RESUMO
Recent preclinical studies have shown that the intake of nonsteroidal anti-inflammatory drugs (NSAIDs) aspirin and naproxen could be an effective intervention strategy against TMPRSS2-ERG fusion-driven prostate tumorigenesis. Herein, as a follow-up mechanistic study, employing TMPRSS2-ERG (fusion) positive tumors and plasma from TMPRSS2-ERG. Ptenflox/flox mice, we profiled the stage specific proteomic changes (focused on inflammatory circulating and prostate tissue/tumor-specific cytokines, chemokines, and growth factors/growth signaling-associated molecules) that contribute to prostate cancer (PCa) growth and progression in the TMPRSS2-ERG fusion-driven mouse model of tumorigenesis. In addition, the association of the protective effects of NSAIDs (aspirin 1400 ppm and naproxen 400 ppm) with the modulation of these specific molecular pathways was determined. A sandwich Elisa based membrane array-proteome profiler identifying 111 distinct signaling molecules was employed. Overall, the plasma and prostate tissue sample analyses identified 54 significant and differentially expressed cytokines, chemokines, and growth factors/growth signaling-associated molecules between PCa afflicted mice (TMPRSS2-ERG. Ptenflox/flox, age-matched noncancerous controls, NSAIDs-supplemented and no-drug controls). Bioinformatic analysis of the array outcomes indicated that the protective effect of NSAIDs was associated with reduced expression of (a) tumor promoting inflammatory molecules (M-CSF, IL-33, CCL22, CCL12, CX3CL1, CHI3L1, and CD93), (b) growth factors- growth signaling-associated molecules (Chemerin, FGF acidic, Flt-3 ligand, IGFBP-5, and PEDF), and (c) tumor microenvironment/stromal remodeling proteins MMP2 and MMP9. Overall, our findings corroborate the pathological findings that protective effects of NSAIDs in TMPSS2-ERG fusion-driven prostate tumorigenesis are associated with antiproliferative and anti-inflammatory effects and possible modulation of the immune cell enriched microenvironment.
Assuntos
Anti-Inflamatórios não Esteroides , Aspirina , Naproxeno , Neoplasias da Próstata , Animais , Masculino , Neoplasias da Próstata/patologia , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Anti-Inflamatórios não Esteroides/farmacologia , Aspirina/farmacologia , Camundongos , Naproxeno/farmacologia , Proteômica/métodos , Inflamação/metabolismo , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Próstata/patologia , Próstata/metabolismo , Próstata/efeitos dos fármacos , Carcinogênese/efeitos dos fármacos , Carcinogênese/genética , Carcinogênese/metabolismo , Proteoma/metabolismo , Humanos , Citocinas/metabolismo , Citocinas/sangueRESUMO
Advances in molecular diagnostics have led to improved diagnosis and molecular understanding of hereditary cancers in the clinic. Improving the management, treatment, and potential prevention of cancers in carriers of predisposing mutations requires preclinical experimental models that reflect the key pathogenic features of the specific syndrome associated with the mutations. Numerous genetically engineered mouse (GEM) models of hereditary cancer have been developed. In this review, we describe the models of Lynch syndrome and hereditary breast and ovarian cancer syndrome, the two most common hereditary cancer predisposition syndromes. We focus on Lynch syndrome models as illustrative of the potential for using mouse models to devise improved approaches to prevention of cancer in a high-risk population. GEM models are an invaluable tool for hereditary cancer models. Here, we review GEM models for some hereditary cancers and their potential use in cancer prevention studies.
Assuntos
Neoplasias Colorretais Hereditárias sem Polipose , Síndrome Hereditária de Câncer de Mama e Ovário , Síndromes Neoplásicas Hereditárias , Humanos , Feminino , Animais , Camundongos , Neoplasias Colorretais Hereditárias sem Polipose/genética , Neoplasias Colorretais Hereditárias sem Polipose/diagnóstico , Predisposição Genética para Doença , Síndromes Neoplásicas Hereditárias/genética , MutaçãoRESUMO
Two potent and selective KRASG12D inhibitors, ERAS-4693 and ERAS-5024, were generated as possible clinical candidates to treat patients harboring G12D mutations in solid tumors. Both molecules exhibited strong anti-tumor activity in the KRASG12D mutant PDAC xenograft mouse models while ERAS-5024 also showed tumor growth inhibition when administered on an intermittent dosing regimen. Acute dose-limiting toxicity consistent with an allergic reaction was observed for both molecules shortly after administration at doses just above those which demonstrated anti-tumor activity, indicative of a narrow therapeutic index. A series of studies were subsequently conducted to identify a common underlying mechanism for the observed toxicity, including CETSA® (Cellular Thermal Shift Assay) as well as several functional off-target screens. Both ERAS-4693 and ERAS-5024 were identified to agonize MRGPRX2 which has been linked to pseudo-allergic reactions. In vivo toxicologic characterization of both molecules included repeat-dose studies in the rat and dog. Dose-limiting toxicities were observed in both species with ERAS-4693 and ERAS-5024 and plasma exposure levels at the maximum tolerated doses were generally below that which caused strong anti-tumor activity, supporting the initial observation of a narrow therapeutic index. Additional overlapping toxicities included a reduction in reticulocytes and clinical pathological changes suggestive of an inflammatory response. Furthermore, increases in plasma histamine were observed in dogs administered ERAS-5024, supporting the hypothesis that MRGPRX2 agonism may be the cause of the pseudo-allergic reaction. This work highlights the importance of balancing both the safety and efficacy of KRASG12D inhibitors as this class of molecules begins to enter clinical development.
