor VIII DNA. Transgene DNA was detected in both local and systemic tissues following oral administration of the chitosan nanoparticles to hemophilia A mice. Functional factor VIII protein was detected in plasma by chromogenic and thrombin generation assays, reaching a peak level of 2-4% FVIII at day 22 after delivery. In addition, a bleeding challenge one month after DNA administration resulted in phenotypic correction in 13/20 mice given 250-600 microg of FVIII DNA in chitosan nanoparticles, compared to 1/13 mice given naked FVIII DNA and 0/6 untreated mice. While further optimization would be required to render this type of delivery system practical for hemophilia A gene therapy, the findings suggest the feasibility of oral, non-viral delivery for gene medicine applications.