Lestaurtinib's antineoplastic activity converges on JAK/STAT signaling to inhibit advanced forms of therapy resistant ovarian cancer.

Ovarian cancer is the deadliest gynecological malignancy, owing to its late-stage diagnosis and high rates of recurrence and resistance following standard-of-care treatment, highlighting the need for novel treatment approaches. Through an unbiased drug screen, we identified the kinase inhibitor, lestaurtinib, as a potent antineoplastic agent for chemotherapy- and PARP-inhibitor (PARPi)-sensitive and -resistant ovarian cancer cells and patient derived xenografts (PDXs). RNA-sequencing revealed that lestaurtinib potently suppressed JAK/STAT signaling and lestaurtinib efficacy was shown to be directly related to JAK/STAT pathway activity in cell lines and PDX models. Most ovarian cancer cells exhibited constitutive JAK/STAT pathway activation and genetic loss of STAT1 and STAT3 resulted in growth inhibition. Lestaurtinib also displayed synergy when combined with cisplatin and olaparib, including in a model of PARPi resistance. In contrast, the most well-known JAK/STAT inhibitor, ruxolitinib, lacked antineoplastic activity against all ovarian cancer cell lines and PDX models tested. This divergent behavior was reflected in the ability of lestaurtinib to block both Y701/705 and S727 phosphorylation of STAT1 and STAT3, whereas ruxolitinib failed to block S727. Consistent with these findings, lestaurtinib additionally inhibited JNK and ERK activity, leading to more complete suppression of STAT phosphorylation. Concordantly, combinatorial treatment with ruxolitinib and a JNK or ERK inhibitor resulted in synergistic antineoplastic effects at dose levels where single agents were ineffective. Taken together, these findings indicate that lestaurtinib, and other treatments that converge on JAK/STAT signaling, are worthy of further pre-clinical and clinical exploration for the treatment of highly aggressive and advanced forms of ovarian cancer. Statement of significance: Lestaurtinib is a novel inhibitor of ovarian cancer, including chemotherapy- and PARPi-resistant models, that acts through robust inhibition of the JAK/STAT pathway and synergizes with standard-of-care agents at clinically relevant concentrations.

Efferocytosis and Bone Dynamics.

PURPOSE OF REVIEW: This review summarizes the recently published scientific evidence regarding the role of efferocytosis in bone dynamics and skeletal health. RECENT FINDINGS: Several types of efferocytes have been identified within the skeleton, with macrophages being the most extensively studied. Efferocytosis is not merely a 'clean-up' process vital for maintaining skeletal homeostasis; it also plays a crucial role in promoting resolution pathways and orchestrating bone dynamics, such as osteoblast-osteoclast coupling during bone remodeling. Impaired efferocytosis has been associated with aging-related bone loss and various skeletal pathologies, including osteoporosis, osteoarthritis, rheumatoid arthritis, and metastatic bone diseases. Accordingly, emerging evidence suggests that targeting efferocytic mechanisms has the potential to alleviate these conditions. While efferocytosis remains underexplored in the skeleton, recent discoveries have shed light on its pivotal role in bone dynamics, with important implications for skeletal health and pathology. However, there are several knowledge gaps and persisting technical limitations that must be addressed to fully unveil the contributions of efferocytosis in bone.

The selective cyclooxygenase-2 inhibitor NS398 ameliorates cisplatin-induced impairments in mitochondrial and cognitive function.

