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The role of store-operated Ca2+ entry (SOCE) in melanoma metastasis is highly controversial. To address this, we here examined UV-dependent metastasis, revealing a critical role for SOCE suppression in melanoma progression. UV-induced cholesterol biosynthesis was critical for UV-induced SOCE suppression and subsequent metastasis, although SOCE suppression alone was both necessary and sufficient for metastasis to occur. Further, SOCE suppression was responsible for UV-dependent differences in gene expression associated with both increased invasion and reduced glucose metabolism. Functional analyses further established that increased glucose uptake leads to a metabolic shift towards biosynthetic pathways critical for melanoma metastasis. Finally, examination of fresh surgically isolated human melanoma explants revealed cholesterol biosynthesis-dependent reduced SOCE. Invasiveness could be reversed with either cholesterol biosynthesis inhibitors or pharmacological SOCE potentiation. Collectively, we provide evidence that, contrary to current thinking, Ca2+ signals can block invasive behavior, and suppression of these signals promotes invasion and metastasis.
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Señalización del Calcio , Melanoma , Calcio/metabolismo , Canales de Calcio/metabolismo , Colesterol , Glucosa , Humanos , Melanoma/genética , Melanoma/metabolismo , Proteína ORAI1/metabolismo , Molécula de Interacción Estromal 1/metabolismoRESUMEN
Pancreatic cancer is typically detected at an advanced stage, and is refractory to most forms of treatment, contributing to poor survival outcomes. The incidence of pancreatic cancer is gradually increasing, linked to an aging population and increasing rates of obesity and pancreatitis, which are risk factors for this cancer. Sources of risk include adipokine signaling from fat cells throughout the body, elevated levels of intrapancreatic intrapancreatic adipocytes (IPAs), inflammatory signals arising from pancreas-infiltrating immune cells and a fibrotic environment induced by recurring cycles of pancreatic obstruction and acinar cell lysis. Once cancers become established, reorganization of pancreatic tissue typically excludes IPAs from the tumor microenvironment, which instead consists of cancer cells embedded in a specialized microenvironment derived from cancer-associated fibroblasts (CAFs). While cancer cell interactions with CAFs and immune cells have been the topic of much investigation, mechanistic studies of the source and function of IPAs in the pre-cancerous niche are much less developed. Intriguingly, an extensive review of studies addressing the accumulation and activity of IPAs in the pancreas reveals that unexpectedly diverse group of factors cause replacement of acinar tissue with IPAs, particularly in the mouse models that are essential tools for research into pancreatic cancer. Genes implicated in regulation of IPA accumulation include KRAS, MYC, TGF-ß, periostin, HNF1, and regulators of ductal ciliation and ER stress, among others. These findings emphasize the importance of studying pancreas-damaging factors in the pre-cancerous environment, and have significant implications for the interpretation of data from mouse models for pancreatic cancer.
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Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Pancreatitis , Ratones , Animales , Neoplasias Pancreáticas/patología , Pancreatitis/patología , Páncreas/patología , Células Acinares/patología , Carcinoma Ductal Pancreático/patología , Microambiente Tumoral , Neoplasias PancreáticasRESUMEN
The chemoresistance of tumor cells is one of the most urgent challenges in modern oncology and in pancreatic cancer, in which this problem is the most prominent. Therefore, the identification of new chemosensitizing co-targets may be a path toward increasing chemotherapy efficacy. In this work, we performed high-performance in vitro knockout CRISPR/Cas9 screening to find potential regulators of the sensitivity of pancreatic cancer. For this purpose, MIA PaCa-2 cells transduced with two sgRNA libraries ("cell cycle/nuclear proteins genes" and "genome-wide") were screened by oxaliplatin and cisplatin. In total, 173 candidate genes were identified as potential regulators of pancreatic cancer cell sensitivity to oxaliplatin and/or cisplatin; among these, 25 genes have previously been reported, while 148 genes were identified for the first time as potential platinum drug sensitivity regulators. We found seven candidate genes involved in pancreatic cancer cell sensitivity to both cisplatin and oxaliplatin. Gene ontology enrichment analysis reveals the enrichment of single-stranded DNA binding, damaged DNA binding pathways, and four associated with NADH dehydrogenase activity. Further investigation and validation of the obtained results by in vitro, in vivo, and bioinformatics approaches, as well as literature analysis, will help to identify novel pancreatic cancer platinum sensitivity regulators.
