Tumor Cell Lines Research Articles

Structurally Diverse Limonoids from Trichilia connaroides and Their Antitumor Activities

Comprehensive SummaryTwelve new limonoids (1—12), named trichilitins A—L, were isolated from the leaves and twigs of Trichilia connaroides, together with ten known compounds (13—22). The structures were elucidated by extensive spectroscopic investigations, X‐ray diffraction analyses, and ECD calculations. Compound 1, which belongs to a unique class of ring B‐seco limonoid, has been identified as 6/7/6/5 tetracyclic due to a key Baeyer‐Villiger oxidation. Compounds 2—7 were identified as ring intact limonoids, while compounds 8—10 were established as ring D‐seco ones, and 11 and 12 were determined to be rearranged ones. All of the compounds were tested for cytotoxicity against three human tumor cell lines (HCT‐116, NCl‐H1975, and SH‐SY5Y). Compounds 6, 7, 13, 14, and 19 exhibited significant cytotoxic effects, especially 7 exhibited significant cytotoxic effects against HCT‐116 with an IC50 value of 0.035 μmol·L–1 and was more active than the positive control, doxorubicin with an IC50 value of 0.20 μmol·L–1. Compound 7 effectively induced apoptosis of HCT‐116, which was associated with S‐phase cell cycle arrest. Furthermore, the Western blot analysis showed that compound 7 could induce cell cycle arrest by promoting the expression levels of p53 and p21.

New vatalanib analogs: Design, synthesis, in silico study and biological evaluation for anticancer activity

In our attempts to improve pharmacokinetics and pharmacodynamics characters of vatalanib, four strategies of the drug design were applied to design and synthesis new quinoxaline based vatalanib analogs. Antiproliferative activities of the synthesized compounds were investigated against two tumor cell lines (HepG2 and HCT‐116) using vatalanib as a positive control. The new candidates showed promising anticancer activity against two tested cancer cell lines with IC50 values ranging from 6.04 ± 0.19 to 100.15 ± 3.26 µM, in particular compounds 11b and 11c which were more potent than vatalanib. The most potent was the 4-benzoylquinoxaline derivative 11b (IC50 = 6.04 ± 0.19 µM and 15.58 ± 0.58 µM) compared to vatalanib (IC50 of 25.27 ± 0.84 µM and 43.91 ± 1.64 µM) against HepG2 and HCT116, respectively. The selectivity indices of 11b were found to be approximately 14 and 5 towards HepG2 and HCT116, respectively. While vatalanib showed selectivity indices of about 5 and 3 towards HepG2 and HCT116, respectively. The most promising compound 11b was further investigated in HepG‐2 cells for its apoptotic effect, cell cycle arrest and in vitro VEGFR-2 inhibitory activity as the main mechanisms of cell death. It was found that 11b can induce apoptosis and arrest the cell cycle at the at G1 phase in addition to, compound 11b showed effective inhibition of VEGFR-2 with IC50 of 0.918 ± 0.033 µM compared to vatalanib (IC50 of 0.265 ± 0.009 µM). Deep computational studies were created for the designed compounds including molecular docking, physicochemical properties, profiling pharmacokinetics, ADMET studies, and toxicity predictions to evaluate the prospective drug candidates. The results of the docking study were consistent with biological testing, furthermore, in silico ADMET study revealed the superior pharmacokinetics characters of the new candidates over vatalanib. So, the current work suggests that the 4-benzoylquinoxaline-1-yl-acetamide derivatives, especially 11b, can serve as lead molecules for development of new effective anticancer drugs.

Expression of myoglobin by tumor cells and its role in progression of malignancy

The review examines available literature data on the expression of myoglobin in various tumors and tumor cell lines of non-muscle nature, and the effect of hypoxia, reactive oxygen and nitrogen species, hormones, growth factors, gender and age on this process. The influence of tumor myoglobin on processes occurring in cells – oxidative stress, inhibition of mitochondrial respiration by nitric oxide and fatty acid metabolism is also analyzed, both in the case of intrinsic endogenous (ectopic) expression of small amounts (~1 µM) of myoglobin and overexpression of the protein (~150 µM) via the myoglobin gene embedded in the tumour cell genome. It is concluded that hypoxia-induced intrinsic expression of low concentrations of myoglobin, due to its ability to utilise reactive oxygen and nitrogen species that can damage tumour cells, ensures their better survival, promoting tumour progression and metastasis. Accordingly, this myoglobin expression is generally associated with a more aggressive tumour type and poor prognosis for the course and outcome of the disease, and may thus serve as a “marker” of an aggressive malignancy. In contrast, artificial overexpression of myoglobin can significantly inhibit tumour development and improve disease course by switching cancer cell metabolism from tumour-specific glycolysis to oxidative phosphorylation inherent in healthy tissue. Myoglobin overexpression may thus be an effective therapeutic tool in oncology.