Assuntos
Hipersensibilidade , Neoplasias Pancreáticas , Humanos , Camundongos , Ratos , Animais , Cães , Proteínas Proto-Oncogênicas p21(ras)/genética , Neoplasias Pancreáticas/patologia , Mutação , Proteínas do Tecido Nervoso , Receptores de Neuropeptídeos/genética , Receptores Acoplados a Proteínas G/genéticaRESUMO
Colorectal cancer (CRC) is the third most prevalent and second deadliest cancer worldwide. In addition, metastasis directly causes up to 90% of all CRC deaths, highlighting the metastatic burden of the disease. Biomarkers such as S100A4 and MACC1 aid in identifying patients with a high risk of metastasis formation. High expression of S100A4 or MACC1 and to a greater extent the combination of both biomarkers is a predictor for metastasis and poor patient survival in CRC. MACC1 is a tumor-initiating and metastasis-promoting oncogene, whereas S100A4 has not been shown to initiate tumor formation but can, nevertheless, promote malignant tumor growth and metastasis formation. Cantharidin is a natural drug extracted from various blister beetle species, and its demethylated analogue norcantharidin has been shown in several studies to have an anti-cancer and anti-metastatic effect in different cancer entities such as CRC, breast cancer, and lung cancer. The impact of the natural compound cantharidin and norcantharidin on S100A4 and MACC1 gene expression, cancer cell migration, motility, and colony formation in vitro was tested. Here, for the first time, we have demonstrated that cantharidin and norcantharidin are transcriptional inhibitors of S100A4 and MACC1 mRNA expression, protein expression, and motility in CRC cells. Our results clearly indicate that cantharidin and, to a lesser extent, its analogue norcantharidin are promising compounds for efficient anti-metastatic therapy targeting the metastasis-inducing genes S100A4 and MACC1 for personalized medicine for cancer patients.
Assuntos
Neoplasias Colorretais , Neoplasias , Humanos , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Cantaridina/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Neoplasias Colorretais/patologia , Proteína A4 de Ligação a Cálcio da Família S100/genética , Transativadores/genética , Transativadores/metabolismoRESUMO
BACKGROUND & AIMS: DNA mismatch repair deficiency drives microsatellite instability (MSI). Cells with MSI accumulate numerous frameshift mutations. Frameshift mutations affecting cancer-related genes may promote tumorigenesis and, therefore, are shared among independently arising MSI tumors. Consequently, such recurrent frameshift mutations can give rise to shared immunogenic frameshift peptides (FSPs) that represent ideal candidates for a vaccine against MSI cancer. Pathogenic germline variants of mismatch repair genes cause Lynch syndrome (LS), a hereditary cancer syndrome affecting approximately 20-25 million individuals worldwide. Individuals with LS are at high risk of developing MSI cancer. Previously, we demonstrated safety and immunogenicity of an FSP-based vaccine in a phase I/IIa clinical trial in patients with a history of MSI colorectal cancer. However, the cancer-preventive effect of FSP vaccination in the scenario of LS has not yet been demonstrated. METHODS: A genome-wide database of 488,235 mouse coding mononucleotide repeats was established, from which a set of candidates was selected based on repeat length, gene expression, and mutation frequency. In silico prediction, in vivo immunogenicity testing, and epitope mapping was used to identify candidates for FSP vaccination. RESULTS: We identified 4 shared FSP neoantigens (Nacad [FSP-1], Maz [FSP-1], Senp6 [FSP-1], Xirp1 [FSP-1]) that induced CD4/CD8 T cell responses in naïve C57BL/6 mice. Using VCMsh2 mice, which have a conditional knockout of Msh2 in the intestinal tract and develop intestinal cancer, we showed vaccination with a combination of only 4 FSPs significantly increased FSP-specific adaptive immunity, reduced intestinal tumor burden, and prolonged overall survival. Combination of FSP vaccination with daily naproxen treatment potentiated immune response, delayed tumor growth, and prolonged survival even more effectively than FSP vaccination alone. CONCLUSIONS: Our preclinical findings support a clinical strategy of recurrent FSP neoantigen vaccination for LS cancer immunoprevention.
