LSD1 inhibition suppresses ASCL1 and de-represses YAP1 to drive potent activity against neuroendocrine prostate cancer.

Lysine-specific demethylase 1 (LSD1 or KDM1A ) has emerged as a critical mediator of tumor progression in metastatic castration-resistant prostate cancer (mCRPC). Among mCRPC subtypes, neuroendocrine prostate cancer (NEPC) is an exceptionally aggressive variant driven by lineage plasticity, an adaptive resistance mechanism to androgen receptor axis-targeted therapies. Our study shows that LSD1 expression is elevated in NEPC and associated with unfavorable clinical outcomes. Using genetic approaches, we validated the on-target effects of LSD1 inhibition across various models. We investigated the therapeutic potential of bomedemstat, an orally bioavailable, irreversible LSD1 inhibitor with low nanomolar potency. Our findings demonstrate potent antitumor activity against CRPC models, including tumor regressions in NEPC patient-derived xenografts. Mechanistically, our study uncovers that LSD1 inhibition suppresses the neuronal transcriptional program by downregulating ASCL1 through disrupting LSD1:INSM1 interactions and de-repressing YAP1 silencing. Our data support the clinical development of LSD1 inhibitors for treating CRPC - especially the aggressive NE phenotype. Statement of Significance: Neuroendocrine prostate cancer presents a clinical challenge due to the lack of effective treatments. Our research demonstrates that bomedemstat, a potent and selective LSD1 inhibitor, effectively combats neuroendocrine prostate cancer by downregulating the ASCL1- dependent NE transcriptional program and re-expressing YAP1.

Androgen receptor activity in prostate cancer dictates efficacy of bipolar androgen therapy through MYC.

Testosterone is the canonical growth factor of prostate cancer but can paradoxically suppress its growth when present at supraphysiological levels. We have previously demonstrated that the cyclical administration of supraphysiological androgen (SPA), termed bipolar androgen therapy (BAT), can result in tumor regression and clinical benefit for patients with castration-resistant prostate cancer. However, predictors and mechanisms of response and resistance have been ill defined. Here, we show that growth inhibition of prostate cancer models by SPA required high androgen receptor (AR) activity and were driven in part by downregulation of MYC. Using matched sequential patient biopsies, we show that high pretreatment AR activity predicted downregulation of MYC, improved clinical response, and prolonged progression-free and overall survival for patients on BAT. BAT induced strong downregulation of AR in all patients, which is shown to be a primary mechanism of acquired resistance to SPA. Acquired resistance was overcome by alternating SPA with the AR inhibitor enzalutamide, which induced adaptive upregulation of AR and resensitized prostate cancer to SPA. This work identifies high AR activity as a predictive biomarker of response to BAT and supports a treatment paradigm for prostate cancer involving alternating between AR inhibition and activation.

Hypertension in pregnancy as an early sex-specific risk factor for cardiovascular diseases: evidence and awareness.

Despite being a physiological condition, human pregnancy is known to cause numerous complications that can endanger the life of the mother and the fetus alike. While the majority of complications are mostly limited within the peripartum period, more and more information is available about persistently higher short- and long-term cardiovascular risk after a pregnancy complicated by a hypertensive disorder. There is evidence that women after gestational hypertension or preeclampsia are more likely to develop arterial hypertension, coronary atherosclerosis, myocardial infarction, stroke, peripheral artery disease, and even diabetes mellitus and venous thromboembolism years after the target pregnancy. This has urged some authors to view hypertensive disorders of pregnancy as a "stress test" for the maternal organism that unmasks latent endothelial dysfunction. An explanation is sought in the presence of common risk factors and underlying pathological pathways with cardiovascular diseases, although a certain etiological mechanism for the development of hypertensive disorders in pregnancy has not been established yet. More attention is needed towards the follow-up of women after a hypertensive pregnancy as it could be an opportunity for early prevention of cardiovascular diseases.

