High-Dose Acetaminophen with Concurrent CYP2E1 Inhibition Has Profound Anticancer Activity without Liver Toxicity.

Acetaminophen (AAP) is metabolized by a variety of pathways such as sulfation, glucuronidation, and fatty acid amide hydrolase-mediated conversion to the active analgesic metabolite AM404. CYP2E1-mediated metabolism to the hepatotoxic reactive metabolite NAPQI (N-acetyl-p-benzoquinone imine) is a minor metabolic pathway that has not been linked to AAP therapeutic benefits yet clearly leads to AAP liver toxicity. N-acetylcysteine (NAC) (an antioxidant) and fomepizole (a CYP2E1 inhibitor) are clinically used for the treatment of AAP toxicity. Mice treated with AAP in combination with fomepizole (plus or minus NAC) were assessed for liver toxicity by histology and serum chemistry. The anticancer activity of AAP with NAC and fomepizole rescue was assessed in vitro and in vivo. Fomepizole with or without NAC completely prevented AAP-induced liver toxicity. In vivo, high-dose AAP with NAC/fomepizole rescue had profound antitumor activity against commonly used 4T1 breast tumor and lewis lung carcinoma lung tumor models, and no liver toxicity was detected. The antitumor efficacy was reduced in immune-compromised NOD-scid IL2Rgammanull mice, suggesting an immune-mediated mechanism of action. In conclusion, using fomepizole-based rescue, we were able to treat mice with 100-fold higher than standard dosing of AAP (650 mg/kg) without any detected liver toxicity and substantial antitumor activity. SIGNIFICANCE STATEMENT: High-dose acetaminophen can be given concurrently with CYP2E1 inhibition to allow for safe dose escalation to levels needed for anticancer activity without detected evidence of toxicity.

Medical Management of Endometriosis in Adolescent and Young Adult Women: A Review of 91 Cases of Biopsy-Confirmed Endometriosis.

OBJECTIVES: To characterise contemporary trends in the hormonal management of endometriosis in adolescent and young adult patients with biopsy-proven endometriosis. METHODS: Retrospective chart review of women aged 14-25 years who underwent laparoscopy for pelvic pain with biopsy-proven endometriosis between January 2011 and September 2020 at an academic tertiary hospital system. The final sample included 91 patients with biopsy-confirmed endometriosis. RESULTS: Combined oral contraceptives (COCs) were the most common initial treatment (64% of patients). Progestin-only formulations (low- and high-dose norethindrone acetate) were offered to younger patients (age 15.9 ± 2.7 years) than those offered COCs (19.9 ± 3.3 years) and levonorgestrel intrauterine devices (LNG-IUDs) (21.9 ± 1.7 years). Current treatments varied widely and included COCs (32%), LNG-IUDs (18%), oral progestins (low- and high-dose norethindrone, medroxyprogesterone) (14%), elagolix (9%), and leuprolide (8%). Oral adjuncts to LNG-IUD were common: usually low- or high-dose norethindrone (37% of patients with an LNG-IUD), but also included progesterone, COCs, and elagolix. CONCLUSIONS: Oral progestins, LNG-IUDs, and COCs were the mainstay of initial treatment. Subsequent treatments varied widely and included COCs, LNG-IUDs, oral progestins, elagolix, leuprolide, and combinations of these agents. We observed that most young women switched between therapies, suggesting that a personalised approach is often used to determine treatment plans among the wide range of options currently available. This study helps define the spectrum of treatment regimens for endometriosis in adolescent females.

AMH independently predicts aneuploidy but not live birth per transfer in IVF PGT-A cycles.

