Function Of Lysine-specific Demethylase 1 Research Articles

Lysine specific demethylase 1 is a molecular driver and therapeutic target in sarcoma.

Sarcomas are a diverse group of tumors with numerous oncogenic drivers, and display varied clinical behaviors and prognoses. This complexity makes diagnosis and the development of new and effective treatments challenging. An incomplete understanding of both cell of origin and the biological drivers of sarcomas complicates efforts to develop clinically relevant model systems and find new molecular targets. Notably, the histone lysine specific demethylase 1 (LSD1) is overexpressed in a number of different sarcomas and is a potential therapeutic target in these malignancies. With the ability to modify histone marks, LSD1 is a key player in many protein complexes that epigenetically regulate gene expression. It is a largely context dependent enzyme, having vastly different and often opposing roles depending on the cellular environment and which interaction partners are involved. LSD1 has been implicated in the development of many different types of cancer, but its role in bone and soft tissue sarcomas remains poorly understood. In this review, we compiled what is known about the LSD1 function in various sarcomas, to determine where knowledge is lacking and to find what theme emerge to characterize how LSD1 is a key molecular driver in bone and soft tissue sarcoma. We further discuss the current clinical landscape for the development of LSD1 inhibitors and where sarcomas have been included in early clinical trials.

Discovery of the antitumor activities of a potent DCN1 inhibitor compound 383 targeting LSD1 in gastric cancer

Dual target compounds have become a hot spot in the treatment of cancer in recent years. Histone lysine specific demethylase 1 (LSD1) is identified as histone demethylase and acts as a key regulator involved in many other cellular activities through its demethylation function. We have reported a triazolo [1,5-α] pyrimidine-based DCN1(defective in cullin neddylation protein 1) inhibitor compound 383 (IC50 = 11 nM) which could selectively inhibit Cullin 3/1 neddylation in MGC-803 cells. In this research, we investigated that compound 383 could target LSD1 and inhibit the biological function of LSD1 in MGC-803 cells (IC50 = 0.53 μM). We found that compound 383 could induce the degradation of LSD1 and inhibit MGC-803 cell proliferation, migration and invasion in a dose-dependent manner. Compound 383 could cause cell cycle arrest at G2/M phase by down-regulating the expression of LSD1. In addition, compound 383 could significantly reverse epithelial-mesenchymal transition (EMT) through increase H3K4me methylation at E-cadherin promotor. Furthermore, the in vivo inhibitory effect of compound 383 without obvious toxicity was confirmed in nude mouse transplanted MGC-803 tumor cells model. Collectively, these results suggest that the DCN1 inhibitor compound 383 exhibits antiproliferative activity in gastric cancer cells by targeting LSD1 which promotes compound 383 as a good starting point for the development of dual-target therapeutics for gastric cancer.

E239K mutation abolishes the suppressive effects of lysine-specific demethylase 1 on migration and invasion of MCF7 cells.

Lysine‐specific demethylase 1 (LSD1) is an important histone demethylase that mediates epithelial to mesenchymal transition (EMT). The E239K mutation of LSD1 was identified in a luminal breast cancer patient from the COSMIC Breast Cancer dataset. To investigate the functional effects of the E239K mutation of LSD1, a stable LSD1 knockdown MCF7 cell line was generated. Rescue with WT LSD1, but not E239K mutated LSD1, suppressed the invasion and migration of the LSD1 knockdown cells, indicating that the E239K mutation abolished the suppressive effects of LSD1 on the invasion and migration of MCF7 cells. Further analysis showed that the E239K mutation abolished LSD1‐mediated invasion and migration of MCF7 cells through downregulation of estrogen receptor α (ERα). Most importantly, the E239K mutation disrupted the interaction between LSD1 and GATA3, which reduced the enrichment of LSD1 at the promoter region of the ERα gene; the reduced enrichment of LSD1 at the promoter region of the ERα gene caused enhanced histone H3K9 methylation, which subsequently suppressed the transcription of the ERα gene. In summary, the E239K mutation abolishes the suppressive function of LSD1 on migration and invasion of breast cancer cells by disrupting the interaction between LSD1 and GATA3.

