Anticancer Therapeutic Target Research Articles

Discovery of a Novel and Potent Cyclin-Dependent Kinase 8/19 (CDK8/19) Inhibitor for the Treatment of Cancer.

The mediator kinases CDK8 and CDK19 control the dynamic transcription of selected genes in response to various signals and have been shown to be hijacked to sustain hyperproliferation by various solid and liquid tumors. CDK8/19 is emerging as a promising anticancer therapeutic target. Here, we report the discovery of compound 12, a novel small molecule CDK8/19 inhibitor. This molecule demonstrated not only decent enzymatic and cellular activities but also remarkable selectivity in CDK and kinome panels. Besides, compound 12 also displayed favorable ADME profiles including low CYP1A2 inhibition, acceptable clearance, and high oral bioavailability in multiple preclinical species. Robust in vivo PD and efficacy studies in mice models further demonstrated its potential use as mono- and combination therapy for the treatment of cancers.

Jolkinolide B synergistically potentiates the antitumor activity of GPX4 inhibitors via inhibiting TrxR1 in cisplatin-resistant bladder cancer cells

Glutathione peroxidase 4 (GPX4) is a promising anticancer therapeutic target; however, the application of GPX4 inhibitors (GPX4i) is limited owing to intrinsic or acquired drug resistance. Hence, understanding the mechanisms underlying drug resistance and discovering molecules that can overcome drug resistance are crucial. Herein, we demonstrated that GPX4i killed bladder cancer cells by inducing lipid reactive oxygen species-mediated ferroptosis and apoptosis, and cisplatin-resistant bladder cancer cells were also resistant to GPX4i, representing a higher half-maximal inhibitory concentration value than that of parent bladder cancer cells. In addition, thioredoxin reductase 1 (TrxR1) overexpression was responsible for GPX4i resistance in cisplatin-resistant bladder cancer cells, and inhibiting TrxR1 restored the sensitivity of these cells to GPX4i. In vitro and in vivo studies revealed that Jolkinolide B (JB), a natural diterpenoid and previously identified as a TrxR1 inhibitor, potentiated the antiproliferative efficacy of GPX4i (RSL3 and ML162) against cisplatin-resistant bladder cancer cells. Furthermore, GPX4 knockdown and inhibition could augment JB-induced paraptosis and apoptosis. Our results suggest that inhibiting TrxR1 can effectively improve GPX4 inhibition-based anticancer therapy. A combination of JB and GPX4i, which is well-tolerated and has several anticancer mechanisms, may serve as a promising therapy for treating bladder cancer.

Abstract 1800: Cytosolic NADK is conditionally essential for folate-dependent nucleotide synthesis in human cells

Abstract The genes that are required for the growth of mammalian cells can depend on an interplay of cell-intrinsic factors and environmental context. However, there is often little investigation into how nutrient availability impacts gene essentiality. Moreover, efforts such as DepMap have catalogued genes that contribute to cancer cell fitness using in vitro CRISPR screens in hundreds of cancer cell lines cultured in traditional media that poorly recapitulate the nutrient availability of human blood. Previously, we developed Human Plasma-Like Medium (HPLM), a physiologic medium designed to better model the nutrient availability of human blood. We recently tested the hypothesis that nutrient availability influences gene essentiality. By performing paired CRISPR-based screens in human blood cancer cells, we identified sets of genes that are differentially required to support cell growth in HPLM versus traditional media such as RPMI. Among the strongest scoring HPLM-essential genes was NAD kinase (NADK), which encodes an enzyme critical for the generation of cytosolic NADPH. NADK has been previously suggested as an anti-cancer therapeutic target, given the central importance of NADPH for enabling proliferative metabolism through macromolecule synthesis and ROS management. Interestingly, NADK has been annotated as an essential gene in only 1% of the cell lines included in DepMap, suggesting that the essentiality of NADK may have been largely masked by the nutrient conditions used to generate the DepMap. To investigate the conditional CRISPR phenotype for NADK, we engineered NADK-knockout cells and found that they showed a 50% stronger growth defect versus control cells in HPLM relative to RPMI. Next, we used localization and activity studies to confirm that cytosolic NAD+ kinase activity is required for the conditionally essential role of NADK. We then employed systematic approaches in media engineering and metabolomics to identify the medium component(s) that contribute to the NADK gene-nutrient interaction and to gain insights into the conditionally essential role of NADK. Through these studies, we traced the cause of NADK essentiality to an inhibition of folate-dependent nucleotide synthesis linked to differential folic acid availability. Finally, we examined the pre-translational potential of NADK as an anti-cancer therapeutic target by demonstrating that the reported NADK inhibitor thionicotinamide largely phenocopies NADK-knockout in a folic-acid dependent manner in the K562 cell line. Collectively, our work improves understanding of how NADK contributes to human cell growth and reveals how the relative importance of cytosolic NAD+ kinase activity can depend on nutrient availability. Citation Format: Kyle M. Flickinger, Kimberly S. Huggler, Jason R. Cantor. Cytosolic NADK is conditionally essential for folate-dependent nucleotide synthesis in human cells [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 1800.