NOTCH1 PEST domain variants are responsive to standard of care treatments despite distinct transformative properties in a breast cancer model.

Activating variants in the PEST region of NOTCH1 have been associated with aggressive phenotypes in human cancers, including triple-negative breast cancer (TNBC). Previous studies suggested that PEST domain variants in TNBC patients resulted in increased cell proliferation, invasiveness, and decreased overall survival. In this study, we assess the phenotypic transformation of activating NOTCH1 variants and their response to standard of care therapies. AAV-mediated gene targeting was used to isogenically incorporate 3 NOTCH1 variants, including a novel TNBC frameshift variant, in two non-tumorigenic breast epithelial cell lines, MCF10A and hTERT-IMEC. Two different variants at the NOTCH1 A2241 site (A2441fs and A2441T) both demonstrated increased transformative properties when compared to a non-transformative PEST domain variant (S2523L). These phenotypic changes include proliferation, migration, anchorage-independent growth, and MAPK pathway activation. In contrast to previous studies, activating NOTCH1 variants did not display sensitivity to a gamma secretase inhibitor (GSI) or resistance to chemotherapies. This study demonstrates distinct transformative phenotypes are specific to a given variant within NOTCH1 and these phenotypes do not correlate with sensitivities or resistance to chemotherapies or GSIs. Although previous studies have suggested NOTCH1 variants may be prognostic for TNBC, our study does not demonstrate prognostic ability of these variants and suggests further characterization would be required for clinical applications.

Post-Hypoxic Cells Promote Metastatic Recurrence after Chemotherapy Treatment in TNBC.

Hypoxia occurs in 90% of solid tumors and is associated with treatment failure, relapse, and mortality. HIF-1α signaling promotes resistance to chemotherapy in cancer cell lines and murine models via multiple mechanisms including the enrichment of breast cancer stem cells (BCSCs). In this work, we utilize a hypoxia fate-mapping system to determine whether triple-negative breast cancer (TNBC) cells that experience hypoxia in the primary tumor are resistant to chemotherapy at sites of metastasis. Using two orthotopic mouse models of TNBC, we demonstrate that cells that experience intratumoral hypoxia and metastasize to the lung and liver have decreased sensitivity to doxorubicin and paclitaxel but not cisplatin or 5-FU. Resistance to therapy leads to metastatic recurrence caused by post-hypoxic cells. We further determined that the post-hypoxic cells that metastasize are enriched in pathways related to cancer stem cell gene expression. Overall, our results show that even when hypoxic cancer cells are reoxygenated in the bloodstream they retain a hypoxia-induced cancer stem cell-like phenotype that persists and promotes resistance and eventually recurrence.

Targeted delivery of cytotoxic proteins to prostate cancer via conjugation to small molecule urea-based PSMA inhibitors.

Prostate cancer cells are characterized by a remarkably low proliferative rate and the production of high levels of prostate-specific proteases. Protein-based toxins are attractive candidates for prostate cancer therapy because they kill cells via proliferation-independent mechanisms. However, the non-specific cytotoxicity of these potent cytotoxins must be redirected to avoid toxicity to normal tissues. Prostate-Specific Membrane Antigen (PSMA) is membrane-bound carboxypeptidase that is highly expressed by prostate cancer cells. Potent dipeptide PSMA inhibitors have been developed that can selectively deliver and concentrate imaging agents within prostate cancer cells based on continuous PSMA internalization and endosomal cycling. On this basis, we conjugated a PSMA inhibitor to the apoptosis-inducing human protease Granzyme B and the potent Pseudomonas exotoxin protein toxin fragment, PE35. We assessed selective PSMA binding and entrance into tumor cell to induce cell death. We demonstrated these agents selectively bound to PSMA and became internalized. PSMA-targeted PE35 toxin was selectively toxic to PSMA producing cells in vitro. Intratumoral and intravenous administration of this toxin produced marked tumor killing of PSMA-producing xenografts with minimal host toxicity. These studies demonstrate that urea-based PSMA inhibitors represent a simpler, less expensive alternative to antibodies as a means to deliver cytotoxic proteins to prostate cancer cells.

