c-Myc Inhibitor Research Articles

C-Myc inhibition and p21 modulation contribute to unsymmetrical bisacridines-induced apoptosis and senescence in pancreatic cancer cells.

Pancreatic cancer (PC) is one of the most aggressive cancers and is the seventh leading cause of cancer-related death worldwide. PC is characterized by rapid progression and resistance to conventional treatments. Mutations in KRAS, CDKN2A, TP53, SMAD4/DPC4, and MYC are major genetic alterations associated with poor treatment outcomes in patients with PC. Therefore, optimizing PC therapy is a tremendous challenge. Unsymmetrical bisacridines (UAs), synthesized by our group, are new promising compounds that have exhibited high cytotoxicity and antitumor activity against several solid tumors, including pancreatic cancer. The cellular effects induced by UAs in PC cells were evaluated by MTT assay (cell growth inhibition), flow cytometry, and fluorescence and light microscopy (cell cycle distribution, apoptosis, and senescence detection). Analysis of the effects of UAs on the levels of proteins (c-Myc, p53, SMAD4, p21, and p16) was performed by Western blotting. Apoptosis was the main triggered mechanism of death after UAs treatment, and induction of the SMAD4 protein can facilitate this process. c-Myc, which is one of the molecular targets of UAs, can participate in the induction of cell death in a p53-independent manner. Moreover, UAs can also induce accelerated senescence through the upregulation of p21. Notably, senescent cells can die via apoptosis after prolonged exposure to UAs. UAs have emerged as potent anticancer agents that induce apoptosis by inhibiting c-Myc protein and triggering cellular senescence in a dose-dependent manner by increasing p21 levels. Thus, UAs exhibit desirable features as promising candidates for future pancreatic anticancer therapies.

NL101 synergizes with the BCL-2 inhibitor venetoclax through PI3K-dependent suppression of c-Myc in acute myeloid leukaemia

BackgroundAcute myeloid leukaemia (AML) comprises a group of heterogeneous and aggressive haematological malignancies with unsatisfactory prognoses and limited treatment options. Treatments targeting B-cell lymphoma-2 (BCL-2) with venetoclax have been approved for patients with AML, and venetoclax-based drug combinations are becoming the standard of care for older patients unfit for intensive chemotherapy. However, the therapeutic duration of either single or combination strategies is limited, and the development of resistance seems inevitable. Therefore, more effective combination regimens are urgently needed.MethodsThe efficacy of combination therapy with NL101, a SAHA-bendamustine hybrid, and venetoclax was evaluated in preclinical models of AML including established cell lines, primary blasts from patients, and animal models. RNA-sequencing and immunoblotting were used to explore the underlying mechanism.ResultsNL101 significantly potentiated the activity of venetoclax in AML cell lines, as evidenced by the enhanced decrease in viability and induction of apoptosis. Mechanistically, the addition of NL101 to venetoclax decreased the stability of the antiapoptotic protein myeloid cell leukaemia-1 (MCL-1) by inhibiting ERK, thereby facilitating the release of BIM and triggering mitochondrial apoptosis. Moreover, the strong synergy between NL101 and venetoclax also relied on the downregulation of c-Myc via PI3K/Akt/GSK3β signalling. The combination of NL101 and venetoclax synergistically eliminated primary blasts from 10 AML patients and reduced the leukaemia burden in an MV4-11 cell-derived xenograft model.ConclusionsOur results encourage the pursuit of clinical trials of combined treatment with NL101 and venetoclax and provide a novel venetoclax-incorporating therapeutic strategy for AML.

NEK6 dampens FOXO3 nuclear translocation to stabilize C-MYC and promotes subsequent de novo purine synthesis to support ovarian cancer chemoresistance

De novo purine synthesis metabolism plays a crucial role in tumor cell survival and malignant progression. However, the specific impact of this metabolic pathway on chemoresistance in ovarian cancer remains unclear. This study aims to elucidate the influence of de novo purine synthesis on chemoresistance in ovarian cancer and its underlying regulatory mechanisms. We analyzed metabolic differences between chemosensitive and chemoresistant ovarian cancer tissues using mass spectrometry-based metabolomics. Cell growth, metabolism, chemoresistance, and DNA damage repair characteristics were assessed in vitro using cell line models. Tumor growth and chemoresistance were assessed in vivo using ovarian cancer xenograft tumors. Intervention of purines and NEK6-mediated purine metabolism on chemoresistance was investigated at multiple levels. Chemoresistant ovarian cancers exhibited higher purine abundance and NEK6 expression. Inhibiting NEK6 led to decreased de novo purine synthesis, resulting in diminished chemoresistance in ovarian cancer cells. Mechanistically, NEK6 directly interacted with FOXO3, contributing to the phosphorylation of FOXO3 at S7 through its kinase activity, thereby inhibiting its nuclear translocation. Nuclear FOXO3 promoted FBXW7 transcription, leading to c-MYC ubiquitination and suppression of de novo purine synthesis. Paeonol, by inhibiting NEK6, suppressed de novo purine synthesis and enhanced chemosensitivity. The NEK6-mediated reprogramming of de novo purine synthesis emerges as a critical pathway influencing chemoresistance in ovarian cancer. Paeonol exhibits the potential to interfere with NEK6, thereby inhibiting chemoresistance.