Chemobrain is a condition that negatively affects cognition in cancer patients undergoing active chemotherapy, as well as following chemotherapy cessation. Chemobrain is also known as chemotherapy-induced cognitive impairment (CICI) and has emerged as a significant medical contingency. There is no therapy to ameliorate this condition, hence identification of novel therapeutic strategies to prevent CICI is of great interest to cancer survivors. Utilizing the platinum-based chemotherapy cisplatin in an investigative approach for CICI, we identified increased expression of cyclooxygenase-2 (COX-2) and prostaglandin E2 (PGE2) in the adult mouse hippocampus, and in human cortical neuron cultures derived from induced pluripotent stem cells (iPSCs). Notably, administration of NS398, a selective COX-2 inhibitor, prevented CICI in vivo without negatively affecting the antitumor efficacy of cisplatin or potentiating tumor growth. Given that dysfunctional mitochondrial bioenergetics plays a prominent role in CICI, we explored the effects of NS398 in cisplatin-induced defects in human cortical mitochondria. We found that cisplatin significantly reduces mitochondrial membrane potential (MMP), increases matrix swelling, causes loss of cristae membrane integrity, impairs ATP production, as well as decreases cell viability and dendrite outgrowth. Pretreatment with NS398 in human cortical neurons attenuated mitochondrial dysfunction caused by cisplatin, while improving cell survival and neurite morphogenesis. These results suggest that aberrant COX-2 inflammatory pathways may contribute in cisplatin-induced mitochondrial damage and cognitive impairments. Therefore, COX-2 signaling may represent a viable therapeutic approach to improve the quality of life for cancer survivors experiencing CICI.

Endoxifen downregulates AKT phosphorylation through protein kinase C beta 1 inhibition in ERα+ breast cancer.

Endoxifen, a secondary tamoxifen metabolite, is a potent antiestrogen exhibiting estrogen receptor alpha (ERα) binding at nanomolar concentrations. Phase I/II clinical trials identified clinical activity of Z-endoxifen (ENDX), in endocrine-refractory metastatic breast cancer as well as ERα+ solid tumors, raising the possibility that ENDX may have a second, ERα-independent, mechanism of action. An unbiased mass spectrometry approach revealed that ENDX concentrations achieved clinically with direct ENDX administration (5 µM), but not low concentrations observed during tamoxifen treatment (<0.1 µM), profoundly altered the phosphoproteome of the aromatase expressing MCF7AC1 cells with limited impact on the total proteome. Computational analysis revealed protein kinase C beta (PKCß) and protein kinase B alpha or AKT1 as potential kinases responsible for mediating ENDX effects on protein phosphorylation. ENDX more potently inhibited PKCß1 kinase activity compared to other PKC isoforms, and ENDX binding to PKCß1 was confirmed using Surface Plasma Resonance. Under conditions that activated PKC/AKT signaling, ENDX induced PKCß1 degradation, attenuated PKCß1-activated AKTSer473 phosphorylation, diminished AKT substrate phosphorylation, and induced apoptosis. ENDX's effects on AKT were phenocopied by siRNA-mediated PKCß1 knockdown or treatment with the pan-AKT inhibitor, MK-2206, while overexpression of constitutively active AKT diminished ENDX-induced apoptosis. These findings, which identify PKCß1 as an ENDX target, indicate that PKCß1/ENDX interactions suppress AKT signaling and induce apoptosis in breast cancer.

A TGF-ß/KLF10 signaling axis regulates atrophy-associated genes to induce muscle wasting in pancreatic cancer.

Cancer cachexia, and its associated complications, represent a large and currently untreatable roadblock to effective cancer management. Many potential therapies have been proposed and tested-including appetite stimulants, targeted cytokine blockers, and nutritional supplementation-yet highly effective therapies are lacking. Innovative approaches to treating cancer cachexia are needed. Members of the Kruppel-like factor (KLF) family play wide-ranging and important roles in the development, maintenance, and metabolism of skeletal muscle. Within the KLF family, we identified KLF10 upregulation in a multitude of wasting contexts-including in pancreatic, lung, and colon cancer mouse models as well as in human patients. We subsequently interrogated loss-of-function of KLF10 as a potential strategy to mitigate cancer associated muscle wasting. In vivo studies leveraging orthotopic implantation of pancreas cancer cells into wild-type and KLF10 KO mice revealed significant preservation of lean mass and robust suppression of pro-atrophy muscle-specific ubiquitin ligases Trim63 and Fbxo32, as well as other factors implicated in atrophy, calcium signaling, and autophagy. Bioinformatics analyses identified Transforming growth factor beta (TGF-ß), a known inducer of KLF10 and cachexia promoting factor, as a key upstream regulator of KLF10. We provide direct in vivo evidence that KLF10 KO mice are resistant to the atrophic effects of TGF-ß. ChIP-based binding studies demonstrated direct binding to Trim63, a known wasting-associated atrogene. Taken together, we report a critical role for the TGF-ß/KLF10 axis in the etiology of pancreatic cancer-associated muscle wasting and highlight the utility of targeting KLF10 as a strategy to prevent muscle wasting and limit cancer-associated cachexia.