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Antineoplásicos/farmacología , Cisplatino/farmacología , Resistencia a Antineoplásicos/efectos de los fármacos , Oxaliplatino/farmacología , Biomarcadores de Tumor , Sistemas CRISPR-Cas , Línea Celular Tumoral , Biología Computacional/métodos , Resistencia a Antineoplásicos/genética , Ensayos de Selección de Medicamentos Antitumorales , Perfilación de la Expresión Génica , Técnicas de Silenciamiento del Gen , Ontología de Genes , Redes Reguladoras de Genes , Humanos , Neoplasias Pancreáticas , Mutaciones Letales SintéticasRESUMEN
BACKGROUND: Treatment with tyrosine kinase inhibitors (TKIs) significantly improves survival of patients with renal cell carcinoma (RCC). However, about one-quarter of the RCC patients are primarily refractory to treatment with TKIs. METHODS: We examined viability of RCC and endothelial cells treated with low-density lipoprotein (LDL) and/or TKIs. Next, we validated the potential role of PI3K/AKT signalling in LDL-mediated TKI resistance. Finally, we examined the effect of a high-fat/high-cholesterol diet on the response of RCC xenograft tumours to sunitinib. RESULTS: The addition of LDL cholesterol increases activation of PI3K/AKT signalling and compromises the antitumour efficacy of TKIs against RCC and endothelial cells. Furthermore, RCC xenograft tumours resist TKIs in mice fed a high-fat/high-cholesterol diet. CONCLUSIONS: The ability of renal tumours to maintain their cholesterol homoeostasis may be a critical component of TKI resistance in RCC patients.
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Carcinoma de Células Renales/tratamiento farmacológico , Colesterol/metabolismo , Interacciones Farmacológicas/genética , Inhibidores de Proteínas Quinasas/administración & dosificación , Animales , Carcinoma de Células Renales/metabolismo , Carcinoma de Células Renales/patología , Línea Celular Tumoral , LDL-Colesterol/administración & dosificación , LDL-Colesterol/metabolismo , Interacciones Farmacológicas/etnología , Elafina/genética , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Femenino , Humanos , Indoles/administración & dosificación , Ratones , Proteínas Proto-Oncogénicas c-akt/genética , Pirroles/administración & dosificación , Transducción de Señal/efectos de los fármacos , Sunitinib , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: The rarity of neuroendocrine malignancies limits the ability to develop new therapies and thus a better understanding of the underlying biology is critical. METHODS: Through a prospective, IRB-approved protocol, patients with neuroendocrine malignancies underwent next-generation sequencing of their tumours to detect somatic mutations (SMs) in 50 cancer-related genes. Clinicopathologic correlation was made among poorly differentiated neuroendocrine carcinomas (NECs/poorly differentiated histology and Ki-67 >20%) and pancreatic neuroendocrine tumours (PanNETs/Ki67 ⩽20%) and non-pancreatic neuroendocrine tumours (NP-NETs/Ki67 ⩽20%). RESULTS: A total of 77 patients were enrolled, with next-generation sequencing results available on 63 patients. Incidence of SMs was 83% (19 out of 23) in poorly differentiated NECs, 45% (5 out of 11) in PanNETs and 14% (4 out of 29) in NP-NETs. TP53 was the most prevalent mutation in poorly differentiated NECs (57%), and KRAS (30%), PIK3CA/PTEN (22%) and BRAF (13%) mutations were also found. Small intestinal neuroendocrine tumours (Ki67 <2%/n=9) did not harbour any mutations. Prevalence of mutations correlated with higher risk of progression within the previous year (32% (low risk) vs 11% (high risk), P=0.01) and TP53 mutation correlated with worse survival (2-year survival 66% vs 97%, P=0.003). CONCLUSIONS: Poorly differentiated NECs have a high mutation burden with potentially targetable mutations. The TP53 mutations are associated with poor survival in neuroendocrine malignancies. These findings have clinical trial implications for choice of therapy and prognostic stratification and warrant confirmation.
