RESUMO
RAS oncogenes (collectively NRAS, HRAS and especially KRAS) are among the most frequently mutated genes in cancer, with common driver mutations occurring at codons 12, 13 and 611. Small molecule inhibitors of the KRAS(G12C) oncoprotein have demonstrated clinical efficacy in patients with multiple cancer types and have led to regulatory approvals for the treatment of non-small cell lung cancer2,3. Nevertheless, KRASG12C mutations account for only around 15% of KRAS-mutated cancers4,5, and there are no approved KRAS inhibitors for the majority of patients with tumours containing other common KRAS mutations. Here we describe RMC-7977, a reversible, tri-complex RAS inhibitor with broad-spectrum activity for the active state of both mutant and wild-type KRAS, NRAS and HRAS variants (a RAS(ON) multi-selective inhibitor). Preclinically, RMC-7977 demonstrated potent activity against RAS-addicted tumours carrying various RAS genotypes, particularly against cancer models with KRAS codon 12 mutations (KRASG12X). Treatment with RMC-7977 led to tumour regression and was well tolerated in diverse RAS-addicted preclinical cancer models. Additionally, RMC-7977 inhibited the growth of KRASG12C cancer models that are resistant to KRAS(G12C) inhibitors owing to restoration of RAS pathway signalling. Thus, RAS(ON) multi-selective inhibitors can target multiple oncogenic and wild-type RAS isoforms and have the potential to treat a wide range of RAS-addicted cancers with high unmet clinical need. A related RAS(ON) multi-selective inhibitor, RMC-6236, is currently under clinical evaluation in patients with KRAS-mutant solid tumours (ClinicalTrials.gov identifier: NCT05379985).
Assuntos
Antineoplásicos , Mutação , Neoplasias , Proteína Oncogênica p21(ras) , Proteínas Proto-Oncogênicas p21(ras) , Animais , Humanos , Camundongos , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Guanosina Trifosfato/metabolismo , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/patologia , Proteína Oncogênica p21(ras)/antagonistas & inibidores , Proteína Oncogênica p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Synthetic lethality through combinatorial targeting DNA damage response (DDR) pathways provides exciting anticancer therapeutic benefit. Currently, the long noncoding RNAs (lncRNAs) have been implicated in tumor drug resistance; however, their potential significance in DDR is still largely unknown. Here, we report that a human lncRNA, CTD-2256P15.2, encodes a micropeptide, named PAR-amplifying and CtIP-maintaining micropeptide (PACMP), with a dual function to maintain CtIP abundance and promote poly(ADP-ribosyl)ation. PACMP not only prevents CtIP from ubiquitination through inhibiting the CtIP-KLHL15 association but also directly binds DNA damage-induced poly(ADP-ribose) chains to enhance PARP1-dependent poly(ADP-ribosyl)ation. Targeting PACMP alone inhibits tumor growth by causing a synthetic lethal interaction between CtIP and PARP inhibitions and confers sensitivity to PARP/ATR/CDK4/6 inhibitors, ionizing radiation, epirubicin, and camptothecin. Our findings reveal that a lncRNA-derived micropeptide regulates cancer progression and drug resistance by modulating DDR, whose inhibition could be employed to augment the existing anticancer therapeutic strategies.