BACKGROUND: While anti-Müllerian hormone (AMH) predicts quantitative IVF outcomes such as oocyte yield, it is not certain whether AMH predicts markers of oocyte quality such as aneuploidy. METHODS: Retrospective case-control analysis of the SART-CORS database, 2014-2016, to determine whether anti-Müllerian hormone (AMH) predicts aneuploidy and live birth in IVF cycles utilizing preimplantation genetic testing for aneuploidy (PGT-A). RESULTS: Of 51,273 cycles utilizing PGT-A for all embryos, 10,878 cycles were included in the final analysis; of these, 2,100 cycles resulted in canceled transfer due to lack of normal embryos and 8,778 cycles resulted in primary FET. AMH levels of cycles with ≥ 1 euploid embryo were greater than those of cycles with no normal embryos, stratifying by number of embryos biopsied (1-2, 3-4, 5-6, and ≥ 7), P < 0.017 for each stratum. Adjusting for age and number of embryos biopsied, AMH was a significant independent predictor of ≥ 1 euploid embryo for all age groups: < 35 yrs (aOR 1.074; 95%CI 1.005-1.163), 35-37 years (aOR 1.085; 95%CI 1.018-1.165) and ≥ 38 years (aOR 1.055; 95%CI 1.020-1.093). In comparative model analysis, AMH was superior to age as a predictor of  ≥ 1 euploid embryo for age groups < 35 years and 35-37 years, but not  ≥ 38 years. Across all cycles, age (aOR 0.945, 95% CI 0.935-0.956) and number of embryos (aOR 1.144, 95%CI 1.127-1.162) were associated with live birth per transfer, but AMH was not (aOR 0.995, 95%CI 0.983-1.008). In the subset of cycles resulting in ≥ 1 euploid embryo for transfer, neither age nor AMH were associated with live birth. CONCLUSIONS: Adjusting for age and number of embryos biopsied, AMH independently predicted likelihood of obtaining ≥ 1 euploid embryo for transfer in IVF PGT-A cycles. However, neither age nor AMH were predictive of live birth once a euploid embryo was identified by PGT-A for transfer. This analysis suggests a predictive role of AMH for oocyte quality (aneuploidy risk), but not live birth per transfer once a euploid embryo is identified following PGT-A.

Altered estradiol-dependent cellular Ca2+ homeostasis and endoplasmic reticulum stress response in Premenstrual Dysphoric Disorder.

Premenstrual Dysphoric Disorder (PMDD) is characterized by debilitating mood symptoms in the luteal phase of the menstrual cycle. Prior studies of affected women have implicated a differential response to ovarian steroids. However, the molecular basis of these patients' differential response to hormone remains poorly understood. We performed transcriptomic analyses of lymphoblastoid cell lines (LCLs) derived from women with PMDD and asymptomatic controls cultured under untreated (steroid-free), estradiol-treated (E2), and progesterone-treated (P4) conditions. Weighted gene correlation network analysis (WGCNA) of transcriptomes identified four gene modules with significant diagnosis x hormone interactions, including one enriched for neuronal functions. Next, in a gene-level analysis comparing transcriptional response to hormone across diagnoses, a generalized linear model identified 1522 genes differentially responsive to E2 (E2-DRGs). Among the top 10 E2-DRGs was a physically interacting network (NUCB1, DST, GCC2, GOLGB1) involved in endoplasmic reticulum (ER)-Golgi function. qRT-PCR validation reproduced a diagnosis x E2 interaction (F(1,24)=7.01, p = 0.014) for NUCB1, a regulator of cellular Ca2+ and ER stress. Finally, we used a thapsigargin (Tg) challenge assay to test whether E2 induces differences in Ca2+ homeostasis and ER stress response in PMDD. PMDD LCLs had a 1.36-fold decrease in Tg-induced XBP1 splicing response compared to controls, and a 1.62-fold decreased response (p = 0.005), with a diagnosis x treatment interaction (F(3,33)=3.51, p = 0.026) in the E2-exposed condition. Altered hormone-dependent in cellular Ca2+ dynamics and ER stress may contribute to the pathophysiology of PMDD.

Cancer Cell-Intrinsic Expression of MHC Class II Regulates the Immune Microenvironment and Response to Anti-PD-1 Therapy in Lung Adenocarcinoma.

MHC class II (MHCII) expression is usually restricted to APC but can be expressed by cancer cells. We examined the effect of cancer cell-specific MHCII (csMHCII) expression in lung adenocarcinoma on T cell recruitment to tumors and response to anti-PD-1 therapy using two orthotopic immunocompetent murine models of non-small cell lung cancer: CMT167 (CMT) and Lewis lung carcinoma (LLC). We previously showed that CMT167 tumors are eradicated by anti-PD1 therapy, whereas LLC tumors are resistant. RNA sequencing analysis of cancer cells recovered from tumors revealed that csMHCII correlated with response to anti-PD1 therapy, with immunotherapy-sensitive CMT167 cells being csMHCII positive, whereas resistant LLC cells were csMHCII negative. To test the functional effects of csMHCII, MHCII expression was altered on the cancer cells through loss- and gain-of-function of CIITA, a master regulator of the MHCII pathway. Loss of CIITA in CMT167 decreased csMHCII and converted tumors from anti-PD-1 sensitive to anti-PD-1 resistant. This was associated with lower levels of Th1 cytokines, decreased T cell infiltration, increased B cell numbers, and decreased macrophage recruitment. Conversely, overexpression of CIITA in LLC cells resulted in csMHCII in vitro and in vivo. Enforced expression of CIITA increased T cell infiltration and sensitized tumors to anti-PD-1 therapy. csMHCII expression was also examined in a subset of surgically resected human lung adenocarcinomas by multispectral imaging, which provided a survival benefit and positively correlated with T cell infiltration. These studies demonstrate a functional role for csMHCII in regulating T cell infiltration and sensitivity to anti-PD-1.