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Biomarcadores de Tumor/metabolismo , Tumores Neuroendocrinos/patología , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Perfilación de la Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Tumores Neuroendocrinos/genética , Proyectos Piloto , PronósticoRESUMEN
Neuroendocrine tumors (NET) and gastrointestinal stromal tumors (GIST) are believed to originate from the cells of Cajal that are randomly dispersed along the aerodigestive tract. Despite their distinct morphologic appearance, NET and GIST may share oncogenic mechanisms. Often presenting in the metastatic setting, treatment options for patients with NET are limited. This case report presents a patient with refractory metastatic NET that did not respond conventional chemotherapy. The patient was treated with a KIF11 inhibitor in a phase I clinical trial and experienced a prolonged and clinically meaningful partial response. On progression at 20 months, the patient's tumor was sequenced to reveal a KIT exon 11 mutation. Institution of imatinib therapy achieved a rapid and sustained antitumor effect with profound clinical benefit. Despite previously reported KIT expression in NET, this is the first documented case of an activating KIT mutation in NET and of successful treatment with both a KIF11 inhibitor and imatinib, each of which was elucidated through molecular profiling of the patient's tumor. Imatinib may be a valuable therapy in NET harboring activating KIT mutations.
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Benzamidas/administración & dosificación , Carcinoma Neuroendocrino/tratamiento farmacológico , Tumores del Estroma Gastrointestinal/tratamiento farmacológico , Piperazinas/administración & dosificación , Proteínas Proto-Oncogénicas c-kit/genética , Pirimidinas/administración & dosificación , Carcinoma Neuroendocrino/genética , Carcinoma Neuroendocrino/patología , Resistencia a Antineoplásicos/genética , Tumores del Estroma Gastrointestinal/genética , Tumores del Estroma Gastrointestinal/patología , Humanos , Mesilato de Imatinib , Cinesinas/antagonistas & inhibidores , Masculino , Persona de Mediana Edad , Transducción de SeñalRESUMEN
Temporally and spatially controlled activation of the Aurora A kinase (AURKA) regulates centrosome maturation, entry into mitosis, formation and function of the bipolar spindle, and cytokinesis. Genetic amplification and mRNA and protein overexpression of Aurora A are common in many types of solid tumor, and associated with aneuploidy, supernumerary centrosomes, defective mitotic spindles, and resistance to apoptosis. These properties have led Aurora A to be considered a high-value target for development of cancer therapeutics, with multiple agents currently in early-phase clinical trials. More recently, identification of additional, non-mitotic functions and means of activation of Aurora A during interphase neurite elongation and ciliary resorption have significantly expanded our understanding of its function, and may offer insights into the clinical performance of Aurora A inhibitors. Here we review the mitotic and non-mitotic functions of Aurora A, discuss Aurora A regulation in the context of protein structural information, and evaluate progress in understanding and inhibiting Aurora A in cancer.
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División Celular , Neoplasias/tratamiento farmacológico , Neoplasias/enzimología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/metabolismo , Animales , Aurora Quinasa A , Aurora Quinasas , Activación Enzimática , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/uso terapéutico , Regulación Neoplásica de la Expresión Génica , Humanos , Modelos Moleculares , Neoplasias/genética , Neoplasias/metabolismo , Conformación Proteica , Mapas de Interacción de Proteínas , Proteínas Serina-Treonina Quinasas/química , Proteínas Serina-Treonina Quinasas/genéticaRESUMEN
In healthy cells, controlled activation of aurora kinases regulates mitosis. Overexpression and hyperactivation of aurora kinases A and B have major roles in tumorigenesis, and can induce aneuploidy and genomic instability. In squamous-cell carcinomas of the head and neck, overexpression of aurora kinase A is associated with decreased survival, and a reduction in aurora kinase A and aurora kinase B expression inhibits cell growth and increases apoptosis. In this Review, we provide an overview of the biological functions of aurora kinases in healthy cells and in cancer cells, and we review small studies and high-throughput datasets that particularly implicate aurora kinase A in the pathogenesis of squamous-cell carcinomas of the head and neck. Early phase trials are beginning to assess the activity of small-molecule inhibitors of aurora kinases. We summarise trials of aurora kinase inhibitors in squamous-cell carcinomas of the head and neck, and discuss directions for future drug combination trials and biomarkers to use with drugs that inhibit aurora kinases.