Assuntos
Antígenos de Neoplasias/farmacologia , Vacinas Anticâncer/farmacologia , Neoplasias Colorretais Hereditárias sem Polipose/tratamento farmacológico , Mutação da Fase de Leitura , Fenômenos Imunogenéticos , Fragmentos de Peptídeos/farmacologia , Adjuvantes Imunológicos/farmacologia , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/imunologia , Vacinas Anticâncer/genética , Vacinas Anticâncer/imunologia , Neoplasias Colorretais Hereditárias sem Polipose/genética , Neoplasias Colorretais Hereditárias sem Polipose/imunologia , Neoplasias Colorretais Hereditárias sem Polipose/patologia , Bases de Dados Genéticas , Modelos Animais de Doenças , Epitopos , Imunidade Celular/efeitos dos fármacos , Imunidade Humoral/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteína 2 Homóloga a MutS/genética , Naproxeno/farmacologia , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/imunologia , Carga Tumoral/efeitos dos fármacos , Microambiente Tumoral , Vacinação , Eficácia de VacinasRESUMO
In the present study, we performed a comparative stage-specific pathological and molecular marker evaluation of TMPRSS2-ERG fusion and PTEN loss-driven (TMPRSS2-ERG. Ptenflox/flox ) versus non-fusion-driven prostate tumorigenesis (Hi-Myc) in mice. Anterior, ventral, and dorsolateral prostates were collected from mice at different ages (or time points post-Cre induction). Results indicated that growth and progression of prostatic intraepithelial lesions to adenocarcinoma stages occurred in both mice models albeit at different rates. In the TMPRSS2-ERG. Ptenflox/flox mice, the initiation of tumorigenesis was slow, but subsequent progression through different stages became increasingly faster. Adenocarcinoma stage was reached early on; however, no high-grade undifferentiated tumors were observed. Conversely, in the Hi-Myc+/- mice, tumorigenesis initiation was rapid; however, progression through different stages was relatively slower and it took a while to reach the more aggressive phenotype stage. Nevertheless, at the advanced stages in the Hi-Myc+/- mice, high-grade undifferentiated tumors were observed compared to the later stage tumors observed in the fusion-driven TMPRSS2-ERG. Ptenflox/flox mice. These results were corroborated by the stage specific-pattern in the molecular expression of proliferation markers (PCNA and c-Myc); androgen receptor (AR); fusion-resultant overexpression of ERG; Prostein (SLC45-A3); and angiogenesis marker (CD-31). Importantly, there was a significant increase in immune cell infiltrations, which increased with the stage of tumorigenesis, in the TMPRSS2-ERG fusion-positive tumors relative to fusion negative tumors. Together, these findings are both novel and highly significant in establishing a working preclinical model for evaluating the efficacy of interventions during different stages of tumorigenesis in TMPRSS2-ERG fusion-driven PCa.
