Striking efficacy of a vaccine targeting TOP2A for triple-negative breast cancer immunoprevention.

Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer that has a poor prognosis. TOP2A is a key enzyme in DNA replication and is a therapeutic target for breast and other cancers. TOP2A-specific Th1-promoting epitopes with optimal binding affinity to MHC II were identified using a combined scoring system. The multi-peptide TOP2A vaccine elicited a robust immunologic response in immunized mice, as demonstrated by the significant production of Th1 cytokines from immunized animals' splenocytes stimulated in vitro with TOP2A peptides. Anti-tumor efficacy of the TOP2A vaccine was demonstrated in a syngeneic TNBC mouse model, in which pre-graft preventive vaccination was associated with significantly decreased tumor growth as compared to adjuvant control. In a genetically engineered mouse (GEM) model of TNBC, vaccinated animals demonstrated a significant reduction in tumor incidence and average tumor volume compared to adjuvant control. Finally, we examined TCR sequences in CD4 tumor Infiltrating lymphocytes (TIL) from vaccinated mice and found that the TIL contained TCR sequences specific to the three vaccine peptides. These data indicate that our newly developed multi-peptide TOP2A vaccine is highly immunogenic, elicits TILs with vaccine specific TCRs, and is highly effective in preventing and intercepting TNBC development and progression in vivo.

Disease-preventive behaviors and subjective well-being in the COVID-19 pandemic.

BACKGROUND: Safety precautions and activity restrictions were common in the early, pre-vaccine phases of the COVID-19 pandemic. We hypothesized that higher levels of participation in potentially risky social and other activities would be associated with greater life satisfaction and perceived meaning in life. At the same time, prosocial COVID-preventive activities such as mask wearing should enhance life satisfaction. METHOD: We assessed the impact of COVID-preventive behaviors on psychological well-being in October 2020. A nationally representative sample of U.S. adults (n = 831) completed a demographic questionnaire, a COVID-related behaviors questionnaire, a Cantril's Ladder item, and the Multidimensional Existential Meaning Scale. Two hierarchical linear models were used to examine the potential impact of COVID-preventive behaviors on life satisfaction and meaning in life while accounting for the influence of demographic factors. RESULTS: The study revealed significant positive relationships between COVID-preventive behaviors and subjective well-being. Wearing a mask was significantly associated with higher life satisfaction, while maintaining social distancing of six feet and avoiding large groups were significantly associated with higher perceived meaning in life. Social activities including dining at restaurants and visiting friends and family were also significantly associated with higher life satisfaction and meaning in life, respectively. CONCLUSION: The study's findings support the conclusion that disease prevention measures such as social distancing and mask wearing do not reduce, and may enhance, subjective well-being during a pandemic. Utilizing the unique context of the COVID-19 pandemic to examine relationships between behavior and subjective well-being, the study also indicates that shallow or medium-depth social activities are likely to be more central to life satisfaction, whereas narrower, deeper social interactions with friends and family are more important to perceived meaning in life.

The proust effect: Scents, food, and nostalgia.

Autobiographical memories activated by the senses, particularly smell and taste, can be among the most potent and influential, an experience labelled the Proust Effect. Contemporary research has helped to explain the physiological, neurological, and psychological reasons underlying this phenomenon. Nostalgic memories triggered by taste and smell are especially self-relevant, arousing, and familiar. These memories have an even more positive emotional profile than nostalgic memories elicited by other means, with individuals reporting lower levels of negative or ambivalent emotions. Scent-evoked and food-evoked nostalgia also confer numerous psychological benefits, including enhanced self-esteem, feelings of social connectedness, and deeper meaning in life. Such memories might be harnessed in clinical or other settings.

Food-evoked nostalgia.

In three studies, we examined food as an elicitor of nostalgia. Study 1 participants visualised eating either a nostalgic or regularly consumed food. Study 2 participants visualised consuming 12 foods. Study 3 participants consumed 12 flavour samples. Following their food experiences, all participants responded to questions regarding the profile of food-evoked nostalgia (i.e. autobiographical relevance, arousal, familiarity, positive and negative emotions) and several psychological functions (i.e. positive affect, self-esteem, social connectedness, meaning in life). Study 2 and 3 participants also reported their state nostalgia. Results revealed that food is a powerful elicitor of nostalgia. Food-evoked nostalgia has a similar contextual profile to previously examined elicitors, but is a predominantly positive emotional experience. Food-evoked nostalgia served multiple psychological functions and predicted greater state nostalgia.