Synthesis, biological evaluation, and in silico studies of lantadene-derived pentacyclic triterpenoids as anticancer agents targeting IKK-β

We report the anticancer activity, structure-activity relationships (SAR), molecular mechanism, and in silico docking studies of nine pentacyclic triterpenoids derived from lantadene A. The NCI-60 cytotoxicity screening of synthesized compounds on 60 human tumor cell lines, representing nine different cancers revealed that compound 6, bearing dual C-3 and C-22 butyryloxy substitutions at the pentacyclic triterpenoid scaffold, displays remarkable potency with mean growth inhibition of 98.7% at 10 μM. The SRB five-dose assay of compound 6 exhibited that it suppresses the growth of all NCI-60 cell lines with a GI50 value in the single-digit micromolar range, except for one. The Western blot analysis revealed that compound 6 inhibits the expression of IKK-β and its down-stream effector NF-κB (p65). Molecular docking analysis of compound 6 with IKK-β showed hydrogen bond interactions with GLN425 and ARG427 and hydrophobic contact with PHE424 residues as the prominent interactions. Molecular dynamics simulation of the docked complex suggested that the complex was fairly stable. Compound 7, possessing the isobutyryloxy groups at the analogous positions of compound 6 was the second-favorable anticancer agent. Conversely, the substitution of the hydroxy group in compounds emerged as the least favorable substitution at both the C-3 and C-22 positions of the pharmacophore. The presence of the 22β-angeloyloxy side-chain, akin to the natural product lantadene A, exhibited moderate favorability for enhanced activity. As a whole, the cytotoxicity data of compounds underscores the preference for linear and extended substitutions at both the C-3 and C-22 positions of the pharmacophore for heightened activity.

Ly6E on tumor cells impairs anti-tumor T-cell responses: a novel mechanism of tumor-induced immune exclusion

BackgroundLymphocyte antigen 6 complex, locus E (Ly6E) has been initially demonstrated to involve in T cell activity and impair viral infectivity. Recently, high expression levels of Ly6E have been reported in tumor microenvironment (TME) of various types of cancers. However, the immunoregulatory mechanism of Ly6E manipulating TME remains unknown.MethodsTCGA database and Kaplan–Meier plotter database were used to evaluate the correlation between Ly6E expression levels and cancer patient survival. After analyzing Ly6E expression levels in human breast cancer tissues and tumor cell lines, we generated Ly6E knockout (KO) and overexpression (OE) mouse cell lines. Cell proliferation ability in vitro and the ability of growth and metastasis in mouse tumor models were compared between KO/OE and wild-type tumor cells. On day 7 after tumor implantation, tumor tissues were separated for flow cytometric assay, bulk RNA sequencing and single-cell RNA sequencing (ScRNA-seq). The role of Ly6E-expressing tumor cell on macrophage was analyzed in vitro.ResultsOur result surprisingly found that high Ly6E expression levels were associated with CD8+ T cell exclusion in tumor tissues and resistance to immunotherapy. Our data showed that knockout of Ly6E in tumor cells prompts tumor regression and inhibits tumor metastases, and Ly6E-OE tumor cells vice versa. The enhanced anti-tumor effect of Ly6E knockout in tumor cells was dependent on T cell response and formed long-lasting memory. The increase in the CD8+ T-cell infiltration into the tumor islet of Ly6E-KO tumors confirmed the role of Ly6E on T cell exclusion. ScRNA-seq analysis showed that M2 macrophages are particularly abundant in the Ly6E-expressing tumor tissues, especially M2-4 macrophage cluster identified by high levels of Arg-1, indicates that Ly6E-expressing tumor cells might restrict T cell infiltration via M2 macrophages. Moreover, in vitro assay showed that cell culture media derived from Ly6E-positive tumor cells promoted macrophage migration and M2 polarization.ConclusionOur study illuminated that Ly6E-expressing tumor cells facilitated the accumulation of M2 macrophages in TME, which contributes to CD8+ T cell exclusion and provides new insights for improving efficacy of cancer immunotherapy.