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Carcinoma de Células Escamosas/etiología , Neoplasias de Cabeza y Cuello/etiología , Proteínas Serina-Treonina Quinasas/fisiología , Aurora Quinasa A , Aurora Quinasa B , Aurora Quinasas , Carcinoma de Células Escamosas/tratamiento farmacológico , Carcinoma de Células Escamosas/enzimología , Ensayos Clínicos como Asunto , Neoplasias de Cabeza y Cuello/tratamiento farmacológico , Neoplasias de Cabeza y Cuello/enzimología , Humanos , Inhibidores de Proteínas Quinasas/uso terapéutico , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/genética , Carcinoma de Células Escamosas de Cabeza y Cuello , TranscriptomaRESUMEN
Extracellular vesicles (EVs) are secreted, cell-derived, membrane-bound compartments implicated in various diseases for their ability to influence distant targets and as carriers of biomarkers. Here, we present a protocol for separating EVs from mammalian pancreatic cancer cells and their characterization using western blot and electron microscopy. We then demonstrate how they are utilized to affect tumor development in a murine model of metastatic pancreatic cancer including a method to quantify hepatic tumor burden in histologic samples. For complete details on the use and execution of this protocol, please refer to Dudgeon et al.1.
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Multiple sclerosis (MS) is a chronic progressive, demyelinating condition whose therapeutic needs are unmet, and whose pathoetiology is elusive. We report that transient receptor potential vanilloid-1 (TRPV1) expressed in a major sensory neuron subset, controls severity and progression of experimental autoimmune encephalomyelitis (EAE) in mice and likely in primary progressive MS. TRPV1-/- B6 congenics are protected from EAE. Increased survival reflects reduced central nervous systems (CNS) infiltration, despite indistinguishable T cell autoreactivity and pathogenicity in the periphery of TRPV1-sufficient and -deficient mice. The TRPV1+ neurovascular complex defining the blood-CNS barriers promoted invasion of pathogenic lymphocytes without the contribution of TRPV1-dependent neuropeptides such as substance P. In MS patients, we found a selective risk-association of the missense rs877610 TRPV1 single nucleotide polymorphism (SNP) in primary progressive disease. Our findings indicate that TRPV1 is a critical disease modifier in EAE, and we identify a predictor of severe disease course and a novel target for MS therapy.
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Encefalomielitis Autoinmune Experimental/fisiopatología , Esclerosis Múltiple/genética , Canales Catiónicos TRPV/fisiología , Traslado Adoptivo , Adulto , Animales , Encéfalo/patología , Encéfalo/fisiología , Encefalomielitis Autoinmune Experimental/genética , Encefalomielitis Autoinmune Experimental/terapia , Femenino , Humanos , Ganglios Linfáticos/citología , Masculino , Ratones , Ratones Transgénicos , Glicoproteína Mielina-Oligodendrócito , Fragmentos de Péptidos , Toxina del Pertussis , Polimorfismo de Nucleótido Simple , Médula Espinal/patología , Médula Espinal/fisiología , Bazo/citologíaRESUMEN
The greater efficacy of DNA-damaging drugs for pancreatic adenocarcinoma (PDAC) relies on targeting cancer-specific vulnerabilities while sparing normal organs and tissues due to their inherent toxicities. We tested LP-184, a novel acylfulvene analog, for its activity in preclinical models of PDAC carrying mutations in the DNA damage repair (DDR) pathways. Cytotoxicity of LP-184 is solely dependent on prostaglandin reductase 1 (PTGR1), so that PTGR1 expression robustly correlates with LP-184 cytotoxicity in vitro and in vivo. Low-passage patient-derived PDAC xenografts with DDR deficiencies treated ex vivo are more sensitive to LP-184 compared with DDR-proficient tumors. Additional in vivo testing of PDAC xenografts for their sensitivity to LP-184 demonstrates marked tumor growth inhibition in models harboring pathogenic mutations in ATR, BRCA1, and BRCA2. Depletion of PTGR1, however, completely abrogates the antitumor effect of LP-184. Testing combinatorial strategies for LP-184 aimed at deregulation of nucleotide excision repair proteins ERCC3 and ERCC4 established synergy. Our results provide valuable biomarkers for clinical testing of LP-184 in a large subset of genetically defined characterized refractory carcinomas. High PTGR1 expression and deleterious DDR mutations are present in approximately one third of PDAC making these patients ideal candidates for clinical trials of LP-184.