Overproduction and purification of Mycobacterium tuberculosis WhiB3 in Escherichia coli is enhanced by co-expression with trigger factor chaperone.

Members of the WhiB-like (Wbl) family of proteins are found in Acintomycetes and are somewhat recalcitrant to overproduction as soluble proteins in the laboratory protein expression workhorse Esherichia coli. The aim of this study was to evaluate the effects of culture conditions and co-expression of the chaperone protein, trigger factor (TF), on the soluble production of recombinant Mycobacterium tuberculosis (Mtb) WhiB3. A pET28a derived expression plasmid coding for His6-WhiB3 was created and the effects of varying the concentration of inducer (IPTG), the timing of induction, the nature of the inducer (auto-induction medium) and the temperature of the cultivation on the production of soluble His6-WhiB3 were tested. Whilst His6-WhiB3 protein was readily detected, the overwhelming majority of the protein was present in the insoluble fraction of cell-free extracts. However, co-expression of the tig from pTf16, coding for TF, increased His6-WhiB3 solubility dramatically, facilitating its isolation by affinity chromatography. Purified His6-WhiB3 was shown to be monomeric, and UV-visible spectra suggested that ∼10% of the isolated protein possessed a [4Fe-4S] cluster. The secondary structural properties of His6-WhiB3 were altered by acquisition of an iron-sulfur cluster. By developing a protocol to readily overproduce and purify WhiB3, this study paves the way for future structure-function experiments.

Psychological effects of reading: the role of nostalgia in re-reading favorite books.

Nostalgia evoked through various experiences (e.g., scents, music) has been shown to enhance emotional well-being and reduce social pain. We propose that reading a familiar book similarly can elicit nostalgia, and provide emotional benefits through narrative transportation beyond that of reading a new book. We tested the relationship between reading new versus familiar books, nostalgia, narrative transportation, and indices of social connectedness. Participants were randomly assigned to re-read a favorite novel, read a new novel of interest, or read a set of newspaper articles. Re-reading elicited greater nostalgia and social connectedness than reading a new novel or newspaper. Narrative transportation and nostalgia fully mediated the effect of reading condition on social connectedness. We discuss implications for our understanding homeostatic nature of nostalgia and mental transportation.

Exploiting Violet-Blue Light to Kill Campylobacter jejuni: Analysis of Global Responses, Modeling of Transcription Factor Activities, and Identification of Protein Targets.

Campylobacter jejuni is a microaerophilic foodborne zoonotic pathogen of worldwide concern as the leading cause of bacterial gastroenteritis. Many strains are increasingly antibiotic resistant and new methods of control are required to reduce food-chain contamination. One possibility is photodynamic inactivation (PDI) using violet-blue (VB) light, to which C. jejuni is highly susceptible. Here, we show that flavin and protoporphyrin IX are major endogenous photosensitizers and that exposure of cells to VB light increases intracellular reactive oxygen species (ROS) to high levels, as indicated by a dichlorodihydrofluorescein reporter. Unusually for an oxygen-respiring bacterium, C. jejuni employs several ROS-sensitive iron-sulfur cluster enzymes in central metabolic pathways; we show that VB light causes rapid inactivation of both pyruvate and 2-oxoglutarate oxidoreductases, thus interrupting the citric acid cycle. Cells exposed to VB light also lose heme from c-type cytochromes, restricting electron transport, likely due to irreversible oxidation of heme-ligating cysteine residues. Evaluation of global gene expression changes by RNAseq and probabilistic modeling showed a two-stage protein damage/oxidative stress response to VB light, driven by specific regulators, including HspR, PerR, Fur, and RacR. Deletion mutant analysis showed that superoxide dismutase and the cytochrome CccA were particularly important for VB light survival and that abolishing repression of chaperones and oxidative stress resistance genes by HcrA, HspR, or PerR increased tolerance to VB light. Our results explain the high innate sensitivity of C. jejuni to VB light and provide new insights that may be helpful in exploiting PDI for novel food-chain interventions to control this pathogen. IMPORTANCE Campylobacteriosis caused by C. jejuni is one of the most widespread zoonotic enteric diseases worldwide and represents an enormous human health and economic burden, compounded by the emergence of antibiotic-resistant strains. New interventions are urgently needed to reduce food-chain contamination. Although UV light is well known to be bactericidal, it is highly mutagenic and problematic for continuous exposure in food production facilities; in contrast, narrow spectrum violet-blue (VB) light is much safer. We confirmed that C. jejuni is highly susceptible to VB light and then identified some of the global regulatory networks involved in responding to photo-oxidative damage. The identification of damaged cellular components underpins efforts to develop commercial applications of VB light-based technologies.