G-protein coupled receptor 5C (GPRC5C) is required for osteoblast differentiation and responds to EZH2 inhibition and multiple osteogenic signals.

Osteoblast differentiation is epigenetically suppressed by the H3K27 methyltransferase EZH2, and induced by the morphogen BMP2 and transcription factor RUNX2. These factors also regulate distinct G protein coupled receptors (GPRCs; e.g., PTH1R, GPR30/GPER1). Because GPRCs transduce many physiological stimuli, we examined whether BMP2 or EZH2 inhibition (i.e., GSK126) regulates other GPRC genes in osteoblasts. RNA-seq screening of >400 mouse GPRC-related genes showed that many GPRCs are downregulated during osteogenic differentiation. The orphan receptor GPRC5C, along with a small subset of other GPRCs, is induced by BMP2 or GSK126 during Vitamin C dependent osteoblast differentiation, but not by all-trans retinoic acid. ChIP-seq analysis revealed that GSK126 reduces H3K27me3 levels at the GPRC5C gene locus in differentiating MC3T3-E1 osteoblasts, consistent with enhanced GPRC5C mRNA expression. Loss of function analyses revealed that shRNA-mediated depletion of GPRC5C decreases expression of bone markers (e.g., BGLAP and IBSP) and mineral deposition in response to BMP2 or GSK126. GPRC5C mRNA was found to be reduced in the osteopenic bones of KLF10 null mice which have compromised BMP2 signaling. GPRC5C mRNA is induced by the bone-anabolic activity of 17ß-estradiol in trabecular but not cortical bone following ovariectomy. Collectively, these findings suggest that GPRC5C protein is a key node in a pro-osteogenic axis that is normally suppressed by EZH2-mediated H3K27me3 marks and induced during osteoblast differentiation by GSK126, BMP2, and/or 17ß-estradiol. Because GPRC5C protein is an understudied orphan receptor required for osteoblast differentiation, identification of ligands that induce GPRC5C signaling may support therapeutic strategies to mitigate bone-related disorders.

A lncRNA from the FTO locus acts as a suppressor of the m6A writer complex and p53 tumor suppression signaling.

N6-methyladenosine (m6A) of mRNAs modulated by the METTL3-METTL14-WTAP-RBM15 methyltransferase complex and m6A demethylases such as FTO play important roles in regulating mRNA stability, splicing, and translation. Here, we demonstrate that FTO-IT1 long noncoding RNA (lncRNA) was upregulated and positively correlated with poor survival of patients with wild-type p53-expressing prostate cancer (PCa). m6A RIP-seq analysis revealed that FTO-IT1 knockout increased mRNA m6A methylation of a subset of p53 transcriptional target genes (e.g., FAS, TP53INP1, and SESN2) and induced PCa cell cycle arrest and apoptosis. We further showed that FTO-IT1 directly binds RBM15 and inhibits RBM15 binding, m6A methylation, and stability of p53 target mRNAs. Therapeutic depletion of FTO-IT1 restored mRNA m6A level and expression of p53 target genes and inhibited PCa growth in mice. Our study identifies FTO-IT1 lncRNA as a bona fide suppressor of the m6A methyltransferase complex and p53 tumor suppression signaling and nominates FTO-IT1 as a potential therapeutic target of cancer.

Krüppel-like factor 10 modulates stem cell phenotypes of pancreatic adenocarcinoma by transcriptionally regulating notch receptors.