Assuntos
Endodesoxirribonucleases , Neoplasias , Peptídeos , Poli ADP Ribosilação , RNA Longo não Codificante , Reparo do DNA , Endodesoxirribonucleases/metabolismo , Humanos , Proteínas dos Microfilamentos/genética , Proteínas dos Microfilamentos/metabolismo , Neoplasias/genética , Neoplasias/metabolismo , Peptídeos/farmacologia , Poli Adenosina Difosfato Ribose/metabolismo , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Poli(ADP-Ribose) Polimerases/genética , Poli(ADP-Ribose) Polimerases/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismoRESUMO
PP2A B55α, encoded by PPP2R2A, acts as a regulatory subunit of the serine/threonine phosphatase PP2A. Despite a frequent loss of heterozygosity of PPP2R2A in cases of non-small cell lung cancer (NSCLC), research on PP2A B55α's functions remains limited and controversial. To investigate the biological roles of PP2A B55α, we conducted bulk RNA-sequencing to assess the impact of PPP2R2A knockdown using two shRNAs in a NSCLC cell line. Gene set enrichment analysis (GSEA) of the RNA-sequencing data revealed significant enrichment of the epithelial-mesenchymal transition (EMT) pathway, with SNAI2 (the gene encoding Slug) emerging as one of the top candidates. Our findings demonstrate that PP2A B55α suppresses EMT, as PPP2R2A deficiency through knockdown or homozygous or hemizygous depletion promotes EMT and metastatic behavior in NSCLC cells, as evidenced by changes in EMT biomarkers, invasion and migration abilities, as well as metastasis in a tail vein assay. Mechanistically, PP2A B55α inhibits EMT by downregulating SNAI2 expression via the GSK3ß-ß-catenin pathway. Importantly, PPP2R2A deficiency also slows cell proliferation by disrupting DNA replication, particularly in PPP2R2A-/- cells. Furthermore, PPP2R2A deficiency, especially PPP2R2A-/- cells, leads to an increase in the cancer stem cell population, which correlates with enhanced resistance to chemotherapy. Overall, the decrease in PP2A B55α levels due to hemizygous/homozygous depletion heightens EMT and the metastatic or stemness/drug resistance potential of NSCLC cells despite their proliferation disadvantage. Our study highlights the significance of PP2A B55α in EMT and metastasis and suggests that targeting EMT/stemness could be a potential therapeutic strategy for treating PPP2R2A-deficient NSCLC.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Transição Epitelial-Mesenquimal , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares , Proteína Fosfatase 2 , Fatores de Transcrição da Família Snail , Transição Epitelial-Mesenquimal/genética , Humanos , Fatores de Transcrição da Família Snail/genética , Fatores de Transcrição da Família Snail/metabolismo , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Proteína Fosfatase 2/genética , Proteína Fosfatase 2/metabolismo , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/metabolismo , Animais , Movimento Celular , Linhagem Celular Tumoral , Glicogênio Sintase Quinase 3 beta/metabolismo , Glicogênio Sintase Quinase 3 beta/genética , Células A549 , Camundongos , Invasividade NeoplásicaRESUMO
Poly (ADP-ribose) Polymerase (PARP) inhibitors (PARPi) have been approved for both frontline and recurrent setting in ovarian cancer with homologous recombination (HR) repair deficiency. However, more than 40% of BRCA1/2-mutated ovarian cancer lack the initial response to PARPi treatment, and the majority of those that initially respond eventually develop resistance. Our previous study has demonstrated that increased expression of aldehyde dehydrogenase 1A1 (ALDH1A1) contributes to PARPi resistance in BRCA2-mutated ovarian cancer cells by enhancing microhomology-mediated end joining (MMEJ) but the mechanism remains unknown. Here, we find that ALDH1A1 enhances the expression of DNA polymerase θ (Polθ, encoded by the POLQ gene) in ovarian cancer cells. Furthermore, we demonstrate that the retinoic acid (RA) pathway is involved in the transcription activation of the POLQ gene. The RA receptor (RAR) can bind to the retinoic acid response element (RARE) located in the promoter of the POLQ gene, promoting transcription activation-related histone modification in the presence of RA. Given that ALDH1A1 catalyzes the biosynthesis of RA, we conclude that ALDH1A1 promotes POLQ expression via the activation of the RA signaling pathway. Finally, using a clinically-relevant patient-derived organoid (PDO) model, we find that ALDH1A1 inhibition by the pharmacological inhibitor NCT-505 in combination with the PARP inhibitor olaparib synergistically reduce the cell viability of PDOs carrying BRCA1/2 mutation and positive ALDH1A1 expression. In summary, our study elucidates a new mechanism contributing to PARPi resistance in HR-deficient ovarian cancer and shows the therapeutic potential of combining PARPi and ALDH1A1 inhibition in treating these patients.