High-Affinity Fully Human Anti-EpCAM Antibody with Biased IL-2 Exhibits Potent Antitumor Activity.

Monoclonal antibodies (mAbs) are widely used in cancer therapy but often show limited efficacy for solid tumors. Enhancing anti-tumor activity by fusing cytokines to tumor-targeting mAbs, which specifically activate immune cells within the tumor microenvironment, represents a promising strategy. However, the optimal design and therapeutic efficacy of antibody-cytokine fusion formats remain unclear. The epithelial cell adhesion molecule (EpCAM), frequently overexpressed in a variety of carcinomas, serves as the target for immunotherapies. In this study, we identified a fully human mAb targeting EpCAM, designated as m801, from a previously constructed phage-displayed fully human antibody library. By fusing m801 with an IL-2 variant (IL-2v) in two configurations, m801.2 (2 anti-EpCAM Fab + 1 IL-2v) and m801.3 (1 anti-EpCAM Fab + 1 IL-2v), we identified m801.2 as the lead candidate due to its superior biophysical properties, including high thermal stability, homogeneity, and low aggregation. Furthermore, m801.2 showed strong binding affinity to EpCAM, with KD values of 0.6 nM, and an EpCAM-expressing tumor cell line, comparable to the original IgG m801. Additionally, m801.2 exhibited IL-2 receptor β subunit (IL-2Rβ)-biased binding activity, with a KD of 27.3 nM, resulting in superior effective T cell activation. In an SW480 xenograft mice model, m801.2 significantly inhibited tumor growth and demonstrated high tolerability. These findings suggest a valuable framework for the future design of immunocytokine therapies.

Synthesis and Cytotoxic Activity of Quinazoline-benzofuran Conjugates.

In order to study on structure-activity relationships of benzofurans. Benzofuran is a kind of natural compound widely existing in nature with pharmacological effects. The development of new anticancer benzofuran derivatives has attracted more and more attention. We have introduced an active quinazoline unit into piperazine-substituted benzofuran, prepared a series of quinazoline-benzofuran compounds, and evaluated cytotoxic activity against a panel of human tumor cell lines by MTT assay. 48 novel quinazoline-substituted benzofuran derivatives have been prepared, and in vitro, cytotoxic activity against five human tumor cell lines was evaluated. The results indicated that some quinazoline-benzofuran conjugates showed selective inhibitory activity against tumor cell lines. We have found that compound 14x displayed excellent cytotoxic activity, which could be considered a potential anticancer agent.

Design, Synthesis, and Biological Evaluation of Some Novel Retinoid Derivatives

Background: As cancer stands as a significant global health concern, many heterocyclic compounds that are more effective in cancer cells than healthy cells are being investigated for their selective anticancer potentials. One such compound is fenretinide, a synthetic derivative of retinoic acid that has a broad spectrum of cytotoxic activity against primary tumor cells, cell lines, and/or xenografts of various cancers. In this context, bexarotene and its derivatives, synthesized from hybridization of the fenretinide, are expected to possess a potential anticancer activity. Objective: The objective of the present study was to investigate the synthesis of novel amid-derived and bexarotene-based compounds, as well as to assess their cytotoxic effects in different cancer cell lines. Methods: This study involved the synthesis of twelve novel retinoid derivatives (6-17) in a six-step process. The cytotoxic activities of these compounds were assessed against various cancer cell lines, such as A549 (human lung carcinoma), HeLa (human cervical cancer), MCF7 (human breast adenocarcinoma), and WiDr (human colon adenocarcinoma). The chemical structures of these compounds were elucidated through their elemental analysis, mass spectrometry (ESI+, ESI-), as well as 1H-NMR and 13C-NMR spectroscopic data. Results: The obtained cell toxicity results indicated that compounds 6, 8, 11, 12, 13, 14, and 17 were found to exhibit the strongest cytotoxic activity in above mentioned cancer cell lines. The IC50 values for active compounds, 11 and 12, were determined as 2.38μM and 2.29μM, respectively. Remarkably, these compounds displayed higher cytotoxic activity in the WiDr cell line related to positive control, camptothecin (CPT). Moreover, compounds 14 and 17 demonstrated very similar level of cytotoxic activity to CPT, indicating their potential for antitumoral applications upon further studies. Conclusion: While compounds 11, 12, 14, and 17 indicated a very comparable anticancer activity to CPT, compounds 6, 8, 11 and 12 showed more selective anticancer effect against cancer cells than noncancerous cells. In accordance with the findings of the present study, they can be evaluated as primary candidates for further studies, specifically as RXRα-targeted anticancer agents.