BACKGROUND: Pancreatic adenocarcinoma (PDAC) is well known for its rapid distant metastasis and local destructive behavior. Loss of Krüppel-like factor 10 (KLF10) contributes to distant migration of PDAC. The role of KLF10 in modulating tumorigenesis and stem cell phenotypes of PDAC is unclear. METHODS: Additional depletion of KLF10 in KC (LSL: KrasG12D; Pdx1-Cre) mice, a spontaneous murine PDAC model, was established to evaluate tumorigenesis. Tumor specimens of PDAC patients were immune-stained of KLF10 to correlate with local recurrence after curative resection. Conditional overexpressing KLF10 in MiaPaCa and stably depleting KLF10 in Panc-1 (Panc-1-pLKO-shKLF10) cells were established for evaluating sphere formation, stem cell markers expression and tumor growth. The signal pathways modulated by KLF10 for PDAC stem cell phenotypes were disclosed by microarray analysis and validated by western blot, qRT-PCR, luciferase reporter assay. Candidate targets to reverse PDAC tumor growth were demonstrated in murine model. RESULTS: KLF10, deficient in two-thirds of 105 patients with resected pancreatic PDAC, was associated with rapid local recurrence and large tumor size. Additional KLF10 depletion in KC mice accelerated progression from pancreatic intraepithelial neoplasia to PDAC. Increased sphere formation, expression of stem cell markers, and tumor growth were observed in Panc-1-pLKO-shKLF10 compared with vector control. Genetically or pharmacologically overexpression of KLF10 reversed the stem cell phenotypes induced by KLF10 depletion. Ingenuity pathway analysis and gene set enrichment analysis showed that Notch signaling molecules, including Notch receptors 3 and 4, were over-expressed in Panc-1-pLKO-shKLF10. KLF10 transcriptionally suppressed Notch-3 and -4 by competing with E74-like ETS transcription factor 3, a positive regulator, for promoter binding. Downregulation of Notch signaling, either genetically or pharmacologically, ameliorated the stem cell phenotypes of Panc-1-pLKO-shKLF10. The combination of metformin, which upregulated KLF10 expression via phosphorylating AMPK, and evodiamine, a non-toxic Notch-3 methylation stimulator, delayed tumor growth of PDAC with KLF10 deficiency in mice without prominent toxicity. CONCLUSIONS: These results demonstrated a novel signaling pathway by which KLF10 modulates stem cell phenotypes in PDAC through transcriptionally regulating Notch signaling pathway. The elevation of KLF10 and suppression of Notch signaling may jointly reduce PDAC tumorigenesis and malignant progression.

Estrogen-regulated miRs in bone enhance osteoblast differentiation and matrix mineralization.

Estrogen signaling is critical for the development and maintenance of healthy bone, and age-related decline in estrogen levels contributes to the development of post-menopausal osteoporosis. Most bones consist of a dense cortical shell and an internal mesh-like network of trabecular bone that respond differently to internal and external cues such as hormonal signaling. To date, no study has assessed the transcriptomic differences that occur specifically in cortical and trabecular bone compartments in response to hormonal changes. To investigate this, we employed a mouse model of post-menopausal osteoporosis (ovariectomy, OVX) and estrogen replacement therapy (ERT). mRNA and miR sequencing revealed distinct transcriptomic profiles between cortical and trabecular bone in the setting of OVX and ERT. Seven miRs were identified as likely contributors to the observed estrogen-mediated mRNA expression changes. Of these, four miRs were prioritized for further study and decreased predicted target gene expression in bone cells, enhanced the expression of osteoblast differentiation markers, and altered the mineralization capacity of primary osteoblasts. As such, candidate miRs and miR mimics may have therapeutic relevance for bone loss resulting from estrogen depletion without the unwanted side effects of hormone replacement therapy and therefore represent novel therapeutic approaches to combat diseases of bone loss.

Disruption of estrogen receptor beta's DNA binding domain impairs its tumor suppressive effects in triple negative breast cancer.