Chronic exposure to mild therapeutic hypothermia prevents cognitive deterioration and neurodegeneration in a rat model of AD‐like pathology

AbstractBackgroundMechanisms of epigenetic regulation modulate gene expression without alterations in the genomic sequence and play important roles in brain development1, synaptic plasticity and memory2. Epigenetic regulation of gene transcription would be an appropriate mechanism for cells to flexibly adapt to the environmental changes. In this context, lysine specific demethylase 1 (LSD1) is an important enzyme that modifies gene transcription3. Recently, it was reported that LSD1 is essential for the regulation of neuronal gene transcriptional programs involved in synaptic plasticity and cognition2–4. Despite, the well‐known function of LSD1 in early development1,4–6, there is some controversy about the specific role of LSD1 in aging. Upon cold exposure LSD1 regulates Brown adipose tissue function and mediates beige adipogenesis to increase thermogenic capacity and prevent changes in body temperature by inducing non‐shivering thermogenesis7. However, the role of repeat cold exposures on brain LSD1 function has never been studied. We investigated whether chronic repeat cold exposures could modulate the hippocampal expression of LSD1 and its potential impact on neurodegeneration and cognitive functions, using non‐transgenic (non‐tg) and transgenic (tg) AD‐like rat model.MethodMcGill‐R‐Thy1‐APP‐tg rats exhibited accumulation of intraneuronal Aβ peptide, display cognitive deficits and reproduce an AD‐like amyloid pathology. Thus,17 months tg and non‐tg rats were expose under therapeutic hypothermia for 2 hours at 4°C daily during 1 month.ResultOur results demonstrated how progressive intraneuronal Aβ accumulation correlates with upregulation of LSD1 at early stages (prior plaques formation) of the AD‐like amyloid pathology, which impact on the alteration of cognitive functions. We showed that chronic mild therapeutic hypothermia modulates hippocampal LSD1 levels, improving synaptic plasticity and preventing cognitive dysfunction.ConclusionTaken together our work suggests that modulating brain LSD1 levels through repeat cold exposures constitute a novel non‐pharmacological therapeutic strategy to prevent neurodegeneration and preserve cognitive functions. (1) Toffolo E, et al. J Neurochem. 2014; (2) Wang J, et al. Nat Neurosci. 2016; (3) Lim CS, Nam HJ, et al. Sci Rep. 201 7; (4) Rusconi F, et al. Trends Neurosci. 2017; (5) Maes T, et al. Epigenomics. 2015; (6) Ciceri F, et al. Curr Pharm Des. 2017; (7) Marcher AB, et al. Cell Rep. 2015.

MicroRNA-155 Participates in the Expression of LSD1 and Proinflammatory Cytokines in Rheumatoid Synovial Cells.

MicroRNA-155 (miRNA-155) is abundant in fibroblast-like synoviocytes (FLS) in rheumatoid arthritis (RA). Lysine-specific demethylase 1 (LSD1) has been found that it can ameliorate the severity of RA. Tumor necrosis factor-alpha, interleukin-1 beta, and interleukin-6 are key proinflammatory cytokines implicated in the pathogenesis of RA. In our study, we investigated whether miRNA-155 participates in the expression of LSD1 and proinflammatory cytokines in rheumatoid synovial cells. First of all, flow cytometry and cell counting kit-8 analysis were employed to explore the apoptosis and proliferation of FLS, respectively. Subsequently, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was applied to probe into the level of miRNA-155 in FLS when stimulated by miRNA-155 molecules. Moreover, RT-qPCR was used to explore the relative LSD1 miRNA expression in FLS when stimulated by miRNA-155 molecules, and Western blot and immunofluorescence assay were applied to probe into the expression level of LSD1. Finally, enzyme-linked immunosorbent assay was employed to analyze the secreting level of proinflammatory cytokines in FLS when stimulated by miRNA-155 molecules. RA-FLS showed a higher apoptosis rate than normal FLS. The cell proliferation of both HFLS and MH7A cells was promoted by miRNA-155 upregulation. Meanwhile, the expression of LSD1 and proinflammatory cytokines in the FLS of RA was also changed by miRNA-155 regulation. In conclusion, miRNA-155 participates in the expression of LSD1 and proinflammatory cytokines in rheumatoid synovial cells. These findings imply a potential function and interaction of miRNA-155 and LSD1.

LSD1 negatively regulates autophagy in myoblast cells by driving PTEN degradation

Lysine-specific demethylase 1 (LSD1) is a well characterized transcriptional regulator functioning on the chromatin to remove mono- and di-methyl groups from lysine 4 or lysine 9 of histone 3 (H3K4 or H3K9). LSD1 also has non-transcriptional activities via targeting non-histone substrates that participate in diverse biological processes. In this report, we determined that LSD1 negatively regulates autophagy in skeletal muscle cells by promoting PTEN degradation in a transcription-independent mechanism. In C2C12 cells, LSD1 inhibition or depletion significantly induced the initiation of autophagy; and autophagy resulted from LSD1 inhibition is associated with AKT/mTORC1 inactivation. Notably, the proteins of PTEN, a prominent repressive AKT modulator, are stabilized by LSD1 inhibition despite a decrease of its mRNA levels. Further data demonstrated that LSD1 interacts with PTEN protein and enhances its ubiquitination and degradation. Together, our findings identify a novel biological function of LSD1 in autophagy, mediated by regulating the stability of PTEN and the activity of AKT/mTORC1.

