RESUMEN
Complex rearrangement patterns and mitotic errors are hallmarks of most pancreatic ductal adenocarcinomas (PDAC), a disease with dismal prognosis despite some therapeutic advances in recent years. DNA double-strand breaks (DSB) bear the greatest risk of provoking genomic instability, and DNA damage repair (DDR) pathways are crucial in preserving genomic integrity following a plethora of damage types. Two major repair pathways dominate DSB repair for safeguarding the genome integrity: non-homologous end joining and homologous recombination (HR). Defective HR, but also alterations in other DDR pathways, such as BRCA1, BRCA2, ATM and PALB2, occur frequently in both inherited and sporadic PDAC. Personalised treatment of pancreatic cancer is still in its infancy and predictive biomarkers are lacking. DDR deficiency might render a PDAC vulnerable to a potential new therapeutic intervention that increases the DNA damage load beyond a tolerable threshold, as for example, induced by poly (ADP-ribose) polymerase inhibitors. The Pancreas Cancer Olaparib Ongoing (POLO) trial, in which olaparib as a maintenance treatment improved progression-free survival compared with placebo after platinum-based induction chemotherapy in patients with PDAC and germline BRCA1/2 mutations, raised great hopes of a substantially improved outcome for this patient subgroup. This review summarises the relationship between DDR and PDAC, the prevalence and characteristics of DNA repair mutations and options for the clinical management of patients with PDAC and DNA repair deficiency.
Asunto(s)
Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/terapia , Reparación del ADN , Antineoplásicos/uso terapéutico , Biomarcadores de Tumor/genética , Daño del ADN , Predicción , Humanos , Ftalazinas/uso terapéutico , Piperazinas/uso terapéuticoRESUMEN
OBJECTIVE: ATM serine/threonine kinase (ATM) is the most frequently mutated DNA damage response gene, involved in homologous recombination (HR), in pancreatic ductal adenocarcinoma (PDAC). DESIGN: Combinational synergy screening was performed to endeavour a genotype-tailored targeted therapy. RESULTS: Synergy was found on inhibition of PARP, ATR and DNA-PKcs (PAD) leading to synthetic lethality in ATM-deficient murine and human PDAC. Mechanistically, PAD-induced PARP trapping, replication fork stalling and mitosis defects leading to P53-mediated apoptosis. Most importantly, chemical inhibition of ATM sensitises human PDAC cells toward PAD with long-term tumour control in vivo. Finally, we anticipated and elucidated PARP inhibitor resistance within the ATM-null background via whole exome sequencing. Arising cells were aneuploid, underwent epithelial-mesenchymal-transition and acquired multidrug resistance (MDR) due to upregulation of drug transporters and a bypass within the DNA repair machinery. These functional observations were mirrored in copy number variations affecting a region on chromosome 5 comprising several of the upregulated MDR genes. Using these findings, we ultimately propose alternative strategies to overcome the resistance. CONCLUSION: Analysis of the molecular susceptibilities triggered by ATM deficiency in PDAC allow elaboration of an efficient mutation-specific combinational therapeutic approach that can be also implemented in a genotype-independent manner by ATM inhibition.
Asunto(s)
Adenocarcinoma/genética , Proteínas de la Ataxia Telangiectasia Mutada/genética , Carcinoma Ductal Pancreático/genética , Recombinación Homóloga , Neoplasias Pancreáticas/genética , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Adenocarcinoma/tratamiento farmacológico , Animales , Apoptosis , Carcinoma Ductal Pancreático/tratamiento farmacológico , Línea Celular Tumoral , Supervivencia Celular , Variaciones en el Número de Copia de ADN , Daño del ADN , Reparación del ADN , Resistencia a Múltiples Medicamentos/genética , Sinergismo Farmacológico , Transición Epitelial-Mesenquimal , Genotipo , Humanos , Ratones , Neoplasias Pancreáticas/tratamiento farmacológico , PronósticoRESUMEN
Current concepts in treating cancer usually neglect individual tumor characteristics such as a given mutational make up. Consequently, a "one-size-fits-all" therapeutic concept may commonly fail in terms of efficacy, evolving drug resistance, and side effects. In times of omics, novel elaborated and personalized approaches emerge for efficiently eradicate cancer cells, while sparing healthy cells. Synthetic lethality-based strategies offer promising opportunities to exploit tumor-specific vulnerabilities and improve tolerability. Furthermore, taking advantage of putative synergistic interaction between synthetic lethal drugs specifically targeting a given tumor genotype, could further enhance efficacy and tolerability, thus preventing drug resistance. Mechanisms of drug resistance in cancers are manifold but critical to assess, in view of restoring drug sensibility. In this chapter, we provide a framework to investigate synthetic lethality and synergistic interactions, as well as drug resistance in cancer cells in vitro.