Hierarchical tumor heterogeneity mediated by cell contact between distinct genetic subclones.

Intratumor heterogeneity is an important mediator of poor outcomes in many cancers, including breast cancer. Genetic subclones frequently contribute to this heterogeneity; however, their growth dynamics and interactions remain poorly understood. PIK3CA and HER2 alterations are known to coexist in breast and other cancers. Herein, we present data that describe the ability of oncogenic PIK3CA mutant cells to induce the proliferation of quiescent HER2 mutant cells through a cell contact-mediated mechanism. Interestingly, the HER2 cells proliferated to become the major subclone over PIK3CA counterparts both in vitro and in vivo. Furthermore, this phenotype was observed in both hormone receptor-positive and -negative cell lines, and was dependent on the expression of fibronectin from mutant PIK3CA cells. Analysis of human tumors demonstrated similar HER2:PIK3CA clonal dynamics and fibronectin expression. Our study provides insight into nonrandom subclonal architecture of heterogenous tumors, which may aid the understanding of tumor evolution and inform future strategies for personalized medicine.

High-sensitivity CRP levels in women with gestational hypertension, preeclampsia and in normotensive pregnant women and its correlations.

INTRODUCTION: Gestational hypertension is a less investigated hypertensive disorder of pregnancy than preeclampsia, but evidence exists of an unfavourable cardiovascular profile for women after such a pregnancy. AIM: To determine serum high-sensitivity C-reactive protein (hs-CRP) levels in women with preeclampsia, gestational hypertension, and in normotensive pregnancy in order to assess the cardiovascular implications and to examine its correlations with some characteristics of women. MATERIALS AND METHODS: Thirty-six women with gestational hypertension, thirty-seven with preeclampsia, and fifty maternal and gestational age-matched controls were included in a single-center prospective clinical-epidemiological study. Serum hs-CRP levels were determined using ELISA method. RESULTS: Significantly higher hs-CRP levels were found in the gestational hypertension group than in the controls (p=0.043), but not in the preeclampsia group (p=0.445). The levels between the two pathological groups did not differ significantly (p=0.247). Odds ratio for hs-CRP levels higher than the provided cut-off was 3.31 (95% CI 1.32-8.29) for the presence of gestational hypertension. In the normotensive pregnant women, the hs-CRP levels had a positive correlation with BSA, pre-pregnancy and current BMI, but such correlations were absent in the hypertensive groups. There were no correlations with the maternal or gestational age, current weight gain in any of the groups or with the highest detected blood pressure in the pathological groups. These levels did not differ according to gravidity, smoking status and smoking during pregnancy. CONCLUSIONS: Elevation of hs-CRP was more pronounced in women with gestational hypertension than in women with preeclampsia, which could indicate a different pathophysiological mechanism and a higher cardiovascular risk for those women.

Ultrasound elastography using a regularized modified error in constitutive equations (MECE) approach: a comprehensive phantom study.

Many of the current techniques in transient elastography, such as shear wave elastography (SWE) assume a dominant planar shear wave propagating in an infinite medium. This underlying assumption, however, can be easily violated in real scenarios in vivo, leading to image artifacts and reconstruction errors. Other approaches that are not bound to planar shear wave assumption, such solutions based on the partial differential equation, can potentially overcome the shortcomings of the conventional SWE. The main objective of this paper is to demonstrate the advantages of the modified error in constitutive equations (MECE) formulation with total variation regularization (MECE + TV) over SWE in reconstructing the elastic moduli of different tissue-mimicking phantoms. Experiments were conducted on phantoms with inclusions of well-defined shapes to study the reconstruction of specific features relevant to practical applications. We compared the performances of MECE + TV and SWE in terms of quantitative metrics to estimate reconstruction accuracy, inclusion shape recovery, edge preservation and edge sharpness, inclusion size representation, and shear elasticity and contrast accuracies. The results indicate that the MECE + TV approach outperforms SWE based on several of these metrics. It is concluded that, with further development, the proposed method may offer elastography reconstructions that are superior to SWE in clinical applications.