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Adenocarcinoma , Oxidorreductasas de Alcohol , Antineoplásicos , Daño del ADN , Neoplasias Pancreáticas , Humanos , Reparación del ADN , Neoplasias Pancreáticas/tratamiento farmacológico , Neoplasias Pancreáticas/genética , Oxidorreductasas de Alcohol/genética , Animales , Antineoplásicos/farmacologíaRESUMEN
Cholesterol dependence is an essential characteristic of pancreatic ductal adenocarcinoma (PDAC). Cholesterol 25-hydroxylase (CH25H) catalyzes monooxygenation of cholesterol into 25-hydroxycholesterol, which is implicated in inhibiting cholesterol biosynthesis and in cholesterol depletion. Here, we show that, within PDAC cells, accumulation of cholesterol was facilitated by the loss of CH25H. Methylation of the CH25H gene and decreased levels of CH25H expression occurred in human pancreatic cancers and was associated with poor prognosis. Knockout of Ch25h in mice accelerated progression of Kras-driven pancreatic intraepithelial neoplasia. Conversely, restoration of CH25H expression in human and mouse PDAC cells decreased their viability under conditions of cholesterol deficit, and decelerated tumor growth in immune competent hosts. Mechanistically, the loss of CH25H promoted autophagy resulting in downregulation of MHC-I and decreased CD8+ T-cell tumor infiltration. Re-expression of CH25H in PDAC cells combined with immune checkpoint inhibitors notably inhibited tumor growth. We discuss additional benefits that PDAC cells might gain from inactivation of CH25H and the potential translational importance of these findings for therapeutic approaches to PDAC. IMPLICATIONS: Loss of CH25H by pancreatic cancer cells may stimulate tumor progression and interfere with immunotherapies.
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Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Esteroide Hidroxilasas , Animales , Humanos , Ratones , Carcinoma Ductal Pancreático/patología , Ratones Noqueados , Neoplasias Pancreáticas/patología , Esteroide Hidroxilasas/metabolismo , Neoplasias PancreáticasRESUMEN
The biology of metastatic pancreatic ductal adenocarcinoma (PDAC) is distinct from that of the primary tumor due to changes in cell plasticity governed by a distinct transcriptome. Therapeutic strategies that target this distinct biology are needed. We detect an upregulation of the neuronal axon guidance molecule Netrin-1 in PDAC liver metastases that signals through its dependence receptor (DR), uncoordinated-5b (Unc5b), to facilitate metastasis in vitro and in vivo. The mechanism of Netrin-1 induction involves a feedforward loop whereby Netrin-1 on the surface of PDAC-secreted extracellular vesicles prepares the metastatic niche by inducing hepatic stellate cell activation and retinoic acid secretion that in turn upregulates Netrin-1 in disseminated tumor cells via RAR/RXR and Elf3 signaling. While this mechanism promotes PDAC liver metastasis, it also identifies a therapeutic vulnerability, as it can be targeted using anti-Netrin-1 therapy to inhibit metastasis using the Unc5b DR cell death mechanism.