The Di-Iron Protein YtfE Is a Nitric Oxide-Generating Nitrite Reductase Involved in the Management of Nitrosative Stress.

Previously characterized nitrite reductases fall into three classes: siroheme-containing enzymes (NirBD), cytochrome c hemoproteins (NrfA and NirS), and copper-containing enzymes (NirK). We show here that the di-iron protein YtfE represents a physiologically relevant new class of nitrite reductases. Several functions have been previously proposed for YtfE, including donating iron for the repair of iron-sulfur clusters that have been damaged by nitrosative stress, releasing nitric oxide (NO) from nitrosylated iron, and reducing NO to nitrous oxide (N2O). Here, in vivo reporter assays confirmed that Escherichia coli YtfE increased cytoplasmic NO production from nitrite. Spectroscopic and mass spectrometric investigations revealed that the di-iron site of YtfE exists in a mixture of forms, including nitrosylated and nitrite-bound, when isolated from nitrite-supplemented, but not nitrate-supplemented, cultures. Addition of nitrite to di-ferrous YtfE resulted in nitrosylated YtfE and the release of NO. Kinetics of nitrite reduction were dependent on the nature of the reductant; the lowest Km, measured for the di-ferrous form, was ∼90 µM, well within the intracellular nitrite concentration range. The vicinal di-cysteine motif, located in the N-terminal domain of YtfE, was shown to function in the delivery of electrons to the di-iron center. Notably, YtfE exhibited very low NO reductase activity and was only able to act as an iron donor for reconstitution of apo-ferredoxin under conditions that damaged its di-iron center. Thus, YtfE is a high-affinity, low-capacity nitrite reductase that we propose functions to relieve nitrosative stress by acting in combination with the co-regulated NO-consuming enzymes Hmp and Hcp.

The Burkholderia cenocepacia iron starvation σ factor, OrbS, possesses an on-board iron sensor.

Burkholderia cenocepacia is an opportunistic pathogen that causes severe infections of the cystic fibrosis (CF) lung. To acquire iron, B. cenocepacia secretes the Fe(III)-binding compound, ornibactin. Genes for synthesis and utilisation of ornibactin are served by the iron starvation (IS) extracytoplasmic function (ECF) σ factor, OrbS. Transcription of orbS is regulated in response to the prevailing iron concentration by the ferric uptake regulator (Fur), such that orbS expression is repressed under iron-sufficient conditions. Here we show that, in addition to Fur-mediated regulation of orbS, the OrbS protein itself responds to intracellular iron availability. Substitution of cysteine residues in the C-terminal region of OrbS diminished the ability to respond to Fe(II) in vivo. Accordingly, whilst Fe(II) impaired transcription from and recognition of OrbS-dependent promoters in vitro by inhibiting the binding of OrbS to core RNA polymerase (RNAP), the cysteine-substituted OrbS variant was less responsive to Fe(II). Thus, the cysteine residues within the C-terminal region of OrbS contribute to an iron-sensing motif that serves as an on-board 'anti-σ factor' in the presence of Fe(II). A model to account for the presence two regulators (Fur and OrbS) that respond to the same intracellular Fe(II) signal to control ornibactin synthesis and utilisation is discussed.

Multi-omic based production strain improvement (MOBpsi) for bio-manufacturing of toxic chemicals.