Abstract 4119: CXCR4/CB2 heterodimer antagonizes CXCR4-mediated metastasis and tumor growth in vivo

Abstract The G-protein-coupled chemokine receptor (GPCR), CXCR4, is ubiquitously expressed in malignant tumor tissues, and controls migration and other tumor survival mechanisms upon interaction with its chemokine agonist, stromal cell-derived 1α (SDF1α). The CXCR4/SDF1α axis is relevant to shaping the tumor microenvironment mainly the metastatic spread of primary tumors to distal organs and immune response; thus, expression of CXCR4 in tumors is vital to tumor aggressiveness, survival probability, and metastasis-associated mortality. CXCR4 signals through a homodimer, but also forms heterodimers with other chemokine and non-chemokine GPCRs. We’ve demonstrated that CXCR4 forms a heterodimer with the GPCR, cannabinoid receptor 2 (CB2), in human breast and prostate cell lines in vitro. The cannabinoid system has earned interest as anticancer therapeutic targets by modulating growth, migration, angiogenesis, etc. In our model, CXCR4-mediated, pro-tumor signaling was reduced by the physical heterodimer, and the dimer was induced by simultaneous treatment with agonists (SDF1α and AM1241) for respective receptors. The resultant heterodimer negatively interfered with migration, cytoskeletal rearrangement, motility associated proteins, and signaling that coordinates movement. Given that cannabinoids have been shown to reduce metastasis and profession of clinical tumors, we hypothesize that a heterodimer of CXCR4 and CB2 disrupts CXCR4-mediated metastasis, and ultimately, decrease tumor progression in vivo.Herein, we tested our hypothesis in an in vivo model of spontaneous metastasis NSG male mice. Luciferase-containing human prostate cancer cells (DU145 RFP/Luc) were implanted subcutaneously. Once tumors reached 40mm3, mice were randomized by volume, weight and age, and divided into 4 treatments: vehicle, SDF1α, AM124, and SDF1α/AM124(combo) for daily intraperitoneal injections. Tumors reaching 400mm3 were excised, and treatments continued for 3 weeks. Spontaneous metastasis was monitored via IVIS imaging at 2- and 3-weeks post-tumor removal. We observed reduced tumor growth, and decreased incidence rates of metastasis to the lung in the combination treatment group, which, based on our in vitro data, is representative of a CXCR4/CB2 heterodimer. This novel definition of heterodimerization could reveal and exploit new mechanisms for CXCR4 regulation in tumor metastasis. The CXCR4/CB2 heterodimer may attenuate responses triggered individually by CXCR4 without toxicity and side effects experienced with current CXCR4 antagonists. Citation Format: Nakea Michelle Pennant, Alifiani B. Hartono, Shiqin Liu, Christopher Massey, Chung Lee, Tanya Stoyanova, Cimona V. Hinton. CXCR4/CB2 heterodimer antagonizes CXCR4-mediated metastasis and tumor growth in vivo [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 4119.