Synthesis, Docking, and Dynamic Studies of New Pyrano‐[3,2‐e]‐Pyrazole Fused 1,2,4‐Triazolo‐[4,3‐c]‐pyrimidine Analogues as Anticancer Agents

AbstractA novel series of dihydropyrazolo‐[4′,3′:5,6]‐pyrano[3,2‐e][1,2,4]triazolo[4,3‐c]‐pyrimidine analogues were designed, synthesized, and their cytotoxic potential against four tumor cell lines assessed. HRMS, 1H NMR, and 13C NMR spectra analysis were used to describe the novel compounds. Compound 9c had noteworthy anticancer activity in vitro against the Hela, MCF‐7, A549, and A2780 cell lines, with IC50 values of 0.98 ± 0.26, 3.11 ± 0.18, 1.24 ± 0.88, and 2.62 ± 0.56 µM, in that order. The molecular docking and dynamics simulation studies revealed the lead molecule's action mechanism with EGFR target protein.

Antitumor activities of anti‑CD44 monoclonal antibodies in mouse xenograft models of esophageal cancer.

CD44 is a type I transmembrane glycoprotein associated with poor prognosis in various solid tumors. Since CD44 plays a critical role in tumor development by regulating cell adhesion, survival, proliferation and stemness, it has been considered a target for tumor therapy. Anti‑CD44 monoclonal antibodies (mAbs) have been developed and applied to antibody‑drug conjugates and chimeric antigen receptor‑T cell therapy. Anti-pan‑CD44 mAbs, C44Mab‑5 and C44Mab‑46, which recognize both CD44 standard (CD44s) and variant isoforms were previously developed. The present study generated a mouse IgG2a version of the anti‑pan‑CD44 mAbs (5‑mG2a and C44Mab‑46‑mG2a) to evaluate the antitumor activities against CD44‑positive cells. Both 5‑mG2a and C44Mab‑46‑mG2a recognized CD44s‑overexpressed CHO‑K1 (CHO/CD44s) cells and esophageal tumor cell line (KYSE770) in flow cytometry. Furthermore, both 5‑mG2a and C44Mab‑46‑mG2a could activate effector cells in the presence of CHO/CD44s cells and exhibited complement-dependent cytotoxicity against both CHO/CD44s and KYSE770 cells. Furthermore, the administration of 5‑mG2a and C44Mab‑46‑mG2a significantly suppressed CHO/CD44s and KYSE770 xenograft tumor development compared with the control mouse IgG2a. These results indicate that 5‑mG2a and C44Mab‑46‑mG2a could exert antitumor activities against CD44‑positive cancers and be a promising therapeutic regimen for tumors.

Deep learning-driven macroscopic AI segmentation model for brain tumor detection via digital pathology: Foundations for terahertz imaging-based AI diagnostics

We used deep learning methods to develop an AI model capable of autonomously delineating cancerous regions in digital pathology images (H&E-stained images). By using a transgenic brain tumor model derived from the TS13-64 brain tumor cell line, we digitized a total of 187 H&E-stained images and annotated the cancerous regions in these images to compile a dataset. A deep learning approach was executed through DEEP:PHI, which abstracts Python coding complexities, thereby simplifying the execution of AI training protocols for users. By employing the Image Crop with Mask technique and patch generation method, we not only maintained an appropriate data class balance but also overcame the challenge of limited computing resources. This approach enabled us to successfully develop an AI training model that autonomously segments cancerous areas. This AI model enables the provision of guiding images for determining cancerous areas with minimal assistance from neuropathologists. In addition, the high-quality, large dataset curated for training using the proposed approach contributes to the development of novel terahertz imaging-based AI cancer diagnosis technologies and accelerates technological advancements.

Copper drives remodeling of metabolic state and progression of clear cell renal cell carcinoma.