Assuntos
Adenocarcinoma , Neoplasias da Próstata , Adenocarcinoma/genética , Animais , Carcinogênese/patologia , Humanos , Masculino , Camundongos , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Próstata/patologia , Neoplasias da Próstata/patologia , Serina Endopeptidases/metabolismo , Regulador Transcricional ERG/genética , Regulador Transcricional ERG/metabolismoRESUMO
OBJECTIVE: Patients with Lynch syndrome (LS) are at markedly increased risk for colorectal cancer. It is being increasingly recognised that the immune system plays an essential role in LS tumour development, thus making an ideal target for cancer prevention. Our objective was to evaluate the safety, assess the activity and discover novel molecular pathways involved in the activity of naproxen as primary and secondary chemoprevention in patients with LS. DESIGN: We conducted a Phase Ib, placebo-controlled, randomised clinical trial of two dose levels of naproxen sodium (440 and 220 mg) administered daily for 6 months to 80 participants with LS, and a co-clinical trial using a genetically engineered mouse model of LS and patient-derived organoids (PDOs). RESULTS: Overall, the total number of adverse events was not different across treatment arms with excellent tolerance of the intervention. The level of prostaglandin E2 in the colorectal mucosa was significantly decreased after treatment with naproxen when compared with placebo. Naproxen activated different resident immune cell types without any increase in lymphoid cellularity, and changed the expression patterns of the intestinal crypt towards epithelial differentiation and stem cell regulation. Naproxen demonstrated robust chemopreventive activity in a mouse co-clinical trial and gene expression profiles induced by naproxen in humans showed perfect discrimination of mice specimens with LS and PDOs treated with naproxen and control. CONCLUSIONS: Naproxen is a promising strategy for immune interception in LS. We have discovered naproxen-induced gene expression profiles for their potential use as predictive biomarkers of drug activity. TRIAL REGISTRATION NUMBER: gov Identifier: NCT02052908.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Quimioprevenção , Neoplasias Colorretais Hereditárias sem Polipose/tratamento farmacológico , Neoplasias Colorretais Hereditárias sem Polipose/imunologia , Naproxeno/farmacologia , Adulto , Idoso , Animais , Anti-Inflamatórios não Esteroides/administração & dosagem , Dinoprostona/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Mucosa Intestinal/metabolismo , Masculino , Camundongos , Pessoa de Meia-Idade , Naproxeno/administração & dosagemRESUMO
Poly[di(carboxylatomethylphenoxy)phosphazene] (PCMP), a new member of polyphosphazene immunoadjuvant family, is synthesized. In vitro assessment of a new macromolecule revealed hydrolytic degradation profile and immunostimulatory activity comparable to its clinical stage homologue PCPP; however, PCMP was characterized by a beneficial reduced sensitivity to the ionic environment. In vivo evaluation of PCMP potency was conducted with human papillomavirus (HPV) virus-like particles (VLPs) based RG1-VLPs vaccine. In contrast with previously reported self-assembly of polyphosphazene adjuvants with proteins, which typically results in the formation of complexes with multimeric display of antigens, PCMP surface modified VLPs in a composition dependent pattern, which at a high polymer-to VLPs ratio led to stabilization of antigenic particles. Immunization experiments in mice demonstrated that PCMP adjuvanted RG1-VLPs vaccine induced potent humoral immune responses, in particular, on the level of highly desirable protective cross-neutralizing antibodies, and outperformed PCPP and Alhydrogel adjuvanted formulations.
Assuntos
Adjuvantes Imunológicos/química , Materiais Biocompatíveis/química , Compostos Organofosforados/química , Infecções por Papillomavirus/prevenção & controle , Vacinas contra Papillomavirus/química , Polímeros/química , Vacinas de Partículas Semelhantes a Vírus/química , Adjuvantes Imunológicos/farmacologia , Animais , Anticorpos Neutralizantes/química , Anticorpos Antivirais/química , Composição de Medicamentos , Liberação Controlada de Fármacos , Feminino , Humanos , Hidrogéis/química , Camundongos Endogâmicos BALB C , Vacinas contra Papillomavirus/farmacologia , Vacinação , Vacinas de Partículas Semelhantes a Vírus/farmacologiaRESUMO
Background The hexapeptide 4A6 (Ac-Thr(tBu)-His(Bzl)-Thr(Bzl)-Nle-Glu(OtBu)-Gly-Bza) was isolated from a peptide library constructed to identify peptide-based transport inhibitors of multidrug resistance (MDR) efflux pumps including P-glycoprotein and Multidrug Resistance-associated Protein 1. 4A6 proved to be a substrate but not an inhibitor of these MDR efflux transporters. In fact, 4A6 and related peptides displayed potent cytotoxic activity via an unknown mechanism. Objective To decipher the mode of cytotoxic activity of 4A6. Methods Screening of 4A6 activity was performed against the NCI60 panel of cancer cell lines. Possible interactions of 4A6 with the 26S proteasome were assessed via proteasome activity and affinity labeling, and cell growth inhibition studies with leukemic cells resistant to the proteasome inhibitor bortezomib (BTZ). Results The NCI60 panel COMPARE analysis revealed that 4A6 had an activity profile overlapping with BTZ. Consistently, 4A6 proved to be a selective and reversible inhibitor of ß5 subunit (PSMB5)-associated chymotrypsin-like activity of the 26S proteasome. This conclusion is supported by several lines of evidence: (i) inhibition of chymotrypsin-like proteasome activity by 4A6 and related peptides correlated with their cell growth inhibition potencies; (ii) 4A6 reversibly inhibited functional ß5 active site labeling with the affinity probe BodipyFL-Ahx3L3VS; and (iii) human myeloid THP1 cells with acquired BTZ resistance due to mutated PSMB5 were highly (up to 287-fold) cross-resistant to 4A6 and its related peptides. Conclusion 4A6 is a novel specific inhibitor of the ß5 subunit-associated chymotrypsin-like proteasome activity. Further exploration of 4A6 as a lead compound for development as a novel proteasome-targeted drug is warranted.