Abstract 5136: Development of a highly specific NAPRT monoclonal antibody for the detection of tumors sensitive to pharmacological inhibition of NAMPT

Abstract Introduction: A key component of targeting synthetic lethality in cancer is identifying specific biomarkers, such as sources of NAD+ synthesis, that differentiate between healthy and diseased tissue. In mammals, NAD+ can be synthesized from nicotinamide via nicotinamide phosphoribosyltransferase (NAMPT) or from nicotinic acid (niacin, NA) via nicotinic acid phosphoribosyltransferase (NAPRT). NAPRT loss has been identified across tumor indications as a synthetic lethal sensitizer to small molecule inhibitors of NAMPT. Given the reliance of cancer cells on NAD+ synthesis, blocking NAD+ production is a compelling target for anti-cancer therapeutics. However, identifying tumors with NAPRT loss has been a challenge as NAPRT can be expressed by as many as 14 transcripts, only some of which code for the active enzyme. This complicates transcript-based analyses and necessitates proteomic assessment. Here, we have developed a mouse monoclonal antibody (4A5D7) to specifically detect active NAPRT and to identify indications enriched for NAPRT loss in patients through an IHC-based assay. We further confirm that loss of NAPRT makes cells susceptible to NAMPT inhibition, and ultimately show that our antibody can identify patient-derived xenograft (PDX) models that respond to NAMPT inhibition. Methods We utilized a hybridoma system to produce antibodies against epitopes in the NAPRT active site; clones were subsequently screened for their ability to selectively recognize active NAPRT. After identifying clone 4A5D7, we validated our proprietary antibody against FFPE specimens across multiple cancer indications (TNBC, NSCLC, SCLC, CRC, gastric, ovarian, and prostate). Stained samples were scored by a pathologist and assessed for percent of positive tumor cells, nuclear and/or cytoplasmic staining, and staining intensity. CCLE and TCGA datasets were mined for indications enriched for suspected NAPRT-loss and sensitivity to NAMPT inhibitors. We then selected cell lines and pan-tumor PDX tissue microarrays (TMAs) from indications of interest and characterized samples by western blot and IHC to determine NAPRT status. Cell lines and PDX tumor tissue were exposed to increasing ranges of NAMPTi in the presence or absence of NAPRT-substrate NA to validate our approach. Results: We validated that our proprietary NAPRT monoclonal antibody clone 4A5D7 only detects functional NAPRT and demonstrated improved performance over previous antibodies. We used our antibody to characterize cell lines and PDX tumor samples, and subsequently show that NAPRT-null cell lines and tumor samples are susceptible to NAMPT inhibition, while samples with functional NAPRT can be rescued by NA. Conclusion: Our proprietary antibody can identify NAPRT null tumors and identify samples that will respond to NAMPT inhibition, a critical advancement for patient selection in clinical trials. Citation Format: Josh Spurrier, Emmanuel S. Burgos, Mark R. Lundquist, Keiichiro Tanaka, Samuel Brook, Matt Gozzi, Jae-Sung Yi, Joseph Edmonds, Mitch Raponi. Development of a highly specific NAPRT monoclonal antibody for the detection of tumors sensitive to pharmacological inhibition of NAMPT [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 5136.

Abstract 2091: Pharmacological inhibition of chaperone-mediated autophagy via NCoR1/RARα targeting is effective against non-small cell lung cancer