Copper (Cu) is a cofactor of cytochrome c oxidase (CuCOX), indispensable for aerobic mitochondrial respiration. This study reveals that advanced clear cell renal cell carcinomas (ccRCCs) accumulate Cu, allocating it to CuCOX. Using a range of orthogonal approaches, including metabolomics, lipidomics, isotope-labeled glucose and glutamine flux analysis, and transcriptomics across tumor samples, cell lines, xenografts, and PDX models, combined with genetic and pharmacological interventions, we explored Cu's role in ccRCC. Elevated Cu levels stimulate CuCOX biogenesis, providing bioenergetic and biosynthetic benefits that promote tumor growth. This effect is complemented by glucose-dependent glutathione production, which facilitates detoxification and mitigates Cu-H2O2 toxicity. Single-cell RNA sequencing (scRNA-seq) and spatial transcriptomics reveal increased oxidative metabolism, altered glutathione and Cu metabolism, and diminished HIF activity during ccRCC progression. Thus, Cu drives an integrated oncogenic remodeling of bioenergetics, biosynthesis, and redox homeostasis, fueling ccRCC growth, which can be targeted for new therapeutic approaches.

Synthesis and Antitumour Activity of Hybrid Compounds Based on 4-Aminophenylarsonic Acid and Spatially Hindered Phenols.

One of the main modern approaches to the creation of effective drugs is the design of new biologically active substances containing two or more pharmacophore groups in their structure. In recent years, there have been many publications on the synthesis and study of biological activity, including antitumour activity, of new organo-arsenic compounds. It is known that spatially hindered phenols can also have antitumor activity, so the synthesis and study of hybrid compounds based on organo-arsenic compounds and spatially hindered phenols is a relevant area of research. In this work, the modification of 4-aminophenylarsonic acid with 3,5-di-tert-butyl-4-hydroxybenzylacetate was carried out. In contrast to a similar transformation of 2-aminophenylarsonic acid, in this case it was possible to obtain both mono- and di-benzyl derivatives of the acid. Using the Zandmeyer method, the oxime isatin containing an arsonic acid fragment in the fifth position of the heterocycle was synthesised. Azo derivatives containing fragments of para-aminophenylarsonic acid and sterically hindered phenols were obtained. 4-((3,5-Di-tert-butyl-2-hydroxyphenyl)diazenyl)phenylarsonic acid was isolated in pure form. At the same time, it was found that the reaction of the diazonium azo salt of 4-aminophenylarsonic acid with 2-hydroxymethyl-4-tert-butylphenol proceeds in two directions. In addition to the classical diazotisation reaction at the 6-position of 2-hydroxymethyl-4-tert-butylphenol, a diazotisation accompanied by a dehydroxymethylation process occurs. The obtained compounds showed cytotoxic activity against human tumor cell lines M-HeLa (cervical epithelioid carcinoma) and HuTu 80 (duodenal adenocarcinoma cells). The most promising is the sodium salt of 4-((3,5-di-tert-butyl-2-hydroxyphenyl)diazenyl)phenylarsonic acid, which is superior to Tamoxifen and 5-fluorouracil in terms of selectivity index towards M-HeLa and HuTu 80 cells.

Growth of Renal Cancer Cell Lines Is Strongly Inhibited by Synergistic Activity of Low-Dosed Amygdalin and Sulforaphane.

Background: Plant derived isolated compounds or extracts enjoy great popularity among cancer patients, although knowledge about their mode of action is unclear. The present study investigated whether the combination of two herbal drugs, the cyanogenic diglucoside amygdalin and the isothiocyanate sulforaphane (SFN), influences growth and proliferation of renal cell carcinoma (RCC) cell lines. Methods: A498, Caki-1, and KTCTL-26 cells were exposed to low-dosed amygdalin (1 or 5 mg/mL), or SFN (5 µM) or to combined SFN-amygdalin. Tumor growth and proliferation were analyzed by MTT, BrdU incorporation, and clone formation assays. Cell cycle phases and cell cycle-regulating proteins were analyzed by flow cytometry and Western blotting, respectively. The effectiveness of the amygdalin-SFN combination was determined using the Bliss independence model. Results: 1 mg/mL amygdalin or 5 µM SFN, given separately, did not suppress RCC cell growth, and 5 mg/mL amygdalin only slightly diminished A498 (but not Caki-1 and KTCTL-26) cell growth. However, already 1 mg/mL amygdalin potently inhibited growth of all tumor cell lines when combined with SFN. Accordingly, 1 mg/mL amygdalin suppressed BrdU incorporation only when given together with SFN. Clonogenic growth was also drastically reduced by the drug combination, whereas only minor effects were seen under single drug treatment. Superior efficacy of co-treatment, compared to monodrug exposure, was also seen for cell cycling, with an enhanced G0/G1 and diminished G2/M phase in A498 cells. Cell cycle regulating proteins were altered differently, depending on the applied drug schedule (single versus dual application) and the RCC cell line, excepting phosphorylated Akt which was considerably diminished in all three cell lines with maximum effects induced by the drug combination. The Bliss independence analysis verified synergistic interactions between amygdalin and SFN. Conclusions: These results point to synergistic effects of amygdalin and SFN on RCC cell growth and clone formation and Akt might be a relevant target protein. The combined use of low-dosed amygdalin and SFN could, therefore, be beneficial as a complementary option to treat RCC. To evaluate clinical feasibility, the in vitro protocol must be applied to an in vivo model.