Triple negative breast cancer (TNBC) is an aggressive sub-type of the disease which accounts for a disproportionately high percentage of breast cancer morbidities and mortalities. For these reasons, a better understanding of TNBC biology is required and the development of novel therapeutic approaches are critically needed. Estrogen receptor beta (ERß) is a reported tumor suppressor that is expressed in approximately 20% of primary TNBC tumors, where it is associated with favorable prognostic features and patient outcomes. Previous studies have shown that ERß mediates the assembly of co-repressor complexes on DNA to inhibit the expression of multiple growth promoting genes and to suppress the ability of oncogenic transcription factors to drive cancer progression. To further elucidate the molecular mechanisms by which ERß elicits its anti-cancer effects, we developed MDA-MB-231 cells that inducibly express a mutant form of ERß incapable of directly binding DNA. We demonstrate that disruption of ERß's direct interaction with DNA abolishes its ability to regulate the expression of well characterized immediate response genes and renders it unable to suppress TNBC cell proliferation. Loss of DNA binding also diminishes the ability of ERß to suppress oncogenic NFκB signaling even though it still physically associates with NFκB and other critical co-factors. These findings enhance our understanding of how ERß functions in this disease and provide a model system that can be utilized to further investigate the mechanistic processes by which ERß elicits its anti-cancer effects.

Deciphering the Role of Klf10 in the Cerebellum.

Recent studies have demonstrated a new role for Klf10, a Krüppel-like transcription factor, in skeletal muscle, specifically relating to mitochondrial function. Thus, it was of interest to analyze additional tissues that are highly reliant on optimal mitochondrial function such as the cerebellum and to decipher the role of Klf10 in the functional and structural properties of this brain region. In vivo (magnetic resonance imaging and localized spectroscopy, behavior analysis) and in vitro (histology, spectroscopy analysis, enzymatic activity) techniques were applied to comprehensively assess the cerebellum of wild type (WT) and Klf10 knockout (KO) mice. Histology analysis and assessment of locomotion revealed no significant difference in Klf10 KO mice. Diffusion and texture results obtained using MRI revealed structural changes in KO mice characterized as defects in the organization of axons. These modifications may be explained by differences in the levels of specific metabolites (myo-inositol, lactate) within the KO cerebellum. Loss of Klf10 expression also led to changes in mitochondrial activity as reflected by a significant increase in the activity of citrate synthase, complexes I and IV. In summary, this study has provided evidence that Klf10 plays an important role in energy production and mitochondrial function in the cerebellum.

Adenosine A2A receptor blockade prevents cisplatin-induced impairments in neurogenesis and cognitive function.

Chemotherapy-induced cognitive impairment (CICI) has emerged as a significant medical problem without therapeutic options. Using the platinum-based chemotherapy cisplatin to model CICI, we revealed robust elevations in the adenosine A2A receptor (A2AR) and its downstream effectors, cAMP and CREB, by cisplatin in the adult mouse hippocampus, a critical brain structure for learning and memory. Notably, A2AR inhibition by the Food and Drug Administration-approved A2AR antagonist KW-6002 prevented cisplatin-induced impairments in neural progenitor proliferation and dendrite morphogenesis of adult-born neurons, while improving memory and anxiety-like behavior, without affecting tumor growth or cisplatin's antitumor activity. Collectively, our study identifies A2AR signaling as a key pathway that can be therapeutically targeted to prevent cisplatin-induced cognitive impairments.

Serum and Soleus Metabolomics Signature of Klf10 Knockout Mice to Identify Potential Biomarkers.