Epigenetic intersection of BDNF Val66Met genotype with premenstrual dysphoric disorder transcriptome in a cross-species model of estradiol add-back.

Premenstrual dysphoric disorder (PMDD) affects over 5% of women, with symptoms similar to anxiety and major depression, and is associated with differential sensitivity to circulating ovarian hormones. Little is known about the genetic and epigenetic factors that increase the risk to develop PMDD. We report that 17ß-estradiol (E2) affects the behavior and the epigenome in a mouse model carrying a single-nucleotide polymorphism of the brain-derived neurotrophic factor gene (BDNF Val66Met), in a way that recapitulates the hallmarks of PMDD. Ovariectomized mice heterozygous for the BDNF Met allele (Het-Met) and their matched wild-type (WT) mice were administered estradiol or vehicle in drinking water for 6 weeks. Using the open field and the splash test, we show that E2 add-back induces anxiety-like and depression-like behavior in Het-Met mice, but not in WT mice. RNA-seq of the ventral hippocampus (vHpc) highlights that E2-dependent gene expression is markedly different between WT mice and Het-Met mice. Through a comparative whole-genome RNA-seq analysis between mouse vHpc and lymphoblastoid cell line cultures from control women and women with PMDD, we discovered common epigenetic biomarkers that transcend species and cell types. Those genes include epigenetic modifiers of the ESC/E(Z) complex, an effector of response to ovarian steroids. Although the BDNF Met genotype intersects the behavioral and transcriptional traits of women with PMDD, we suggest that these similarities speak to the epigenetic factors by which ovarian steroids produce negative behavioral effects.

Comparing genome versus proteome-based identification of clinical bacterial isolates.

Whole-genome sequencing (WGS) is gaining importance in the analysis of bacterial cultures derived from patients with infectious diseases. Existing computational tools for WGS-based identification have, however, been evaluated on previously defined data relying thereby unwarily on the available taxonomic information.Here, we newly sequenced 846 clinical gram-negative bacterial isolates representing multiple distinct genera and compared the performance of five tools (CLARK, Kaiju, Kraken, DIAMOND/MEGAN and TUIT). To establish a faithful 'gold standard', the expert-driven taxonomy was compared with identifications based on matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) analysis. Additionally, the tools were also evaluated using a data set of 200 Staphylococcus aureus isolates.CLARK and Kraken (with k =31) performed best with 626 (100%) and 193 (99.5%) correct species classifications for the gram-negative and S. aureus isolates, respectively. Moreover, CLARK and Kraken demonstrated highest mean F-measure values (85.5/87.9% and 94.4/94.7% for the two data sets, respectively) in comparison with DIAMOND/MEGAN (71 and 85.3%), Kaiju (41.8 and 18.9%) and TUIT (34.5 and 86.5%). Finally, CLARK, Kaiju and Kraken outperformed the other tools by a factor of 30 to 170 fold in terms of runtime.We conclude that the application of nucleotide-based tools using k-mers-e.g. CLARK or Kraken-allows for accurate and fast taxonomic characterization of bacterial isolates from WGS data. Hence, our results suggest WGS-based genotyping to be a promising alternative to the MS-based biotyping in clinical settings. Moreover, we suggest that complementary information should be used for the evaluation of taxonomic classification tools, as public databases may suffer from suboptimal annotations.

Development of syngeneic murine cell lines for use in immunocompetent orthotopic lung cancer models.