Abstract LB-A14: Combinational therapy by targeting LSD1 and BRD4 in prostate cancer

Abstract As a nuclear transcriptional factor, androgen receptor (AR) plays a significant role in prostate cancer (PCa) development. Androgen deprivation therapy (ADT) blunting the AR activity is the mainstream strategy for treating PCa. Despite the initial effectiveness of ADTs, most cases will relapse to a lethal stage termed castration-resistant prostate cancer (CRPC), where AR regains activity and drives tumor growth. This unmet clinical challenge to further target AR has led to a number of clinical investigations of targeting some of the key components in AR signaling. Epigenetic regulators are promising targets as they are critical players for rewiring the AR network during PCa progression. Lysine specific demethylase 1 (LSD1), an important epigenetic modifier that is well known for its repressor function through demethylating histone 3 lysine 4 (H3K4), can also function as an activator. In PCa cells, our epigenomic profiling study showed that LSD1 resides at active chromatin regions marked by high H3K27ac and FOXA1. Further genome-wide studies showed that inhibiting LSD1 disrupts AR cistrome and impairs its transcriptional output. Our mechanistic studies show that the activator function of LSD1 is through controlling the chromatin binding of FOXA1, which is a critical pioneer factor for AR. Importantly, at the AR-regulated enhancers, we observed that LSD1 inhibition also caused a rapid and broad decrease of H3K27ac signal, which directs Bromodomain-containing protein 4 (BRD4) chromatin binding. BRD4 belongs to BET (bromodomain and extra terminal domain) family, which plays an essential role in promoting the initiation and elongation steps in gene transcription process. The most credited function of BRD4 is its regulation of oncogene c-MYC via binding to the highly active super enhancers. Moreover, the oncogenic function of BRD4 in PCa has been well attributed to its coactivator role with AR. Multiple inhibitors targeting BRD4 have been developed and shown early promising clinical outcomes, however the drug resistance also emerges in the preclinical models. Herein, we reason that targeting LSD1 and BRD4 together could further impair the AR action and achieve better anti-tumor effect. To examine whether there is a synergistic effect, we employed inhibitors that have been tested in clinical trials: ORY-1001 and I-BET762, which target LSD1 and BRD4 respectively. We found that the combinational treatment lead to a more effective reduction of AR transcriptional activity than either single treatment. The proliferation of AR-positive LNCaP PCa cell line and CWR22RV1 CRPC cell line which expresses high levels of C-terminal truncated AR splicing variants (AR-Vs) was also more profoundly inhibited by combined therapy. Besides disrupting the full-length AR cistrome, LSD1 inhibition also impairs the activity of AR-V7, a major form of AR-Vs that drives developing ADT resistance in CRPC. However, the function of BRD4 in regulating AR-V7 activity remains to be elucidated. Therefore, we tested the combined treatment in CWR22RV1 cells under hormone-depleted condition. By targeting LSD1 and BRD4 together, AR-V7 regulated gene expression was significantly reduced. Currently, we are investigating the genome-wide effect of targeting both epigenetic regulators on AR and AR-V7. To provide clinical evidence, we are testing this combined treatment in patient-derived xenograft models, which recapitulate the CRPC by harboring high levels of AR and AR-Vs and ETS gene translocation. Citation Format: Muqing Li, Dong Han, zifeng wang, shuai gao, Changmeng Cai. Combinational therapy by targeting LSD1 and BRD4 in prostate cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr LB-A14. doi:10.1158/1535-7163.TARG-19-LB-A14