Anti-Warburg Mechanism of Ginsenoside F2 in Human Cervical Cancer Cells via Activation of miR193a-5p and Inhibition of β-Catenin/c-Myc/Hexokinase 2 Signaling Axis.

Though Ginsenoside F2 (GF2), a protopanaxadiol saponin from Panax ginseng, is known to have an anticancer effect, its underlying mechanism still remains unclear. In our model, the anti-glycolytic mechanism of GF2 was investigated in human cervical cancer cells in association with miR193a-5p and the β-catenin/c-Myc/Hexokinase 2 (HK2) signaling axis. Here, GF2 exerted significant cytotoxicity and antiproliferation activity, increased sub-G1, and attenuated the expression of pro-Poly (ADPribose) polymerase (pro-PARP) and pro-cysteine aspartyl-specific protease (procaspase3) in HeLa and SiHa cells. Consistently, GF2 attenuated the expression of Wnt, β-catenin, and c-Myc and their downstream target genes such as HK2, pyruvate kinase isozymes M2 (PKM2), and lactate dehydrogenase A (LDHA), along with a decreased production of glucose and lactate in HeLa and SiHa cells. Moreover, GF2 suppressed β-catenin and c-Myc stability in the presence and absence of cycloheximide in HeLa cells, respectively. Additionally, the depletion of β-catenin reduced the expression of c-Myc and HK2 in HeLa cells, while pyruvate treatment reversed the ability of GF2 to inhibit β-catenin, c-Myc, and PKM2 in GF2-treated HeLa cells. Notably, GF2 upregulated the expression of microRNA139a-5p (miR139a-5p) in HeLa cells. Consistently, the miR139a-5p mimic enhanced the suppression of β-catenin, c-Myc, and HK2, while the miR193a-5p inhibitor reversed the ability of GF2 to attenuate the expression of β-catenin, c-Myc, and HK2 in HeLa cells. Overall, these findings suggest that GF2 induces apoptosis via the activation of miR193a-5p and the inhibition of β-catenin/c-Myc/HK signaling in cervical cancer cells.

C-Myc Inhibition Induces an Additive Effect with Cyclophosphamide in Acute Lymphoblastic Leukemia

Background: Cellular-myelocytomatosis (c-Myc), an oncoprotein and a transcription factor, is involved in several essential cellular processes. The c-Myc expression level is highly regulated in normal cells. It has been proved that c-Myc expression is deregulated in malignant cells due to rearrangements and mutations. The overexpression of this molecule is also reported to be present in acute lymphoblastic leukemia (ALL) as well, which is correlated with an unfavorable response to treatment, poor prognosis, and decreased overall survival. The upregulation of c-Myc results in increased proliferation, cell growth, and survival of ALL cells. Hence, making it an ideal target for leukemia treatment. This study evaluates the effect of c-Myc silencing combined with cyclophosphamide treatment, an FDA-approved chemotherapeutic. Methods: Peripheral blood and bone marrow samples (mononuclear cells) were derived from eleven ALL patients. To silence c-Myc, small interfering-RNA (siRNA)-lipofectamine was used. The efficacy of gene silencing was assessed by the qRT-PCR test. Next, the effect of c-Myc silencing combined with cyclophosphamide treatment in ALL primary cells was evaluated using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) test. Results: ALL cells were successfully transfected with c-Myc-siRNA. Also, treating cells with cyclophosphamide exerted a slight fall in the c-Myc mRNA level. The MTT test revealed that following the inhibition of c-Myc by siRNA, the viability of primary ALL cells decreased in response to cyclophosphamide treatment. Also, it was discovered that silencing c-Myc with siRNA combined with cyclophosphamide treatment significantly inhibits the growth of primary ALL cells compared to cyclophosphamide monotherapy. Conclusion: c-Myc possesses high potential in the treatment of several cancers. Our findings add ALL to this category as well. Silencing c-Myc sensitizes ALL cells to cyclophosphamide treatment and can help with the better treatment of the afflicted individuals.