BACKGROUND: Immunocompetent animal models are required to study tumor-host interactions, immunotherapy, and immunotherapeutic combinations, however the currently available immunocompetent lung cancer models have substantial limitations. While orthotopic models potentially help fill this gap, the utility of these models has been limited by the very small number of murine lung cancer cell lines capable of forming orthotopic tumors in immunocompetent C57BL/6 hosts. METHODS: Primary lung tumors with specific genetic alterations were created in C57BL/6 background mice. These tumors were then passaged through other animals to increase tumorigenicity and select for the ability to grow in a non-self animal. Once tumors demonstrated growth in a non-self host, cell lines were established. Successful cell lines were evaluated for the ability to produce orthotopic lung tumors in immunocompetent hosts. RESULTS: We produced six murine lung cancer lines capable of orthotopic lung tumor formation in immunocompetent C57BL/6 animals. These lines demonstrate the expected genetic alterations based on their primary tumor genetics. CONCLUSIONS: These novel cell lines will be useful for evaluating tumor-host interactions, the impact of specific oncogenic alterations on the tumor microenvironment, and immunotherapeutic approaches. This method of generating murine lines capable of orthotopic growth can likely be applied to other tumors and will broaden the applicability of pre-clinical testing of immunotherapeutic treatment regimens.

Rapid and tunable post-translational coupling of genetic circuits.

One promise of synthetic biology is the creation of genetic circuitry that enables the execution of logical programming in living cells. Such 'wet programming' is positioned to transform a wide and diverse swathe of biotechnology ranging from therapeutics and diagnostics to water treatment strategies. Although progress in the development of a library of genetic modules continues apace, a major challenge for their integration into larger circuits is the generation of sufficiently fast and precise communication between modules. An attractive approach is to integrate engineered circuits with host processes that facilitate robust cellular signalling. In this context, recent studies have demonstrated that bacterial protein degradation can trigger a precise response to stress by overloading a limited supply of intracellular proteases. Here we use protease competition to engineer rapid and tunable coupling of genetic circuits across multiple spatial and temporal scales. We characterize coupling delay times that are more than an order of magnitude faster than standard transcription-factor-based coupling methods (less than 1 min compared with ∼20-40 min) and demonstrate tunability through manipulation of the linker between the protein and its degradation tag. We use this mechanism as a platform to couple genetic clocks at the intracellular and colony level, then synchronize the multi-colony dynamics to reduce variability in both clocks. We show how the coupled clock network can be used to encode independent environmental inputs into a single time series output, thus enabling frequency multiplexing (information transmitted on a common channel by distinct frequencies) in a genetic circuit context. Our results establish a general framework for the rapid and tunable coupling of genetic circuits through the use of native 'queueing' processes such as competitive protein degradation.

Transdermal estradiol for postpartum depression: results from a pilot randomized, double-blind, placebo-controlled study.

Postpartum depression (PPD) is a common complication following delivery, though evidence-based treatment options are limited. This study explores the feasibility and efficacy of outpatient PPD treatment with transdermal estradiol (TE). In a pilot, double-blind, placebo-controlled trial, women with PPD were randomized to receive transdermal 17ß-estradiol (100 mcg/day) or placebo patch. Over 6 weeks, women completed weekly ratings on the Beck Depression Inventory (BDI), Edinburgh Postnatal Depression Scale (EPDS), and Hamilton Depression Scale (HAM-D). Primary outcome measures were treatment response (> 50% decrease from baseline BDI) and remission (BDI < 10) at 6 weeks, and secondary outcome measures included severity on all scales at weeks 3 and 6. Of 12 recruited women, 6 received TE and 6 received placebo. By week 6, 5 women receiving TE responded to treatment and 4 showed symptom remission, compared to 2 responders and 1 remitter in the placebo group. This difference was not significant (p = 0.24). In a mixed-model of BDI ratings, TE was associated with a 9.2 point decrease at 3 weeks (95%CI - 19.5 to + 1.0, p = 0.074) and a 10.5 point decrease at 6 weeks (95%CI - 21.0-0.0, p = 0.049) compared to placebo, though these differences did not survive multiple comparisons correction. Analogous effects were found for HAM-D but not EPDS scores. Interestingly, no significant difference in plasma estradiol levels existed between groups. We were unable to demonstrate a significant therapeutic benefit of TE compared with placebo in PPD. Although limited by under-recruitment and loss to follow-up, our results suggest TE is a feasible option for outpatient PPD management, with preliminary evidence (based on secondary outcomes) for efficacy. Therapeutic effects may be seen as early as 3 weeks and may not directly depend on peripheral measures of estradiol.

Case report: extreme coronary calcifications and hypomagnesemia in a patient with a 17q12 deletion involving HNF1B.