Dissecting LSD1‐Dependent Neuronal Maturation in the Olfactory Epithelium

AbstractNeurons in the olfactory epithelium (OE) each express a single dominant olfactory receptor (OR) allele from among roughly 1,000 different OR genes. While monogenic and monoallelic OR expression has been appreciated for over two decades, regulators of this process are still being described; most recently, epigenetic modifiers have been of high interest as silent OR genes are decorated with transcriptionally repressive trimethylated histone 3 lysine 9 (H3K9me3) whereas active OR genes are decorated with transcriptionally activating trimethylated histone 3 lysine 4 (H3K4me3). The lysine specific demethylase 1 (LSD1) demethylates at both of these lysine residues and has been shown to disrupt neuronal maturation and OR expression in the developing embryonic OE. Despite the growing literature on LSD1 expression in the OE, a complete characterization of the timing of LSD1 expression relative to neuronal maturation and of the function of LSD1 in the adult OE have yet to be reported. To fill this gap, the present study determined that LSD1 (1) is expressed in early dividing cells before OR expression and neuronal maturation and decreases at the time of OR stabilization; (2) colocalizes with the repressor CoREST (also known as RCOR1) and histone deacetylase 2 in these early dividing cells; and (3) is required for neuronal maturation during a distinct time window between activating reserve stem cells (horizontal basal cells) and Neurogenin1 (+) immediate neuronal precursors. Thus, this study clarifies the role of LSD1 in olfactory neuronal maturation.

LSD1 modulates the non-canonical integrin \u03b23 signaling pathway in non-small cell lung carcinoma cells

The epigenetic writer lysine-specific demethylase 1 (LSD1) is aberrantly upregulated in many cancer types and its overexpression correlates with poor survival and tumor progression. In this study, we analysed LSD1 function in non-small cell lung cancer adenocarcinomas. Expression profiling of 182 cases of lung adenocarcinoma proved a significant correlation of LSD1 overexpression with lung adenocarcinoma progression and metastasis. KRAS-mutated lung cancer cell clones were stably silenced for LSD1 expression. RNA-seq and comprehensive pathway analysis revealed, that genes related to a recently described non-canonical integrin β3 pathway, were significantly downregulated by LSD1 silencing. Hence, invasion and self-renewal capabilities were strongly decreased. Notably, this novel defined LSD1/integrin β3 axis, was also detected in human lung adenocarcinoma specimens. Furthermore, the linkage of LSD1 to an altered expression pattern of lung-lineage specific transcription factors and genes, which are involved in alveolar epithelial differentiation, was demonstrated. Thus, our findings point to a LSD1-integrin β3 axis, conferring attributes of invasiveness and tumor progression to lung adenocarcinoma.

Dissecting LSD1‐Dependent Neuronal Maturation in the Olfactory Epithelium

Neurons in the olfactory epithelium (OE) each express a single dominant olfactory receptor (OR) allele from among roughly 1,000 different OR genes. While monogenic and monoallelic OR expression has been appreciated for over two decades, regulators of this process are still being described; most recently, epigenetic modifiers have been of high interest as silent OR genes are decorated with transcriptionally repressive trimethylated histone 3 lysine 9 (H3K9me3) whereas active OR genes are decorated with transcriptionally activating trimethylated histone 3 lysine 4 (H3K4me3). The lysine specific demethylase 1 (LSD1) demethylates at both of these lysine residues and has been shown to disrupt neuronal maturation and OR expression in the developing embryonic OE. Despite the growing literature on LSD1 expression in the OE, a complete characterization of the timing of LSD1 expression relative to neuronal maturation and of the function of LSD1 in the adult OE have yet to be reported. To fill this gap, the present study determined that LSD1 (1) is expressed in early dividing cells before OR expression and neuronal maturation and decreases at the time of OR stabilization; (2) colocalizes with the repressor CoREST (also known as RCOR1) and histone deacetylase 2 in these early dividing cells; and (3) is required for neuronal maturation during a distinct time window between activating reserve stem cells (horizontal basal cells) and Neurogenin1 (+) immediate neuronal precursors. Thus, this study clarifies the role of LSD1 in olfactory neuronal maturation.

Abstract 4704: The evaluation of INCB059872, an FAD-directed inhibitor of LSD1, in preclinical models of human small cell lung cancer