Assuntos
Antineoplásicos/farmacologia , Oligopeptídeos/farmacologia , Inibidores de Proteassoma/farmacologia , Animais , Apoptose/efeitos dos fármacos , Bortezomib/farmacologia , Linhagem Celular , Resistencia a Medicamentos Antineoplásicos , Humanos , Camundongos , Biblioteca de PeptídeosRESUMO
BACKGROUND: Activated anaplastic lymphoma kinase (ALK) gene fusions are recurrent events in a small fraction of colorectal cancers (CRCs), although these events have not yet been exploited as in other malignancies. METHODS: We detected ALK protein expression by immunohistochemistry and gene rearrangements by fluorescence in situ hybridisation in the ALKA-372-001 phase I study of the pan-Trk, ROS1, and ALK inhibitor entrectinib. One out of 487 CRCs showed ALK positivity with a peculiar pattern that prompted further characterisation by targeted sequencing using anchored multiplex PCR. RESULTS: A novel ALK fusion with the carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase (CAD) gene (CAD-ALK fusion gene) was identified. It resulted from inversion within chromosome 2 and the fusion of exons 1-35 of CAD with exons 20-29 of ALK. After failure of previous standard therapies, treatment of this patient with the ALK inhibitor entrectinib resulted in a durable objective tumour response. CONCLUSIONS: We describe the novel CAD-ALK rearrangement as an oncogene and provide the first evidence of its drugability as a new molecular target in CRC.
Assuntos
Antineoplásicos/uso terapêutico , Aspartato Carbamoiltransferase/genética , Benzamidas/uso terapêutico , Carbamoil Fosfato Sintase (Glutamina-Hidrolizante)/genética , Neoplasias Colorretais/tratamento farmacológico , Di-Hidro-Orotase/genética , Rearranjo Gênico , Indazóis/uso terapêutico , Receptores Proteína Tirosina Quinases/genética , Quinase do Linfoma Anaplásico , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Feminino , Humanos , Pessoa de Meia-IdadeRESUMO
Targeted quantification of DNA methylation allows for interrogation of the most informative loci across many samples quickly and cost-effectively. Here we report improved bisulfite padlock probes (BSPPs) with a design algorithm to generate efficient padlock probes, a library-free protocol that dramatically reduces sample-preparation cost and time and is compatible with automation, and an efficient bioinformatics pipeline to accurately obtain both methylation levels and genotypes from sequencing of bisulfite-converted DNA.
Assuntos
Sondas de DNA/química , Sondas de DNA/genética , DNA/química , DNA/genética , Reação em Cadeia da Polimerase/métodos , Análise de Sequência de DNA/métodos , Sulfitos/química , Sequência de Bases , Biblioteca Gênica , Dados de Sequência MolecularRESUMO
The US National Cancer Institute (NCI) 60 human tumour cell line anticancer drug screen (NCI60) was developed in the late 1980s as an in vitro drug-discovery tool intended to supplant the use of transplantable animal tumours in anticancer drug screening. This screening model was rapidly recognized as a rich source of information about the mechanisms of growth inhibition and tumour-cell kill. Recently, its role has changed to that of a service screen supporting the cancer research community. Here I review the development, use and productivity of the screen, highlighting several outcomes that have contributed to advances in cancer chemotherapy.
Assuntos
Antineoplásicos/uso terapêutico , Ensaios de Seleção de Medicamentos Antitumorais/história , Animais , Antineoplásicos/história , História do Século XX , História do Século XXI , Humanos , National Institutes of Health (U.S.)/história , National Institutes of Health (U.S.)/organização & administração , Células Tumorais Cultivadas , Estados UnidosRESUMO
Human embryonic stem cells (hESCs) hold great promise for regenerative medicine because they can undergo unlimited self-renewal and retain the capability to differentiate into all cell types in the body. Although numerous genes/proteins such as Oct4 and Gata6 have been identified to play critical regulatory roles in self-renewal and differentiation of hESC, the majority of the regulators in these cellular processes and more importantly how these regulators co-operate with each other and/or with epigenetic modifications are still largely unknown. We propose here a systematic approach to integrate genomic and epigenomic data for identification of direct regulatory interactions. This approach allows reconstruction of cell-type-specific transcription networks in embryonic stem cells (ESCs) and fibroblasts at an unprecedented scale. Many links in the reconstructed networks coincide with known regulatory interactions or literature evidence. Systems-level analyses of these networks not only uncover novel regulators for pluripotency and differentiation, but also reveal extensive interplays between transcription factor binding and epigenetic modifications. Especially, we observed poised enhancers characterized by both active (H3K4me1) and repressive (H3K27me3) histone marks that contain enriched Oct4- and Suz12-binding sites. The success of such a systems biology approach is further supported by experimental validation of the predicted interactions.