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Carcinoma Ductal Pancreático , Neoplasias Hepáticas , Neoplasias Pancreáticas , Humanos , Netrina-1 , Retinoides , Células Estrelladas Hepáticas/metabolismo , Línea Celular Tumoral , Neoplasias Pancreáticas/patología , Carcinoma Ductal Pancreático/patología , Neoplasias Hepáticas/metabolismo , Receptores de Netrina , Proteínas de Unión al ADN , Factores de Transcripción , Proteínas Proto-Oncogénicas c-etsRESUMEN
In pancreatic ductal adenocarcinoma (PDAC), the fibroblastic stroma constitutes most of the tumor mass and is remarkably devoid of functional blood vessels. This raises an unresolved question of how PDAC cells obtain essential metabolites and water-insoluble lipids. We have found a critical role for cancer-associated fibroblasts (CAFs) in obtaining and transferring lipids from blood-borne particles to PDAC cells via trogocytosis of CAF plasma membranes. We have also determined that CAF-expressed phospholipid scramblase anoctamin 6 (ANO6) is an essential CAF trogocytosis regulator required to promote PDAC cell survival. During trogocytosis, cancer cells and CAFs form synapse-like plasma membranes contacts that induce cytosolic calcium influx in CAFs via Orai channels. This influx activates ANO6 and results in phosphatidylserine exposure on CAF plasma membrane initiating trogocytosis and transfer of membrane lipids, including cholesterol, to PDAC cells. Importantly, ANO6-dependent trogocytosis also supports the immunosuppressive function of pancreatic CAFs towards cytotoxic T cells by promoting transfer of excessive amounts of cholesterol. Further, blockade of ANO6 antagonizes tumor growth via disruption of delivery of exogenous cholesterol to cancer cells and reverses immune suppression suggesting a potential new strategy for PDAC therapy.
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Toll-like receptors are potent activators of the innate immune system and generate signals leading to the initiation of the adaptive immune response that can be utilized for therapeutic purposes. We tested the hypothesis that combined treatment with a Toll-like receptor agonist and an antitumor monoclonal antibody is effective and induces host-protective antitumor immunity. C57BL/6 human mutated HER2 (hmHER2) transgenic mice that constitutively express kinase-deficient human HER2 under control of the CMV promoter were established. These mice demonstrate immunological tolerance to D5-HER2, a syngeneic human HER2-expressing melanoma cell line. This human HER2-tolerant model offers the potential to serve as a preclinical model to test both antibody therapy and the immunization potential of human HER2-targeted therapeutics. Here, we show that E6020, a Toll-like receptor-4 (TLR4) agonist effectively boosted the antitumor efficacy of the monoclonal antibody trastuzumab in immunodeficient C57BL/6 SCID mice as well as in C57BL/6 hmHER2 transgenic mice. E6020 and trastuzumab co-treatment resulted in significantly greater inhibition of tumor growth than was observed with either agent individually. Furthermore, mice treated with the combination of trastuzumab and the TLR4 agonist were protected against rechallenge with human HER2-transfected tumor cells in hmHER2 transgenic mouse strains. These findings suggest that combined treatment with trastuzumab and a TLR4 agonist not only promotes direct antitumor effects but also induces a host-protective human HER2-directed adaptive immune response, indicative of a memory response. These data provide an immunological rationale for testing TLR4 agonists in combination with antibody therapy in patients with cancer.