Abstract Methylated histone marks on H3K4 and H3K9 are generally coupled with transcriptional activation and repression, respectively. Altered levels of these histone methylation marks lead to abnormal gene expression and are associated with oncogenesis. Lysine specific demethylase 1 (LSD1) catalyzes demethylation of mono and di-methylated lysine 4 and 9 of histone H3 via an FAD-dependent redox-process. Deregulated LSD1 activity perturbs normal gene expression and can lead to cellular transformation. In particular, the function of LSD1 has been reported to maintain stem cell-like gene expression patterns in various cancers, most notably small cell lung cancer (SCLC), a cancer type that is characterized by a highly undifferentiated state and with high unmet medical need. INCB059872 is a potent, selective and orally bioavailable inhibitor of LSD1 that achieves inhibitory activity through the formation of covalent FAD-adducts. The effect of INCB059872 in SCLC cell lines was assessed in vitro and in vivo. In these studies, the proliferation of a panel of SCLC cell lines was inhibited by INCB059872, with EC50 values ranging from 47 to 377 nM. Non-tumorigenic cells, such as IL-2 stimulated T cells from normal donors, by contrast, were significantly less sensitive with IC50 values > 10 μM. Oral administration of INCB059872 on once daily (QD) and alternative day (QoD) dosing regimens inhibited tumor growth in the NCI-H526 and NCI-H1417 human SCLC xenograft models. Consistent with previous reports, INCB059872 treatment in these models led to the induction of the FEZ1 and UMODL1 genes relative to vehicle-treated animals, as part of a gene signature in SCLC cell lines that is predictive of LSD1 responsiveness. Moreover, serum levels of the neuroendocrine marker pro-GRP were markedly reduced at all efficacious dosing regimens in the NCI-H1417 human SCLC xenograft model, suggesting that serum pro-GRP levels could be used as a surrogate pharmacodynamic marker of LSD1 inhibition. Currently, the efficacy of INCB059872 in combination with standard of care therapies for SCLC is under evaluation in these preclinical models. These data support the clinical evaluation of INCB059872 in patients with SCLC. Citation Format: Sang Hyun Lee, Xuesong Mike Liu, Melody Diamond, Valerie Dostalik, Margaret Favata, Chunhong He, Liangxing Wu, Richard Wynn, Wenqing Yao, Gregory Hollis, Reid Huber, Peggy Scherle, Bruce Ruggeri. The evaluation of INCB059872, an FAD-directed inhibitor of LSD1, in preclinical models of human small cell lung cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4704.

LSD1 Interacts with Zfp516 to Promote UCP1 Transcription and Brown Fat Program.

Zfp516, a brown fat (BAT)-enriched and cold-inducible transcription factor, promotes transcription of UCP1 and other BAT-enriched genes for non-shivering thermogenesis. Here, we identify lysine-specific demethylase 1 (LSD1) as a direct binding partner of Zfp516. We show that, through interaction with Zfp516, LSD1 is recruited to UCP1 and other BAT-enrichedgenes, such as PGC1α, to function as a coactivator by demethylating H3K9. We also show that LSD1 is induced during brown adipogenesis and that LSD1 and its demethylase activity is required for the BAT program. Furthermore, we show that LSD1 ablation in mice using Myf5-Cre alters embryonic BAT development. Moreover, BAT-specific deletion of LSD1 via the use of UCP1-Cre impairs the BAT program and BAT development, making BAT resemble WAT, reducing thermogenic activity and promoting obesity. Finally, we demonstrate an invivo requirement of the Zfp516-LSD1 interaction for LSD1 function in BAT gene activation.

New insight into LSD1 function in human cortical neurogenesis.

The cerebral cortex of primates has evolved massively and intricately in comparison to that of other species. Accumulating evidence indicates that this is caused by changes in cell biological features of neural stem cells (NSCs), which differentiate into neurons and glial cells during development. The fate of NSCs during rodent cortical development is stringently regulated by epigenetic factors, such as histone modification enzymes, but the role of these factors in human corticogenesis is largely unknown. We have recently discovered that a lysine-specific demethylase 1 (LSD1), which catalyzes the demethylation of methyl groups in the histone tail, plays a unique role in human fetal NSCs (hfNSCs). We show that, unlike the role previously reported in mice, LSD1 in hfNSCs is necessary for neuronal differentiation and controls the expression of HEYL, one of the NOTCH target genes, by modulating the methylation level of histones on its promoter region. Interestingly, LSD1-regulation of Heyl expression is not observed in mouse NSCs. Furthermore, we first demonstrated that HEYL is able to maintain the undifferentiated state of hfNSCs. Our findings provide a new insight indicating that LSD1 may be a key player in the development and characterization of the evolved cerebral cortex.

Lysine‐specific demethylase‐1 contributes to malignant behavior by regulation of invasive activity and metabolic shift in esophageal cancer

Lysine-specific demethylase-1 (LSD1) removes the methyl groups from mono- and di-methylated lysine 4 of histone H3. Previous studies have linked LSD1 to malignancy in several human tumors, and LSD1 is considered to epigenetically regulate the energy metabolism genes in adipocytes and hepatocellular carcinoma. This study investigates the function of LSD1 in the invasive activity and the metabolism of esophageal cancer cells. We investigated whether LSD1 immunohistochemical expression levels are related to clinical and pathological features, including the maximum standard uptake value in fluorodeoxyglucose positron emission tomography assay. The influence of LSD1 on cell proliferation, invasion and glucose uptake was evaluated in vitro by using specific small interfering RNA for LSD1, and an LSD1 inhibitor. We also evaluated two major energy pathways (glycolytic pathway and mitochondrial respiration) by measuring the extracellular acidification rate (ECAR) and the oxygen consumption rate (OCR) with an extracellular flux analyzer. High LSD1 immunohistochemical expression was significantly associated with high tumor stage, lymphovascular invasion, poor prognosis, and high maximum standard uptake value in esophageal cancer patients. In the in vitro analysis, LSD1 knockdown significantly suppressed the invasive activity and glucose uptake of cancerous cells, reduced their ECAR and increased their OCR and OCR/ECAR. LSD1 may contribute to malignant behavior by regulating the invasive activity and metabolism, activating the glycolytic pathway and inhibiting the mitochondrial respiration of esophageal cancer cells. The results support LSD1 as a potential therapeutic target.

