LSD1 in drug discovery: From biological function to clinical application.

Lysine-specific demethylase 1 (LSD1) is a flavin adenine dinucleotide (FAD) dependent monoamine oxidase (MAO) that erases the mono-, and dimethylation of histone 3 lysine 4 (H3K4), resulting in the suppression of target gene transcriptions. Besides, it can also demethylate some nonhistone substrates to regulate their biological functions. As reported, LSD1 is widely upregulated and plays a key role in several kinds of cancers, pharmacological or genetic ablation of LSD1 in cancer cells suppresses cell aggressiveness by several distinct mechanisms. Therefore, numerous LSD1 inhibitors, including covalent and noncovalent, have been developed and several of them have entered clinical trials. Herein, we systemically reviewed and discussed the biological function of LSD1 in tumors, lymphocytes as well as LSD1-targeting inhibitors in clinical trials, hoping to benefit the field of LSD1 and its inhibitors.

New drug approvals for 2022: Synthesis and clinical applications.

The U.S. Food and Drug Administration has approved a total of 37 new drugs in 2022, which are composed of 20 chemical entities and 17 biologics. In particular, 20 chemical entities, including 17 small molecule drugs, 1 radiotherapy, and 2 diagnostic agents, provide privileged scaffolds, breakthrough clinical benefits, and a new mechanism of action for the discovery of more potent clinical candidates. The structure-based drug development with clear targets and fragment-based drug development with privileged scaffolds have always been the important modules in the field of drug discovery, which could easily bypass the patent protection and bring about improved biological activity. Therefore, we summarized the relevant valuable information about clinical application, mechanism of action, and chemical synthesis of 17 newly approved small molecule drugs in 2022. We hope this timely and comprehensive review could bring about creative and elegant inspiration on the synthetic methodologies and mechanism of action for the discovery of new drugs with novel chemical scaffolds and extended clinical indications.

LSD1: an emerging face in altering the tumor microenvironment and enhancing immune checkpoint therapy.

Dysregulation of various cells in the tumor microenvironment (TME) causes immunosuppressive functions and aggressive tumor growth. In combination with immune checkpoint blockade (ICB), epigenetic modification-targeted drugs are emerging as attractive cancer treatments. Lysine-specific demethylase 1 (LSD1) is a protein that modifies histone and non-histone proteins and is known to influence a wide variety of physiological processes. The dysfunction of LSD1 contributes to poor prognosis, poor patient survival, drug resistance, immunosuppression, etc., making it a potential epigenetic target for cancer therapy. This review examines how LSD1 modulates different cell behavior in TME and emphasizes the potential use of LSD1 inhibitors in combination with ICB therapy for future cancer research studies.

Myofibroblast Deficiency of LSD1 Alleviates TAC-Induced Heart Failure.

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Lysine demethylase LSD1 delivered via small extracellular vesicles promotes gastric cancer cell stemness.

Several studies have examined the functions of nucleic acids in small extracellular vesicles (sEVs). However, much less is known about the protein cargos of sEVs and their functions in recipient cells. This study demonstrates the presence of lysine-specific demethylase 1 (LSD1), which is the first identified histone demethylase, in the culture medium of gastric cancer cells. We show that sEVs derived from gastric cancer cells and the plasma of patients with gastric cancer harbor LSD1. The shuttling of LSD1-containing sEVs from donor cells to recipient gastric cancer cells promotes cancer cell stemness by positively regulating the expression of Nanog, OCT4, SOX2, and CD44. Additionally, sEV-delivered LSD1 suppresses oxaliplatin response of recipient cells in vitro and in vivo, whereas LSD1-depleted sEVs do not. Taken together, we demonstrate that LSD1-loaded sEVs can promote stemness and chemoresistance to oxaliplatin. These findings suggest that the LSD1 content of sEV could serve as a biomarker to predict oxaliplatin response in gastric cancer patients.

Inhibition of lysine-specific demethylase 1 enhances the sensitivity of the chemotherapeutic drug doxorubicin in gastric cancer cell.