Robust systematic approaches for the metabolic engineering of cell factories remain elusive. The available models for predicting phenotypical responses and mechanisms are incomplete, particularly within the context of compound toxicity that can be a significant impediment to achieving high yields of a target product. This study describes a Multi-Omic Based Production Strain Improvement (MOBpsi) strategy that is distinguished by integrated time-resolved systems analyses of fed-batch fermentations. As a case study, MOBpsi was applied to improve the performance of an Escherichia coli cell factory producing the commodity chemical styrene. Styrene can be bio-manufactured from phenylalanine via an engineered pathway comprised of the enzymes phenylalanine ammonia lyase and ferulic acid decarboxylase. The toxicity, hydrophobicity, and volatility of styrene combine to make bio-production challenging. Previous attempts to create styrene tolerant E. coli strains by targeted genetic interventions have met with modest success. Application of MOBpsi identified new potential targets for improving performance, resulting in two host strains (E. coli NST74ΔaaeA and NST74ΔaaeA cpxPo) with increased styrene production. The best performing re-engineered chassis, NST74ΔaaeA cpxPo, produced ∼3 × more styrene and exhibited increased viability in fed-batch fermentations. Thus, this case study demonstrates the utility of MOBpsi as a systematic tool for improving the bio-manufacturing of toxic chemicals.

Perceptions of free will and self-control in a medically relevant treatment recommendation scenario.

Do provider perceptions of patient free will and treatment related self-control influence treatment recommendations and do such perceptions differ due to race? If so, such bias may be a mechanism for racial disparities in medical treatment recommendations. We hypothesized: (1) greater perceived patient free will would indirectly effect treatment recommendations for patients through increased perceived patient treatment related self-control; (2) participants would perceive greater free will for a hypothetical racial ingroup patient than outgroup patient; and (3) such effect would be exacerbated by greater levels of racial identity and racial bias. A 2 (Participant: Black vs. White) x 2 (Target: Black vs. White) x Continuous (Racial Identity/Racial Bias) between-subjects design supported hypothesis 1. Perceived patient free will predicted more rigorous treatment recommendations treatment related self-control. No evidence was found in support of hypotheses 2 and 3. Using a novel experimental design, this work demonstrates the importance of free will and self-control perceptions.

The telomere length landscape of prostate cancer.

Replicative immortality is a hallmark of cancer, and can be achieved through telomere lengthening and maintenance. Although the role of telomere length in cancer has been well studied, its association to genomic features is less well known. Here, we report the telomere lengths of 392 localized prostate cancer tumours and characterize their relationship to genomic, transcriptomic and proteomic features. Shorter tumour telomere lengths are associated with elevated genomic instability, including single-nucleotide variants, indels and structural variants. Genes involved in cell proliferation and signaling are correlated with tumour telomere length at all levels of the central dogma. Telomere length is also associated with multiple clinical features of a tumour. Longer telomere lengths in non-tumour samples are associated with a lower rate of biochemical relapse. In summary, we describe the multi-level integration of telomere length, genomics, transcriptomics and proteomics in localized prostate cancer.

Making America Great Again? National Nostalgia's Effect on Outgroup Perceptions.

Nostalgia is a fond longing for the past that has been shown to increase feelings of meaning, social connectedness, and self-continuity. Although nostalgia for personal memories provides intra- and interpersonal benefits, there may be negative consequences of group-based nostalgia on the perception and acceptance of others. The presented research examined national nostalgia (a form of collective nostalgia), and its effects on group identification and political attitudes in the United States. In a sample of US voters (N = 252), tendencies to feel personal and national nostalgia are associated with markedly different emotional and attitudinal profiles. Higher levels of national nostalgia predicted both positive attitudes toward President Trump and racial prejudice, though there was no evidence of such relationships with personal nostalgia. National nostalgia most strongly predicted positive attitudes toward president Trump among those high in racial prejudice. Furthermore, nostalgia's positive relationship with racial prejudice was partially mediated by perceived outgroup threat. Results from this study will help us better understand how the experience of national nostalgia can influence attitudes and motivate political behavior.

Why Can't I Resist Those "Puppy Dog" (or "Kitty Cat") Eyes? A Study of Owner Attachment and Factors Associated with Pet Obesity.