Assuntos
Diferenciação Celular/genética , Células-Tronco Embrionárias/metabolismo , Redes Reguladoras de Genes , Células-Tronco Pluripotentes/metabolismo , Fatores de Transcrição/metabolismo , Animais , Sítios de Ligação , Linhagem Celular , Células-Tronco Embrionárias/citologia , Elementos Facilitadores Genéticos , Epigênese Genética , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Fator 3 de Transcrição de Octâmero/genética , Regiões Promotoras GenéticasRESUMO
Modulation of DNA repair proteins by small molecules has attracted great interest. An in vitro helicase activity screen was used to identify molecules that modulate DNA unwinding by Werner syndrome helicase (WRN), mutated in the premature aging disorder Werner syndrome. A small molecule from the National Cancer Institute Diversity Set designated NSC 19630 [1-(propoxymethyl)-maleimide] was identified that inhibited WRN helicase activity but did not affect other DNA helicases [Bloom syndrome (BLM), Fanconi anemia group J (FANCJ), RECQ1, RecQ, UvrD, or DnaB). Exposure of human cells to NSC 19630 dramatically impaired growth and proliferation, induced apoptosis in a WRN-dependent manner, and resulted in elevated γ-H2AX and proliferating cell nuclear antigen (PCNA) foci. NSC 19630 exposure led to delayed S-phase progression, consistent with the accumulation of stalled replication forks, and to DNA damage in a WRN-dependent manner. Exposure to NSC 19630 sensitized cancer cells to the G-quadruplex-binding compound telomestatin or a poly(ADP ribose) polymerase (PARP) inhibitor. Sublethal dosage of NSC 19630 and the chemotherapy drug topotecan acted synergistically to inhibit cell proliferation and induce DNA damage. The use of this WRN helicase inhibitor molecule may provide insight into the importance of WRN-mediated pathway(s) important for DNA repair and the replicational stress response.
Assuntos
Dano ao DNA/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Exodesoxirribonucleases/antagonistas & inibidores , Maleimidas/farmacologia , RecQ Helicases/antagonistas & inibidores , Adenosina Trifosfatases/antagonistas & inibidores , Adenosina Trifosfatases/metabolismo , Apoptose/efeitos dos fármacos , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Replicação do DNA/efeitos dos fármacos , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Inibidores Enzimáticos/química , Exodesoxirribonucleases/genética , Exodesoxirribonucleases/metabolismo , Células HeLa , Histonas/metabolismo , Humanos , Immunoblotting , Maleimidas/química , Estrutura Molecular , Oxazóis/farmacologia , Antígeno Nuclear de Célula em Proliferação/metabolismo , RecQ Helicases/genética , RecQ Helicases/metabolismo , Fase S/efeitos dos fármacos , Estresse Fisiológico/efeitos dos fármacos , Fatores de Tempo , Inibidores da Topoisomerase I/farmacologia , Topotecan/farmacologia , Helicase da Síndrome de WernerRESUMO
Cancer prevention and early detection, the first two of the eight primary goals of the National Cancer Plan released in April 2023, are at the forefront of the nation's strategic efforts to reduce cancer incidence and mortality. The Division of Cancer Prevention (DCP) of the NCI is the federal government's principal component devoted to promoting and supporting innovative cancer prevention research. Recent advances in tumor immunology, cancer immunotherapy, and vaccinology strongly suggest that the host immune system can be effectively harnessed to elicit protective immunity against the development of cancer, that is, cancer immunoprevention. Cancer immunoprevention may be most effective if the intervention is given before or early in the carcinogenic process while the immune system remains relatively uncompromised. DCP has increased the emphasis on immunoprevention research in recent years and continues to expand program resources and interagency collaborations designed to facilitate research in the immunoprevention field. These resources support a wide array of basic, translational, and clinical research activities, including discovery, development, and validation of biomarkers for cancer risk assessment and early detection (Early Detection Research Network), elucidation of biological and pathophysiological mechanistic determinants of precancer growth and its control (Translational and Basic Science Research in Early Lesions), spatiotemporal multiomics characterization of precancerous lesions (Human Tumor Atlas Network/Pre-Cancer Atlas), discovery of immunoprevention pathways and immune targets (Cancer Immunoprevention Network), and preclinical and clinical development of novel agents for immunoprevention and interception (Cancer Prevention-Interception Targeted Agent Discovery Program, PREVENT Cancer Preclinical Drug Development Program, and Cancer Prevention Clinical Trials Network).