AIM: Lysine-Specific Demethylase 1 (LSD1) inhibitors have been developed and reached the clinic, but its effect in combination with cytotoxic chemotherapy is unclear. Here, we investigated the anti-tumor effect of LSD1 inhibitor GSK-LSD1 and its anti-tumor effect with the DNA damage drug doxorubicin (DOX) in gastric cancer (GC) cells. METHODS: Cells were treated with different concentrations of GSK-LSD1 to examine the anti-tumor effect versus cell viability by MTT and cell cycle arrest by flow cytometry. To explore whether LSD1 inhibitors can increase the anti-tumor effect of DNA damage drugs, cells were treated with DOX for 48 h after pretreatment with GSK-LSD1 for 48 h. Cell viability was detected by MTT and apoptosis-related proteins were examined by Western blot. Furthermore, anti-tumor efficacy of combination GSK-LSD1 with DOX was also measured in MGC-803 xenografts model in nude mice. RESULTS: The results showed that LSD1 was highly expressed in GC cell lines. Inhibition of LSD1 has a weak effect on cell viability and cell cycle. Moreover, LSD1 inhibitors pretreatment could significantly increase the anti-tumor effect of DOX. Further study found that inhibition of LSD1 can significantly enhance DOX-induced the apoptosis, accompanied by down-regulation of antiapoptotic Bcl-2 expression and up-regulation of proapoptotic Bax expression. We also confirmed that inhibition of LSD1 can sensitize the anti-tumor effect of DOX in vivo. CONCLUSION: Our findings suggest that the LSD1 inhibitor GSK-LSD1 has a weak inhibitory effect on the viability and cell cycle of GC cells, but can enhance the sensitivity of DOX.

Generation, secretion and degradation of cancer immunotherapy target PD-L1.

Cancer immunotherapy is a rapidly developing and effective method for the treatment of a variety of malignancies in recent years. As a significant immune checkpoint, programmed cell death 1 ligand 1 (PD-L1) and its receptor programmed cell death protein 1 (PD-1) play the most significant role in cancer immune escape and cancer immunotherapy. Though PD-L1 have become an important target for drug development and there have been various approved drugs and clinic trials targeting it, and various clinical response rate and adverse reactions prevent many patients from benefiting from it. In recent years, combination trials have become the main direction of PD-1/PD-L1 antibodies development. Here, we summarized PD-L1 biofunctions and key roles in various cancers along with the development of PD-L1 inhibitors. The regulators that are involved in controlling PD-L1 expression including post-translational modification, mRNA level regulation as well as degradation and exosome secretory pathway of PD-L1 were focused. This systematic summary may provide comprehensive understanding of different regulations on PD-L1 as well as a broad prospect for the search of the important regulator of PD-L1. The regulatory factors of PD-L1 can be potential targets for immunotherapy and increase strategies of immunotherapy in combination.

LSD1 deletion decreases exosomal PD-L1 and restores T-cell response in gastric cancer.