Abstract 966: LSD1 functions as a global androgen receptor coactivator and is a therapeutic target in prostate cancer

Abstract Androgen receptor (AR) plays a pivotal role in prostate cancer (PCa) and remains critical for more aggressive castration-resistant PCa (CRPC) through transactivation of multiple genes. Although CRPC is generally responsive to further CYP17A1 inhibition (abiraterone) or more potent AR antagonist (enzalutamide) treatments, patients treated with these agents still relapse within 1-2 years, and high levels of AR and AR regulated genes in many of these relapsed tumors indicate that AR activity has again been restored. Therefore, there remains a need for further novel AR targeted therapies. Initially identified as a component in the REST complex that functions as a transcriptional suppressor, Lysine Specific Demethylase 1 (LSD1) removes the methyl group from histone 3 lysine 4 (H3K4me1 and 2), and turns off the marks of active transcription. Our recent genome-wide integrated analysis of LSD1 and AR chromatin bindings and gene expression profiles indicates that LSD1 is broadly associated with AR regulated enhancers, and LSD1 functions as a coactivator on AR-stimulated genes through demethylating novel histone or nonhistone substrates in addition to H3K4me. LSD1 is also associated with FOXA1 at AR regulated enhancer sites, and an LSD1-FOXA1 interaction enhances binding of both proteins at these sites. These results clearly demonstrated LSD1 as a therapeutic target in prostate cancer. We then accessed if LSD1 enzymatic activity is required for its global AR coactivator function. AR transcription activity is significantly impaired in cells overexpressing a catalytic-deficient LSD1 mutant. In addition, this mutant also decreased FOXA1 binding on AR-regulated enhancers. We then selected of a panel of LSD1 inhibitors (a few of them have been used in clinical trials for lung cancer and leukemia) to assess the effects on AR activity and PCa cell growth. Inhibition of LSD1 consistently impaired AR activity through abrogating acetylated H3K27 at AR-regulated enhancers. LSD1 inhibitor also decreased FOXA1 binding prior to androgen stimulation. While most of inhibitors showed relatively low efficacy in PCa cells (>50μM), LSD1-C12 is highly potent and specific at μM range to dramatically decrease AR activity and PCa cell growth. Overall, our study suggests that the AR coactivator function of LSD1 depends on its enzymatic activity and targeting LSD1 with more specific and potent inhibitors is a promising therapeutic approach to treat PCa. Furthermore, understanding the mechanism of LSD1 coactivator function will potentially advance PCa therapy by targeting LSD1. Citation Format: Shuai Gao, Yanfei Gao, Hansen He, Myles Brown, Steve Balk, Changmeng Cai. LSD1 functions as a global androgen receptor coactivator and is a therapeutic target in prostate cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 966. doi:10.1158/1538-7445.AM2015-966

A Systematic Review of Histone Lysine‐Specific Demethylase 1 and Its Inhibitors

Histone lysine-specific demethylase 1 (LSD1) is the first discovered and reported histone demethylase by Dr. Shi Yang's group in 2004. It is classified as a member of amine oxidase superfamily, the common feature of which is using the flavin adenine dinucleotide (FAD) as its cofactor. Since it is located in cell nucleus and acts as a histone methylation eraser, LSD1 specifically removes mono- or dimethylated histone H3 lysine 4 (H3K4) and H3 lysine 9 (H3K9) through formaldehyde-generating oxidation. It has been indicated that LSD1 and its downstream targets are involved in a wide range of biological courses, including embryonic development and tumor-cell growth and metastasis. LSD1 has been reported to be overexpressed in variety of tumors. Inactivating LSD1 or downregulating its expression inhibits cancer-cell development. LSD1 targeting inhibitors may represent a new insight in anticancer drug discovery. This review summarizes recent studies about LSD1 and mainly focuses on the basic physiological function of LSD1 and its involved mechanisms in pathophysiologic conditions, as well as the development of LSD1 inhibitors as potential anticancer therapeutic agents.