BACKGROUND: 17q12 deletion syndrome encompasses a broad constellation of clinical phenotypes, including renal magnesium wasting, maturity-onset diabetes of the young (MODY), renal cysts, genitourinary malformations, and neuropsychiatric illness. Manifestations outside of the renal, endocrine, and nervous systems have not been well described. CASE PRESENTATION: We report a 62-year-old male referred to the Undiagnosed Diseases Program (UDP) at the National Institutes of Health (NIH) who presented with persistent hypermagnesiuric hypomagnesemia and was found to have a 17q12 deletion. The patient exhibited several known manifestations of the syndrome, including severe hypomagnesemia, renal cysts, diabetes and cognitive deficits. Coronary CT revealed extensive coronary calcifications, with a coronary artery calcification score of 12,427. Vascular calcifications have not been previously reported in this condition. We describe several physiologic mechanisms and a review of literature to support the expansion of the 17q12 deletion syndrome to include vascular calcification. CONCLUSION: Extensive coronary and vascular calcifications may be an extension of the 17q12 deletion phenotype, particularly if hypomagnesemia and hyperparathyroidism are prevalent. In patients with 17q12 deletions involving HNF1B, hyperparathyroidism and hypomagnesemia may contribute to significant cardiovascular risk.

Expression Profiling of Macrophages Reveals Multiple Populations with Distinct Biological Roles in an Immunocompetent Orthotopic Model of Lung Cancer.

Macrophages represent an important component of the tumor microenvironment and play a complex role in cancer progression. These cells are characterized by a high degree of plasticity, and they alter their phenotype in response to local environmental cues. Whereas the M1/M2 classification of macrophages has been widely used, the complexity of macrophage phenotypes has not been well studied, particularly in lung cancer. In this study we employed an orthotopic immunocompetent model of lung adenocarcinoma in which murine lung cancer cells are directly implanted into the left lobe of syngeneic mice. Using multimarker flow cytometry, we defined and recovered several distinct populations of monocytes/macrophages from tumors at different stages of progression. We used RNA-seq transcriptional profiling to define distinct features of each population and determine how they change during tumor progression. We defined an alveolar resident macrophage population that does not change in number and expresses multiple genes related to lipid metabolism and lipid signaling. We also defined a population of tumor-associated macrophages that increase dramatically with tumor and selectively expresses a panel of chemokine genes. A third population, which resembles tumor-associated monocytes, expresses a large number of genes involved in matrix remodeling. By correlating transcriptional profiles with clinically prognostic genes, we show that specific monocyte/macrophage populations are enriched in genes that predict outcomes in lung adenocarcinoma, implicating these subpopulations as critical determinants of patient survival. Our data underscore the complexity of monocytes/macrophages in the tumor microenvironment, and they suggest that distinct populations play specific roles in tumor progression.

Deletion of 5-Lipoxygenase in the Tumor Microenvironment Promotes Lung Cancer Progression and Metastasis through Regulating T Cell Recruitment.

Eicosanoids, including PGs, produced by cyclooxygenases (COX), and leukotrienes, produced by 5-lipoxygenase (5-LO) have been implicated in cancer progression. These molecules are produced by both cancer cells and the tumor microenvironment (TME). We previously reported that both COX and 5-LO metabolites increase during progression in an orthotopic immunocompetent model of lung cancer. Although PGs in the TME have been well studied, less is known regarding 5-LO products produced by the TME. We examined the role of 5-LO in the TME using a model in which Lewis lung carcinoma cells are directly implanted into the lungs of syngeneic WT mice or mice globally deficient in 5-LO (5-LO-KO). Unexpectedly, primary tumor volume and liver metastases were increased in 5-LO-KO mice. This was associated with an ablation of leukotriene (LT) production, consistent with production mainly mediated by the microenvironment. Increased tumor progression was partially reproduced in global LTC4 synthase KO or mice transplanted with LTA4 hydrolase-deficient bone marrow. Tumor-bearing lungs of 5-LO-KO had decreased numbers of CD4 and CD8 T cells compared with WT controls, as well as fewer dendritic cells. This was associated with lower levels of CCL20 and CXL9, which have been implicated in dendritic and T cell recruitment. Depletion of CD8 cells increased tumor growth and eliminated the differences between WT and 5-LO mice. These data reveal an antitumorigenic role for 5-LO products in the microenvironment during lung cancer progression through regulation of T cells and suggest that caution should be used in targeting this pathway in lung cancer.