BACKGROUND: Histone lysine-specific demethylase 1 (LSD1) expression has been shown to be significantly elevated in gastric cancer (GC) and may be associated with the proliferation and metastasis of GC. It has been reported that LSD1 repressed tumor immunity through programmed cell death 1 ligand 1 (PD-L1) in melanoma and breast cancer. The role of LSD1 in the immune microenvironment of GC is unknown. METHODS: Expression LSD1 and PD-L1 in GC patients was analyzed by immunohistochemical (IHC) and Western blotting. Exosomes were isolated from the culture medium of GC cells using an ultracentrifugation method and characterized by transmission electronic microscopy (TEM), nanoparticle tracking analysis (NTA), sucrose gradient centrifugation, and Western blotting. The role of exosomal PD-L1 in T-cell dysfunction was assessed by flow cytometry, T-cell killing and enzyme-linked immunosorbent assay (ELISA). RESULTS: Through in vivo exploration, mouse forestomach carcinoma (MFC) cells with LSD1 knockout (KO) showed significantly slow growth in 615 mice than T-cell-deficient BALB/c nude mice. Meanwhile, in GC specimens, expression of LSD1 was negatively correlated with that of CD8 and positively correlated with that of PD-L1. Further study showed that LSD1 inhibited the response of T cells in the microenvironment of GC by inducing the accumulation of PD-L1 in exosomes, while the membrane PD-L1 stayed constant in GC cells. Using exosomes as vehicles, LSD1 also obstructed T-cell response of other cancer cells while LSD1 deletion rescued T-cell function. It was found that while relying on the existence of LSD1 in donor cells, exosomes can regulate MFC cells proliferation with distinct roles depending on exosomal PD-L1-mediated T-cell immunity in vivo. CONCLUSION: LSD1 deletion decreases exosomal PD-L1 and restores T-cell response in GC; this finding indicates a new mechanism with which LSD1 may regulate cancer immunity in GC and provides a new target for immunotherapy against GC.

Enhancement of anticancer drug sensitivity in multidrug resistance cells overexpressing ATP-binding cassette (ABC) transporter ABCC10 by CP55, a synthetic derivative of 5-cyano-6-phenylpyrimidin.

ATP-binding cassette (ABC) transporter C10 (ABCC10), also named multidrug resistance protein 7 (MRP7), is a member of ABC transporter superfamily and has been revealed to transport a wide range of chemotherapeutic agents including taxanes, epothilone B, Vinca alkaloids, and anthracyclines. In our previous study, a 5-cyano-6-phenylpyrimidin derivative CP55 was synthesized and found significantly reversal effect of multidrug resistance (MDR) mediated by ABCB1. In this study, we found CP55 also efficiently reversed MDR mediated by ABCC10. Our in vitro study showed that co-treatment with CP55 significantly increased the efficacy of ABCC10-substrate anticancer drugs in MDR cells overexpressing ABCC10. Furthermore, we showed that treatment with CP55 increased the intracellular accumulation of [3H]-labeled anticancer drugs and in-turn decreasing drug efflux by inhibiting the transport activity, without altering ABCC10 protein ex-pression level or cellular localization. Potential CP55-ABCC10 interactions were predicted via docking analysis using human ABCC10 homology model and obtained high docking score. Therefore, CP55 represents a promising therapeutic agent in the combinational treatment of chemo-resistant cancer related to ABCC10.

Chemical molecular-based approach to overcome multidrug resistance in cancer by targeting P-glycoprotein (P-gp).

Multidrug resistance (MDR) remains one of the major impediments for efficacious cancer chemotherapy. Increased efflux of multiple chemotherapeutic drugs by transmembrane ATP-binding cassette (ABC) transporter superfamily is considered one of the primary causes for cancer MDR, in which the role of P-glycoprotein (P-gp/ABCB1) has been most well-established. The clinical co-administration of P-gp drug efflux inhibitors, in combination with anticancer drugs which are P-gp transport substrates, was considered to be a treatment modality to surmount MDR in anticancer therapy by blocking P-gp-mediated multidrug efflux. Extensive attempts have been carried out to screen for sets of nontoxic, selective, and efficacious P-gp efflux inhibitors. In this review, we highlight the recent achievements in drug design, characterization, structure-activity relationship (SAR) studies, and mechanisms of action of the newly synthetic, potent small molecules P-gp inhibitors in the past 5 years. The development of P-gp inhibitors will increase our knowledge of the mechanisms and functions of P-gp-mediated drug efflux which will benefit drug discovery and clinical cancer therapeutics where P-gp transporter overexpression has been implicated in MDR.

Cullin-RING Ligases as Promising Targets for Gastric Carcinoma Treatment.