Shikonin Suppresses Cell Tumorigenesis in Gastric Cancer Associated with the Inhibition of c-Myc and Yap-1.

The study aimed to study the potential roles and mechanisms of shikonin in gastric cancer by network pharmacology and biological experiments. The key genes and targets of shikonin in gastric cancer were predicted by network pharmacology and molecular docking study. The effect of shikonin on the proliferation, migration, and invasion of gastric cancer cells was detected by the CCK8 method, and wound healing and transwell assays. The expression levels of c-Myc and Yap-1 were detected via western blotting in gastric cancer cells after shikonin intervention. The results of network pharmacology revealed the key target genes of shikonin on gastric cancer cells to be c-Myc, Yap-1, AKT1, etc. GO and KEGG analysis showed regulation of cell migration, proliferation, adhesion, and other biological processes, including the PI3K-Akt signaling pathway, HIF-1 signaling pathway, necroptosis, and other cancer pathways. Molecular docking showed shikonin to be most closely combined with protooncogenes c-Myc and Yap-1. In vitro experiments showed that the proliferation rate, migration, and invasion ability of the gastric cancer cell group decreased significantly after shikonin intervention for 24h. The expression levels of c-Myc and Yap-1 in gastric cancer cells were found to be significantly decreased after shikonin intervention. This study showed protooncogenes c-Myc and Yap-1 to be the core target genes of shikonin on gastric cancer cells. Shikonin may suppress gastric cancer cells by inhibiting the protooncogenes c-Myc and Yap-1. This suggests that shikonin may be a good candidate for the treatment of gastric cancer.

Topical application of phenformin ameliorates the psoriasis-like inflammatory response via the inhibition of c-Myc expression in keratinocytes

BackgroundPsoriasis is a chronic inflammatory skin disease characterized by a complex pathogenesis involving various types of cells and cytokines. Among those, the pro-inflammatory cytokine IL-23/IL-17A axis plays a crucial role in the development and rapid progression of psoriasis. Phenformin, a derivative of metformin and a member of the biguanide class of drugs, exhibits superior anti-inflammatory and anti-tumor efficacy compared to metformin. However, the potential role of phenformin in anti-psoriatic skin inflammation has not been explored. MethodsIn this study, we utilized a mouse model of psoriasis and an in vitro model using human keratinocytes to investigate whether phenformin can suppress psoriasis-like inflammatory responses. ResultsOur results demonstrate that the topical application of phenformin significantly inhibited acute skin inflammatory responses in the psoriasis mouse model induced by imiquimod (IMQ). Additionally, phenformin suppressed the expression of psoriasis-related cytokines IL-17, IL-23, IL-8, and S100A8/S100A9 in an in vitro psoriatic keratinocyte model induced by IMQ. Furthermore, we found that IMQ-induced psoriatic skin and IMQ-treated keratinocytes exhibited high expression of the c-Myc gene, which was downregulated by phenformin. The c-Myc inhibitor JQ1 similarly inhibited the psoriatic inflammatory response and the expression of psoriasis-related cytokines in both in vitro and in vivo models. Conclusionphenformin ameliorates the psoriasis-like inflammatory response by inhibiting c-Myc expression in keratinocytes, suggesting its potential as a topical drug for the treatment of psoriasis.

Therapeutic Targeting of the GLS1-c-Myc Positive Feedback Loop Suppresses Glutaminolysis and Inhibits Progression of Head and Neck Cancer.

Head and neck squamous cell carcinoma (HNSCC) is addicted to glutaminolysis. Targeting this metabolic dependency has emerged as a potential therapeutic approach for HNSCC. In this study, we conducted a bioinformatic analysis of The Cancer Genome Atlas HNSCC cohort that revealed a robust correlation between expression of MYC (encoding the protein c-Myc) and glutaminase 1 (GLS1), which catalyzes the first step in glutaminolysis. Intriguingly, disruption of GLS1 signaling in HNSCC cells by genetic depletion or CB-839 treatment resulted in a reduction in c-Myc protein stability via a ubiquitin-specific peptidase 1-dependent ubiquitin-proteasome pathway. On the other hand, c-Myc directly binds to the promoter region of GLS1 and upregulates its transcription. Notably, the GLS1-c-Myc pathway enhanced acetyl-coenzyme A carboxylase-dependent Slug acetylation, prompting cancer cell invasion and metastasis. Thus, the GLS1-c-Myc axis emerged as a positive feedback loop critical for driving the aggressiveness of HNSCC. Therapeutically, combining CB-839 with the c-Myc inhibitor MYCi975 strongly suppressed GLS1-c-Myc signaling, resulting in a superior antitumor effect compared with either single agent in an orthotopic mouse model of HNSCC. These findings hold promise for the development of effective therapies for patients with HNSCC, addressing an urgent need arising from the significant incidence and high metastatic rate of the disease. Significance: GLS1 and c-Myc form a positive feedback loop that promotes head and neck cancer metastasis and can be targeted as a promising therapeutic strategy for this disease.

Polyamine and EIF5A hypusination downstream of c-Myc confers targeted therapy resistance in BRAF mutant melanoma

BackgroundBRAF inhibitors are widely employed in the treatment of melanoma with the BRAF V600E mutation. However, the development of resistance compromises their therapeutic efficacy. Diverse genomic and transcriptomic alterations are found in BRAF inhibitor resistant melanoma, posing a pressing need for convergent, druggable target that reverse therapy resistant tumor with different resistance mechanisms.MethodsCRISPR-Cas9 screens were performed to identify novel target gene whose inhibition selectively targets A375VR, a BRAF V600E mutant cell line with acquired resistance to vemurafenib. Various in vitro and in vivo assays, including cell competition assay, water soluble tetrazolium (WST) assay, live-dead assay and xenograft assay were performed to confirm synergistic cell death. Liquid Chromatography-Mass Spectrometry analyses quantified polyamine biosynthesis and changes in proteome in vemurafenib resistant melanoma. EIF5A hypusination dependent protein translation and subsequent changes in mitochondrial biogenesis and activity were assayed by O-propargyl-puromycin labeling assay, mitotracker, mitoSOX labeling and seahorse assay. Bioinformatics analyses were used to identify the association of polyamine biosynthesis with BRAF inhibitor resistance and poor prognosis in melanoma patient cohorts.ResultsWe elucidate the role of polyamine biosynthesis and its regulatory mechanisms in promoting BRAF inhibitor resistance. Leveraging CRISPR-Cas9 screens, we identify AMD1 (S-adenosylmethionine decarboxylase 1), a critical enzyme for polyamine biosynthesis, as a druggable target whose inhibition reduces vemurafenib resistance. Metabolomic and proteomic analyses reveal that polyamine biosynthesis is upregulated in vemurafenib-resistant cancer, resulting in enhanced EIF5A hypusination, translation of mitochondrial proteins and oxidative phosphorylation. We also identify that sustained c-Myc levels in vemurafenib-resistant cancer are responsible for elevated polyamine biosynthesis. Inhibition of polyamine biosynthesis or c-Myc reversed vemurafenib resistance both in vitro cell line models and in vivo in a xenograft model. Polyamine biosynthesis signature is associated with poor prognosis and shorter progression free survival after BRAF/MAPK inhibitor treatment in melanoma cohorts, highlighting the clinical relevance of our findings.ConclusionsOur findings delineate the molecular mechanisms involving polyamine-EIF5A hypusination-mitochondrial respiration pathway conferring BRAF inhibitor resistance in melanoma. These targets will serve as effective therapeutic targets that can maximize the therapeutic efficacy of existing BRAF inhibitors.

Comprehensive analysis of microRNAs modulated by histone deacetylase inhibitors identifies microRNA-7-5p with anti-myeloma effect.

Basic research to expand treatment options for multiple myeloma is greatly needed due to the refractory nature of the disease. Histone deacetylase (HDAC) inhibitors, which are epigenetic regulators, are attractive but have limited applications. MicroRNAs (miRNAs), which are also epigenetic regulators, are important molecules that may lead to future therapeutic breakthroughs. In this study, we comprehensively searched for miRNAs that are altered by HDAC inhibitors in myeloma cells. We identified miR-7-5p (miR-7) as a miRNA downregulated by HDAC inhibitors. Transfection of myeloma cell lines with miR-7 suppressed cell proliferation, induced apoptosis, and enhanced the effects of the HDAC inhibitor panobinostat. Expression of miR-7 was downregulated by c-Myc inhibition, but upregulated by bortezomib. Comprehensive examination of miR-7 targets revealed four candidates: SLC6A9, LRRC59, EXOSC2, and PSME3. Among these, we focused on PSME3, an oncogene involved in proteasome capacity in myeloma cells. PSME3 knockdown increases myeloma cell death and panobinostat sensitivity. In conclusion, miR-7, which is downregulated by HDAC inhibitors, is a tumor suppressor that targets PSME3. This miR-7 downregulation may be involved in HDAC inhibitor resistance. In addition, combinations of anti-myeloma drugs that complement changes in miRNA expression should be considered.

The Selective SIRT3 Inhibitor 3-TYP Represses Primary Myeloma Growth by Reducing c-Myc Stability.

Multiple myeloma is a hematological cancer that can be treated but remains incurable. With the advancement of science and technology, more drugs have been developed for myeloma chemotherapy that greatly improve the quality of life of patients. However, relapse remains a serious problem puzzling patients and doctors. Thus, developing more highly active and specific inhibitors is urgent for myeloma-targeted therapy. In this study, we identified the SIRT3 inhibitor 3-TYP (3-(1H-1,2,3-triazol-4-yl) pyridine) after screening a histone modification compound library, which showed high cytotoxicity and induced DNA damage in myeloma cells. Furthermore, the inhibitory effect of 3-TYP in our xenograft tumor studies also confirmed that compound 3-TYP could inhibit primary myeloma growth by reducing c-Myc protein stability by decreasing c-Myc Ser62 phosphorylation levels. Taken together, the results of our study identified 3-TYP as a novel c-Myc inhibitor, which could be a potential chemotherapeutic agent to target multiple myeloma.

Anti-Leukemic Effects of Small Molecule Inhibitor of c-Myc (10058-F4) on Chronic Myeloid Leukemia Cells.

As one of the main molecules in BCR-ABL signaling, c-Myc acts as a pivotal key in disease progression and disruption of long-term remission in patients with CML. To clarify the effects of c-Myc inhibition in CML, we examined the anti-tumor property of a well-known small molecule inhibitor of c-Myc 10058-F4 on K562 cell line. This experimental study was conducted in K562 cell line for evaluation of cytotoxic activity of 10058-F4 using Trypan blue and MTT assays. Flow cytometry and Quantitative RT-PCR analysis were also conducted to determine its mechanism of action. Additionally, Annexin/PI staining was performed for apoptosis assessment. The results of Trypan blue and MTT assay demonstrated that inhibition of c-Myc, as shown by suppression of c-Myc expression and its associated genes PP2A, CIP2A, and hTERT, could decrease viability and metabolic activity of K562 cells, respectively. Moreover, a robust elevation in cell population in G1-phase coupled with up-regulation of p21 and p27 expression shows that 10058-F4 could hamper cell proliferation, at least partly, through induction of G1 arrest. Accordingly, we found that 10058-F4 induced apoptosis via increasing Bax and Bad; In contrast, no significant alterations were observed NF-KB pathway-targeted anti-apoptotic genes in the mRNA levels. Notably, disruption of the NF-κB pathway with bortezomib as a common proteasome inhibitor sensitized K562 cells to the cytotoxic effect of 10058-F4, substantiating the fact that the NF-κB axis functions probably attenuate the K562 cells sensitivity to c-Myc inhibition. It can be concluded from the results of this study that inhibition of c-Myc induces anti-neoplastic effects on CML-derived K562 cells as well as increases the efficacy of imatinib. For further insight into the safety and effectiveness of 10058-F4 in CML, in vivo studies will be required.

DSG2 and c-MYC Interact to Regulate the Expression of ADAM17 and Promote the Development of Cervical Cancer.

To explore the effect of DSG2 on the growth of cervical cancer cells and its possible regulatory mechanism. The expression levels and survival prognosis of DSG2 and ADAM17 in cervical squamous cell carcinoma tissues and adjacent normal tissues were analyzed by bioinformatics. CCK-8 assay, colony formation assay and Transwell assay were used to detect the effects of DSG2 on the proliferative activity, colony formation ability and migration ability of SiHa and Hela cells. The effect of DSG 2 on the level of ADAM17 transcription and translation was detected by qPCR and Western blot experiments. The interaction between DSG2 and c-MYC was detected by immunocoprecipitation. c-MYC inhibitors were used in HeLa cells overexpressing DSG2 to analyze the effects of DSG2 and c-MYC on proliferation, colony formation and migration of Hela cells, as well as the regulation of ADAM17 expression. DSG2 was highly expressed in cervical squamous cell carcinoma compared with normal tissues (P<0.05), and high DSG2 expression suggested poor overall survival (P<0.05). After DSG2 knockdown, the proliferative activity, colony formation and migration ability of SiHa and Hela cells were significantly decreased (P<0.05). Compared with adjacent normal tissues, ADAM17 was highly expressed in cervical squamous cell carcinoma (P<0.05), and high ADAM17 expression suggested poor overall survival in cervical cancer patients (P<0.05). The results of immunocoprecipitation showed the interaction between DSG2 and c-MYC. Compared with DSG2 overexpression group, DSG2 overexpression combined with c-MYC inhibition group significantly decreased cell proliferation, migration and ADAM17 expression (P < 0.05). DSG2 is highly expressed in cervical cancer, and inhibition of DSG2 expression can reduce the proliferation and migration ability of cervical cancer cells, which may be related to the regulation of ADAM17 expression through c-MYC interaction.

Development, synthesis and validation of improved c-Myc/Max inhibitors.

The pathophysiological foundations of various diseases are often subject to alteration through the utilization of small compounds, rendering them invaluable tools for the exploration and advancement of novel therapeutic strategies. Within the scope of this study, we meticulously curated a diverse library of novel small compounds meticulously designed to specifically target the c-Myc/Max complex. We conducted invitro examinations of novel c-Myc inhibitors across a spectrum of cancer cell lines, including PANC1 (pancreatic adenocarcinoma), MCF7 (breast carcinoma), DU-145 (prostate carcinoma), and A549 (lung cancer). The initial analysis involved a 25 μM dose, which enabled the identification of potent anticancer compounds effective against a variety of tumour types. We identified c-Myc inhibitors with remarkable potency, featuring IC50 values as low as 1.6 μM and up to 40 times more effective than the reference molecule in diminishing cancer cell viability. Notably, c-Myc-i7 exhibited exceptional selectivity, displaying 37-fold and 59-fold preference for targeting prostate and breast cancers, respectively, over healthy cells. Additionally, we constructed drug-likeness models. This study underscores the potential for invitro investigations of various tumour types using novel c-Myc inhibitors to yield ground-breaking and efficacious anticancer compounds.

Design, synthesis, and activity evaluation of 2-iminobenzimidazoles as c-Myc inhibitors for treating multiple myeloma

Multiple myeloma (MM) is a plasma cell malignancy that remains incurable and poses a significant threat to global public health. The multifunctional transcription factor c-Myc plays a crucial role in various cellular processes and is closely associated with MM progression. As part of the basic-helix-loop-helix-leucine zipper (bHLHZip) family, c-Myc forms heterodimers with its obligate partner Max, binds to the Enhancer-box (E-box) of DNA, and ultimately co-regulates gene expression. Therefore, impeding the capacity for heterodimerization to bind to DNA represents a favored strategy in thwarting c-Myc transcription. In this study, we first synthesized a series of novel 2-iminobenzimidazole derivatives and further estimated their potential anti-MM activity. Notably, among all the derivatives, 5b and 5d demonstrated remarkable inhibitory activity against RPMI-8226 and U266 cells, with IC50 values of 0.85 μM and 0.97 μM for compound 5b, and 0.96 μM and 0.89 μM for compound 5d. Western blot and dual-luciferase reporter assays demonstrated that compounds 5b and 5d effectively suppressed both c-Myc protein expression and transcriptional activity of the c-Myc promoter in RPMI-8226 and U266 cells. Furthermore, these compounds induced apoptosis and G1 cell cycle arrest in the aforementioned MM cells. Molecular docking studies revealed that 5b and 5d exhibited strong binding affinity to the interface between c-Myc/Max and E-box of DNA. Taken together, our findings suggest that further investigations are warranted for potential therapeutic applications of 5b and 5d for c-Myc-related diseases.

A novel combined therapeutic strategy of Nano-EN-IR@Lip mediated photothermal therapy and stem cell inhibition for gastric cancer

Recurrence and metastasis of gastric cancer is a major therapeutic challenge for treatment. The presence of cancer stem cells (CSCs) is a major obstacle to the success of current cancer therapy, often leading to treatment resistance and tumor recurrence and metastasis. Therefore, it is important to develop effective strategies to eradicate CSCs. In this study, we developed a combined therapeutic strategy of photothermal therapy (PTT) and gastric cancer stem cells (GCSCs) inhibition by successfully synthesizing nanoliposomes loaded with IR780 (photosensitizer) and EN4 (c-Myc inhibitor). The nanocomposites are biocompatible and exhibit superior photoacoustic (PA) imaging properties. Under laser irradiation, IR780-mediated PTT effectively and rapidly killed tumor cells, while EN4 synergistically inhibited the self-renewal and stemness of GCSCs by suppressing the expression and activity of the pluripotent transcription factor c-Myc, preventing the tumor progression of gastric cancer. This Nano-EN-IR@Lip is expected to be a novel clinical nanomedicine for the integration of gastric cancer diagnosis, treatment and prevention.