Assuntos
Antineoplásicos , Vacinas Anticâncer , Neoplasias , Humanos , Vacinas Anticâncer/uso terapêutico , Imunoterapia , Neoplasias/prevenção & controle , BiomarcadoresRESUMO
The concept of green chemoprevention was introduced in 2012 by Drs. Jed Fahey and Thomas Kensler as whole-plant foods and/or extract-based interventions demonstrating cancer prevention activity. Refining concepts and research demonstrating proof-of-principle approaches are highlighted within this review. Early approaches included extensively investigated whole foods, including broccoli sprouts and black raspberries showing dose-responsive effects across a range of activities in both animals and humans with minimal or no apparent toxicity. A recent randomized crossover trial evaluating the detoxification of tobacco carcinogens by a broccoli seed and sprout extract in the high-risk cohort of current smokers highlights the use of a dietary supplement as a potential next-generation green chemoprevention or green cancer prevention approach. Challenges are addressed, including the selection of dose, duration and mode of delivery, choice of control group, and standardization of the plant food or extract. Identification and characterization of molecular targets and careful selection of high-risk cohorts for study are additional important considerations when designing studies. Goals for precision green cancer prevention include acquiring robust evidence from carefully controlled human studies linking plant foods, extracts, and compounds to modulation of targets for cancer risk reduction in individual cancer types.
Assuntos
Neoplasias , Animais , Humanos , Neoplasias/prevenção & controle , Quimioprevenção , Suplementos NutricionaisRESUMO
BACKGROUND: Lynch syndrome is a hereditary cancer predisposition syndrome caused by germline mutations in DNA mismatch repair genes, which lead to high microsatellite instability and frameshift mutations at coding mononucleotide repeats in the genome. Recurrent frameshift mutations in these regions are thought to play a central role in the increased risk of various cancers, but no biomarkers are currently available for the surveillance of high microsatellite instability-associated cancers. METHODS: A frameshift mutation-based biomarker panel was developed and validated by targeted next-generation sequencing of supernatant DNA from cultured high microsatellite instability colorectal cancer cells. This panel supported selection of 122 frameshift mutation targets as potential biomarkers. This biomarker panel was then tested using matched tumor, adjacent normal tissue, and buffy coat samples (53 samples) and blood-derived cell-free DNA (cfDNA) (38 samples) obtained from 45 high microsatellite instability and mismatch repair-deficient patients. We also sequenced cfDNA from 84 healthy participants to assess background noise. RESULTS: Recurrent frameshift mutations at coding mononucleotide repeats were detectable not only in tumors but also in cfDNA from high microsatellite instability and mismatch repair-deficient patients, including a Lynch syndrome carrier, with a varying range of target detection (up to 85.2%), whereas they were virtually undetectable in healthy participants. Receiver operating characteristic curve analysis showed high sensitivity and specificity (area under the curve = 0.94) of the investigated panel. CONCLUSIONS: We demonstrated that frameshift mutations can be detected in cfDNA from high microsatellite instability and mismatch repair-deficient patients and asymptomatic carriers. The 122-target frameshift mutation panel described here has promise as a tool for improved surveillance of high microsatellite instability and mismatch repair-deficient patients, with the potential to reduce the frequency of invasive screening methods for this high-cancer-risk cohort.