RESUMO
Rationale: The mitogen-activated protein kinase pathway (MAPK) is one of the major cancer-driving pathways found in non-small cell lung cancer (NSCLC) patients. ERK inhibitors (ERKi) have been shown to be effective in NSCLC patients with MAPK pathway mutations. However, like other MAPK inhibitors, ERKi rarely confers complete and durable responses. The mechanism of tumor relapse after ERKi treatment is yet defined. Methods: To best study the mechanism of tumor relapse after ERK inhibitor treatment in NSCLC patients, we treated various NSCLC cell lines and patient-derived xenograft (PDX) with ERK inhibitors and evaluated the enrichment of cancer stem cell (CSC) population. We then performed a Next-generation sequencing (NGS) to identify potential pathways that are responsible for the CSC enrichment. Further, the involvement of specific pathways was examined using molecular and cellular methods. Finally, we investigated the therapeutic benefits of ERKi treatment combined with JAK/STAT pathway inhibitor using cellular and xenograft NSCLC models. Results: We found that ERKi treatment expands the CSC population in NSCLC cells through enhanced epithelial-to-mesenchymal transition (EMT)-mediated cancer cell dedifferentiation. Mechanistically, ERK inactivation induces EMT via pSTAT3-mediated upregulation of Slug, in which, upregulation of miR-204 and downregulation of SPDEF, a transcription repressor of Slug, are involved. Finally, the JAK/STAT pathway inhibitor Ruxolitinib blocks the ERK inactivation-induced EMT and CSC expansion, as well as the tumor progression in xenograft models after ERKi treatment. Conclusions: This study revealed a potential tumor relapse mechanism of NSCLC after ERK inhibition through the unintended activation of the EMT program, ascertained the pSTAT-miR-204-SPDEF-Slug axis, and provided a promising combination inhibitor approach to prevent tumor relapse in patients.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , MicroRNAs , Humanos , Carcinoma Pulmonar de Células não Pequenas/patologia , Neoplasias Pulmonares/patologia , Janus Quinases/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Transdução de Sinais , Fatores de Transcrição STAT/metabolismo , Recidiva Local de Neoplasia/genética , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Fatores de Transcrição/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , MicroRNAs/farmacologia , Regulação Neoplásica da Expressão GênicaRESUMO
Epithelial ovarian cancer is the most lethal malignancy of the female reproductive tract. A healthy ovary expresses both Estrogen Receptor α (ERα) and ß (ERß). Given that ERα is generally considered to promote cell survival and proliferation, thereby, enhancing tumor growth, while ERß shows a protective effect against the development and progression of tumors, the activation of ERß by its agonists could be therapeutically beneficial for ovarian cancer. Here, we demonstrate that the activation of ERß using a newly developed ERß agonist, OSU-ERb-12, can impede ovarian cancer cell expansion and tumor growth in an ERα-independent manner. More interestingly, we found that OSU-ERb-12 also reduces the cancer stem cell (CSC) population in ovarian cancer by compromising non-CSC-to-CSC conversion. Mechanistically, we revealed that OSU-ERb-12 decreased the expression of Snail, a master regulator of the epithelial-to-mesenchymal transition (EMT), which is associated with de novo CSC generation. Given that ERα can mediate EMT and facilitate maintenance of the CSC subpopulation and that OSU-ERb-12 can block the transactivity of ERα, we conclude that OSU-ERb-12 reduces the CSC subpopulation by inhibiting EMT in an ERα-dependent manner. Taken together, our data indicate that the ERß agonist OSU-ERb-12 could be used to hinder tumor progression and limit the CSC subpopulation with the potential to prevent tumor relapse and metastasis in patients with ovarian cancer.