The Mortality After Release from Incarceration Consortium (MARIC): Protocol for a multi-national, individual participant data meta-analysis.

INTRODUCTION: More than 30 million adults are released from incarceration globally each year. Many experience complex physical and mental health problems, and are at markedly increased risk of preventable mortality. Despite this, evidence regarding the global epidemiology of mortality following release from incarceration is insufficient to inform the development of targeted, evidence-based responses. Many previous studies have suffered from inadequate power and poor precision, and even large studies have limited capacity to disaggregate data by specific causes of death, sub-populations or time since release to answer questions of clinical and public health relevance. OBJECTIVES: To comprehensively document the incidence, timing, causes and risk factors for mortality in adults released from prison. METHODS: We created the Mortality After Release from Incarceration Consortium (MARIC), a multi-disciplinary collaboration representing 29 cohorts of adults who have experienced incarceration from 11 countries. Findings across cohorts will be analysed using a two-step, individual participant data meta-analysis methodology. RESULTS: The combined sample includes 1,337,993 individuals (89% male), with 75,795 deaths recorded over 9,191,393 person-years of follow-up. CONCLUSIONS: The consortium represents an important advancement in the field, bringing international attention to this problem. It will provide internationally relevant evidence to guide policymakers and clinicians in reducing preventable deaths in this marginalized population. KEY WORDS: Mortality; incarceration; prison; release; individual participant data meta-analysis; consortium; cohort.

Microparticle Encapsulation of a Prostate-targeted Biologic for the Treatment of Liver Metastases in a Preclinical Model of Castration-resistant Prostate Cancer.

PRX302 is a highly potent, mutant bacterial pore-forming biologic protoxin engineered for selective activation by PSA, a serine protease expressed by benign and malignant prostate epithelial cells. Although being developed as a local therapy for benign prostatic hyperplasia and localized prostate cancer, PRX302 cannot be administered systemically as a treatment for metastatic disease due to binding to ubiquitously expressed glycosylphosphatidylinositol (GPI)-anchored proteins, which leads to poor accumulation within the tumor microenvironment. To overcome this limitation, poly-lactic-co-glycolic acid (PLGA) microparticles encapsulating the protoxin were developed, which are known to accumulate in the liver, a major site of metastasis for prostate cancer and other solid tumors. A highly sensitive and reproducible sandwich ELISA to quantify PRX302 released from microparticles was developed. Utilizing this assay, PRX302 release from different microparticle formulations was assessed over multiple days. Hemolysis assays documented PSA-dependent pore formation and lytic potential (i.e., function) of the released protoxin. MTT assays demonstrated that conditioned supernatant from PRX302-loaded, but not blank (i.e., unloaded), PLGA microparticles was highly cytotoxic to PC3 and DU145 human prostate cancer cells in the presence of exogenous PSA. Microparticle encapsulation prevented PRX302 from immediately interacting with GPI-anchored proteins as demonstrated in a competition assay, which resulted in an increased therapeutic index and significant antitumor efficacy following a single dose of PRX302-loaded microparticles in a preclinical model of prostate cancer liver metastasis with no obvious toxicity. These results document that PRX302 released from PLGA microparticles demonstrate in vivo antitumor efficacy in a clinically relevant preclinical model of metastatic prostate cancer.

Correction: Disruption of the retinoblastoma pathway by small interfering RNA and ectopic expression of the catalytic subunit of telomerase lead to immortalization of human ovarian surface epithelial cells.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Analyzing acoustoelastic effect of shear wave elastography data for perfused and hydrated soft tissues using a macromolecular network inspired model.