Abstract Dysregulation of autophagy, the lysosomal degradation of intracellular components, has been universally described in cancer. While activity of macroautophagy changes in a context-dependent manner in cancers, the upregulation of chaperone-mediated autophagy (CMA) is well-described in all solid tumors studied to date. Upregulated CMA directly contributes to both tumor growth and protection against anti-oncogenic interventions in cancers. Selective degradation of anti-oncogenic and pro-apoptotic proteins, such as mutant p53 and PUMA, misfolded proteins, and glycolytic enzymes by CMA allows for increased survival, reduced cell death, increased stress resistance, and enhancement of the Warburg effect in cancer cells, respectively. As such, inhibition of CMA is an attractive target for anti-cancer therapeutics, but the field currently lacks protein targets specific to CMA and selective CMA inhibitors. We have previously demonstrated the regulatory effect of retinoic acid receptor alpha (RARα) and its coregulator, nuclear receptor corepressor 1 (NCoR1), on CMA through transcriptional upregulation of the CMA effector proteins (Bourdenx et al. Cell 2021, Gomez-Sintes et al, Nat. Commun. 2022). Here, we identified a high NCoR1/RARα expression ratio in human non-small cell lung cancer (NSCLC) tumors compared to healthy lung tissue. We further illustrated a dependency on NCoR1 expression and its interaction with RARα to maintain high CMA activity in NSCLC cell lines. Provided the reliance on NCoR1 repression of RARα for high CMA activity in NSCLC, we hypothesized that small molecule targeting of the druggable NCoR1/RARα interaction to disrupt the interaction may result in suppression of CMA effector proteins and CMA inhibition. Through a series of in-silico screens for predicted NCoR1/RARα targeting, and functional screens for CMA inhibition, we identified CIM7, a first-in-class potent and selective CMA inhibitor. We validated CIM7 function through binding to RARα and subsequent disruption of NCoR1 binding, resulting in alterations to CMA-related gene expression. CIM7 inhibits CMA in vivo with no observed toxicity in normal tissues. We additionally demonstrated the therapeutic potential of CIM7 through its ability to reduce tumor growth in vivo in mouse NSCLC xenografts. Our findings support the NCoR1/RARα interaction as a major regulator of CMA in NSCLC and demonstrate the potential of pharmacologic CMA inhibition as an anti-cancer therapeutic strategy. Citation Format: Mericka McCabe, Rabia R. Khawaja, Rajanya Bhattacharyya, Antonio Diaz, Thomas P. Garner, Kristen Lindenau, Rebecca Sereda, Olaya Santiago-Fernández, Ana Maria Cuervo, Evripidis Gavathiotis. Pharmacological inhibition of chaperone-mediated autophagy via NCoR1/RARα targeting is effective against non-small cell lung cancer [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 2091.

AURKB promotes bladder cancer progression by deregulating the p53 DNA damage response pathway via MAD2L2

BackgroundBladder cancer (BC) is the most common urinary tract malignancy. Aurora kinase B (AURKB), a component of the chromosomal passenger protein complex, affects chromosomal segregation during cell division. Mitotic arrest-deficient 2-like protein 2 (MAD2L2) interacts with various proteins and contributes to genomic integrity. Both AURKB and MAD2L2 are overexpressed in various human cancers and have synergistic oncogenic effects; therefore, they are regarded as emerging therapeutic targets for cancer. However, the relationship between these factors and the mechanisms underlying their oncogenic activity in BC remains largely unknown. The present study aimed to explore the interactions between AURKB and MAD2L2 and how they affect BC progression via the DNA damage response (DDR) pathway.MethodsBioinformatics was used to analyze the expression, prognostic value, and pro-tumoral function of AURKB in patients with BC. CCK-8 assay, colony-forming assay, flow cytometry, SA-β-gal staining, wound healing assay, and transwell chamber experiments were performed to test the viability, cell cycle progression, senescence, and migration and invasion abilities of BC cells in vitro. A nude mouse xenograft assay was performed to test the tumorigenesis ability of BC cells in vivo. The expression and interaction of proteins and the occurrence of the senescence-associated secretory phenotype were detected using western blot analysis, co-immunoprecipitation assay, and RT-qPCR.ResultsAURKB was highly expressed and associated with prognosis in patients with BC. AURKB expression was positively correlated with MAD2L2 expression. We confirmed that AURKB interacts with, and modulates the expression of, MAD2L2 in BC cells. AURKB knockdown suppressed the proliferation, migration, and invasion abilities of, and cell cycle progression in, BC cells, inducing senescence in these cells. The effects of AURKB knockdown were rescued by MAD2L2 overexpression in vitro and in vivo. The effects of MAD2L2 knockdown were similar to those of AURKB knockdown. Furthermore, p53 ablation rescued the MAD2L2 knockdown-induced suppression of BC cell proliferation and cell cycle arrest and senescence in BC cells.ConclusionsAURKB activates MAD2L2 expression to downregulate the p53 DDR pathway, thereby promoting BC progression. Thus, AURKB may serve as a potential molecular marker and a novel anticancer therapeutic target for BC.