Systems biology definition of the core proteome of metabolism and expression is consistent with high-throughput data.

Finding the minimal set of gene functions needed to sustain life is of both fundamental and practical importance. Minimal gene lists have been proposed by using comparative genomics-based core proteome definitions. A definition of a core proteome that is supported by empirical data, is understood at the systems-level, and provides a basis for computing essential cell functions is lacking. Here, we use a systems biology-based genome-scale model of metabolism and expression to define a functional core proteome consisting of 356 gene products, accounting for 44% of the Escherichia coli proteome by mass based on proteomics data. This systems biology core proteome includes 212 genes not found in previous comparative genomics-based core proteome definitions, accounts for 65% of known essential genes in E. coli, and has 78% gene function overlap with minimal genomes (Buchnera aphidicola and Mycoplasma genitalium). Based on transcriptomics data across environmental and genetic backgrounds, the systems biology core proteome is significantly enriched in nondifferentially expressed genes and depleted in differentially expressed genes. Compared with the noncore, core gene expression levels are also similar across genetic backgrounds (two times higher Spearman rank correlation) and exhibit significantly more complex transcriptional and posttranscriptional regulatory features (40% more transcription start sites per gene, 22% longer 5'UTR). Thus, genome-scale systems biology approaches rigorously identify a functional core proteome needed to support growth. This framework, validated by using high-throughput datasets, facilitates a mechanistic understanding of systems-level core proteome function through in silico models; it de facto defines a paleome.

Survival outcomes and molecular drivers of testicular cancer in hispanic men.

OBJECTIVE: To examine survival outcomes and molecular drivers in testis cancer among Hispanic men using a large national sample and molecular database. METHODS: We reviewed the SEER registry for testicular cancer from 2000 to 2020. Cox proportional hazards models were used to examine the relationship between race/ethnicity and cancer-specific survival (CSS) by tumor type (seminoma vs. nonseminomatous germ cell tumors [NSGCT]). All models were adjusted for demographic, socioeconomic, and treatment variables. We accessed somatic mutations for testicular cancers through AACR Project GENIE v13.1 and compared mutational frequencies by ethnicity. RESULTS: Our cohort consisted of 43,709 patients (23.3% Hispanic) with median follow-up 106 months (interquartile range: 45-172). Compared to Non-Hispanic Whites (NWH), Hispanics presented at a younger age but with more advanced disease. Hispanics experienced worse CSS for NSGCT (HR 1.7, 95% CI: 1.5-2.0, P < 0.01) but not seminoma. Somatic mutation data was available for 699 patients. KIT and KRAS mutations occurred in 24.2% and 16.9% of seminoma patients (n = 178), respectively. TP53 and KRAS mutations occurred in 12.1% and 7.9% of NSGCT patients (n = 521), respectively. No differences in mutational frequencies were observed between ethnic groups. There was significant heterogeneity in primary ancestral group for Hispanic patients with available data (n = 53); 14 (26.4%) patients had primary Native American ancestry and 30 (56.6%) had primary European ancestry. CONCLUSIONS: Cancer-specific survival is worse for Hispanic men with non-seminoma of the testicle. Somatic mutation analysis suggests no differences by ethnicity, though genetic ancestry is heterogeneous among patients identifying as Hispanic.

LRRK2 kinase inhibition reverses G2019S mutation-dependent effects on tau pathology progression.