Shear wave elastography (SWE) has enhanced our ability to non-invasively make in vivo measurements of tissue elastic properties of animal and human tissues. Recently, researchers have taken advantages of acoustoelasticity in SWE to extract nonlinear elastic properties from soft biological tissues. However, most investigations of the acoustoelastic effects of SWE data (AE-SWE) rely on classic hyperelastic models for rubber-like (dry) materials. In this paper, we focus solely on understanding acoustoelasticity in soft hydrated tissues using SWE data and propose a straightforward approach to modeling the constitutive behavior of soft tissue that has a direct microstructural/macromolecular interpretation. Our approach incorporates two constitutive features relevant to biological tissues into AE-SWE: static dilation of the medium associated with nonstructural components (e.g. tissue hydration and perfusion) and finite extensibility derived from an ideal network of biological filaments. We evaluated the proposed method using data from an in-house tissue-mimicking phantom experiment, and ex vivo and in vivo AE-SWE data available in the SWE literature. In conclusion, predictions made by our approach agreed well with measurements obtained from phantom, ex vivo and in vivo tissue experiments.

A comparison of hyperelastic constitutive models applicable to shear wave elastography (SWE) data in tissue-mimicking materials.

Shear wave elastography (SWE) techniques have received substantial attention in recent years. Strong experimental data in SWE suggest that shear wave speed changes significantly due to the known acoustoelastic effect (AE). This presents both challenges and opportunities toward in vivo characterization of biological soft tissues. In this work, under the framework of continuum mechanics, we model a tissue-mimicking material as a homogeneous, isotropic, incompressible, hyperelastic material. Our primary objective is to quantitatively and qualitatively compare experimentally measured acoustoelastic data with model-predicted outcomes using multiple strain energy functions. Our analysis indicated that the classic Neo-Hookean and Mooney-Rivlin models are inadequate for modeling the AE in tissue-mimicking materials. However, a subclass of strain energy functions containing both high-order/exponential term(s) and second-order invariant dependence showed good agreement with experimental data. Based on data investigated, we also found that discrepancies may exist between parameters inversely estimated from uniaxial compression and SWE data. Overall, our findings may improve our understanding of clinical SWE results.

Drp1/Fis1-mediated mitochondrial fragmentation leads to lysosomal dysfunction in cardiac models of Huntington's disease.

Huntington's disease (HD) is a fatal hereditary neurodegenerative disorder, best known for its clinical triad of progressive motor impairment, cognitive deficits and psychiatric disturbances, is caused by CAG-repeat expansion in exon 1 of Huntingtin (HTT). However, in addition to the neurological disease, mutant HTT (mHTT), which is ubiquitously expressed in all tissues, impairs other organ systems. Not surprisingly, cardiovascular dysautonomia as well as the deterioration of circadian rhythms are among the earliest detectable pathophysiological changes in individuals with HD. Mitochondrial dysfunction in the brain and skeletal muscle in HD has been well documented, as the disease progresses. However, not much is known about mitochondrial abnormalities in the heart. In this study, we describe a role for Drp1/Fis1-mediated excessive mitochondrial fission and dysfunction, associated with lysosomal dysfunction in H9C2 expressing long polyglutamine repeat (Q73) and in human iPSC-derived cardiomyocytes transfected with Q77. Expression of long polyglutamine repeat led to reduced ATP production and mitochondrial fragmentation. We observed an increased accumulation of damaged mitochondria in the lysosome that was coupled with lysosomal dysfunction. Importantly, reducing Drp1/Fis1-mediated mitochondrial damage significantly improved mitochondrial function and cell survival. Finally, reducing Fis1-mediated Drp1 recruitment to the mitochondria, using the selective inhibitor of this interaction, P110, improved mitochondrial structure in the cardiac tissue of R6/2 mice. We suggest that drugs focusing on the central nervous system will not address mitochondrial function across all organs, and therefore will not be a sufficient strategy to treat or slow down HD disease progression.