Gastric carcinoma has serious morbidity and mortality, which seriously threats human health. The studies on gastrointestinal cell biology have shown that the ubiquitination modification that occurs after protein translation plays an essential role in the pathogenesis of gastric carcinoma. Protein ubiquitination is catalyzed by E3 ubiquitin ligase and can regulate various substrate proteins in different cellular pathways. Cullin-RING E3 ligase (CRLs) is a representative of the E3 ubiquitin ligase family, which requires cullin (CUL) neddylation modification for activation to regulate homeostasis of ~20% of cellular proteins. The substrate molecules regulated by CRLs are often involved in many cell progressions such as cell cycle progression, cell apoptosis, DNA damage and repair. Given that CRLs play an important role in modulation of biological activities, so targeting a certain CULs member neddylation may be an attractive strategy for selectively controlling the cellular proteins levels to achieve the goal of cancer treatment. In this review, we will discuss the roles of CULs and Ring protein in gastric carcinoma and summarize the current neddylation modulators for gastric carcinoma treatment.

Systematic Review and Meta-Analysis of Lysine-Specific Demethylase 1 Expression as a Prognostic Biomarker of Cancer Survival and Disease Progression.

BACKGROUND: Numerous studies on the prognostic significance of lysine-specific demethylase 1 (LSD1) up-regulation in tumors have different outcomes. The inconsistency originated from various studies looking into the association between LSD1 and tumor cells has prompted the decision of this quantitative systematic review to decipher how up-regulated LSD1 and overall survival (OS) or recurrence-free survival (RFS) or disease-free survival (DFS) are linked in tumor patients. METHODS: Articles were searched from online databases such as Embase, Web of Science Core, PubMed, Google Scholar, and Scopus. The extraction of the hazard ratios (HR) with their 95% confidence intervals (CIs) was attained and survival data of 3151 tumor patients from 17 pieces of related research were used for this meta-analysis. RESULTS: To shed light on the link between LSD1 up-regulation and the prognosis of diverse tumors, the pooled hazard ratios (HRs) with their 95% confidence intervals (CIs) were determined. In this meta-analysis, it was observed that LSD1 up-regulation is linked with poor OS (HR = 2.08, 95% CI: 1.66-2.61, P < .01) and RFS (HR = 3.09, 95% CI: 1.81-5.26, P < .01) in tumor patients. However, LSD1 up-regulation was not linked to DFS (HR = 1.49, 95% CI: .83-2.69, P = .18) in tumor patients. The subcategory examination grouped by tumor type and ethnicity showed that LSD1 up-regulation was linked with a poor outcome in the esophageal tumor and hepatocellular carcinoma and Asian patients, respectively. For clinical-pathological factors, up-regulated LSD1 was significantly linked with Lymph node status. CONCLUSION: Despite the shortfall of the present work, this meta-analysis proposes that LSD1 up-regulation may be a prognostic biomarker for patients with tumors including esophageal tumors and hepatocellular carcinoma. We propose that large-scale studies are vital to substantiate these outcomes.

Discovery of new tranylcypromine derivatives as highly potent LSD1 inhibitors.

Tranylcypromine (TCP)-based structural modifications lead to the discovery of new LSD1 inhibitors, of which compounds 26b and 29b effectively inhibit LSD1 with the IC50 values of 17 and 11 nM, respectively and also show good selectivity over MAO-B. Mechanistic studies showed that compound 29b concentration-dependently induced H3K4me1/2 accumulation in LSD1 overexpressed MGC-803 cells and also inhibited metastasis of MGC-803 cells. Collectively, both compounds could be promising lead compounds for further investigation.

Designed, synthesized and biological evaluation of proteolysis targeting chimeras (PROTACs) as AR degraders for prostate cancer treatment.