BACKGROUND: Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common cause of familial Parkinson's disease (PD). These mutations elevate the LRRK2 kinase activity, making LRRK2 kinase inhibitors an attractive therapeutic. LRRK2 kinase activity has been consistently linked to specific cell signaling pathways, mostly related to organelle trafficking and homeostasis, but its relationship to PD pathogenesis has been more difficult to define. LRRK2-PD patients consistently present with loss of dopaminergic neurons in the substantia nigra but show variable development of Lewy body or tau tangle pathology. Animal models carrying LRRK2 mutations do not develop robust PD-related phenotypes spontaneously, hampering the assessment of the efficacy of LRRK2 inhibitors against disease processes. We hypothesized that mutations in LRRK2 may not be directly related to a single disease pathway, but instead may elevate the susceptibility to multiple disease processes, depending on the disease trigger. To test this hypothesis, we have previously evaluated progression of α-synuclein and tau pathologies following injection of proteopathic seeds. We demonstrated that transgenic mice overexpressing mutant LRRK2 show alterations in the brain-wide progression of pathology, especially at older ages. METHODS: Here, we assess tau pathology progression in relation to long-term LRRK2 kinase inhibition. Wild-type or LRRK2G2019S knock-in mice were injected with tau fibrils and treated with control diet or diet containing LRRK2 kinase inhibitor MLi-2 targeting the IC50 or IC90 of LRRK2 for 3-6 months. Mice were evaluated for tau pathology by brain-wide quantitative pathology in 844 brain regions and subsequent linear diffusion modeling of progression. RESULTS: Consistent with our previous work, we found systemic alterations in the progression of tau pathology in LRRK2G2019S mice, which were most pronounced at 6 months. Importantly, LRRK2 kinase inhibition reversed these effects in LRRK2G2019S mice, but had minimal effect in wild-type mice, suggesting that LRRK2 kinase inhibition is likely to reverse specific disease processes in G2019S mutation carriers. Additional work may be necessary to determine the potential effect in non-carriers. CONCLUSIONS: This work supports a protective role of LRRK2 kinase inhibition in G2019S carriers and provides a rational workflow for systematic evaluation of brain-wide phenotypes in therapeutic development.

Sex Differences in Lung Cancer.

Sex disparities in the incidence and mortality of lung cancer have been observed since cancer statistics have been recorded. Social and economic differences contribute to sex disparities in lung cancer incidence and mortality, but evidence suggests that there are also underlying biological differences that contribute to the disparity. This review summarizes biological differences which could contribute to the sex disparity. Sex hormones and other biologically active molecules, tumor cell genetic differences, and differences in the immune system and its response to lung cancer are highlighted. How some of these differences contribute to disparities in the response to therapies, including cytotoxic, targeted, and immuno-therapies, is also discussed. We end the study with a discussion of our perceived future directions to identify the key biological differences which could contribute to sex disparities in lung cancer and how these differences could be therapeutically leveraged to personalize lung cancer treatment to the individual sexes.

Crosstalk between miRNAs and DNA Methylation in Cancer.

miRNAs are some of the most well-characterized regulators of gene expression. Integral to several physiological processes, their aberrant expression often drives the pathogenesis of both benign and malignant diseases. Similarly, DNA methylation represents an epigenetic modification influencing transcription and playing a critical role in silencing numerous genes. The silencing of tumor suppressor genes through DNA methylation has been reported in many types of cancer and is associated with tumor development and progression. A growing body of literature has described the crosstalk between DNA methylation and miRNAs as an additional layer in the regulation of gene expression. Methylation in miRNA promoter regions inhibits its transcription, while miRNAs can target transcripts and subsequently regulate the proteins responsible for DNA methylation. Such relationships between miRNA and DNA methylation serve an important regulatory role in several tumor types and highlight a novel avenue for potential therapeutic targets. In this review, we discuss the crosstalk between DNA methylation and miRNA expression in the pathogenesis of cancer and describe how miRNAs influence DNA methylation and, conversely, how methylation impacts the expression of miRNAs. Finally, we address how these epigenetic modifications may be leveraged as biomarkers in cancer.

A-to-I edited miR-411-5p targets MET and promotes TKI response in NSCLC-resistant cells.

Non-small cell lung cancer (NSCLC) patients carrying an epidermal growth factor receptor (EGFR) mutation have an initial favorable clinical response to the tyrosine kinase inhibitors (TKIs). Unfortunately, rapid resistance occurs mainly because of genetic alterations, including amplification of the hepatocyte growth factor receptor (MET) and its abnormal activity. The RNA post-transcriptional modifications that contribute to aberrant expression of MET in cancer are largely under-investigated and among them is the adenosine-to-inosine (A-to-I) RNA editing of microRNAs. A reduction of A-to-I editing in position 5 of miR-411-5p has been identified in several cancers, including NSCLC. In this study, thanks to cancer-associated gene expression analysis, we assessed the effect of the edited miR-411-5p on NSCLC cell lines. We found that edited miR-411-5p directly targets MET and negatively affects the mitogen-activated protein kinases (MAPKs) pathway. Considering the predominant role of the MAPKs pathway on TKIs resistance, we generated NSCLC EGFR mutated cell lines resistant to TK inhibitors and evaluated the effect of edited miR-411-5p overexpression. We found that the edited miR-411-5p reduces proliferation and induces apoptosis, promoting EGFR TKIs response in NSCLC-resistant cells.