Supplementary Data 1-5 from Reciprocal Relationship between O6-Methylguanine-DNA Methyltransferase P140K Expression Level and Chemoprotection of Hematopoietic Stem Cells

Since the initial COVID-19 outbreak, researchers from a variety of scientific disciplines have sought to understand the factors influencing the evolution of cases and fatalities worldwide. This study proposes a two-stage econometric modeling approach that first identifies the speed of transmission followed by an examination of socioeconomic, demographic, health, epidemiological, climate, pollution, and political factors as potential drivers of COVID-19 spread across waves and counties in the United States. Utilizing daily data on confirmed cases and deaths from 3014 counties across 48 contiguous states and the District of Columbia from March 2020 to March 2022, we find that the trajectory of the pandemic and vaccination uptake patterns were influenced by a complex interplay of demographic characteristics, socioeconomic factors, health determinants, and policy interventions that differential affected viral transmissibility, mortality outcomes, and immunization efficacy across the six distinct pandemic waves. This analysis provides insights into pandemic dynamics across distinct waves and geographic regions.

Maternal depression may have a significant impact on multiple aspects of parenting throughout childhood, leading to negative outcomes for children. However, the impact of depression on parenting perceptions, practices, and involvement may differ among mothers, particularly when protective factors are present. Using data from the Fragile Families and Child Wellbeing Study, we compared three aspects of parenting (perceptions, practices, and involvement) between depressed and non-depressed mothers, and then investigated the influence of interpersonal supports and community resources on these aspects of the parenting, specifically for depressed mothers. Findings indicated that mothers with depression were equally involved with the child at home, but were more likely to hold negative perceptions of parenting. Furthermore, interpersonal supports and community resources had an indirect impact on parenting practices and involvement through perceptions of parenting, highlighting the importance of interventions for mothers with depression that address the appraisal and expectations of parenting stress and competencies.

BACKGROUND:Disasters are increasing in frequency and intensity due to climate change. Youth are the largest and most vulnerable group exposed to disasters. More evidence is needed regarding how youth mental health and health behaviors vary based on disaster exposure, how mental health influences health behaviors, and how diverse groups of youth may be differentially affected. METHODS:Using data from the Youth Risk Behavior Survey and Federal Emergency Management Agency, we examined the impact of Hurricane Sandy (2012) on mental health (sadness, suicidality) and health behaviors (substance use, physical activity, screen time, sexual behaviors) of 240,365 youth. Difference-in-differences regression analyses evaluated pre-disaster (2005-2011) and post-disaster (2013-2019) changes. RESULTS:There were 83,442 (34.7 %) adolescents (51.2 % female, 18.1 % non-Hispanic White) located in 6 districts exposed to Hurricane Sandy and 156,923 (65.3 %) adolescents located in 13 non-exposed districts. Exposure was associated with differences in substance use, physical (in)activity, and risky sexual behaviors, but not mental health outcomes. Mental health did not moderate the association between disaster exposure and health behaviors. Hispanic adolescents and those identifying as Other races/ethnicities were most vulnerable to post-disaster negative health outcomes. LIMITATIONS:Study limitations include questionnaire design, cross-sectional data, sampling, and possible response biases. CONCLUSIONS:Findings provide critical information about youth mental health and health behaviors following disasters, as well as how youth may differ in their responses. Findings provide guidance for early identification and resource allocation for youth at higher risk for post-disaster health issues.

In an extension of the standard spatial autoregressive (SAR) model, Aquaro, Bailey and Pesaran (ABP; 2021, https://doi.org/10.1002/jae.2792) introduced a SAR panel model that allows one to produce heterogeneous point estimates for each spatial unit. Their methodology has been implemented as the Stata routine hetsar (Belotti, 2021, Statistical Sofware Components S458926). As the COVID-19 pandemic has evolved in the U.S. since its first outbreak in February 2020 with following resurgences of multiple widespread and severe waves of the pandemic, the level of interactions between geographic units (for example, states and counties) has differed greatly over time in terms of the prevalence of the disease. Applying ABP’s HETSAR model to 2020 and 2021 COVID-19 data outcomes (confirmed case and death rates) at the state level, we extend our previous spatial econometric analysis (Baum and Henry, 2020, Boston College Working Papers in Economics 1009) on socioeconomic and demographic factors influencing the spatial spread of COVID-19 confirmed case and death rates in the U.S.