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Neoplasias , Mutaciones Letales Sintéticas , Sinergismo Farmacológico , Humanos , Mutación , Neoplasias/genética , Flujo de TrabajoRESUMEN
As one of the deadliest cancers, pancreatic ductal adenocarcinoma (PDAC) requires sophisticated model systems to dissect disease onset, progression, and therapy resistance, as well as to personalize therapy. In recent years, patient- and pluripotent stem cell-derived organoids have become state-of-the-art systems to refine existing therapeutic strategies and deepen our knowledge of disease pathophysiology.
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Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/terapia , Humanos , Organoides/patología , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patología , Neoplasias Pancreáticas/terapia , Neoplasias PancreáticasRESUMEN
Despite intensive research and progress in personalized medicine, pancreatic ductal adenocarcinoma remains one of the deadliest cancer entities. Pancreatic duct-like organoids (PDLOs) derived from human pluripotent stem cells (PSCs) or pancreatic cancer patient-derived organoids (PDOs) provide unique tools to study early and late stage dysplasia and to foster personalized medicine. However, such advanced systems are neither rapidly nor easily accessible and require an in vivo niche to study tumor formation and interaction with the stroma. Here, the establishment of the porcine urinary bladder (PUB) is revealed as an advanced organ culture model for shaping an ex vivo pancreatic niche. This model allows pancreatic progenitor cells to enter the ductal and endocrine lineages, while PDLOs further mature into duct-like tissue. Accordingly, the PUB offers an ex vivo platform for earliest pancreatic dysplasia and cancer if PDLOs feature KRASG12D mutations. Finally, it is demonstrated that PDOs-on-PUB i) resemble primary pancreatic cancer, ii) preserve cancer subtypes, iii) enable the study of niche epithelial crosstalk by spiking in pancreatic stellate and immune cells into the grafts, and finally iv) allow drug testing. In summary, the PUB advances the existing pancreatic cancer models by adding feasibility, complexity, and customization at low cost and high flexibility.
Asunto(s)
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Células Madre Pluripotentes , Animales , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patología , Humanos , Organoides/patología , Neoplasias Pancreáticas/patología , Porcinos , Vejiga Urinaria , Neoplasias PancreáticasRESUMEN
Real-time isolation, propagation, and pharmacotyping of patient-derived pancreatic cancer organoids (PDOs) may enable treatment response prediction and personalization of pancreatic cancer (PC) therapy. In our methodology, PDOs are isolated from 54 patients with suspected or confirmed PC in the framework of a prospective feasibility trial. The drug response of single agents is determined by a viability assay. Areas under the curves (AUC) are clustered for each drug, and a prediction score is developed for combined regimens. Pharmacotyping profiles are obtained from 28 PDOs (efficacy 63.6%) after a median of 53 days (range 21-126 days). PDOs exhibit heterogeneous responses to the standard-of-care drugs, and are classified into high, intermediate, or low responder categories. Our developed prediction model allows a successful response prediction in treatment-naïve patients with an accuracy of 91.1% for first-line and 80.0% for second-line regimens, respectively. The power of prediction declines in pretreated patients (accuracy 40.0%), particularly with more than one prior line of chemotherapy. Progression-free survival (PFS) is significantly longer in previously treatment-naïve patients receiving a predicted tumor sensitive compared to a predicted tumor resistant regimen (mPFS 141 vs. 46 days; p = 0.0048). In conclusion, generation and pharmacotyping of PDOs is feasible in clinical routine and may provide substantial benefit.