Attachment theory posits that patterns of interaction derived from the attachment system provide a starting point for understanding how people both receive and provide care. Extending this theory to human-animal interactions provides insights into how human psychology affects pets, such as pet obesity. The goal of this study was to determine how attachment anxiety and avoidance might contribute to pet obesity. We assessed 563 pet owners' attachment-related anxiety and avoidance, as well as additional attachment-related constructs (emotional rejection, evaluation concern, caregiving, and attentiveness to a pet). We also assessed various factors associated with pet obesity, including weight, body condition, daily treats, and daily interaction. The results indicate that dog owners high in attachment anxiety are concerned about how their pet may evaluate them, leading to more caregiving and attentiveness that results in more treats given per day, and a larger body condition (but not weight). In addition, owners high in attachment avoidance may seek to downplay the possibility of the dog negatively evaluating them, thus providing more negligent care. These findings suggest that attachment plays a unique role in shaping the pet-caregiver relationship and influences various elements that contribute to pet obesity, particularly in dogs. As such, the findings may lend a novel perspective to strategies for reducing pet obesity and provide a framework for future research into pet health.

Analysis of Kytococcus sedentarius Strain Isolated from a Dehumidifier Operating in a University Lecture Theatre: Systems for Aerobic Respiration, Resisting Osmotic Stress, and Sensing Nitric Oxide.

A strain of Kytococcus sedentarius was isolated from a dehumidifier operating in a university lecture theatre. Genome analysis and phenotypic characterisation showed that this strain, K. sedentarius MBB13, was a moderately halotolerant aerobe with a branched aerobic electron transport chain and genes that could contribute to erythromycin resistance. The major compatible solute was glycine betaine, with ectoine and proline being deployed at higher osmolarities. Actinobacteria possess multiple WhiB-like (Wbl) regulatory proteins, and K. sedentarius MBB13 has four (WhiB1, WhiB2, WhiB3, and WhiB7). Wbls are iron-sulfur proteins that regulate gene expression through interactions with RNA polymerase sigma factors and/or other regulatory proteins. Bacterial two-hybrid analyses suggested that WhiB1 and WhiB2, but not WhiB3 and WhiB7, interact with the C-terminal domain of the major sigma factor, σA; no interaction was detected between any of the Wbl proteins and the only alternative sigma factors, σB, σH, or σJ. The interaction between σA and WhiB1 or WhiB2 was disrupted in a heterologous system under growth conditions that produce nitric oxide and the iron-sulfur clusters of the isolated WhiB1 and WhiB2 proteins reacted with nitric oxide. Thus, K. sedentarius strain exhibits the major phenotypic characteristics of the type strain and a comprehensive examination of the interactions between its four Wbl proteins and four sigma factors suggested that the Wbl proteins all operate through interaction with σA.

The past as a resource for the bereaved: nostalgia predicts declines in distress.

Nostalgia, a sentimental longing for one's past, can serve as a resource for individuals coping with discomforting experiences. The experience of bereavement poses psychological and physical risks. In a longitudinal study, we examined whether dispositional nostalgia predicted reductions in distress associated with the death of a loved one. Undergraduate students (N = 133) provided information regarding their loss (time elapsed since loss, expectedness) and levels of initial grief, nostalgia, and distress (hyperarousal, intrusion, avoidance) at three time points over a one-month period (Times 2 and 3 occurred one week and one month after the initial session, respectively). Individuals experiencing higher nostalgia reported a decrease in intrusive thoughts across time, whereas those experiencing lower nostalgia reported no change in intrusive thoughts across time. Hyperarousal (physical symptoms, negative feelings) decreased across time among individuals with higher initial grief who experienced greater nostalgia, but increased across time among those with higher initial grief who experienced lesser nostalgia. No changes occurred in avoidance. Nostalgia can palliate bereavement.

Coupling of Peptidoglycan Synthesis to Central Metabolism in Mycobacteria: Post-transcriptional Control of CwlM by Aconitase.

Mycobacterium tuberculosis causes human tuberculosis, and a better understanding of its biology is required to identify vulnerabilities that might be exploited in developing new therapeutics. The iron-sulfur cluster of the essential M. tuberculosis central metabolic enzyme, aconitase (AcnA), disassembles when exposed to oxidative/nitrosative stress or iron chelators. The catalytically inactive apo-AcnA interacts with a sequence resembling an iron-responsive element (IRE) located within the transcript of another essential protein, CwlM, a regulator of peptidoglycan synthesis. A Mycobacterium smegmatis cwlM conditional mutant complemented with M. tuberculosis cwlM with a disrupted IRE is unable to recover from combinations of oxidative, nitrosative, and iron starvation stresses. An equivalent M. tuberculosis cwlM conditional mutant complemented with the cwlM gene lacking a functional IRE exhibits a growth defect in THP-1 macrophages. It appears that AcnA acts to couple peptidoglycan synthesis and central metabolism, and disruption of this coupling potentially leaves mycobacteria vulnerable to attack by macrophages.