The transcription factor Krüppel-like factor 10 (Klf10), also known as Tieg1 for TGFß (Inducible Early Gene-1) is known to control numerous genes in many cell types that are involved in various key biological processes (differentiation, proliferation, apoptosis, inflammation), including cell metabolism and human disease. In skeletal muscle, particularly in the soleus, deletion of the Klf10 gene (Klf10 KO) resulted in ultrastructure fiber disorganization and mitochondrial metabolism deficiencies, characterized by muscular hypertrophy. To determine the metabolic profile related to loss of Klf10 expression, we analyzed blood and soleus tissue using UHPLC-Mass Spectrometry. Metabolomics analyses on both serum and soleus revealed profound differences between wild-type (WT) and KO animals. Klf10 deficient mice exhibited alterations in metabolites associated with energetic metabolism. Additionally, chemical classes of aromatic and amino-acid compounds were disrupted, together with Krebs cycle intermediates, lipids and phospholipids. From variable importance in projection (VIP) analyses, the Warburg effect, citric acid cycle, gluconeogenesis and transfer of acetyl groups into mitochondria appeared to be possible pathways involved in the metabolic alterations observed in Klf10 KO mice. These studies have revealed essential roles for Klf10 in regulating multiple metabolic pathways whose alterations may underlie the observed skeletal muscle defects as well as other diseases.

Surveillance mammography after treatment for male breast cancer.

PURPOSE: To identify the practice patterns related to use of surveillance mammography in male breast cancer (MaBC) survivors. METHODS: Using administrative claims data from OptumLabs Data Warehouse, we identified men who underwent surgery for breast cancer during 2007-2017. We calculated the proportion of men who had at least one mammogram (a) within 13 months for all patients and (b) within 24 months amongst those who maintained their insurance coverage for at least that length of time after surgery. Multivariate logistic regression modeling was used to identify factors associated with mammography within each timeframe. RESULTS: Out of 729 total MaBC survivors, 209 (29%) underwent mammography within 13 months after surgery. Among those who had lumpectomy, 41% underwent mammography, whereas among those who had mastectomy, 27% had mammography. Amongst 526 men who maintained consistent insurance coverage for 24 months after surgery, 215 (41%) underwent mammography at least once during that 24-month period. In this cohort, the proportion who had at least one mammogram during the 24-month period was 49% after lumpectomy and 40% after mastectomy. In a multivariate logistic regression model, more recent diagnosis (2015+) and older age at diagnosis were associated with lower odds of undergoing mammography, while receipt of radiation was associated with higher odds of undergoing mammography. CONCLUSIONS: Although recent ASCO guidelines recommend surveillance mammography after lumpectomy, a minority of MaBC survivors undergo surveillance mammography, even after lumpectomy. This is likely due to the paucity of data regarding the true benefits and harms of surveillance/screening mammography for MaBC.

Estrogen receptor beta repurposes EZH2 to suppress oncogenic NFκB/p65 signaling in triple negative breast cancer.

Triple Negative Breast Cancer (TNBC) accounts for 15-20% of all breast cancer cases, yet is responsible for a disproportionately high percentage of breast cancer mortalities. Thus, there is an urgent need to identify novel biomarkers and therapeutic targets based on the molecular events driving TNBC pathobiology. Estrogen receptor beta (ERß) is known to elicit anti-cancer effects in TNBC, however its mechanisms of action remain elusive. Here, we report the expression profiles of ERß and its association with clinicopathological features and patient outcomes in the largest cohort of TNBC to date. In this cohort, ERß was expressed in approximately 18% of TNBCs, and expression of ERß was associated with favorable clinicopathological features, but correlated with different overall survival outcomes according to menopausal status. Mechanistically, ERß formed a co-repressor complex involving enhancer of zeste homologue 2/polycomb repressive complex 2 (EZH2/PRC2) that functioned to suppress oncogenic NFκB/RELA (p65) activity. Importantly, p65 was shown to be required for formation of this complex and for ERß-mediated suppression of TNBC. Our findings indicate that ERß+ tumors exhibit different characteristics compared to ERß- tumors and demonstrate that ERß functions as a molecular switch for EZH2, repurposing it for tumor suppressive activities and repression of oncogenic p65 signaling.

Krüppel-like factor 10 regulates the contractile properties of skeletal muscle fibers in mice.