RESUMEN
Pancreatic ductal adenocarcinoma (PDAC) still presents with a dismal prognosis despite intense research. Better understanding of cellular homeostasis could identify druggable targets to improve therapy. Here we propose RAD50-interacting protein 1 (RINT1) as an essential mediator of cellular homeostasis in PDAC. In a cohort of resected PDAC, low RINT1 protein expression correlated significantly with better survival. Accordingly, RINT1 depletion caused severe growth defects in vitro associated with accumulation of DNA double-strand breaks (DSB), G2 cell cycle arrest, disruption of Golgi-endoplasmic reticulum homeostasis, and cell death. Time-resolved transcriptomics corroborated by quantitative proteome and interactome analyses pointed toward defective SUMOylation after RINT1 loss, impairing nucleocytoplasmic transport and DSB response. Subcutaneous xenografts confirmed tumor response by RINT1 depletion, also resulting in a survival benefit when transferred to an orthotopic model. Primary human PDAC organoids licensed RINT1 relevance for cell viability. Taken together, our data indicate that RINT1 loss affects PDAC cell fate by disturbing SUMOylation pathways. Therefore, a RINT1 interference strategy may represent a new putative therapeutic approach. SIGNIFICANCE: These findings provide new insights into the aggressive behavior of PDAC, showing that RINT1 directly correlates with survival in patients with PDAC by disturbing the SUMOylation process, a crucial modification in carcinogenesis.
Asunto(s)
Carcinoma Ductal Pancreático , Proteínas de Ciclo Celular/fisiología , Reparación del ADN/genética , Neoplasias Pancreáticas , Sumoilación , Animales , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patología , Línea Celular Tumoral , Estudios de Cohortes , Daño del ADN/genética , Femenino , Homeostasis/genética , Humanos , Ratones , Ratones Desnudos , Ratones Transgénicos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Procesamiento Proteico-Postraduccional/genética , Sumoilación/genéticaRESUMEN
Somatic cell reprogramming and tissue repair share relevant factors and molecular programs. Here, Dickkopf-3 (DKK3) is identified as novel factor for organ regeneration using combined transcription-factor-induced reprogramming and RNA-interference techniques. Loss of Dkk3 enhances the generation of induced pluripotent stem cells but does not affect de novo derivation of embryonic stem cells, three-germ-layer differentiation or colony formation capacity of liver and pancreatic organoids. However, DKK3 expression levels in wildtype animals and serum levels in human patients are elevated upon injury. Accordingly, Dkk3-null mice display less liver damage upon acute and chronic failure mediated by increased proliferation in hepatocytes and LGR5+ liver progenitor cell population, respectively. Similarly, recovery from experimental pancreatitis is accelerated. Regeneration onset occurs in the acinar compartment accompanied by virtually abolished canonical-Wnt-signaling in Dkk3-null animals. This results in reduced expression of the Hedgehog repressor Gli3 and increased Hedgehog-signaling activity upon Dkk3 loss. Collectively, these data reveal Dkk3 as a key regulator of organ regeneration via a direct, previously unacknowledged link between DKK3, canonical-Wnt-, and Hedgehog-signaling.
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Proteínas Adaptadoras Transductoras de Señales/genética , Reprogramación Celular/genética , Reprogramación Celular/fisiología , Genómica/métodos , Organogénesis/genética , Organogénesis/fisiología , Animales , Modelos Animales de Enfermedad , Ratones , Ratones Endogámicos C57BL , Regeneración/genética , Regeneración/fisiologíaRESUMEN
Personalized medicine in treating pancreatic ductal adenocarcinoma (PDAC) is still in its infancy, albeit PDAC-related deaths are projected to rise over the next decade. Only recently, maintenance therapy with the PARP inhibitor olaparib showed improved progression-free survival in germline BRCA1/2-mutated PDAC patients after platinum-based induction for the first time. Transferability of such a concept to other DNA damage response (DDR) genes remains unclear. Here, we conducted a placebo-controlled, three-armed preclinical trial to evaluate the efficacy of multi-DDR interference (mDDRi) as maintenance therapy vs. continuous FOLFIRINOX treatment, implemented with orthotopically transplanted ATM-deficient PDAC cell lines. Kaplan-Meier analysis, cross-sectional imaging, histology, and in vitro analysis served as analytical readouts. Median overall survival was significantly longer in the mDDRi maintenance arm compared to the maintained FOLFIRINOX treatment. This survival benefit was mirrored in the highest DNA-damage load, accompanied by superior disease control and reduced metastatic burden. In vitro analysis suggests FOLFIRINOX-driven selection of invasive subclones, erased by subsequent mDDRi treatment. Collectively, this preclinical trial substantiates mDDRi in a maintenance setting as a novel therapeutic option and extends the concept to non-germline BRCA1/2-mutant PDAC.