Abstract 1361: The clinicopathological significance of LSD1 in esophageal cancers

Abstract Background: Lysine-specific demethylase 1 (LSD1) is the first discovered histone demethylase which specially removes H3K4me1/2 through a flavin-adenine-dinucleotide (FAD). Previous studies have indicated that LSD1 is associated with cell proliferation, cell invasion and epithelial-mesenchymal transition in human malignant tumors. Recently, it is accepted that LSD1 epigenetically regulates energy-expenditure genes in adipocytes depending on the cellular FAD availability. As a PET scan with a FDG tracer can detect cancers, the method is exceedingly useful for tumor staging or therapeutic effect determination of esophageal cancers. The aim of this study is to investigate the expression and function of LSD1 in esophageal cancer cell-lines and, to analyze the relationships between the LSD1 expression and clinicopathological data including SUVmax (standardized uptake value). Methods: The influence of LSD1 on cell proliferation and invasion was evaluated by knockdown experiments. Using 113 resected esophageal cancer specimens, we evaluated the LSD1 expression by immunohistochemistry and devided them into two groups; a low expression group (LEG) and a high expression group (HEG). We compared these two groups and analyzed the relationships between LSD1 expression and clinicopathological data. Results: In vitro analysis, knockdown of LSD1 did not impact cell proliferation, but suppressed cell invasion. In the immunohistochemical study, the HEG's age was significantly higher (P=0.021), and this group had esophageal cancers at lower locations (P=0.058). HEG was significantly correlated with the T-stage (P=0.020), lymph and blood vessels invasion (P=0.020, P=0.001), and a higher SUVmax (P=0.0088) and lower overall survival (P=0.0017). Conclusions: Our study demonstrated the relationships between the expression of LSD1 and T-stage, lymph and blood vessels invasion, and the SUVmax in esophageal cancers. It is suggested that LSD1 has a potential to impact the malignancy of the esophageal cancer, and may be a therapeutic target in patients with esophageal cancers. Citation Format: Keisuke Kosumi, Yoshifumi Baba, Ryuichi Karashima, Satoshi Ida, Yu Imamura, Takatsugu Ishimoto, Shiro Iwagami, Yasuo Sakamoto, Yuji Miyamoto, Naoya Yoshida, Akihisa Sakamoto, Shinjiro Hino, Mitsuyoshi Nakao, Hideo Baba. The clinicopathological significance of LSD1 in esophageal cancers. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1361. doi:10.1158/1538-7445.AM2014-1361

Antagonistic actions of Rcor proteins regulate LSD1 activity and cellular differentiation.

Lysine-specific demethylase 1 (LSD1) demethylates nucleosomal histone H3 lysine 4 (H3K4) residues in collaboration with the corepressor CoREST/REST corepressor 1 (Rcor1) and regulates cell fates by epigenetically repressing gene targets. The balanced regulation of this demethylase, if any, is however unknown. We now demonstrate the actions of two other Rcor paralogs, Rcor2 and Rcor3, in regulating LSD1 enzymatic activity and biological function in hematopoietic cells. All three Rcor proteins interact with LSD1 and with the erythro-megakaryocytic transcription factor growth factor independence (Gfi)1b; however, whereas Rcor2, like Rcor1, facilitates LSD1-mediated nucleosomal demethylation, Rcor3 competitively inhibits this process. Appending the SANT2 domain of Rcor1 to Rcor3 confers the ability to facilitate LSD1-mediated demethylation on the chimeric Rcor protein. Consistent with their biochemical activities, endogenous Rcor1, Rcor2, and LSD1 promote differentiation, whereas Rcor3 opposes these processes. Recruitment of Rcor3 to cognate gene targets by Gfi1b and LSD1 leads to inhibition of H3K4 demethylation of chromatin and transcriptional derepression of these loci. Remarkably, profound alterations in Rcor1/3 levels during erythroid versus megakaryocytic differentiation potentiate antagonistic outcomes. In mature erythroid cells, a strong upsurge in Rcor3 and a sharp decline in Rcor1 levels counteract LSD1/Rcor1/2-mediated differentiation. In contrast, the opposite changes in Rcor1/3 levels in megakaryocytes favor differentiation and likely maintain homeostasis between these lineages. Overall, our results identify Rcor3 as a natural inhibitor of LSD1 and highlight a dual mechanism of regulating the enzymatic activity and restraining the epigenetic impact of this robust demethylase during hematopoietic differentiation.

Abstract 1805: LSD1 regulation of SKP2 expression in prostate cancer.