Natural product-inspired molecules for covalent inhibition of SHP2 tyrosine phosphatase

Natural products have been playing indispensable roles in the development of lifesaving drug molecules. They are also valuable sources for covalent protein modifiers. However, they often are scarce in nature and have complex chemical structures, which are limiting their further biomedical development. Thus, natural product-inspired small molecules which still contain the essence of the parent natural products but are readily available and amenable for structural modification, are important and desirable in searching for lead compounds for various disease treatment. Inspired by the complex and diverse ent-kaurene diterpenoids with significant biological activities, we have created a synthetically accessible and focused covalent library by incorporating the bicyclo[3.2.1]octane α-methylene ketone, which is considered as the pharmacophore of ent-kaurene diterpenoids, as half of the structure, and replacing the other half with much less complex but more druglike scaffolds. From this library, we have identified and characterized selective covalent inhibitors of protein tyrosine phosphatase SHP2, an important anti-cancer therapeutic target. The success of this study demonstrated the importance of creating and evaluating natural product-inspired library as well as their application in targeting challenging disease targets.

Biological Significance of EphB4 Expression in Cancer.

Eph receptors and their Eph receptor-interacting (ephrin) ligands comprise a vital cell communication system with several functions. In cancer cells, there was evidence of bilateral Eph receptor signaling with both tumor-suppressing and tumor-promoting actions. As a member of the Eph receptor family, EphB4 has been linked to tumor angiogenesis, growth, and metastasis, which makes it a viable and desirable target for drug development in therapeutic applications. Many investigations have been conducted over the last decade to elucidate the structure and function of EphB4 in association with its ligand ephrinB2 for its involvement in tumorigenesis. Although several EphB4-targeting drugs have been investigated, and some selective inhibitors have been evaluated in clinical trials. This article addresses the structure and function of the EphB4 receptor, analyses its possibility as an anticancer therapeutic target, and summarises knowledge of EphB4 kinase inhibitors. To summarise, EphB4 is a difficult but potential treatment option for cancers.

Homeostatic status of thyroid hormones and brain water movement as determinant factors in biology of cerebral gliomas: a pilot study using a bioinformatics approach.

The expression and localization of the water channel transporters, aquaporins (AQPs), in the brain are substantially modified in gliomas during tumorigenesis, cell migration, edema formation, and resolution. We hypothesized that the molecular changes associated with AQP1 and AQP4 in the brain may potentially be anticancer therapeutic targets. To test this hypothesis, a bioinformatics analysis of publicly available data from international consortia was performed. We used RNA-seq as an experimental strategy and identified the number of differential AQP1 and AQP4 transcript expressions in glioma tissue compared to normal brain tissue. AQPs genes are overexpressed in patients with glioma. Among the glioma subtypes, AQP1 and AQP4 were overexpressed in astrocytoma (low-grade glioma) and classical (high-grade glioma). Overall survival analysis demonstrated that both AQP genes can be used as prognostic factors for patients with low-grade glioma. Additionally, we observed a correlation between the expression of genes involved in the tyrosine and thyroid hormone pathways and AQPs, namely: PNMT, ALDH1A3, AOC2, HGDATP1B1, ADCY5, PLCB4, ITPR1, ATP1A3, LRP2, HDAC1, MED24, MTOR, and ACTB1 (Spearman's coefficient = geq 0.20 and p-value = ≤ 0.05). Our findings indicate that the thyroid hormone pathways and AQPs 1 and 4 are potential targets for new anti-tumor drugs and therapeutic biomarkers for malignant gliomas.

Mechanical activation and expression of HSP27 in epithelial ovarian cancer

Understanding the complex biomechanical tumor microenvironment (TME) is of critical importance in developing the next generation of anti-cancer treatment strategies. This is especially true in epithelial ovarian cancer (EOC), the deadliest of the gynecologic cancers due to recurrent disease or chemoresistance. However, current models of EOC progression provide little control or ability to monitor how changes in biomechanical parameters alter EOC cell behaviors. In this study, we present a microfluidic device designed to permit biomechanical investigations of the ovarian TME. Using this microtissue system, we describe how biomechanical stimulation in the form of tensile strains upregulate phosphorylation of HSP27, a heat shock protein implicated in ovarian cancer chemoresistance. Furthermore, EOC cells treated with strain demonstrate decreased response to paclitaxel in the in vitro vascularized TME model. The results provide a direct link to biomechanical regulation of HSP27 as a mediator of EOC chemoresistance, possibly explaining the failure of such therapies in some patients. The work presented here lays a foundation to elucidating mechanobiological regulation of EOC progression, including chemoresistance and could provide novel targets for anti-cancer therapeutics.

CBX2 Deletion Suppresses Growth and Metastasis of Colorectal Cancer by Mettl3-p38/ERK MAPK Signalling Pathway.