As a continuation of our research on developing potent and potentially safe androgen receptor (AR) degrader, a series of novel proteolysis targeting chimeras (PROTACs) containing the phthalimide degrons with different linker were designed, synthesized and evaluated for their AR degradation activity against LNCaP (AR+) cell line. Most of the synthesized compounds displayed moderate to satisfactory AR binding affinity and might lead to antagonist activity against AR. Among them, compound A16 exhibited the best AR binding affinity (85%) and degradation activity against AR. Due to the strong fluorescence properties of pomalidomide derivatives, B10 was found to be effectively internalized and visualized in LNCaP (AR + ) cells than PC-3 (AR-) cells. Moreover, the molecular docking of A16 with AR and the active site of DDB1-CRBN E3 ubiquitin ligase complex provides guidance to design new PROTAC degrons targeting AR for prostate cancer therapy. These results represent a step toward the development of novel and improved AR PROTACs.

Gramine-based structure optimization to enhance anti-gastric cancer activity.

Gramine is a natural indole alkaloid with a wide range of biological activities, but its anti-gastric cancer activity is poor. Herein, a pharmacophore fusion strategy was adopted to design and synthesize a new series of indole-azole hybrids on the structural basis of gramine. Based on our previous studies, different nitrogen-containing five-membered heterocyclic rings and terminal alkyne group were introduced into the indole-based scaffold to investigate their effect on improving the anti-gastric cancer activity of gramine derivatives. Structure-activity relationship (SAR) studies highlighted the role played by terminal alkyne in enhancing the inhibitory effect, and compound 16h displayed the best antiproliferative activity against gastric cancer MGC803 cells with IC50 value of 3.74 µM. Further investigations displayed compound 16h could induce mitochondria-mediated apoptosis, and caused cell cycle arrest at G2/M phase. Besides, compound 16h could inhibit the metastasis ability of MGC803 cells. Our studies may provide a new strategy for structural optimization of gramine to enhance anti-gastric cancer activity, and provide a potential candidate for the treatment of gastric cancer.

Overcome the tumor immunotherapy resistance by combination of the HDAC6 inhibitors with antitumor immunomodulatory agents.

Tumor immunotherapy is currently subject of intense scientific and clinical developments. In previous decade, therapists used natural immune system from the human body to treat several diseases. Although tumor immune disease is a big challenge, combinatorial therapeutic strategy has been succeeded to show the clinical significance. In this context, we discuss the HDAC6 and tumor immune diseases relationship. Also, we summarized the current state of knowledge that based on the combination treatments of the HDAC6 inhibitors (HDAC6is) with antitumor immunomodulatory agents. We observed that, the combination therapies slow down the tumor immune diseases by blocking the aggresome and proteasome pathway. The combination therapy was able to reduce M2 macrophage and increasing PD-L1 blockade sensitivity. Most importantly, multiple combinations of HDAC6is with other agents may consider as potential strategies to treat tumor immune diseases, by reducing the side effects and improve efficacy for the future clinical development.

Design and synthesis of new indole containing biaryl derivatives as potent antiproliferative agents.

A new series of indole containing biaryl derivatives were designed and synthesized, and further biological evaluations of their antiproliferative activity against cancer cell lines (MGC-803 and TE-1 cells) were also conducted. Of these synthesized biaryls, compound 4-methyl-2-((5-methyl-[1,2,4]triazolo[1,5-a]pyrimidin-7-yl)methyl)quinazoline (23) performed as the most potent antiproliferative agent that inhibited cell viability of MGC-803 cells with an IC50 value of 8.28 µM. In addition, investigation of mechanism exhibited that the compound 4-methyl-2-((5-methyl-[1,2,4]triazolo[1,5-a]pyrimidin-7-yl)methyl)quinazoline (23) could inhibit the expression of c-Myc and glycolysis related proteins, decrease the ATP and lactate production, and further induce apoptosis by activating the AMP-activated protein kinase (AMPK) and p53 signaling pathways.

Jaridon 6, a new diterpene from Rabdosia rubescens (Hemsl.) Hara, can display anti-gastric cancer resistance by inhibiting SIRT1 and inducing autophagy.