Abstract Prostate carcinogenesis is a complicated process involving both genetic and epigenetic changes. Aberrant expression of LSD1 (Lysine-specific demethylase 1) has been reported in human prostate cancers. Although high expression of LSD1 is implicated in prostate tumorigenesis, the underlying molecular mechanisms are not fully understood. Here we reported that LSD1 promotes proliferation and migration of prostate cancer cells via up-regulation of SKP2 expression. Knockdown of LSD1 using small interfering RNA (siRNA) decreases SKP2 expression and inhibits prostate cancer cell proliferation and migration. Promoter luciferase reporter and chromatin immunoprecipitation (ChIP) experiments show that LSD1 regulates SKP2 expression by controlling the levels of H3K4 methylation in SKP2 promoter and this function of LSD1 is mediated through its demethylase activity. As demonstrated by immunohistochemistry, LSD1 and SKP2 proteins positively correlate with each other and their expression is associated with cell proliferation in human prostate cancer tissues. Our data not only identify SKP2 as a direct regulated target gene of LSD1 but also reveal that SKP2 plays an important role in contributing LSD1 mediated prostate cancer cell proliferation and migration, suggesting that the LSD1-SKP2 axis can serve as therapeutic targets for treatment of prostate cancers. Citation Format: Liya Ding, Haojie Huang. LSD1 regulation of SKP2 expression in prostate cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1805. doi:10.1158/1538-7445.AM2013-1805

Abstract 673: Targeting LSD1-HDACs crosstalk as a potential therapeutic strategy for triple negative breast cancer cells.

Abstract Triple negative breast cancer (TNBC) is the most aggressive type of breast tumor and one of the major clinical hurdles encountered in the development of specific treatment for this disease is lack of effective targeted therapy. Our recent studies demonstrate that lysine-specific demethylase 1 (LSD1) and histone deacetylases (HDACs) closely interact in controlling gene expression and growth of TNBC cells. However the underlying mechanisms are largely unknown. Here we show that knockdown of LSD1 expression (LSD1-KD) via RNA interference (RNAi) inhibits mRNA levels of most HDAC isozymes in a TNBC cell line, MDA-MB-231. HDAC5-targeted RNAi induces the most significant accumulation of H3K4me2, a specific substrate of LSD1, implicating a necessity for HDAC5 in LSD1 function. Combined treatment with LSD1 inhibitor, pargyline, and HDAC inhibitor, vorinostat, leads to superior growth inhibition and apoptosis in TNBC cells, but exhibits additive or antagonistic effect on growth inhibition in non-TNBC counterparts as well as non-tumorigenic mammary epithelial cells. Moreover, LSD1-KD augments vorinostat-induced re-expression of a subset of aberrantly silenced genes such as NR4A1, PCDH1, RGS16, BIK, and E-Cadherin whose re-expression may be tumor suppressive in TNBC cells, but not in other breast cancer subtypes. The re-expression of these genes is associated with increased H3K4me2 and acetyl-H3K9 marks at gene promoters, indicative of active chromatin. Integrative analysis of a panel of 36 human breast cancer cell lines of distinct subtypes shows that the expression of these genes is more profoundly suppressed in TNBC cells compared with other breast cancer subtypes. Genome-wide microarray study in MDA-MB-231 cells identifies a group of tumor suppressor genes (CDKN1A, CDKN1C, CRABP2, TNFAIP3, TP53TG1 and ING1) whose expression is induced by vorinostat and significantly enhanced by LSD1-KD. We also show that concurrent depletion of RGS16 by siRNA reduces overall cytotoxicity of vorinostat and blocks the re-expression of E-Cadherin, CDKN1C and ING1 in LSD1 deficient MDA-MB-231 cells. Furthermore, co-treatment with RGS16 siRNA reverses the down-regulation of NF-KB expression induced by combined inhibition of LSD1 and HDACs, suggesting a crucial role of RGS16 in controlling key apoptotic pathways in response to combination therapy in TNBC cells. Taken together, these results provide novel mechanistic insight into how LSD1 and HDACs modulate each other to enhance therapeutic efficacy of vorinostat in TNBC; and also suggest that inhibition of LSD1-HDACs axis-mediated histone modifications can be a potential therapeutic strategy to improve treatment of TNBC. This work is funded by: Breast Cancer Research Foundation, UPCI CCSG Shared Facilities & the Samuel and Emma Winters Foundation. Citation Format: Shauna Vasilatos, Tiffany A. Katz, Steffi Oesterreich, Nancy E. Davidson, Yi Huang. Targeting LSD1-HDACs crosstalk as a potential therapeutic strategy for triple negative breast cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 673. doi:10.1158/1538-7445.AM2013-673