Colorectal cancer (CRC) seriously endangers human health owing to its high morbidity and mortality. Previous studies have suggested that high expression of CBX2 may be associated with poor prognosis in CRC patients. However, its functional role in CRC remains to be elucidated. Herein, we found that CBX2 overexpression in colorectal cancer tissue compared with adjacent tissues. Additionally, forest maps and the nomogram model indicated that elevated CBX2 expression was an independent prognostic factor in CRC. Moreover, we confirmed that the deletion of CBX2 markedly suppressed the proliferation and migration of CRC cells in vitro and in vivo. Furthermore, downregulation of CBX2 promotes CRC cell apoptosis and hinders the cell cycle. Mechanistically, our data demonstrated that deletion of CBX2 inhibited the MAPK signaling pathway by regulating the protein levels of Mettl3. In conclusion, our study demonstrated that CBX2 is a vital tumor suppressor in CRC and could be a promising anti-cancer therapeutic target.

Synthesis and biological evaluation of selenopyrimidine derivatives bearing thiazolyl moiety

The aim of this study is to explain how new thiazole and selenophene moieties were obtained from 2-amino-4-(4-methylthiazol-2-yl)selenophene-3-carbonitrile (3) and to assess the antioxidant and cytotoxic properties of these new compounds. Compound 3 is prepared and subjected to react with each of CS2, malononitrile in EtOH/TEA, ECA, a mixture of HCl and AcOH, formamide, formic acid, malononitrile in the presence of EtONa to give their corresponding selenopyrimidine and selenopyridine derivatives 4–10. Moreover, compound 3 reacted with thiourea, hydrazine hydrate, CS2 in NaOH/DMF, and phenyl isothiocyanate to afford selenolo[2,3-d]pyrimidines 11–17 and related compounds. The recently synthesized compounds were tested for their DPPH radical scavenging and cytotoxic properties against the HePG-2 and MCF-7 cell lines. Compounds 5 and 11 demonstrated potential radical scavenging activity, but compounds 5 and 6 exhibited the strongest cytotoxic action against the examined cell lines. According to molecular docking data, the compounds under research are intriguing candidates for the development of new anticancer therapeutic targets.

The Golgi Apparatus as an Anticancer Therapeutic Target.

Although the discovery of the Golgi apparatus (GA) was made over 125 years ago, only a very limited number of therapeutic approaches have been developed to target this complex organelle. The GA serves as a modification and transport center for proteins and lipids and also has more recently emerged as an important store for some ions. The dysregulation of GA functions is implicated in many cellular processes associated with cancer and some GA proteins are indeed described as cancer biomarkers. This dysregulation can affect protein modification, localization, and secretion, but also cellular metabolism, redox status, extracellular pH, and the extracellular matrix structure. Consequently, it can directly or indirectly affect cancer progression. For these reasons, the GA is an appealing anticancer pharmacological target. Despite this, no anticancer drug specifically targeting the GA has reached the clinic and few have entered the clinical trial stage. Advances in nanodelivery approaches may help change this scenario by specifically targeting tumor cells and/or the GA through passive, active, or physical strategies. This article aims to examine the currently available anticancer GA-targeted drugs and the nanodelivery strategies explored for their administration. The potential benefits and challenges of modulating and specifically targeting the GA function in the context of cancer therapy are discussed.

Phenotypic profiling of solute carriers characterizes serine transport in cancer

Serine is a vital amino acid in tumorigenesis. While cells can perform de novo serine synthesis, most transformed cells rely on serine uptake to meet their increased biosynthetic requirements. Solute carriers (SLCs), a family of transmembrane nutrient transport proteins, are the gatekeepers of amino acid acquisition and exchange in mammalian cells and are emerging as anticancer therapeutic targets; however, the SLCs that mediate serine transport in cancer cells remain unknown. Here we perform an arrayed RNAi screen of SLC-encoding genes while monitoring amino acid consumption and cell proliferation in colorectal cancer cells using metabolomics and high-throughput imaging. We identify SLC6A14 and SLC25A15 as major cytoplasmic and mitochondrial serine transporters, respectively. We also observe that SLC12A4 facilitates serine uptake. Dual targeting of SLC6A14 and either SLC25A15 or SLC12A4 diminishes serine uptake and growth of colorectal cancer cells in vitro and in vivo, particularly in cells with compromised de novo serine biosynthesis. Our results provide insight into the mechanisms that contribute to serine uptake and intracellular handling.