Design, Synthesis and Biological Activity Evaluation of β-Carboline Derivatives Containing Nitrogen Heterocycles.

A series of β-carboline derivatives containing nitrogen heterocycles were designed and synthesized. All compounds were screened for their antitumor activity against four human tumor cell lines (A549, K562, PC-3, T47D). Notably, compound N-(4-(morpholinomethyl)phenyl)-2-((5-(1-(3,4,5-trimethoxyphenyl)-9H-pyrido[3,4-b]indol-3-yl)-1,3,4-oxadiazol-2-yl)thio)acetamide (8q) exhibited significant inhibitory activity against PC-3 cells with an IC50 value of 9.86 µM. Importantly, compound 8q effectively suppressed both the proliferation and migration of PC-3 cells. Mechanistic studies revealed that compound 8q induced cell apoptosis and caused the accumulation of reactive oxygen species (ROS), leading to cell cycle arrest in the G0/G1 phase in PC-3 cells.

Five Unreported Ketone Compounds-Penicrustones A-E-From the Endophytic Fungus Penicillium crustosum.

Five unreported ketone compounds-penicrustones A-E-were isolated from the solid fermentation of the endophytic fungus Penicillium crustosum. Their structures were elucidated on the basis of extensive spectroscopic analysis. Their absolute configurations were determined via ECD calculations and single-crystal X-Ray crystallography. All compounds were evaluated for their antimicrobial and antitumor activities. Compounds 4 and 5 showed moderate inhibitory effects on Micrococcus luteus, with MIC values of 12.5 and 25.0 μg/mL, respectively. In addition to this, compound 4 also showed cytotoxicity on tumor cell lines KTC-1 and Hela, with IC50 values of 4.28 and 4.64 μg/mL, respectively.

Ferroptosis-related gene signature for predicting prognosis and identifying potential therapeutic drug in EGFR wild-type lung adenocarcinoma

Epidermal growth factor receptor wild type lung adenocarcinoma (EGFRWT LUAD) still has limited treatment options and unsatisfactory clinical outcomes. Ferroptosis, as a form of cell death, has been reported to play a dual role in regulating tumor cell survival. In this study, we constructed a 3-ferroptosis-gene signature, FeSig, and verified its accuracy and efficacy in predicting EGFRWT LUAD prognosis at both the RNA and protein levels. Patients with higher FeSig scores were found to have worse clinical outcomes. Additionally, we explored the relationship between FeSig and tumor microenvironment, revealing that enhanced interactions between fibroblasts and tumor cells in FeSighigh patients causing tumor resistance to ferroptosis. To address this challenge, we screened potential drugs from NCI-60 (The US National Cancer Institute 60 human tumour cell line anticancer drug screen) and Connectivity map database, ultimately identifying 6-mercatopurine (6-MP) as a promising candidate. Both in vitro and in vivo experiments demonstrated its efficacy in treating FeSighigh EGFRWT LUAD tumor models. In summary, we develop a novel FeSig for predicting prognosis and guiding drug application.

Novel Thiazole-Fused [4,5-g] or [5,4-g]Quinazolin-8-ones and Their Quinazoline Analogues: Synthesis and Biological Evaluation.