RESUMO
DNA lesion bypass facilitates DNA synthesis across bulky DNA lesions, playing a critical role in DNA damage tolerance and cell survival after DNA damage. Assessing lesion bypass efficiency in the cell is important to better understanding of the mechanism of carcinogenesis and chemoresistance. Here we developed a chromatin immunoprecipitation (ChIP)-based method to measure lesion bypass activity across cisplatin-induced intrastrand crosslinks in cancer cells. DNA lesion bypass enables the replication to continue in the presence of replication blocks. Thus, the successful lesion bypass should result in the coexistence of DNA lesions and the newly synthesized DNA fragment opposite to this lesion. Using ChIP, we precipitated the cisplatin-induced intrastrand crosslinks, and quantitated the precipitated newly synthesized DNA that was labeled with BrdU. We validated this method on ovarian cancer cells with inhibited TLS activity. We then applied this method to show that ovarian cancer stem cells exhibit high lesion bypass activity relative to bulk cancer cells from the same cell line. In conclusion, this novel ChIP-based lesion bypass assay can detect the extent to which cisplatin-induced DNA lesions are bypassed in live cells. Our study may be applied more broadly to the study of other DNA lesions, as specific antibodies to these specific lesions are available.
Assuntos
DNA Polimerase Dirigida por DNA , DNA , Imunoprecipitação da Cromatina , DNA/metabolismo , Dano ao DNA , Reparo do DNA , Replicação do DNA , DNA Polimerase Dirigida por DNA/metabolismoRESUMO
The Ras subfamily of small GTPases is mutated in â¼30% of human cancers and represents compelling yet challenging anticancer drug targets owing to their flat protein surface. We previously reported a bicyclic peptidyl inhibitor, cyclorasin B3, which binds selectively to Ras-GTP with modest affinity and blocks its interaction with downstream effector proteins in vitro but lacks cell permeability or biological activity. In this study, optimization of B3 yielded a potent pan-Ras inhibitor, cyclorasin B4-27, which binds selectively to the GTP-bound forms of wild-type and mutant Ras isoforms (KD = 21 nM for KRasG12V-GppNHp) and is highly cell-permeable and metabolically stable (serum t1/2 > 24 h). B4-27 inhibits Ras signaling in vitro and in vivo by blocking Ras from interacting with downstream effector proteins and induces apoptosis of Ras-mutant cancer cells. When administered systemically (i.v.), B4-27 suppressed tumor growth in two different mouse xenograft models at 1-5 mg/kg of daily doses.
Assuntos
Antineoplásicos/farmacologia , Peptídeos Cíclicos/química , Peptídeos Cíclicos/farmacologia , Proteínas ras/antagonistas & inibidores , Animais , Antineoplásicos/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Descoberta de Drogas , Humanos , Masculino , Camundongos , Camundongos Nus , Isoformas de Proteínas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Black spot disease, which is caused by the pathogenic fungal Ceratocystis fimbriata, seriously affects the production of sweet potato and its quality during postharvest storage. In this study, the preliminary identification of the rhizosphere actinomycete strain SPS-33, and its antifungal activity of volatiles in vitro and in vivo was investigated. Based on morphological identification and phylogenetic analysis of the 16S rRNA gene sequence, strain SPS-33 was identified as Streptomyces lavendulae. Volatile organic compounds (VOCs) emitted by SPS-33 inhibited mycelial growth and sporulation of C. fimbriata in vitro and also induced a series of observable hyphae morphological changes. In an in vivo pathogenicity assay, exposure to SPS-33 significantly decreased the lesion diameter and water loss rate in sweet potato tuberous roots (TRs) inoculated with C. fimbriata. It increased the antioxidant enzymes' activities of peroxidase, catalase, and superoxide dismutase as well as decreased malondialdehyde and increased total soluble sugar. In the VOC profile of SPS-33 detected by a headspace solid-phase micro extraction (HS-SPME) and gas chromatography-mass spectrometry (GC-MS), heptadecane, tetradecane, and 3-methyl-1-butanol were the most abundant compounds. 2-Methyl-1-butanol, 3-methyl-1-butanol, pyridine, and phenylethyl alcohol showed strong antifungal effects against C. fimbriata. These findings suggest that VOCs from S. lavendulae SPS-33 have the potential for pathogen C. fimbriata control in sweet potato postharvest storage by fumigant action.