In an extension of the standard spatial autoregressive (SAR) model, Aquaro, Bailey and Pesaran (ABP; 2021, https://doi.org/10.1002/jae.2792) introduced a SAR panel model that allows one to produce heterogeneous point estimates for each spatial unit. Their methodology has been implemented as the Stata routine hetsar (Belotti, 2021, Statistical Sofware Components S458926). As the COVID-19 pandemic has evolved in the U.S. since its first outbreak in February 2020 with following resurgences of multiple widespread and severe waves of the pandemic, the level of interactions between geographic units (for example, states and counties) has differed greatly over time in terms of the prevalence of the disease. Applying ABP’s HETSAR model to 2020 and 2021 COVID-19 data outcomes (confirmed case and death rates) at the state level, we extend our previous spatial econometric analysis (Baum and Henry, 2020, Boston College Working Papers in Economics 1009) on socioeconomic and demographic factors influencing the spatial spread of COVID-19 confirmed case and death rates in the U.S.

Purpose: Given the rapid expansion of recreational marijuana legalization (RML) polices, it is essential to assess whether such policies are associated with shifts in the use of marijuana and other substances, particularly for adolescents, who are uniquely susceptible to negative repercussions of marijuana use. This analysis seeks to provide greater generalizability, specificity, and methodological rigor than limited prior evidence. Methods: Youth Risk Behavior Survey data from 47 states from 1999 to 2017 assessed marijuana, alcohol, cigarette, and e-cigarette use among adolescents (14-18 & thorn; years; N = 1,077,938). Associations between RML and adolescent past-month substance use were analyzed using quasi experimental difference-in-differences zero-inflated negative binomial models. Results: Controlling for other state substance policies, year and state fixed effects, and adolescent demographic characteristics, models found that RML was not associated with a significant shift in the likelihood of marijuana use but predicted a small significant decline in the level of marijuana use among users (incidence rate ratio = .844, 95% confidence interval [.720-.989]) and a small increase in the likelihood of any e-cigarette use (odds ratio of zero use = .647, 95% confidence interval [.515-.812]). Patterns did not vary over adolescent age or sex, with minimal differences across racial/ethnic groups. Conclusions: Results suggest minimal short-term effects of RML on adolescent substance use, with small declines in marijuana use and increase in the likelihood of any e-cigarette use. Given the delayed rollout of commercial marijuana sales in RML states and rapid expansion of RML policies, ongoing assessment of the consequences for adolescent substance use and related health and behavioral repercussions is essential. (c) 2020 Society for Adolescent Health and Medicine. All rights reserved.

Retroviral insertional mutagenesis (RIM) is both a relevant risk in gene therapy and a powerful tool for identifying genes that enhance the competitiveness of repopulating hematopoietic stem and progenitor cells (HSPCs). However, focusing only on the gene closest to the retroviral vector insertion site (RVIS) may underestimate the effects of RIM, as dysregulation of distal and/or multiple genes by a single insertion event was reported in several studies. As a proof of concept, we examined the common insertion site (CIS) Bcl-x(L), which revealed seven genes located within +/- 150 kb from the RVIS for our study. We confirmed that Bcl-x(L) enhanced the competitiveness of HSPCs, whereas the Bcl-x(L) neighbor Id1 hindered HSPC long-term repopulation. This negative influence of Id1 could be counteracted by co-expressing Bcl-x(L). Interestingly, >90% of early reconstituted myeloid cells were found to originate from transduced HSPCs upon simultaneous overexpression of Bcl-x(L) and Id1, which implies that Bcl-x(L) and Id1 can collaborate to rapidly replenish the myeloid compartment under stress conditions. To directly compare the competitiveness of HSPCs conveyed by multiple transgenes, we developed a multiple competitor competitive repopulation (MCCR) assay to simultaneously screen effects on HSPC repopulating capacity in a single mouse. The MCCR assay revealed that multiple genes within a CIS can have positive or negative impact on hematopoiesis. Furthermore, these data highlight the importance of studying multiple genes located within the proximity of an insertion site to understand complex biological effects, especially as the number of gene therapy patients increases.