Background/Objectives: In connection with previous work on V-shaped polycyclic thiazolo[5,4-f]quinazolin-9-one and [5,4-f]quinazoline derivatives that can modulate the activity of various kinases, the synthesis of straight thiazole-fused [4,5-g] or [5,4-g]quinazolin-8-ones and quinazoline derivatives hitherto undescribed was envisioned. Methods: An innovative protocol allowed to obtain the target structures. The synthesis of inverted thiazolo[4,5-h] and [5,4-h]quinazolin-8-one derivatives was also explored with the aim of comparing biological results. The compounds obtained were tested against a representative panel of eight mammalian protein kinases of human origin: CDK9/CyclinT, Haspin, Pim-1, GSK-3β, CK-1ε, JAK3, CLK1 and DYRK1A. Results and Conclusions: The results obtained show that the novel linear thiazoloquinazolines are not kinase inhibitors. The cytotoxicity of these newly synthesized compounds was assessed against seven representative tumor cell lines (human cancers: Huh7-D12, Caco-2, HCT-116, MCF-7, MDA-MB-231, MDA-MB-468 and PC-3). The majority of these novel molecules proved capable of inhibiting the growth of the tested cells. The 7-Benzyl-8-oxo-7,8-dihydrothiazolo[5,4-g]quinazolinones 5b and 6b are the most potent, with IC50 values in the micromolar range. None of these compounds showed toxicity against normal cells. A larger program of investigations will be launched to investigate the real potential interest of such compounds in anticancer applications.

Naphthoindole-2-carboxamides as a lipophilic chemotype of hetarene-anthraquinones potent against P-gp resistant tumor cells

The acquisition of multidrug resistance (MDR) to chemotherapy is a major obstacle to successful cancer treatment. Aiming to improve the potency of anthraquinone-derived antitumor compounds against MDR cancer cells, we employed a rational design approach to develop new heteroarene-fused anthraquinones. Shifting the carboxamide group in the naphtho[2,3-f]indole scaffold from the 3-position to 2 increased the lipophilicity and P-glycoprotein (P-gp) binding of the derivatives, potentially enhancing their ability to circumvent P-gp-mediated MDR. To validate the computations, we developed a scheme for heterocyclization into esters of naphtho[2,3-f]indole-2-carboxylic acid, based on the 5-endo-dig cyclization of 2-alkynyl-3-amino-1,4-dimethoxyanthraquinone under mild basic conditions using tetra-n-butylammonium fluoride (TBAF). The synthesized naphthoindole-2-carboxamides, particularly compound 1a bearing (S)-3-aminopyrrolidine in the carboxamide fragment, demonstrated the highest antiproliferative activity. Most importantly, 1a suppressed the growth of the P-gp-positive K562/4 leukemia tumor cell line (resistance index = 2.4), while its 3-isomer LCTA-2640 and Dox did not (RI = 125 and 140, respectively). Studies of intracellular uptake and distribution showed that 1a, unlike its 3-substituted isomer, effectively accumulated in resistant tumor cells, confirming the correlation between in silico and experimental data. The lead compound 1a interacts with DNA duplex and inhibits topoisomerase 1 but does not induce oxidative stress. Treatment with 1a increases the population of apoptotic cells in both K562 and K562/4 sublines, regardless of the cell cycle phase. Taken together, this work provides an interesting example of how a little modification in chemical structure can lead to striking differences in antitumor properties. In conclusion, we have identified a potent class of compounds that offer distinct advantages in combating resistant tumor cells.

Endogenous EWSR1-FLI1 degron alleles enable control of fusion oncoprotein expression in tumor cell lines and xenografts.

Pediatric malignancies frequently harbor chromosomal translocations that induce expression of fusion oncoproteins. The EWSR1-FLI1 fusion oncoprotein acts as a neomorphic transcription factor and is the dominant genetic driver of Ewing's sarcoma. Interrogation of the mechanisms by which EWSR1-FLI1 drives tumorigenesis has been limited by a lack of model systems to precisely and selectively control its expression in patient-derived cell lines and xenografts. Here, we report the generation of a panel of patient-derived EWS cell lines in which inducible protein degrons were engineered into the endogenous EWSR1-FLI1 locus. These alleles enabled rapid and efficient depletion of EWSR1-FLI1. Complete suppression of EWSR1-FLI1 induced a reversible cell cycle arrest at the G1-S checkpoint, and we identified a core set of transcripts downstream of EWSR1-FLI1 across multiple cell lines and degron systems. Additionally, depletion of EWSR1-FLI1 potently suppressed tumor growth in xenograft models validating efforts to directly target EWSR1-FLI1 in Ewing's sarcoma.