Tumor resistance is the main cause of treatment failure and is associated with many tumor factors. Jaridon 6, a new diterpene extracted from Rabdosia rubescens (Hemsl.) Hara, which has been previously extracted by our research team, has been tested having more obvious advantages in resistant tumor cells. However, its mechanism is unclear. In this study, we studied the effect and the specific mechanism of Jaridon 6 in resistant gastric cancer cells. Cytotoxicity test, colony test, western blotting, and nude test verified the anti-drug resistance ability of Jaridon 6 in the MGC803/PTX and MGC803/5-Fu cells. Jaridon 6 has shown obvious inhibitory effects in the sirtuin 1 (SIRT1) enzyme test. Transmission electron microscopy and immunofluorescence tests further proved the autophagic action of Jaridon 6. Jaridon 6 could inhibit the proliferation of the resistant gastric cancer cell in vivo and in vitro. Jaridon 6 inhibited SIRT1 enzyme and induced autophagy by inhibiting the phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) pathway. Thus, it may be considered for treating gastric cancer resistance by individual or combined administration, as an SIRT1 inhibitor and autophagy inducer.

Skp2 in the ubiquitin-proteasome system: A comprehensive review.

The ubiquitin-proteasome system (UPS) is a complex process that regulates protein stability and activity by the sequential actions of E1, E2 and E3 enzymes to influence diverse aspects of eukaryotic cells. However, due to the diversity of proteins in cells, substrate selection is a highly critical part of the process. As a key player in UPS, E3 ubiquitin ligases recruit substrates for ubiquitination specifically. Among them, RING E3 ubiquitin ligases which are the most abundant E3 ubiquitin ligases contribute to diverse cellular processes. The multisubunit cullin-RING ligases (CRLs) are the largest family of RING E3 ubiquitin ligases with tremendous plasticity in substrate specificity and regulate a vast array of cellular functions. The F-box protein Skp2 is a component of CRL1 (the prototype of CRLs) which is expressed in many tissues and participates in multiple cellular functions such as cell proliferation, metabolism, and tumorigenesis by contributing to the ubiquitination and subsequent degradation of several specific tumor suppressors. Most importantly, Skp2 plays a pivotal role in a plethora of cancer-associated signaling pathways. It enhances cell growth, accelerates cell cycle progression, promotes migration and invasion, and inhibits cell apoptosis among others. Hence, targeting Skp2 may represent a novel and attractive strategy for the treatment of different human cancers overexpressing this oncogene. In this review article, we summarized the known roles of Skp2 both in health and disease states in relation to the UPS.

OP16 induces deadly autophagy and apoptosis of cells by inhibiting Akt in esophageal squamous cell carcinoma.

BACKGROUND: OP16, a derivative of the novel ent-kaurene diterpenoid compound separated from Rabdosia rubescens, has been confirmed for its efficacy and safety in the treatment of esophageal squamous cell carcinoma (ESCC) in our previous study. However, the precise mechanisms of tumor lethality mediated by OP16 have not yet been fully characterized. AIMS: To investigate the effects and molecular mechanism of OP16 on autophagy and apoptosis of ESCC cells. METHODS: Effects and mechanism of OP16 on autophagy of ESCC cells were first detected by Western blot, immunofluorescence, mRFP-GFP-LC3 adenovirus infection and transmission electron microscope. Next, function of autophagy and apoptosis induced by OP16 on cell death was investigated by flow cytometry and CCK-8 assay. Finally, molecular mechanism of OP16 affecting autophagy and apoptosis of ESCC cells was explored by Western blot. RESULTS: We demonstrated that OP16 could induce autophagy by promoting the formation of autophagosome and autolysosome, and promote autophagic cell death in ESCC. Furthermore, we also found that OP16 could promote cell apoptosis by activating mitochondria apoptosis pathway in ESCC. Finally, we demonstrated that OP16 affecting autophagy and mitochondria apoptosis pathway was mediated by phosphorylation of Akt. CONCLUSION: Our data show that OP16 could promote cell death through affecting autophagy and mitochondria apoptosis pathway mediated by Akt in ESCC, which enriches the theoretical mechanism of anti-tumor effects of OP16 and provides a basis for treatment of OP16 on ESCC.