The association of claudin-18 and androgen receptor expression in prostatic carcinoma: an immunohistochemical study

BackgroundClaudin-18 (CLDN18) is a recently identified anticancer therapeutic target with promising results for various gastrointestinal malignancies. The role of CLDN18 in prostatic carcinoma has not been investigated. The aim of this study was to investigate CLDN18 and androgen receptor (AR) expression in prostatic carcinoma and to link these findings with other clinicopathological characteristics. This retrospective study was carried out on 120 cases of prostatic lesions, including 100 cases of prostatic carcinoma and 20 cases of benign prostatic hyperplasia. The immunohistochemical staining technique was used to evaluate the expression of both CLDN18 and AR in prostatic carcinoma in relation to clinicopathological parameters.ResultsCLDN18 expression was completely absent in benign prostatic tissue, while it was detected in the membrane of 30 (30%) of studied carcinoma cases, with a statistically significant difference (p = 0.046). In contrast to other variables, a statistically significant relationship was identified between CLDN18 expression and Gleason Grade group (p = 0.000), stage (p = 0.03), and nodal metastasis (p = 0.000). The expression of the androgen receptor was detected in the nucleus of 96 (96%) of the cancer cases under study, with no statistically significant difference between the studied groups (p = 0.427). A statistically significant relation was found between AR expression and Gleason Grade group (p = 0.03) and stage (p = 0.01), while no relation with other variables was detected. AR expression and CLDN18 expression were shown to be statistically significantly correlated (p = 0.002).ConclusionsCLDN18 was expressed in prostatic carcinoma and correlated with an adverse tumor outcome. CLDN18 may be regulated by AR. CLDN18 could be a candidate therapeutic marker for the treatment of prostatic carcinoma.

First-in-Human Study with Preclinical Data of BCL-2/BCL-xL Inhibitor Pelcitoclax in Locally Advanced or Metastatic Solid Tumors.

B-cell lymphoma-extra-large (BCL-xL) regulates apoptosis and is an attractive anticancer therapeutic target. However, BCL-xL inhibition also kills mature platelets, hampering clinical development. Using an innovative prodrug strategy, we have developed pelcitoclax (APG-1252), a potent, dual BCL-2 and BCL-xL inhibitor. Aims of this study were to characterize the antitumor activity and safety of pelcitoclax and explore its underlying mechanisms of action (MOA). Cell line-derived xenograft and patient-derived xenograft (PDX) models were tested to evaluate antitumor activity and elucidate MOA. Subjects (N = 50) with metastatic small-cell lung cancer and other solid tumors received intravenous pelcitoclax once or twice weekly. Primary outcome measures were safety and tolerability; preliminary efficacy (responses every 2 cycles per RECIST version 1.1) represented a secondary endpoint. Pelcitoclax exhibited strong BAX/BAK‒dependent and caspase-mediated antiproliferative and apoptogenic activity in various cancer cell lines. Consistent with cell-based apoptogenic activity, pelcitoclax disrupted BCL-xL:BIM and BCL-xL:PUMA complexes in lung and gastric cancer PDX models. Levels of BCL-xL complexes correlated with tumor growth inhibition by pelcitoclax. Combined with taxanes, pelcitoclax enhanced antitumor activity by downregulating antiapoptotic protein myeloid cell leukemia-1 (MCL-1). Importantly, pelcitoclax was well tolerated and demonstrated preliminary therapeutic efficacy, with overall response and disease control rates of 6.5% and 30.4%, respectively. Most common treatment-related adverse events included transaminase elevations and reduced platelets that were less frequent with a once-weekly schedule. Our data demonstrate that pelcitoclax has antitumor activity and is well tolerated, supporting its further clinical development for human solid tumors, particularly combined with agents that downregulate MCL-1.