INTRODUCTION/AIMS: Klf10 is a member of the Krüppel-like family of transcription factors, which is implicated in mediating muscle structure (fiber size, organization of the sarcomere), muscle metabolic activity (respiratory chain), and passive force. The aim of this study was to further characterize the roles of Klf10 in the contractile properties of skeletal muscle fibers. METHODS: Fifty-two single fibers were extracted from female wild-type (WT) and Klf10 knockout (KO) oxidative (soleus) and glycolytic (extensor digitorum longus [EDL]) skinned muscles. Each fiber was immersed successively in relaxing (R), washing (W), and activating (A) solutions. Calcium was included in the activating solution to induce a maximum contraction of the fiber. The maximum force (Fmax ) was measured and normalized to the cross-sectional area to obtain the maximum stress (Stressmax ). After a steady state in contraction was reached, a quick stretch-release was performed; the force at the maximum stretch (Fstretch ) was measured and the stiffness was assessed. RESULTS: Deletion of the Klf10 gene induced changes in the contractile parameters (Fmax , Stressmax , Stiffness), which were lower and higher for soleus and EDL fibers compared with littermates, respectively. These measurements also revealed changes in the proportion and resistance of attached cross-bridges. DISCUSSION: Klf10 plays a major role in the homeostasis of the contractile behavior of skeletal muscle fibers in a muscle fiber type-specific manner. These findings further implicate important roles for Klf10 in skeletal muscle function and shed new light on understanding the molecular processes regulating the contractility of skeletal muscle fibers.

Endoxifen, an Estrogen Receptor Targeted Therapy: From Bench to Bedside.

The selective estrogen receptor (ER) modulator, tamoxifen, is the only endocrine agent with approvals for both the prevention and treatment of premenopausal and postmenopausal estrogen-receptor positive breast cancer as well as for the treatment of male breast cancer. Endoxifen, a secondary metabolite resulting from CYP2D6-dependent biotransformation of the primary tamoxifen metabolite, N-desmethyltamoxifen (NDT), is a more potent antiestrogen than either NDT or the parent drug, tamoxifen. However, endoxifen's antitumor effects may be related to additional molecular mechanisms of action, apart from its effects on ER. In phase 1/2 clinical studies, the efficacy of Z-endoxifen, the active isomer of endoxifen, was evaluated in patients with endocrine-refractory metastatic breast cancer as well as in patients with gynecologic, desmoid, and hormone-receptor positive solid tumors, and demonstrated substantial oral bioavailability and promising antitumor activity. Apart from its potent anticancer effects, Z-endoxifen appears to result in similar or even greater bone agonistic effects while resulting in little or no endometrial proliferative effects compared with tamoxifen. In this review, we summarize the preclinical and clinical studies evaluating endoxifen in the context of breast and other solid tumors, the potential benefits of endoxifen in bone, as well as its emerging role as an antimanic agent in bipolar disorder. In total, the summarized body of literature provides compelling arguments for the ongoing development of Z-endoxifen as a novel drug for multiple indications.

Nicotinamide Mononucleotide Prevents Cisplatin-Induced Cognitive Impairments.

Chemotherapy-induced cognitive impairment (CICI) is often reported as a neurotoxic side effect of chemotherapy. Although CICI has emerged as a significant medical problem, meaningful treatments are not currently available due to a lack of mechanistic understanding underlying CICI pathophysiology. Using the platinum-based chemotherapy cisplatin as a model for CICI, we show here that cisplatin suppresses nicotinamide adenine dinucleotide (NAD+) levels in the adult female mouse brain in vivo and in human cortical neurons derived from induced pluripotent stem cells in vitro. Increasing NAD+ levels through nicotinamide mononucleotide (NMN) administration prevented cisplatin-induced abnormalities in neural progenitor proliferation, neuronal morphogenesis, and cognitive function without affecting tumor growth and antitumor efficacy of cisplatin. Mechanistically, cisplatin inhibited expression of the NAD+ biosynthesis rate-limiting enzyme nicotinamide phosphoribosyl transferase (Nampt). Selective restoration of Nampt expression in adult-born neurons was sufficient to prevent cisplatin-induced defects in dendrite morphogenesis and memory function. Taken together, our findings suggest that aberrant Nampt-mediated NAD+ metabolic pathways may be a key contributor in cisplatin-induced neurogenic impairments, thus causally leading to memory dysfunction. Therefore, increasing NAD+ levels could represent a promising and safe therapeutic strategy for cisplatin-related neurotoxicity. SIGNIFICANCE: Increasing NAD+ through NMN supplementation offers a potential therapeutic strategy to safely prevent cisplatin-induced cognitive impairments, thus providing hope for improved quality of life in cancer survivors. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/81/13/3727/F1.large.jpg.