Asunto(s)
Proteínas de la Ataxia Telangiectasia Mutada/genética , Carcinoma Ductal Pancreático/tratamiento farmacológico , Quimioterapia de Mantención/métodos , Neoplasias Pancreáticas/tratamiento farmacológico , Ftalazinas/uso terapéutico , Piperazinas/uso terapéutico , Inhibidores de Poli(ADP-Ribosa) Polimerasas/uso terapéutico , Animales , Antineoplásicos/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Proteínas de la Ataxia Telangiectasia Mutada/deficiencia , Proteína BRCA2/deficiencia , Proteína BRCA2/genética , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/mortalidad , Carcinoma Ductal Pancreático/patología , Línea Celular Tumoral , Inestabilidad Cromosómica , Daño del ADN , Femenino , Fluorouracilo/uso terapéutico , Regulación Neoplásica de la Expresión Génica , Humanos , Irinotecán/uso terapéutico , Leucovorina/uso terapéutico , Ratones , Ratones Desnudos , Persona de Mediana Edad , Oxaliplatino/uso terapéutico , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/mortalidad , Neoplasias Pancreáticas/patología , Poli(ADP-Ribosa) Polimerasas , Análisis de Supervivencia , Ubiquitina-Proteína Ligasas/deficiencia , Ubiquitina-Proteína Ligasas/genética , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The original version of this article contained an error in the name of one of the co-authors (Kayvan Mohkam). This has been corrected in the PDF and HTML versions.
RESUMEN
Pancreatic ductal adenocarcinoma (PDAC) is one of the solid tumors with the poorest prognosis. The stroma of this tumor is abundant and composed of extracellular matrix and stromal cells (including cancer-associated fibroblasts and immune cells). Nerve fibers invading this stroma represent a hallmark of PDAC, involved in neural remodeling, which participates in neuropathic pain, cancer cell dissemination and tumor relapse after surgery. Pancreatic cancer-associated neural remodeling is regulated through functional interplays mediated by physical and molecular interactions between cancer cells, nerve cells and surrounding Schwann cells, and other stromal cells. In the present study, we show that Schwann cells (glial cells supporting peripheral neurons) can enhance aggressiveness (migration, invasion, tumorigenicity) of pancreatic cancer cells in a transforming growth factor beta (TGFß)-dependent manner. Indeed, we reveal that conditioned medium from Schwann cells contains high amounts of TGFß able to activate the TGFß-SMAD signaling pathway in cancer cells. We also observed in human PDAC samples that high levels of TGFß signaling activation were positively correlated with perineural invasion. Secretome analyses by mass spectrometry of Schwann cells and pancreatic cancer cells cultured alone or in combination highlighted the central role of TGFß in neuro-epithelial interactions, as illustrated by proteomic signatures related to cell adhesion and motility. Altogether, these results demonstrate that Schwann cells are a meaningful source of TGFß in PDAC, which plays a crucial role in the acquisition of aggressive properties by pancreatic cancer cells.
RESUMEN
BACKGROUND & AIMS: Transforming growth factor beta (TGFß) acts either as a tumor suppressor or as an oncogene, depending on the cellular context and time of activation. TGFß activates the canonical SMAD pathway through its interaction with the serine/threonine kinase type I and II heterotetrameric receptors. Previous studies investigating TGFß-mediated signaling in the pancreas relied either on loss-of-function approaches or on ligand overexpression, and its effects on acinar cells have so far remained elusive. METHODS: We developed a transgenic mouse model allowing tamoxifen-inducible and Cre-mediated conditional activation of a constitutively active type I TGFß receptor (TßRICA) in the pancreatic acinar compartment. RESULTS: We observed that TßRICA expression induced acinar-to-ductal metaplasia (ADM) reprogramming, eventually facilitating the onset of KRASG12D-induced pre-cancerous pancreatic intraepithelial neoplasia. This phenotype was characterized by the cellular activation of apoptosis and dedifferentiation, two hallmarks of ADM, whereas at the molecular level, we evidenced a modulation in the expression of transcription factors such as Hnf1ß, Sox9, and Hes1. CONCLUSIONS: We demonstrate that TGFß pathway activation plays a crucial role in pancreatic tumor initiation through its capacity to induce ADM, providing a favorable environment for KRASG12D-dependent carcinogenesis. Such findings are highly relevant for the development of early detection markers and of potentially novel treatments for pancreatic cancer patients.