We present the new Stata command radf to compute several tests for explosive behaviour in time series. The command implements the right-tail augmented Dickey and Fuller (1979) (ADF) unit root test, and its further developments based on supremum statistics derived from ADF-type regressions estimated using rolling windows, recursive windows (Phillips, Wu and Yu 2011), and recursive flexible windows (Phillips, Shi and Yu 2015). The command allows for the number of lags of the dependent variable in the test regression to be either specified by the user or endogenously determined using a data-dependent procedure. The use of the command is illustrated with an empirical example.

As the COVID-19 pandemic has progressed in the U.S., hotspots have been shifting geographically over time to suburban and rural counties showing a high prevalence of the disease. We analyze daily U.S. county-level variations in COVID-19 confirmed case counts to evaluate the spatial dependence between neighboring counties. We find strong evidence of county-level socioeconomic factors influencing the spatial spread. We show the potential of combining spatial econometric techniques and socioeconomic factors in assessing the spatial effects of COVID-19 among neighboring counties.

Objective: Although maternal depression is a risk factor for poor child health and well-being, health outcomes may vary, and children may still do well even in the context of risk. Thus, the aims of this study were to explore heterogeneity in outcomes of child physical health and psychosocial well-being and examine differences based on experiences of maternal depression. Method: Data were from 3,211 child–mother dyads from the Fragile Families and Child Wellbeing study. We used latent profile analyses to generate multidimensional profiles of child health and well-being and used multinomial logistic regression analyses to explore differences in profile membership based on the child’s experiences of maternal depression. Results: Analyses revealed five distinct profiles of varying child health and well-being, contrasting the traditional dichotomy of well versus unwell and highlighting the potential for both positive and negative outcomes to occur simultaneously. Although depression was associated with an increased risk of poor health and well-being, the majority of children experiencing maternal depression also experienced some indicators of positive health and well-being. Conclusion: Findings support the need for ongoing depression screening in mothers throughout childhood and emphasize the importance of using a strengths-based approach when working with high-risk populations.

STEM-focused industries are critical to the innovation-driven economy. As many firms are running short of STEM workers, international immigrants are increasingly recognized as a potential for high-tech job recruitment. This paper studies STEM occupations in Sweden 2011–2015 and tests hypotheses on new recruitment and the economic impact of foreign STEM workers. The empirical analysis shows that the probability that a new employee is a STEM immigrant increases with the share of STEM immigrants already employed, while the marginal effect on average firm wages is positively associated with the share of immigrant STEM workers. We also document heterogeneity in the results, suggesting that European migrants are more attractive for new recruitment, but non-EU migrants have the largest impact on wage determination.

1Institute for Tumor Biology and Experimental Therapy, Georg-Speyer-Haus, Frankfurt, Germany 2Division of Hematology/Oncology, Boston Children’s Hospital, Boston, Massachusetts, USA. 3Harvard Medical School, Boston, Massachusetts, USA 4Hannover Medical School, Institute of Experimental Hematology, Hannover, Germany 5Centre for Immunodeficiency, UCL Institute of Child Health, and Great Ormond Street Hospital for Children NHS Trust London, UK 6Genethon, Evry, France 7LOEWE-Research Group, Paul-Ehrlich-Institute, Langen, Germany 8Hannover Medical School, Cluster of Excellence REBIRTH, Hannover, Germany