Mechanism underlying the effect of Pulsatilla decoction in hepatocellular carcinoma treatment: a network pharmacology and in vitro analysis

BackgroundCurrently, hepatocellular carcinoma (HCC) is associated with a poor prognosis. Moreover, there exist limited strategies for treating HCC. Pulsatilla decoction (PD), a traditional Chinese medicine formula, has been used to treat inflammatory bowel disease and several cancer types. Accordingly, we explored the mechanism of PD in HCC treatment via network pharmacology and in vitro experiments.MethodsOnline databases were searched for gene data, active components, and potential target genes associated with HCC development. Subsequently, bioinformatics analysis was performed using protein–protein interaction and Network Construction and Kyoto Encyclopedia of Genes and Genomes (KEGG) to screen for potential anticancer components and therapeutic targets of PD. Finally, the effect of PD on HCC was further verified by in vitro experiments.ResultsNetwork pharmacological analysis revealed that 65 compounds and 180 possible target genes were associated with the effect of PD on HCC. These included PI3K, AKT, NF-κB, FOS, and NFKBIA. KEGG analysis demonstrated that PD exerted its effect on HCC mainly via the PI3K-AKT, IL-17, and TNF signaling pathways. Cell viability and cell cycle experiments revealed that PD could significantly inhibit cancer cell proliferation and kill HCC cells by inducing apoptosis. Furthermore, western blotting confirmed that apoptosis was mediated primarily via the PI3K-AKT, IL-17, and TNF signaling pathways.ConclusionTo the best of our knowledge, this is the first study to elucidate the molecular mechanism and potential targets of PD in the treatment of HCC using network pharmacology.

Uncovering the Contrasting Binding Behavior of Plant Flavonoids Fisetin and Morin Having Subsidiary Hydroxyl Groups (-OH) with HRAS1 and HRAS2 i-Motif DNA Structures: Decoding the Structural Alterations and Positional Influences.

Research on the interactions of naturally existing flavonoids with various noncanonical DNA such as i-motif (IM) DNA structures is helpful in comprehending the molecular basis of binding mode as well as providing future direction for the application and invention of novel effective therapeutic drugs. IM DNA structures have been identified as prospective anticancer therapeutic targets, and flavonoids are smaller molecules with a variety of health-promoting attributes, including anticancer activities. The extensive investigation comprising a series of techniques reveals the contrasting mode of the binding behavior of fisetin and morin with various IM DNA structures. We have discovered that structural alterations of hydroxyl groups located at different places of aromatic rings influence flavonoid's reactivity. This minor structural alteration appears to be critical for fisetin and morin's capacity to interact differentially with HRAS1 and HRAS2 IM DNA. Hence, fisetin appears to be an efficient ligand for HRAS1 and morin is considered to be an efficient ligand for HRAS2 IM DNA. This novel exploration opens up the possibility of employing the strategy for regulation of gene expression in cancerous cells. Our finding also reveals the flavonoid-mediated specific interaction with IM DNA while pointing toward tangible strategies for drug discovery and other essential cellular functions.

MiR-4284 Functions as a Tumor Suppressor in Renal Cell Carcinoma Cells by Targeting Glutamate Decarboxylase 1.

MicroRNAs (miRNAs) play a crucial role as oncogenic or tumor suppressors in the pathogenesis and progression of tumors. However, few studies have investigated the exact role of miR-4284 in renal cell carcinoma (RCC). We aimed to investigate the role of miR-4284 as a tumor suppressor in renal cancer cell lines. A498 and Caki-1 were transfected with miR-4284. The Cell Counting Kit-8, colony formation, apoptosis assays, and quantitative reverse transcription-polymerase chain reaction were used to evaluate tumor growth-inhibiting functions. The wound-healing, transwell, and sphere-formation assays were conducted to investigate tumorigenic characteristics. The potential target genes of miR-4284 were predicted and experimentally verified. A xenograft experiment was performed to estimate the tumor-growth-suppressive function of miR-4284. miR-4284 overexpression suppressed proliferation, induced apoptosis, and suppressed tumorigenic features of renal cancer cells. Glutamate decarboxylase 1 (GAD1) was directly targeted by miR-4284. A xenograft mouse model injected with Caki-1 cells transfected with miR-4284 showed significantly decreased tumor growth rate and volume. miR-4284 affected tumor growth, metastasis, and apoptosis of renal cancer cells in vitro and in vivo. These findings highlight the potential of miR-4284 as a target for anticancer miRNA therapeutics in RCC.