The estrogen receptor-alpha S118P variant does not affect breast cancer incidence or response to endocrine therapies.

PURPOSE: Estrogen receptor-alpha (ER) is a therapeutic target of ER-positive (ER+) breast cancers. Although ER signaling is complex, many mediators of this pathway have been identified. Specifically, phosphorylation of ER at serine 118 affects responses to estrogen and therapeutic ligands and has been correlated with clinical outcomes in ER+ breast cancer patients. We hypothesized that a newly described germline variant (S118P) at this residue would drive cellular changes consistent with breast cancer development and/or hormone resistance. METHODS: Isogenic human breast epithelial cell line models harboring ER S118P were developed via genome editing and characterized to determine the functional effects of this variant. We also examined the frequency of ER S118P in a case-control study (N = 536) of women with and without breast cancer with a familial risk. RESULTS: In heterozygous knock-in models, the S118P variant demonstrated no significant change in proliferation, migration, MAP Kinase pathway signaling, or response to the endocrine therapies tamoxifen and fulvestrant. Further, there was no difference in the prevalence of S118P between women with and without cancer relative to population registry databases. CONCLUSIONS: This study suggests that the ER S118P variant does not affect risk for breast cancer or hormone therapy resistance. Germline screening and modification of treatments for patients harboring this variant are likely not warranted.

PSA-selective activation of cytotoxic human serine proteases within the tumor microenvironment as a therapeutic strategy to target prostate cancer.

Prostate cancer is the most diagnosed malignancy and the second leading cause of cancer-related death in American men. While localized therapy is highly curative, treatments for metastatic prostate cancer are largely palliative. Thus, new innovative therapies are needed to target metastatic tumors. Prostate-Specific Antigen (PSA) is a chymotrypsin-like protease with a unique substrate specificity that is secreted by both normal and malignant prostate epithelial cells. Previous studies demonstrated the presence of high levels (µM-mM) of enzymatically active PSA is present in the extracellular fluid of the prostate cancer microenvironment. Because of this, PSA is an attractive target for a protease activated pro-toxin therapeutic strategy. Because prostate cancers typically grow very slowly, a strategy employing a proliferation-independent cytotoxic payload is preferred. Recently, it was shown that the human protease Granzyme B (GZMB), at low micromolar concentrations in the extracellular space, can cleave an array of extracellular matrix (ECM) proteins thus perturbing cell growth, signaling, motility, and integrity. It is also well established that other human proteases such as trypsin can induce similar effects. Because both enzymes require N-terminal proteolytic activation, we propose to convert these proteins into PSA-activated cytotoxins. In this study, we examine the enzymatic and cell targeting parameters of these PSA-activated cytotoxic serine proteases. These pro-enzymes were activated robustly by PSA and induced ECM damage that led to the death of prostate cancer cells in vitro thus supporting the potential use of this strategy as means to target metastatic prostate cancers.

Great Expectations: HIV Risk Behaviors and Misperceptions of Low HIV Risk among Incarcerated Men.

Incarcerated populations have relatively high HIV prevalence but little has been reported about their aggregate HIV risk behaviors or perceptions of risk. A random selection of HIV-negative men (n = 855) entering a US state prison system were surveyed to assess five risk behaviors and his self-perceived HIV risk. Using multivariate logistic regression, we identified factors associated with having elevated actual but low perceived risk (EALPR). Of the 826 men with complete data, 88% were at elevated risk. While 64% of the sample had risk perceptions concordant with their actual risk, 14% had EALPR (with the remainder at low actual but high perceived risk). EALPR rates were lower in those with a pre-incarceration HIV test but higher for those with a negative prison entry HIV test. HIV testing counseling should assess for discordance between actual and perceived risk and communicate the continued risk of HIV despite a negative result.