RESUMO
Poly (ADP-ribose) polymerase (PARP) inhibitors (PARPi) are approved to treat recurrent ovarian cancer with BRCA1 or BRCA2 mutations, and as maintenance therapy for recurrent platinum-sensitive ovarian cancer (BRCA wild-type or mutated) after treatment with platinum. However, the acquired resistance against PARPi remains a clinical hurdle. Here, we demonstrated that PARP inhibitor (olaparib)-resistant epithelial ovarian cancer (EOC) cells exhibited an elevated aldehyde dehydrogenase (ALDH) activity, mainly contributed by increased expression of ALDH1A1 due to olaparib-induced expression of BRD4, a member of bromodomain and extraterminal (BET) family protein. We also revealed that ALDH1A1 enhanced microhomology-mediated end joining (MMEJ) activity in EOC cells with inactivated BRCA2, a key protein that promotes homologous recombination (HR) by using an intrachromosomal MMEJ reporter. Moreover, NCT-501, an ALDH1A1-selective inhibitor, can synergize with olaparib in killing EOC cells carrying BRCA2 mutation in both in vitro cell culture and the in vivo xenograft animal model. Given that MMEJ activity has been reported to be responsible for PARPi resistance in HR-deficient cells, we conclude that ALDH1A1 contributes to the resistance to PARP inhibitors via enhancing MMEJ in BRCA2-/- ovarian cancer cells. Our findings provide a novel mechanism underlying PARPi resistance in BRCA2-mutated EOC cells and suggest that inhibition of ALDH1A1 could be exploited for preventing and overcoming PARPi resistance in EOC patients carrying BRCA2 mutation.
Assuntos
Família Aldeído Desidrogenase 1/metabolismo , Carcinoma Epitelial do Ovário/tratamento farmacológico , Carcinoma Epitelial do Ovário/genética , Reparo do DNA , Proteínas Nucleares/metabolismo , Neoplasias Ovarianas/tratamento farmacológico , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Retinal Desidrogenase/metabolismo , Família Aldeído Desidrogenase 1/antagonistas & inibidores , Família Aldeído Desidrogenase 1/genética , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Proteína BRCA1/genética , Proteína BRCA2/genética , Carcinoma Epitelial do Ovário/metabolismo , Carcinoma Epitelial do Ovário/patologia , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Reparo do DNA por Junção de Extremidades , Resistencia a Medicamentos Antineoplásicos , Sinergismo Farmacológico , Feminino , Humanos , Camundongos , Camundongos Nus , Mutação , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Ftalazinas/administração & dosagem , Ftalazinas/farmacologia , Piperazinas/administração & dosagem , Piperazinas/farmacologia , Inibidores de Poli(ADP-Ribose) Polimerases/administração & dosagem , Retinal Desidrogenase/antagonistas & inibidores , Retinal Desidrogenase/genética , Teofilina/administração & dosagem , Teofilina/farmacologia , Fatores de Transcrição/metabolismo , Transfecção , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Cancer stem cells (CSC) play a central role in cancer metastasis and development of drug resistance. miRNA are important in regulating CSC properties and are considered potential therapeutic targets. Here we report that miR-328-3p (miR-328) is significantly upregulated in ovarian CSC. High expression of miR-328 maintained CSC properties by directly targeting DNA damage binding protein 2, which has been shown previously to inhibit ovarian CSC. Reduced activity of ERK signaling in ovarian CSC, mainly due to a low level of reactive oxygen species, contributed to the enhanced expression of miR-328 and maintenance of CSC. Inhibition of miR-328 in mouse orthotopic ovarian xenografts impeded tumor growth and prevented tumor metastasis. In summary, our findings provide a novel mechanism underlying maintenance of the CSC population in ovarian cancer and suggest that targeted inhibition of miR-328 could be exploited for the eradication of CSC and aversion of tumor metastasis in ovarian cancer. SIGNIFICANCE: These findings present inhibition of miR-328 as a novel strategy for efficient elimination of CSC to prevent tumor metastasis and recurrence in patients with epithelial ovarian cancer.