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Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Melanoma Experimental/tratamiento farmacológico , Receptor Toll-Like 4/agonistas , Animales , Anticuerpos Monoclonales Humanizados/administración & dosificación , Protocolos de Quimioterapia Combinada Antineoplásica/inmunología , Línea Celular Tumoral , Humanos , Inmunohistoquímica , Melanoma Experimental/enzimología , Melanoma Experimental/genética , Melanoma Experimental/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones SCID , Ratones Transgénicos , Receptor ErbB-2/biosíntesis , Receptor ErbB-2/genética , Receptor ErbB-2/inmunología , Receptor Toll-Like 4/inmunología , TrastuzumabRESUMEN
BACKGROUND: Pathologic response to preoperative therapy is increasingly recognized as an important prognostic factor in solid tumors. The impact of pathologic response on survival in pancreatic adenocarcinoma is not well established. METHODS: Data on 135 consecutive patients treated with chemoradiation followed by pancreatectomy for adenocarcinoma of the pancreatic head and/or body between July 1987 and May 2009 were reviewed. Histopathologic examination was performed in 107 patients to determine pathologic response, defined as minor (<50% fibrosis relative to residual neoplastic cells), partial (50-94% fibrosis), or major (95-100% fibrosis). RESULTS: Minor, partial, and major pathologic response rates were 17% (n = 18), 64% (n = 68), and 19% (n = 21), including a 7% (n = 8) complete pathologic response rate. Pathologic response correlated with R0 resection (P = 0.019), negative lymph nodes (P = 0.006), and smaller tumor size (P = 0.001). Median survival rates by pathologic response were as follows: 17 months [95% confidence interval (CI), 0-36 months] for minor response, 20 months (95% CI, 17-23 months) for partial response, and 66 months (95% CI, 8-124 months) for major response (minor versus partial response, P = not significant; partial versus major response, P < 0.001). On multivariate analysis, major pathologic response was the only factor significantly associated with improved survival (P = 0.025; hazard ratio, 2.26). CONCLUSIONS: Major pathologic response to preoperative therapy occurs in a minority of patients with pancreatic adenocarcinoma and is independently associated with prolonged survival.
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Adenocarcinoma/mortalidad , Adenocarcinoma/patología , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Pancreatectomía , Neoplasias Pancreáticas/mortalidad , Neoplasias Pancreáticas/patología , Adenocarcinoma/terapia , Adulto , Anciano , Anciano de 80 o más Años , Terapia Combinada , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Recurrencia Local de Neoplasia/mortalidad , Recurrencia Local de Neoplasia/patología , Recurrencia Local de Neoplasia/terapia , Estadificación de Neoplasias , Neoplasias Pancreáticas/terapia , Pronóstico , Dosificación Radioterapéutica , Tasa de SupervivenciaRESUMEN
Preparation of single-cell suspension from primary tumor tissue can provide a valuable resource for functional, genetic, proteomic, and tumor microenvironment studies. Here, we describe an effective protocol for mouse pancreatic tumor dissociation with further processing of tumor suspension for single-cell RNA sequencing analysis of cellular populations. We further provide an outline of the bioinformatics processing of the data and clustering of heterogeneous cellular populations comprising pancreatic tumors using Common Workflow Language (CWL) pipelines within user-friendly Scientific Data Analysis Platform (https://SciDAP.com). For complete details on the use and execution of this protocol, please refer to Gabitova-Cornell et al. (2020).
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Biología Computacional/métodos , Neoplasias Pancreáticas , Análisis de Secuencia de ARN/métodos , Análisis de la Célula Individual/métodos , Animales , Femenino , Masculino , Ratones , Páncreas/citología , Páncreas/patología , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Programas InformáticosRESUMEN
We investigated the contributions of platelet microRNAs (miRNAs) to the rate of growth and regulation of gene expression in primary ectopic tumors using mouse models. We previously identified an inhibitory role for platelets in solid tumor growth, mediated by tumor infiltration of platelet microvesicles (microparticles) which are enriched in platelet-derived miRNAs. To investigate the specific roles of platelet miRNAs in tumor growth models, we implanted pancreatic ductal adenocarcinoma cells as a bolus into mice with megakaryocyte-/platelet-specific depletion of mature miRNAs. We observed an ~50% increase in the rate of growth of ectopic primary tumors in these mice compared to controls including at early stages, associated with reduced apoptosis in the tumors, in particular in tumor cells associated with platelet microvesicles-which were depleted of platelet-enriched miRNAs-demonstrating a specific role for platelet miRNAs in modulation of primary tumor growth. Differential expression RNA sequencing of tumor cells isolated from advanced primary tumors revealed a broad cohort of mRNAs modulated in the tumor cells as a function of host platelet miRNAs. Altered genes comprised 548 up-regulated transcripts and 43 down-regulated transcripts, mostly mRNAs altogether spanning a variety of growth signaling pathways-notably pathways related to epithelial-mesenchymal transition-in tumor cells from platelet miRNA-deleted mice compared with those from control mice. