Panel Of Cancer Cells Research Articles

Synthesis, computational analysis, and anti-proliferative evaluation of novel bithienylbenzamidine derivatives: Towards cytostatic cancer therapeutics

Our research unveils the synthesis and characterization of four novel monocationic 2,5-diarylbithiophene derivatives 4a-d, prepared through Suzuki cross-coupling reactions and subsequent amidine formation. Density Functional Theory (DFT) calculations provide unprecedented insights into the electronic properties and structure-activity relationships of these compounds. In vitro screening against the NCI-60 cancer cell panel demonstrates remarkable anti-proliferative activities. Compound 4c emerges as a standout candidate, exhibiting potent growth inhibition across multiple cancer types, with GI50 values as low as 0.36 µM against certain ovarian cancer lines. Intriguingly, our research suggests a potential shift from purely cytotoxic towards more cytostatic behavior, potentially offering improved therapeutic windows and reduced side effects. Molecular docking studies provide evidence for the compounds' interaction with human carbonic anhydrase IX, hinting at a possible mechanism of action. This comprehensive study not only introduces a promising new class of anti-cancer compounds but also provides a rich foundation of structural, electronic, and biological data to fuel future research. The bithienylbenzamidine derivatives, particularly compound 4c, represent exciting candidates for further development in the ongoing battle against cancer.

New Cytotoxic α‐Aminoacylamide and bis‐1,5‐Disubstituted Tetrazole Adducts From Amino‐Diterpene Molecules by Ugi Reaction

ABSTRACTIn search for new molecules of diterpene origin with promising anticancer activity, two amino‐derivatives (methyl maleopimarate aminoimide and methyl 1β,13‐epoxydihydroquinopimarate C4‐hydrazone) were involved in the 4‐component Ugi reaction (Ugi‐4CR) and pseudo‐7‐component azido‐Ugi condensation (azido‐Ugi‐7CR) to afford a series of adducts holding α‐aminoacylamide and bis‐1,5‐disubstituted tetrazole substituents. The NCI‐60 cancer cell panel screening revealed diterpene‐type Ugi adducts 2, 5, and 6 with strong antiproliferative potency with GI50 in range of 1.2–15.4 μM. The high positive correlations with standard anticancer drugs suggest microtubules or progesterone and androgen receptors as possible targets of the synthesized compounds.

Anticancer Activity of Imidazolyl Gold(I/III) Compounds in Non-Small Cell Lung Cancer Cell Lines.

Lung cancer is a leading cause of cancer-related death worldwide that needs updated therapies to contrast both the serious side effects and the occurrence of drug resistance. A panel of non-small cell lung cancer (NSCLC) cells were herein employed as cancer models. Eight structurally related gold(I) and gold(III) complexes with NHC and halides or triphenylphosphane ligands were investigated as lung cancer cell growth inhibitors. As expected, gold compounds with PPh3 were found to be more cytotoxic than homoleptic [(NHC)2-Au(I)]X or heteroleptic NHC-Au(I)X or NHC-Au(III)X3 complexes. Mixed ligand gold(I) compounds exhibiting the linear NHC-AuPPh3 (compound 7) or the trigonal NHC-Au(Cl)PPh3 (compound 8) arrangements at the central metal were found to be the best lung cancer cytotoxic compounds. Analysis of the TrxR residual activity of the treated cells revealed that these compounds efficiently inhibit the most accredited molecular target for gold compounds, the TrxR, with compound 8 reaching more than 80% activity reduction in lung cells. Some of the current cancer lung therapy protocols consist of specific lung cancer cell cytotoxic agents combined with antifolate drugs; interestingly, the herein gold compounds are both TrxR and antifolate inhibitors. The human DHFR was inhibited with IC50 ranging between 10-21 µM, depending on substrate concentrations, proceeding by a likely allosteric mechanism only for compound 8.

Expanded profiling of WD repeat domain 5 inhibitors reveals actionable strategies for the treatment of hematologic malignancies

WD40 Repeat Domain 5 (WDR5) is a highly conserved nuclear protein that recruits MYC oncoprotein transcription factors to chromatin to stimulate ribosomal protein gene expression. WDR5 is tethered to chromatin via an arginine-binding cavity known as the "WIN" site. Multiple pharmacological inhibitors of the WDR5-interaction site of WDR5 (WINi) have been described, including those with picomolar affinity and oral bioavailability in mice. Thus far, however, WINi have only been shown to be effective against a number of rare cancer types retaining wild-type p53. To explore the full potential of WINi for cancer therapy, we systematically profiled WINi across a panel of cancer cells, alone and in combination with other agents. We report that WINi are unexpectedly active against cells derived from both solid and blood-borne cancers, including those with mutant p53. Among hematologic malignancies, we find that WINi are effective as a single agent against leukemia and diffuse large B cell lymphoma xenograft models, and can be combined with the approved drug venetoclax to suppress disseminated acute myeloid leukemia in vivo. These studies reveal actionable strategies for the application of WINi to treat blood-borne cancers and forecast expanded utility of WINi against other cancer types.

Ribosomal RNA transcription governs splicing through ribosomal protein RPL22.

Ribosome biosynthesis is a cancer vulnerability executed by targeting RNA polymerase I (Pol I) transcription. We developed advanced, specific Pol I inhibitors to identify drivers of this sensitivity. By integrating multi-omics features and drug sensitivity data from a large cancer cell panel, we discovered that RPL22 frameshift mutation conferred Pol I inhibitor sensitivity in microsatellite instable cancers. Mechanistically, RPL22 directly interacts with 28S rRNA and mRNA splice junctions, functioning as a splicing regulator. RPL22 deficiency, intensified by 28S rRNA sequestration, promoted the splicing of its paralog RPL22L1 and p53 negative regulator MDM4. Chemical and genetic inhibition of rRNA synthesis broadly remodeled mRNA splicing controlling hundreds of targets. Strikingly, RPL22-dependent alternative splicing was reversed by Pol I inhibition revealing a ribotoxic stress-initiated tumor suppressive pathway. We identify a mechanism that robustly connects rRNA synthesis activity to splicing and reveals their coordination by ribosomal protein RPL22.

A new isoxazolyl-urea derivative induces apoptosis, paraptosis, and ferroptosis by modulating MAPKs in pancreatic cancer cells

MAPK pathway regulates the major events including cell division, cell death, migration, invasion, and angiogenesis. Small molecules that modulate the MAPK pathway have been demonstrated to impart cytotoxicity in cancer cells. Herein, the synthesis of a new isoxazolyl-urea derivative (QR-4) has been described and its effect on the growth of pancreatic cancer cells has been investigated. QR-4 reduced the cell viability in a panel of pancreatic cancer cells with minimal effect on normal hepatocytes. QR-4 induced the cleavage of PARP and procaspase-3, reduced the expression of antiapoptotic proteins, increased SubG1 cells, and annexin V/PI-stained cells indicating the induction of apoptosis. QR-4 also triggered paraptosis as witnessed by the reduction of mitochondrial membrane potential, decrease in the expression of Alix, increase in the levels of ATF4 and CHOP, and enhanced ER stress. QR-4 also modulated ferroptosis-related events such as elevation in iron levels, alteration in GSH/GSSG ratio, and increase in the expression of TFRC with a parallel decrease in the expression of GPX4 and SLC7A11. The mechanistic approach revealed that QR-4 increases the phosphorylation of all three forms of MAPKs (JNK, p38, and ERK). Independent application of specific inhibitors of these MAPKs resulted in a partial reversal of QR-4-induced effects. Overall, these reports suggest that a new isoxazolyl-urea imparts cell death via apoptosis, paraptosis, and ferroptosis by regulating the MAPK pathway in pancreatic cancer cells.

Multiaction Pt(IV) Complexes: Cytotoxicity in Ovarian Cancer Cell Lines and Mechanistic Studies.

Ovarian cancer has the worst case-to-fatality ratio of all gynecologic malignancies. The main reasons for the high mortality rate are relapse and the development of chemoresistance. In this paper, the cytotoxic activity of two new multiaction platinum(IV) derivatives of cisplatin and oxaliplatin in a panel of ovarian cancer cells is reported. Cis,cis,trans-[Pt(NH3)2Cl2(IPA)(DCA)] (1) and trans-[Pt(DACH)(OX)(IPA)(DCA)] (2) (IPA = indole-3-propionic acid, DCA = dichloroacetate, DACH = 1R,2R-1,2-diaminocyclohexane, OX = oxalate) were synthesized and characterized by elemental analysis, ESI-MS, FT-IR, and 1H, 13C, and195Pt NMR spectroscopy. The biological activity was evaluated in A2780, PEA1, PEA2, SKOV3, SW626, and OVCAR3 cells. Both complexes are potent cytotoxins. Remarkably, complex 2 is 14 times more active in OVCAR3 cells than cisplatin and is able to overcome cisplatin resistance in PEA2 and A2780cis cells, which are models of post-treatment patient-developed and laboratory-induced resistance. This complex also shows activity in 3D cancer models of the A2780 cells. Mechanistic studies revealed that the complexes induce apoptosis via DNA damage and ROS generation.

Synthesis and Systematic Investigation of Lepidiline A and Its Gold(I), Silver(I), and Copper(I) Complexes Using In Vitro Cancer Models and Multipotent Stem Cells.

The imidazole alkaloid lepidiline A from the root of Lepidium meyenii has a moderate to low in vitro anticancer effect. Our aim was to extend cytotoxicity investigations against a panel of cancer cells, including multidrug-resistant cancer cells, and multipotent stem cells. Lepidiline A is a N-heterocyclic carbene precursor, therefore a suitable ligand source for metal complexes. Thus, we synthesized lepidiline A and its copper(I), gold(I), and silver(I) complexes and tested them against ovarian, gastrointestinal, breast, and uterine cancer cells and bone marrow-derived and adipose-derived mesenchymal stem cells. Lepidiline A and its copper complex demonstrated moderate cytotoxicity, while silver and gold complexes exhibited significantly enhanced and consistent cytotoxicity against both cancer and stem cell lines. ABCB1 in the multidrug-resistant uterine sarcoma line conferred significant resistance against lepidiline A and the copper-lepidiline A complex, but not against the silver and gold complexes. Our results indicate that only the copper complex induced a significant and universal increase in the production of reactive oxygen species within cells. In summary, binding of metal ions to lepidiline A results in enhanced cytotoxicity with the nature of the metal ion playing a critical role in determining its properties.

Antiplatelet drug ticagrelor suppresses triple negative breast cancer metastasis by targeting PI3K

Metastatic recurrence is still a major challenge in breast cancer treatment. Patients with triple negative breast cancer (TNBC) develop early recurrence and relapse more frequently. Due to the lack of specific therapeutic targets, new targeted therapies for TNBC are urgently needed. Phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) pathway is one of the active pathways involved in chemoresistance and survival of TNBC, being considered as a potential target for TNBC treatment. Our present study identified ticagrelor, an anti-platelet drug, as a pan-PI3K inhibitor with potent inhibitory activity against four isoforms of class I PI3K. At doses normally used in clinic, ticagrelor showed weak cytotoxicity against a panel of breast cancer cells, but significantly inhibited the migration, invasion and the actin cytoskeleton organization of human TNBC MDA-MB-231 and SUM-159PT cells. Mechanistically, ticagrelor effectively inhibited PI3K downstream mTOR complex 1 (mTORC1) and mTORC2 signaling by targeting PI3K and decreased the protein expression of epithelial-mesenchymal transition (EMT) markers. In vivo, ticagrelor significantly suppressed tumor cells lung metastasis in 4T1 tumor bearing BALB/c mice model and experimental lung metastasis model which was established by tail vein injection of GFP-labeled MDA-MB-231 cells. The above data demonstrated that ticagrelor can inhibit the migration and invasion of TNBC both in vitro and in vivo by targeting PI3K, suggesting that ticagrelor, a pan-PI3K inhibitor, might represent a promising therapeutic agent for the treatment of metastatic TNBC.

Sphaeropsidin A C15-C16 Cross-Metathesis Analogues with Potent Anticancer Activity.

We recently discovered that sphaeropsidin A (SphA), a fungal metabolite from Diplodia cupressi, overcomes apoptosis resistance in cancer cells by inducing cellular shrinkage by impairing regulatory volume increase. Previously, we prepared a pyrene-conjugated derivative of SphA by a cross-metathesis reaction involving the phytotoxin's C15,C16-alkene. This derivative's evaluation in a cancer cell panel revealed a significant increase in potency, with the IC50 values 5-10× lower than those displayed by the original natural product. Herein, we describe the preparation and anticancer evaluation of fifteen novel C15,C16-alkene cross-metathesis analogues in which the pyrene moiety was replaced with other aromatic or non-aromatic hydrophobic groups. The idea for this replacement was to prepare a family of compounds that would not be predicted to be mutagenic compared with the original pyrene analogue. We predict several of our new compounds to be non-mutagenic, while retaining the high potency of the original pyrene-containing analogues. Examples of these potential lead compounds included those containing pentamethylphenyl and triphenylethylene pendant groups. As an additional feature of the current investigation, we prepared several deuterated pyrene-containing compounds to overcome intellectual property issues associated with non-patentability of the original pyrene derivative.

Synthesis and cytotoxicity of novel 6,8,9-trisubstituted purine analogs against liver cancer cells

A series of novel 6-(substituted phenyl piperazine)-8-(4-substituted phenyl)-9-cyclopentyl purines, 10–51, were synthesized by a four-step synthesis, achieving an overall yield of about 43 %. The reaction conditions were effectively optimized, and the final products were obtained with high purity and yield in all synthesis steps. The synthesized nucleobases were evaluated for their in vitro cytotoxic activities on selected human cancer cell lines (HUH7 (liver), HCT116 (colon), and MCF7 (breast)) using the Sulforhodamine B (SRB) assay. Among these analogs, compounds bearing 4-trifluoromethyl phenyl (19, 20 and 21), 4-methoxy phenyl (27) and 4-fluoro phenyl (34) substitutions at C-8 of purine were the most potent, and they were also analyzed in drug-resistance and drug-sensitive hepatocellular cancer cell (HCC) panels. Compound 19 displayed remarkable anticancer activities (IC50 = 2.9–9.3 μM) against Huh7, FOCUS, SNU475, SNU182, HepG2, and Hep3B cells compared to the positive control, Fludarabine. Additionally, the pharmacological properties and toxicity profiles of the molecules were investigated computationally by the Swiss-ADME and Pro-Tox II online tools, respectively. Results showed that our compounds have favorable physicochemical characteristics for oral bioavailability and do not reveal any toxicity endpoints such as carcinogenicity, immunotoxicity, mutagenicity, or cytotoxicity.

Exploring anti-cancer constituents and cytotoxic effects of Feijoa sellowiana fruit extract on breast cancer

Breast cancer remains a leading cause of mortality among women, necessitating heightened attention and innovative treatment approaches. Given the heterogeneous nature of breast cancer, exploring novel therapeutic avenues is crucial. Natural products, with their potential to offer less aggressive alternatives to conventional chemotherapy, have garnered interest. In this study, the potential cytotoxic effect of Feijoa sellowiana fruit extract (FE) was investigated on a panel of human breast cancer cells. GC-MS analysis was performed to identify the active constituents present in the FE extract and MTT analysis was conducted to evaluate the cytotoxicity of FE against breast cancer cells. Results showed a strong efficacy of FE against MDA-MB-453 and MDA-MB-231 cell lines. The cytotoxicity was evident after a 24-hour treatment duration for both lines. It was observed that the two cell lines in which the FE extract was most effective belonged to the triple-negative breast cancer category. The viability of MCF-7 cells decreased to 23.2% after 72 hours of exposure to 1000 μg/mL FE, and this decline was also noticeable at lower concentrations. Conversely, the BT-474 cell line displayed the least susceptibility, with a viability of 43.9% even at the highest concentration of 1000 μg/mL FE. These findings underscore FE's targeted efficacy against triple-negative breast cancer cells, indicating its promise as an alternative avenue to tackle this formidable cancer subtype.

RAS-RAF-miR-296-3p signaling axis increases Rad18 expression to augment radioresistance in pancreatic and thyroid cancers

Oncogenic RAS and RAF signaling has been implicated in contributing to radioresistance in pancreatic and thyroid cancers. In this study, we sought to better clarify molecular mechanisms contributing to this effect. We discovered that miRNA 296–3p (miR-296–3p) is significantly correlated with radiosensitivity in a panel of pancreatic cancer cells, and miR-296–3p is highly expressed in normal cells, but low in cancer cell lines. Elevated expression of miR-296–3p increases radiosensitization while decreasing the expression of the DNA repair enzyme RAD18 in both pancreatic and thyroid cancer cells. RAD18 is overexpressed in both pancreatic and thyroid tumors compared to matched normal controls, and high expression of RAD18 in tumors is associated with poor prognostic features. Modulating the expression of mutant KRAS in pancreatic cancer cells or mutant BRAF in thyroid cancer cells demonstrates a tight regulation of RAD18 expression in both cancer types. Depletion of RAD18 results in DNA damage and radiation-induced cell death. Importantly, RAD18 depletion in combination with radiotherapy results in marked and sustained tumor regression in KRAS mutant pancreatic cancer orthotopic tumors and BRAF mutant thyroid heterotopic tumors. Overall, our findings identify a novel coordinated RAS/RAF-miR-296-3p-RAD18 signaling network in pancreatic and thyroid cancer cells, which leads to enhanced radioresistance.

Abstract LB031: Discovery and Characterization of HH100937, a potent and selective SOS1 inhibitor demonstrates synergistic efficacy in combination with KRAS/MAPK therapies

Abstract Besides marketed inhibitors targeting KRAS G12C, novel therapies are needed to effectively treat cancer patients harboring other KRAS mutations. SOS1, a guanine nucleotide-exchange factors (GEFs), plays a critical role in catalyzing the conversion of KRAS from its GDP- to GTP-bound form, regardless of KRAS mutation status and represents a promising therapeutic target for all KRAS-driven tumors. Up to date, several SOS1 inhibitors have entered early phase clinical trials. But a more potent SOS1 inhibitor with improved drug-like properties is still needed to prove efficacy as monotherapy or in combination with other drugs targeting the KRAS/MAPK pathway.We discovered a potent SOS1 inhibitor, HH100937 that effectively disrupted the interaction between SOS1 and wild-type KRAS or with G12C, G12D or G12V mutations. HH100937 exhibited lower IC50s than BI-3406 and MTRX0902 in blocking ERK phosphorylation and cancer cell proliferation, leading to better anti-tumor efficacies in xenografts than reference compounds at same doses. HH100937 also showed synergistic effect with AMG510 in cancer cells with KRAS G12C mutation but refractory to AMG510. The combination of HH100937 and AMG510 in xenografts led to tumor regression in all mice, while some tumors did not response well to AMG510 monotherapy. Our SOS1 inhibitor also exhibited synergistic effect with HH2710, a clinical-stage ERK inhibitor, in a broad panel of cancer cells with KRAS mutations.The pharmacological potency of HH100937 was highly selective over SOS2, other kinases tested or other targets in a safety panel of 47 targets. HH100937 was well tolerated in 28-day DRF study in rats and dogs up to 250 mg/kg QD and 30 mg/kg QD, respectively. The AUC of drug exposure in DRF studies was approximately three to six times higher than that at the efficacious dose. HH100937 also showed favorable DMPK properties in mice, rats and dogs, and acceptable safety profiles in vitro.Taken together, HH100937 is a highly potent and selective SOS1 inhibitor with better anti-tumor activities than some SOS1 inhibitors at clinical stage. It can be developed as mono therapy or in combination with other KRAS/MAPK targeting therapies. Citation Format: Qing Xie, Dongsheng Li, Shujuan Jiang, Jie Zhu, Songda Yu, Maozhi Yang, Yalei Cai, Lei Liu, Bing Yu, Wangyang Tu, Yixiang Zhang, Leping Li. Discovery and Characterization of HH100937, a potent and selective SOS1 inhibitor demonstrates synergistic efficacy in combination with KRAS/MAPK therapies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB031.

The Combination of Afatinib With Dasatinib or Miransertib Results in Synergistic Growth Inhibition of Stomach Cancer Cells.

Of various human epidermal growth factor receptor (HER) inhibitors, only the anti-HER2 monoclonal antibody (mAb) Herceptin/trastuzumab and the antibody-drug conjugate trastuzumab deruxtecan (T-Dxd) has been approved for the treatment of patients with stomach cancer. However, the duration of response may be short in many patients, with tumor heterogeneity being one contributing factor. We investigated the effect of various types of targeted agents on growth in vitro and migration of a panel of human stomach cancer cells (HSCCLs) and the impact of cell proliferation rate on the anti-tumor activities of these agents. We also investigated the association between the cell surface expression of the HER family members, hepatocyte growth factor receptor (c-Met), anaplastic lymphoma kinase (ALK)7 and cancer stem cell markers CD44 and CD133, and the response to the targeted agents. Of the 18 agents examined, the cyclin dependent kinase (CDK) 1/2/5/9 inhibitor dinaciclib was the most effective and inhibited the growth of all human HSCCLs at 50% inhibitory concentration (IC50) values between 9 nM to 23 nM. Of various HER inhibitors, the irreversible pan-HER family inhibitors (e.g., afatinib) were more effective than the reversible dual epidermal growth factor receptor (EGFR)/HER2 tyrosine kinase inhibitor (TKI) lapatinib and the EGFR-specific TKI erlotinib in inhibiting the growth of HSCCLs. Of agents targeting different downstream cell signaling molecules, dasatinib targeting Ab1/Src/C-Kit, trametinib targeting MERK1/2 and miransertib targeting AKT1/2/3 inhibited growth of majority of HSCCLs, with the IC50 values ranging from 2 nM to 7 µM. Many of these agents were more effective in inhibiting the growth of HSCCLs when they were proliferating at a slower rate. Treatment with neratinib, afatinib, dinaciclib, dasatinib, stattic, miransertib and paclitaxel significantly inhibited migration of stomach cancer cells. Interestingly, treatment with a combination of afatinib and dasatinib or afatinib and miransertib resulted in synergistic and additive growth inhibition of stomach cancer cells. These results suggest that treatment with a combination of these agents may be of therapeutic value in stomach cancer and warrants further investigations.

Abstract 3273: Tirapazamine sensitizes homologous recombination-proficient cancers by enhancing DNA damage andferroptosis

Abstract Despite its success in HR-deficient tumors, the application of PARP inhibitors (PARPis) in HR-proficient cancers is very limited. Since the majority of cancers are HR-proficient, there is a need for developing strategies to extend the use of PARPis to HR-proficient cancers. We investigated whether the hypoxic cytotoxin, tirapazamine (TPZ) sensitizes HR-proficient cancer cells to PARPis under normoxia and hypoxia (2% O2), and found that TPZ enhanced PARPi cell killing to a greater extent under hypoxia compared to normoxia. Mechanistically, this enhanced killing was due to the induction of DNA double-strand breaks and free radicals that lead to ferroptotic cell death. To determine the role of hypoxia in the anti-tumor effect of the combination treatment, we examined the efficacy of TPZ and PARPi treatment in hypoxic spheroids and subcutaneous xenograft models derived from a panel of HR-proficient human non-small cell lung cancer (NSCLC) cells. We found that PARPis combined with TPZ significantly inhibited the growth of the hypoxic NSCLC spheroids. In addition, the combination treatment significantly inhibited the growth of A549 and CORL105 xenografts that possess significant hypoxic fractions, but did not significantly inhibit the growth of well-oxygenated Calu3 and H3122 xenografts. Taken together, this study demonstrates that TPZ can sensitize HR-proficient tumor cells to PARPi through the introduction of DNA strand breaks and free radicals, suggesting that TPZ treatment can broaden the usage of PARPi to HR-proficient cancers. Citation Format: Yanyan Jiang, Hala Estephan, Yizhou Wang, Ftoon Aljarbou, Nuria V. Lopera, Eui Jung Moon, Ester M. Hammond, Amato J. Giaccia. Tirapazamine sensitizes homologous recombination-proficient cancers by enhancing DNA damage andferroptosis [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 3273.

Abstract 3341: Targeting CXCR4 via the small molecule inhibitor NMX1 as a therapeutic strategy to treat high-risk malignancies

Abstract Introduction: Cytokines are secreted by various cells of the immune system, endothelium, and stroma that play a critical role in cell survival. The activation of C-X-C chemokine receptor type 4 (CXCR4) by its ligand SDF-1 leads to signaling by the AKT and mTOR pathways. CXCR4 is highly expressed in several types of cancer and contributes to tumor growth, metastasis, and resistance to therapy. Interleukin 11 (IL-11) is a cytokine that often drives disease progression, as well as contributing to tumor immune evasion. NMX1 (NovoMedix) is a novel oral small molecule that inhibits mTOR signaling, CXCR4 expression, IL-11 secretion, and activates AMPK. In animal models of breast cancer, NMX1 effectively prevented tumor growth and metastasis, while also protecting from doxorubicin-induced cardiotoxicity. In this study, we investigated the anticancer activity of NMX1 in preclinical models of high-risk pediatric and adult malignancies. Methods: Cell viability assays using alamar blue were performed in a panel of pediatric and adult cancer cells, including leukemia, brain, breast, lung, neuroblastoma, and sarcoma. Cells were treated with increasing concentrations of NMX1 ranging from 250 nM to 16 µM for 96 hours, followed by determination of half maximal inhibitory concentrations (IC50). To validate target modulation, the level of IL-11 and CXCR4 was determined by ELISA and immunoblot, respectively. Protein kinase activity was assessed by phosphorylation levels. Induction of apoptosis was confirmed by caspase-mediated PARP cleavage. To identify effective drug combinations, NMX1-treated cells were screened with a comprehensive drug library. Synergy was evaluated in the top candidates by dose-response matrices and SynergyFinder. Safety and tolerability was tested in animal models. Results: NMX1 treatment induced cytotoxicity at micromolar concentrations in diverse types of cancer, with IC50 values ranging from 0.2-8.7 µM and 0.4-4.4 µM for leukemia and solid tumor cells, respectively. NMX1 induced cancer cell death by PARP cleavage and apoptosis. Mechanistically, CXCR4 expression, mTOR signaling, and IL-11 secretion were decreased upon treatment with NMX1, validating effective target modulation. The drug combination screen identified several candidates in a tumor type-dependent manner. Synergistic effects were observed using NMX1 combined with clinically relevant concentrations of rucaparib, linsitinib, and crizotinib, which target PARP, IGFR, and c-MET, respectively. Conclusions: Our study shows antitumor activity by NMX1 through inhibition of CXCR4, IL-11, and mTOR signaling, providing an effective therapeutic strategy for high-risk and refractory malignancies. Preclinical data on feasible drug combinations and biological correlates of activity support development of early phase clinical studies involving NMX1. Citation Format: Patrick Sipila, Son Tran, Chunfen Zhang, Laura G. Corral, Kyle W. Chan, Cathy A. Swindlehurst, Aru Narendran. Targeting CXCR4 via the small molecule inhibitor NMX1 as a therapeutic strategy to treat high-risk malignancies [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 3341.

Abstract 2117: Elucidation of the fates of CDK2 inhibited aneuploid and residual lung cancers

Abstract Genomic instability is a hallmark of cancer. Unlike normal diploid cells, cancer cells exhibit abnormal centrosome numbers and chromosomal instability. Mitosis with supernumerary centrosomes yields progeny with aberrant chromosome segregation and aneuploidy. While most progeny cells with abnormal chromosomes are eliminated through apoptosis, some have distinct fates with a growth advantage. These cancer cells can acquire resistance to antineoplastic agents. This confers an aggressive tumor biology and unfavorable clinical outcomes. Cyclin-dependent kinase 2 (CDK2) regulates cell cycle progression and the centrosome cycle. We previously reported that CDK2 inhibition prevents supernumerary centrosome clustering, causing multipolar mitosis and anaphase catastrophe in lung and other aneuploid cancers. This was associated with residual transplanted tumors in mice. To elucidate cell fates of progeny with multipolar mitosis after CDK2/9 inhibition with CYC065 (0.2 µM) treatment, a panel of murine and human lung cancer cells underwent time-lapse fluorescent microscopy. This elucidated multipolar mitotic events within the progeny. Intriguingly, distinct outcomes occurred after mitosis including formation of multipolar and multinucleated cells. Cell death of progeny is the predominant fate, but some progeny survive despite continuous CDK2 inhibition. Surprisingly, some of these cells fuse together. Those multinucleated cells can undergo multiple cell cycles without successful cytokinesis. To confirm these effects were through CDK2 inhibition, the selective CDK2 inhibitor Tagtociclib (PF-07104091, at the 2 µM dosage) was used. This treatment statistically-significantly increased multipolar lung cancer cells and increased apoptotic death during time-lapse fluorescent microscopy. Findings were independently validated by CDK2 shRNA knockdown. Focused ion beam scanning electron microscopy (FIB-SEM) and immunofluorescent staining revealed the ultrastructure of multinucleated cells. Insights into tumor biology came from intravital imaging of transplanted lung cancer cells in mice. Multipolar and multinuclear cells followed CDK2 antagonism. Incucyte® Live-Cell Analysis System and Artificial Intelligence-based imaging determined distinct growth responses in aneuploid as compared to non-aneuploid lung cancer cells after CDK2 inhibitor treatments. In summary, these findings are translationally relevant. CDK2 inhibition of aneuploid lung cancers yields distinct cell fates. This is linked to resistance to CDK2 antagonism and to formation of residual in vivo tumors. Citation Format: Liliya Tyutyunyk, Zibo Chen, Xiuxia Liu, Yeap Ng, Aayush Bhatawadekar, Kedar Narayan, Roberto Weigert, Xi Liu, Ethan Dmitrovsky. Elucidation of the fates of CDK2 inhibited aneuploid and residual lung cancers [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 2117.

Abstract 3340: RMC-5127, a first-in-class, orally bioavailable mutant-selective RASG12V(ON) inhibitor is central nervous system (CNS)-penetrant and demonstrates anti-tumor activity in a preclinical intracranial xenograft model

Abstract Patients with advanced cancers harboring activating mutations in RAS, particularly those with non-small cell lung cancer, often develop brain metastases leading to increased morbidity and mortality. KRASG12V is the second most frequent RAS mutation in RAS-addicted cancers, including pancreatic cancer (34 %), colorectal cancer (21 %), and non-small cell lung cancer (19%). RMC-5127 is an orally bioavailable, mutant-selective tri-complex inhibitor of the GTP-bound (ON) form of RASG12V. RMC-5127 non-covalently binds to an abundant intracellular chaperone protein cyclophilin A (CypA), resulting in a binary complex that engages RASG12V(ON) to form a high-affinity tri-complex that sterically inhibits RAS binding to effectors. RMC-5127 drove deep suppression of RAS pathway activity, inhibited cell proliferation, and induced apoptosis in a panel of KRASG12V mutant human cancer cells in vitro but only caused submaximal inhibition in the panel of K/N/HRAS wildtype cancer cells, indicative of selectivity for KRASG12V over K/N/HRAS wildtype. Repeated oral dosing of RMC-5127 resulted in profound and durable anti-tumor activity in subcutaneous CDX and PDX models of KRASG12V mutant NSCLC, PDAC, and CRC in vivo. Dose-dependent exposure of RMC-5127 was observed in the whole brain of naïve mice, indicating the compound is brain penetrant. An intracranial xenograft model of KRASG12V tumors was established in immunodeficient mice to assess the CNS anti-tumor activity of RMC-5127. The intracranial tumor exposure of RMC-5127 was comparable with that observed in the whole brains of naïve mice and was sufficient to drive robust pharmacodynamic responses in the brain tumor. Moreover, RMC-5127 exhibited profound and durable anti-tumor activity in the intracranial model, with tumor regressions at well-tolerated doses. The anti-tumor activity of RMC-5127 in intracranial tumors was consistent with that in subcutaneous tumors at equivalent tumor exposures. In conclusion, these preclinical data demonstrate that RMC-5127 is a CNS-penetrant RASG12V(ON) inhibitor and support further study to determine the potential to benefit patients with advanced RASG12V-mutated cancers, including those that have developed or are at risk of developing brain metastases. Citation Format: Zhe Chen, Andre Eriksson, Bianca Lee, Jay Dinglasan, Nilufar Montazer, Jim Cregg, Anne Edwards, Kate Sanders, Jacqueline A. Smith, David Wildes, Mallika Singh, Zhican Wang, Jingjing Jiang. RMC-5127, a first-in-class, orally bioavailable mutant-selective RASG12V(ON) inhibitor is central nervous system (CNS)-penetrant and demonstrates anti-tumor activity in a preclinical intracranial xenograft model [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 3340.

Abstract 640: In vivo rescue of p53 tumor suppressor function with ADGN-531 across Pan-p53 alterations

Abstract p53 tumor suppressor mutations result in loss of function of p53 in approximately 50% of all human cancers.TP53 missense, nonsense mutations and p53 deletion are present in about 50%, 12% and 5% of all p53 driven cancers, respectively. To date there is no effective treatment for rescuing p53 function in cancers using a pan-p53 approach. The use of ADGN-531 is a new potent strategy combining mRNA with a tumor selective peptide nanocarrier to rescue of p53 tumor suppressor function. ADGN-531 contains proprietary p53-mRNA complexed with proprietary short amphipathic peptides in the form of nanoparticles. ADGN-531 was evaluated on 20 different cancer cell lines harboring various p53 mutations. In-vivo efficacy of IV-administered ADGN-531 nanoparticles was evaluated on colorectal SW403 (p53 nonsense/KRASG12V), osteosarcoma SaOs2 (p53 null) and NSCLC NCI-H358 (p53 null/KRASG12C) mouse xenografts at a dose of 0.25-1.0 mg/kg weekly. ADGN-531/sotorasib synergy was evaluated in vitro on NCI-H358 (p53 null/KRASG12C) and Mia-PACA (p53R48W/KRASG12C) cells and in vivo on NCI-H358 mouse xenograft. ADGN-531 markedly inhibits the growth of a large panel of cancer cells harboring p53-missense or nonsense mutations or p53-deletion, by rescuing p53 transcriptional activity and subsequent expression of its target proteins. ADGN-531 induces cell cycle arrest in G1 due to p21 upregulation and apoptosis following BAX and PUMA activation. IV-administration of ADGN-531 resulted in dose responsive tumor growth inhibition in SaOs2, SW403 and H358 xenografts independent of KRAS status. We demonstrated a strong synergy in a NSCLC H358 xenograft (tumor regression 80%) when restoring P53 function with ADGN-531 (0.5mg/kg, weekly) along with inhibition of KRASG12C mutant using sotorasib (10mg/kg, daily). ADGN-531 potentiated the effect of sotorasib on sotorasib resistant cells harboring high level of KRASG12C in permanent active state. ADGN-531 treatments are well tolerated, no sign of clinical toxicity was detected after single or repeated administrations. ADGN-531 is effective in rescuing p53 function both in vitro and in vivo across a wide range of p53 alterations. Our study provides a proof-of-concept that restoration of tumor suppressor function by ADGN-531 could be used as a single agent therapy in P53 altered tumors independent of other driver mutations e.g., KRAS, or with other therapies for potent combinatorial cancer treatment. Citation Format: Gilles Divita, Audrey Grunenberger, Elodie Czuba, Veronique Josserand, Melanie Guidetti, Neil Desai. In vivo rescue of p53 tumor suppressor function with ADGN-531 across Pan-p53 alterations [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 640.