Evolving MRSA: High-level ß-lactam resistance in Staphylococcus aureus is associated with RNA Polymerase alterations and fine tuning of gene expression.

Most clinical MRSA (methicillin-resistant S. aureus) isolates exhibit low-level ß-lactam resistance (oxacillin MIC 2-4 µg/ml) due to the acquisition of a novel penicillin binding protein (PBP2A), encoded by mecA. However, strains can evolve high-level resistance (oxacillin MIC ≥256 µg/ml) by an unknown mechanism. Here we have developed a robust system to explore the basis of the evolution of high-level resistance by inserting mecA into the chromosome of the methicillin-sensitive S. aureus SH1000. Low-level mecA-dependent oxacillin resistance was associated with increased expression of anaerobic respiratory and fermentative genes. High-level resistant derivatives had acquired mutations in either rpoB (RNA polymerase subunit ß) or rpoC (RNA polymerase subunit ß') and these mutations were shown to be responsible for the observed resistance phenotype. Analysis of rpoB and rpoC mutants revealed decreased growth rates in the absence of antibiotic, and alterations to, transcription elongation. The rpoB and rpoC mutations resulted in decreased expression to parental levels, of anaerobic respiratory and fermentative genes and specific upregulation of 11 genes including mecA. There was however no direct correlation between resistance and the amount of PBP2A. A mutational analysis of the differentially expressed genes revealed that a member of the S. aureus Type VII secretion system is required for high level resistance. Interestingly, the genomes of two of the high level resistant evolved strains also contained missense mutations in this same locus. Finally, the set of genetically matched strains revealed that high level antibiotic resistance does not incur a significant fitness cost during pathogenesis. Our analysis demonstrates the complex interplay between antibiotic resistance mechanisms and core cell physiology, providing new insight into how such important resistance properties evolve.

A Phase II Single Arm Pilot Study of the CHK1 Inhibitor Prexasertib (LY2606368) in BRCA Wild-Type, Advanced Triple-Negative Breast Cancer.

LESSONS LEARNED: Monotherapy with prexasertib demonstrated modest activity in BRCA wild-type, recurrent triple-negative breast cancer, highlighting the unmet need for combination treatment strategies. Neutropenia, anemia, and thrombocytopenia are common with the use of prexasertib but are manageable with supportive care measures. Prophylactic use of granulocyte colony stimulating factor should be considered to avoid dose reductions or treatment delays. Pharmacodynamic studies showed prexasertib treatment induced DNA damage in peripheral immune cells. BACKGROUND: Cell cycle checkpoint kinase 1 (CHK1) is a major G2/M cell cycle regulator in tumors with p53 dysfunction, such as triple-negative breast cancer (TNBC). We hypothesized the second-generation CHK1 inhibitor, prexasertib, would yield clinical activity in sporadic TNBC. METHODS: This single arm, phase II trial evaluated prexasertib at 105 mg/m2 IV every 2 weeks in patients with metastatic/recurrent TNBC. The primary endpoint was overall response rate (ORR). RESULTS: All nine patients enrolled were germline BRCA wild-type (BRCAwt) and had at least one prior treatment. One partial response (PR) was observed (ORR of 11.1%). Four patients experienced stable disease. The median progression-free survival (PFS) was 86 days (range 17 to 159 days). Grade 3/4 treatment-related adverse events included afebrile neutropenia (n = 8; 88.9%), anemia (n = 3; 33.3%), and thrombocytopenia (n = 1; 11.1%). Pharmacodynamic studies showed prexasertib treatment induced DNA damage in peripheral immune cells and demonstrated a decrease in activated/reinvigorated CD8 T cells; however, the one patient with a PR showed evidence of T-cell recovery. CONCLUSION: Prexasertib monotherapy had modest clinical efficacy in BRCAwt TNBC. Further studies of prexasertib in combination with other agents are needed.