The recently discovered CRISPR/Cas9 system is widely used in basic research and is a useful tool for disease modeling and gene editing therapies. However, long-term expression of DNA-modifying enzymes can be associated with cytotoxicity and is particularly unwanted in clinical gene editing strategies. Because current transient expression methods may still suffer from cytotoxicity and/or low efficiency, we developed non-integrating retrovirus-based CRISPR/Cas9 all-in-one particles for targeted gene knockout. By redirecting the gammaretroviral packaging machinery, we transiently delivered Streptococcus pyogenes Cas9 (SpCas9) mRNA and single-guide RNA transcripts into various (including primary) cell types. Spatiotemporal co-delivery of CRISPR/Cas9 components resulted in efficient disruption of a surrogate reporter gene, as well as functional knockout of endogenous human genes CXCR4 and TP53. Although acting in a hit-and-run fashion, knockout efficiencies of our transient particles corresponded to 52%-80% of those obtained from constitutively active integrating vectors. Stable SpCas9 overexpression at high doses in murine NIH3T3 cells caused a substantial G0/G1 arrest accompanied by reduced cell growth and metabolic activity, which was prevented by transient SpCas9 transfer. In summary, the non-integrating retrovirus-based vector particles introduced here allow efficient and dose-controlled delivery of CRISPR/Cas9 components into target cells.

The variety of gene therapy vectors for a multitude of different diseases has increased tremendously over the years. However, a number of patients that underwent gene therapy in different trials developed hematological malignancy caused by integration of the provirus in the vicinity of proto-oncogenes. These severe adverse advents prompted intense research efforts towards safer gene therapy, leading to the removal of the long terminal repeat enhancer elements and the use of internal promoters in retroviral vectors. Still, a bottleneck of transition from basic research to clinical application is the test for safety of integrating retro- and lentiviral vectors.

We model the time series of credit default swap (CDS) spreads on sovereign debt in the Eurozone, allowing for stochastic volatility and examining the effects of country-specific and systemic shocks. A weekly volatility series is produced from daily quotations on 11 Eurozone countries: CDS for 2009–2010. Using Stata's gmm command, we construct a highly nonlinear model of the evolution of realized volatility when subjected to both idiosyncratic and systemic shocks. Evaluation of the quality of the fit for the 24 moment conditions is produced by a Mata auxiliary routine. This model captures many of the features of these financial markets during a turbulent period in the recent history of the single currency. We find that systemic volatility shocks increase returns on virtuous borrowers' CDS while reducing returns for the most troubled countries' obligations.