Abstract 660: OPN-6602, a potent dual EP300/CBP bromodomain inhibitor, targets multiple myeloma through concomitant suppression of IRF4 and c-MYC

Abstract Multiple myeloma (MM) is an aggressive hematological cancer derived from malignant plasma cells in the bone marrow. E1A-binding protein (EP300) and CREB-binding protein (CBP) are transcription coactivators that contain both epigenetic writer histone acetyltransferase (HAT) and reader bromo- (BRD) domains that regulate transcription of genes via chromatin remodeling. EP300/CBP inhibition causes cell cycle arrest and apoptosis in MM through suppression of interferon regulatory factor (IRF4) and concomitant repression of c-MYC, rendering EP300/CBP as a novel target for MM. OPN-6602 is a potent, selective, and orally active small molecule dual EP300/CBP inhibitor discovered through structure guided drug design. Its binding interaction exhibits low nM activity on EP300 (Kd= 1.5 nM) and CREBBP (Kd= 1 nM), with &amp;gt; 200-fold less activity against BRD4. OPN-6602 binding to EP300 is particularly potent by Surface Plasmon Resonance analysis, displaying a Kd of 0.87 nM, with a fast association rate (1.5 E+06/Ms) and a slow dissociation rate (1.3 E-03/s) with a residence time of 13 min. It potently inhibits cell growth in MM cell lines OPM-2 (IC50 ~ 4 nM) and MM.1S (IC50 ~ 47 nM) as well as histone acetylation (IC50 ~ 2 nM) via BRD intramolecular modulation of HAT activity. OPN-6602 demonstrated potent inhibition of H3K27Ac in both LK2 (CBP-deficient lung cancer cells, IC50 = 11 nM) and in OPM-2 (IC50 = 1.6 nM). OPN-6602 also potently inhibited H3K27ac expression in PBMCs with IC50s of 0.75 and 1.4 nM, respectively. The effects of OPN-6602 with dexamethasone (dex) or lenalidomide (len) on the growth of OPM-2 and MM1.S cells were evaluated in a matrix combination study. As single agents, maximal growth inhibition reached 60-75% following dex treatment and only 30-40% following len treatment. In vitro combination studies utilizing a systematic assessment of a broad range of OPN-6602 concentrations in combination with dex or len demonstrated strong synergy. Synergy was observed between OPN-6602 at 3.7 nM and len through a concentration range from 0.027 to 20 µM. Synergy was observed with 1.2 or 3.7 nM of OPN-6602 in combo with dex concentrations from 0.055 to 40 µM. In the OPM-2 xenograft model, OPN-6602 inhibits tumor growth by 30-111% at doses from 2.5-60 mg/kg QD. This growth inhibition correlated with downregulation of IRF4 and cMYC in tumor samples analyzed by QPCR analysis. The pharmacokinetic profile is favorable for oral QD dose administration due to a high Cmax (19 µg/mL at 24 mg/kg) and short half-life (T1/2 = 1.5 hr). A first-in-human study of OPN-6602 in patients with MM is planned for 2024. Citation Format: Bernice Matusow, Wayne Spevak, Chao Zhang, Yan Ma, Rafe Shellooe, James Tsai, Pei Pei Li, Pan Yu Chen, Gaston Habets, Christine Nichols, Parmveer Singh, Kerry Inokuchi, Jackie Walling, Jason Halladay, Gideon Bollag. OPN-6602, a potent dual EP300/CBP bromodomain inhibitor, targets multiple myeloma through concomitant suppression of IRF4 and c-MYC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 660.

Abstract 3039: Pan cancer therapeutic inhibition of oncogenic c-MYC by the engineered destabilized 3’UTR of c-MYC

Abstract c-MYC is a master transcription factor that is over expressed in more than 70% of human cancers. There is no clinically approved direct inhibitor of c-MYC. Recently, we reported the development of the engineered destabilized 3’UTR of MYC constructs which is a novel mRNA overwriting technology through which we directly destabilized and degraded c-MYC. Here we report that we have extended this technology in directly targeting and degrading c-MYC in multiple ethnically diverse triple negative breast cancer cell lines, ovarian cancers, neuroendocrine prostate cancers and pancreatic cancers. The constructs are specific, titratable, and directly engaged the c-MYC target validated by RNA transcript technologies. We have performed head-to-head IC50 dose dependent titration comparison with the engineered destabilized 3’UTR of c-MYC constructs and chemotherapies (DNA damaging agents, microtubules inhibiting drugs, alkylating agents) as well as immune checkpoint inhibitor. We found evidence that suggests that the engineered destabilized 3’UTR of c-MYC is likely to be superior to these therapies in pan cancers driven by c-MYC. Citation Format: Chidiebere U. Awah. Pan cancer therapeutic inhibition of oncogenic c-MYC by the engineered destabilized 3’UTR of c-MYC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3039.

Abstract 7577: A new cancer therapy strategy: Synthetic lethality induced by inhibition of mTOR and cMYC and JNK activation

Abstract Cancer cells are characterized by aberrant signal transduction and cell cycle control pathways that lead to unchecked cell proliferation. Uncontrolled cancer cell proliferation, as well as immune suppressive macrophage proliferation, contribute to tumor growth and spread. Effort to suppress tumor cell growth using Inhibitors of PI3K or MEK signaling pathways can initially inhibit tumor growth, but resistance develops due to upregulation of the alternative pathway. We identified a novel series of allosteric tyrosine kinase receptor inhibitors that induce synthetic lethality in AML, lung carcinoma, pancreatic carcinoma, and other tumor cells and in proliferating macrophages in vitro and in vivo by simultaneously inhibiting cMyc and Tor and activating the stress activated kinase JNK, leading to p53 mediated cell cycle arrest. Select kinase inhibitors with a unique positioning of a single side chain inhibited CDK8/19 and thereby Myc while stimulating phosphorylation and activation of JNK and p38. JNK activation promoted sustained ATF-2 and cJun-mediated p53 and Chk1/Chk2 activation, thereby promoting cell cycle arrest in G2/M and ultimately, apoptosis. These inhibitors potently suppressed tumor growth in vivo by halting both tumor cell and macrophage proliferation in G2/M. They synergized with anti-PD-1 and anti-CTLA-4 and led to tumor eradication and immunological memory formation. These studies identify novel allosteric inhibitors with potent synthetically lethal anti-tumor activity. Citation Format: Hui Chen, Jason Rivera, Marc Paradise, Zihan Xu, Giuliana Mognol, Judith Varner. A new cancer therapy strategy: Synthetic lethality induced by inhibition of mTOR and cMYC and JNK activation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7577.

Abstract 7581: Nicotine promotes the proliferation and metastasis of non-small cell lung cancer through c-Myc/EZH2 pathway

Abstract Objective: Lung cancer, particularly non-small cell lung cancer (NSCLC), continues to be a predominant contributor to global cancer-related mortality, with smoking emerging as a significant risk factor. Nicotine, which is one of the main chemicals from the tobacco smoke, is identified as carcinogen for lung cancer, especially for squamous carcinoma (LUSC). This study aims to investigates the intricate molecular mechanisms by which nicotine facilitates proliferation and migration in NSCLC through the c-Myc/EZH2 signaling pathway. Methods: Clinical tissue specimens were procured from surgical patients with lung cancer for immunohistochemical staining. Additionally, public databases were scrutinized to compare EZH2 expression between smokers and non-smokers. NSCLC cell lines underwent treatment with nicotine in a time- and concentration-dependent manner, and the impact on EZH2 was corroborated through Western Blot, Real-time PCR, and immunofluorescence staining. Proliferation and migration assays, encompassing CCK8 experiments, colony formation assays, EdU staining, and Transwell assays, were conducted to discern the effects of nicotine on NSCLC. Experimental techniques such as si-RNA and CHIP were deployed to investigate changes in c-Myc-mediated regulation of EZH2, elucidating the intricate role of the c-Myc/EZH2 pathway in nicotine-induced NSCLC proliferation and migration. Results: Smokers exhibited heightened EZH2 expression in tumor tissue compared to non-smokers. Nicotine treatment induced a concentration- and time-dependent upregulation of EZH2 in NSCLC cell lines. In 10 μM nicotine-treated NSCLC cells, concurrent treatment with 10 μM DZNep (EZH2 inhibitor) or si-EZH2 significantly augmented proliferation and migration. Moreover, the upregulation of EZH2 correlated with increased c-Myc expression, and siRNA-mediated c-Myc inhibition resulted in a corresponding reduction in EZH2 expression. Nicotine was identified as a promoter of NSCLC proliferation and migration through the c-Myc/EZH2 signaling pathway. This study establishes a robust association between nicotine and EZH2 in NSCLC cells, unveiling the novel regulatory axis of c-Myc/EZH2. Combined treatment with c-Myc and EZH2 inhibitors demonstrated efficacy in suppressing nicotine-induced promotion of NSCLC proliferation and migration. Conclusion: In summary, this study elucidates the mechanisms through which nicotine propels NSCLC development via the c-Myc/EZH2 pathway, providing a theoretical underpinning for future NSCLC treatment paradigms. The combined use of c-Myc and EZH2 inhibitors emerges as a promising therapeutic strategy, poised to inhibit nicotine-induced NSCLC proliferation and migration, offering enhanced treatment modalities for patients. Citation Format: Chen Ding, Hua Huang, Chen Chen, Yongwen Li, Hongyu Liu, Jun Chen. Nicotine promotes the proliferation and metastasis of non-small cell lung cancer through c-Myc/EZH2 pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7581.