Development and Characterization of Novel Endoxifen-Resistant Breast Cancer Cell Lines Highlight Numerous Differences from Tamoxifen-Resistant Models.

Despite the availability of drugs that target ERα-positive breast cancer, resistance commonly occurs, resulting in relapse, metastasis, and death. Tamoxifen remains the most commonly-prescribed endocrine therapy worldwide, and "tamoxifen resistance" has been extensively studied. However, little consideration has been given to the role of endoxifen, the most abundant active tamoxifen metabolite detected in patients, in driving resistance mechanisms. Endoxifen functions differently from the parent drug and other primary metabolites, including 4-hydroxy-tamoxifen (4HT). Many studies have shown that patients who extensively metabolize tamoxifen into endoxifen have superior outcomes relative to patients who do not, supporting a primary role for endoxifen in driving tamoxifen responses. Therefore, "tamoxifen resistance" may be better modeled by "endoxifen resistance" for some patients. Here, we report the development of novel endoxifen-resistant breast cancer cell lines and have extensively compared these models to 4HT and fulvestrant (ICI)-resistant models. Endoxifen-resistant cells were phenotypically and molecularly distinct from 4HT-resistant cells and more closely resembled ICI-resistant cells overall. Specifically, endoxifen resistance was associated with ERα and PR loss, estrogen insensitivity, unique gene signatures, and striking resistance to most FDA-approved second- and third-line therapies. Given these findings, and the importance of endoxifen in the efficacy of tamoxifen therapy, our data indicate that endoxifen-resistant models may be more clinically relevant than existing models and suggest that a better understanding of endoxifen resistance could substantially improve patient care. IMPLICATIONS: Here we report on the development and characterization of the first endoxifen-resistant models and demonstrate that endoxifen resistance may better model tamoxifen resistance in a subset of patients.

Ultrasound image processing to estimate the structural and functional properties of mouse skeletal muscle.

Noninvasive imaging techniques are increasingly used for monitoring muscle behavior in mice. However, muscle is a complex tissue that exhibits different properties under passive and active conditions. In addition to structural properties, it is also important to analyze functional characteristics. At present, such information can be obtained with ultrasound elastography. However, this technique is poorly used for small rodent models (mice and gerbils). Thus, this study aims at establish referent hindlimb muscle data, and experimental guidelines, for wild-type (WT) control mice as well as the TIEG1 knockout (KO) mouse model that is known to exhibit skeletal muscle defects. Ultrasound was performed with the Aixplorer machine using a SLH 20-6 linear transducer probe (2.8 cm footprint). A region of interest (ROI) was placed around a superficial group of muscles. Subsequently, from the B-mode image, a classification of all the muscles and ultrasound biomarkers such as echo intensity and texture anisotropy have been determined. The influence of the gain setting (from 40% to 70%) was analyzed on these parameters. Moreover, the elasticity (E) was also measured within the ROI. This study provides a suitable methodology for collecting experimental data: 1) the correct range of gain (between 50% and 70%) to apply for the ultrasound measurement of muscle structure, 2) the structural and functional referent data for a group of healthy muscles, 3) the gray scale index, the texture anisotropy and the elasticity (ETIEG1 KO = 36.1 ± 10.3 kPa, EWT = 44.4 ± 13.4 kPa) parameters, which were obtained for a group of muscles as a function of genotype.