Molecular hallmarks of T-ALL are the aberrant activation of NOTCH signaling and high activity of the PI3K-AKT-mTOR pathway. Upregulated mTOR and NOTCH have been linked to the resistance of T-ALL to chemotherapy as well as to high frequencies of leukemia-initiating cells. Hence, the mTOR and the NOTCH pathways are promising therapeutic targets in T-ALL. However, clinical success of the mTOR inhibitor Rapamycin in acute leukemia has been disappointing. Similar results have been observed in mouse models of T-ALL treated with Notch inhibitors. To investigate the impact of mTOR and Notch inhibition in a genetically complex T-ALL, we developed an aggressive murine T-ALL model, driven by tyrosine kinase signaling, loss of Pten, Cux1-haploinsufficiency and constitutive Notch signaling. In vitro, T-ALL blasts were highly sensitive to inhibition of AKT, mTOR and Notch signaling. We transplanted the leukemias into secondary recipients and initiated treatment with Rapamycin after the onset of leukemia. Rapamycin significantly prolonged survival of the animals (placebo: 27 days, Rapamycin 49 days, p<0.001). Eventually, all Rapamycin treated animals succumbed to the T-ALL that extensively infiltrated the bone marrow and solid organs despite continuous drug administration. When Rapamycin-resistant blasts were explanted and cultured in petri dishes they again became susceptible to Rapamycin, demonstrating a context-dependent resistance rather than outgrowth of intrinsically resistant clones. Gene set enrichment analysis revealed that Rapamycin-resistant T-ALL in vivo upregulated genetic networks associated with cell-cell interactions. Stromal cell support from OP9-cells as well as from mesenchymal stem cells recapitulated the in vivo effect and induced resistance to mTOR and Notch-inhibition in T-ALL blasts. Coating the tissue culture wells with Collagen, Fibronectin, Retronectin or Matrigel, did not elicit resistance. By using trans-well assays we show that the stroma-induced resistance was dependent on direct cell-cell interactions. Immunoblots and PhosFlow probing the mTORC1/C2 and Notch pathway demonstrated an identical drug effect on their intracellular targets in resistant T-ALL blasts cultured on stroma cells and susceptible cells in suspension. Since the number of molecules potentially involved in cell-to-cell contacts is vast, we focused on central nodes that organize this process in order to find a potentially druggable target that is critically involved in stroma-induced resistance. Transcriptome profiling pointed towards upregulation of Rac-associated pathways. We determined the activation of Rac1 by PAK-pull down assays in T-ALL blasts grown in suspension or on stromal cells. We observed an increase (FC=1.96 ± 0.58, p=0.04) in activated Rac1 in the T-ALL blasts in contact with a stromal layer. To determine whether Rac activation plays a role in stroma-induced resistance, we devised a strategy to abrogate Rac signaling in T-ALL blasts, but not in the stromal cells, since inhibition of Rac in stromal cells by the Rac-inhibitor NSC23766 led to the their detachment. Furthermore, Rac1,2 and 3 can be functionally redundant, making knock down experiments using shRNAs challenging. The Clostridium difficile serotype F strain 1470 produces toxin B isoform (TcdBF), that selectively glucosylates and inactivates Rac(1,2,3). We pretreated T-ALL blasts with TcdBF and observed a dose-dependent functional inhibition of Rac GTPases monitored by dephosphorylation of the Rac effector kinase pS144/141-PAK-1/2. T-ALL blasts were then incubated for 5 hours with increasing toxin doses, washed 3 times and incubated in toxin-free medium. Eighteen hours after the end of the exposure to the toxins, Rac was still inhibited. Strikingly, in the TcdBF-pretreated T-ALL, the stroma-induced resistance effect was abrogated and clusters of apoptotic cells were clearly visible (>2 fold reduction of the input, p=0.002). In contrast, the carrier-treated T-ALL exhibited resistance to the inhibitors on stroma (>10 fold expansion of the input, p<0.0003). Altogether, we identify the Rac-GTPases as a nexus of stroma-induced drug resistance and show that inhibition of Rac and mTOR is synthetically lethal to T-ALL blasts T-ALL blasts that are in contact with stromal cells, paving the way to augment the effectiveness of small molecule inhibitors in acute leukemia.

Successful application of gene therapy strategies may require stringently regulated transgene expression. Along this line, we describe a doxycycline (Dox)-inducible ‘all-in-one’ lentiviral vector design using the pTET-T11 (TII) minimal-promoter and a reverse transactivator protein (rtTA2S-M2) driven by the phosphoglycerate kinase promoter allowing for tight regulation of transgene expression (Lv.TII vectors). Vector design was evaluated in human hematopoietic cells in the context of cytidine deaminase (hCDD)-based myeloprotective gene therapy. Upon Dox administration, a rapid (16–24 h) and dose-dependent (>0.04 μg ml −1 Dox) onset of transgene expression was detected in Lv.TII.CDD gene-modified K562 cells as well as in primary human CD34 + hematopoietic cells. Importantly, in both cell models low background transgene expression was observed in the absence of Dox. Functionality of Dox-inducible hCDD expression was demonstrated by >10-fold increase in cytosine arabinoside (1-β- d -arabinofuranosylcytosine, Ara-C) resistance of Lv.TII.CDD-transduced K562 cells. In addition, Lv.TII.CDD-transduced CD34 + -derived myeloid cells were protected from up to 300 n m Ara-C (control affected from 50 n m onwards). These data clearly demonstrate the suitability of our self-inactivating lentiviral vector to induce robust, tightly regulated transgene expression in human hematopoietic cells with minimal background activity and highlight the potential of our construct in myeloprotective gene therapy strategies.