Assuntos
Biomarcadores Tumorais , Neoplasias Colorretais Hereditárias sem Polipose , Mutação da Fase de Leitura , Instabilidade de Microssatélites , Humanos , Neoplasias Colorretais Hereditárias sem Polipose/genética , Neoplasias Colorretais Hereditárias sem Polipose/diagnóstico , Neoplasias Colorretais Hereditárias sem Polipose/sangue , Biomarcadores Tumorais/sangue , Biomarcadores Tumorais/genética , Masculino , Feminino , Pessoa de Meia-Idade , Adulto , Idoso , Reparo de Erro de Pareamento de DNA/genética , Sequenciamento de Nucleotídeos em Larga Escala , Curva ROC , Estudos de Casos e Controles , Sensibilidade e EspecificidadeRESUMO
In diploid mammalian genomes, parental alleles can exhibit different methylation patterns (allele-specific DNA methylation, ASM), which have been documented in a small number of cases except for the imprinted regions and X chromosomes in females. We carried out a chromosome-wide survey of ASM across 16 human pluripotent and adult cell lines using Illumina bisulfite sequencing. We applied the principle of linkage disequilibrium (LD) analysis to characterize the correlation of methylation between adjacent CpG sites on single DNA molecules, and also investigated the correlation between CpG methylation and single nucleotide polymorphisms (SNPs). We observed ASM on 23% approximately 37% heterozygous SNPs in any given cell line. ASM is often cell-type-specific. Furthermore, we found that a significant fraction (38%-88%) of ASM regions is dependent on the presence of heterozygous SNPs in CpG dinucleotides that disrupt their methylation potential. This study identified distinct types of ASM across many cell types and suggests a potential role for CpG-SNP in connecting genetic variation with the epigenome.
Assuntos
Ilhas de CpG/genética , Metilação de DNA/fisiologia , Genoma Humano/genética , Polimorfismo de Nucleotídeo Único/fisiologia , Adulto , Alelos , Sequência de Bases , Linhagem Celular , Metilação de DNA/genética , Análise Mutacional de DNA , Feminino , Frequência do Gene , Humanos , Desequilíbrio de LigaçãoRESUMO
Epidermal growth factor receptor (EGFR) mutations occur in about 50% of lung adenocarcinomas in Asia and about 15% in the US. EGFR mutation-specific inhibitors have been developed and made significant contributions to controlling EGFR mutated non-small cell lung cancer. However, resistance frequently develops within 1 to 2 years due to acquired mutations. No effective approaches that target mutant EGFR have been developed to treat relapse following tyrosine kinase inhibitor (TKI) treatment. Vaccination against mutant EGFR is one area of active exploration. In this study, we identified immunogenic epitopes for the common EGFR mutations in humans and formulated a multi-peptide vaccine (Emut Vax) targeting the EGFR L858R, T790M, and Del19 mutations. The efficacy of the Emut Vax was evaluated in both syngeneic and genetic engineered EGFR mutation-driven murine lung tumor models with prophylactic settings, where the vaccinations were given before the onset of the tumor induction. The multi-peptide Emut Vax effectively prevented the onset of EGFR mutation-driven lung tumorigenesis in both syngeneic and genetically engineered mouse models (GEMMs). Flow cytometry and single-cell RNA sequencing were conducted to investigate the impact of Emut Vax on immune modulation. Emut Vax significantly enhanced Th1 responses in the tumor microenvironment and decreased suppressive Tregs to enhance anti-tumor efficacy. Our results show that multi-peptide Emut Vax is effective in preventing common EGFR mutation-driven lung tumorigenesis, and the vaccine elicits broad immune responses that are not limited to anti-tumor Th1 response.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Humanos , Animais , Camundongos , Receptores ErbB , Mutação , Inibidores de Proteínas Quinases , Recidiva Local de Neoplasia , Carcinogênese , Transformação Celular Neoplásica , Microambiente TumoralRESUMO
The continuing emergence of SARS-CoV-2 variants has highlighted the need to identify additional points for viral inhibition. Ribosome inactivating proteins (RIPs), such as MAP30 and Momordin which are derived from bitter melon (Momordica charantia), have been found to inhibit a broad range of viruses. MAP30 has been shown to potently inhibit HIV-1 with minimal cytotoxicity. Here we show that MAP30 and Momordin potently inhibit SARS-CoV-2 replication in A549 human lung cells (IC50 ~ 0.2 µM) with little concomitant cytotoxicity (CC50 ~ 2 µM). Both viral inhibition and cytotoxicity remain unaltered by appending a C-terminal Tat cell-penetration peptide to either protein. Mutation of tyrosine 70, a key residue in the active site of MAP30, to alanine completely abrogates both viral inhibition and cytotoxicity, indicating the involvement of its RNA N-glycosylase activity. Mutation of lysine 171 and lysine 215, residues corresponding to those in Ricin which when mutated prevented ribosome binding and inactivation, to alanine in MAP30 decreased cytotoxicity (CC50 ~ 10 µM) but also the viral inhibition (IC50 ~ 1 µM). Unlike with HIV-1, neither Dexamethasone nor Indomethacin exhibited synergy with MAP30 in the inhibition of SARS-CoV-2. From a structural comparison of the two proteins, one can explain their similar activities despite differences in both their active-sites and ribosome-binding regions. We also note points on the viral genome for potential inhibition by these proteins.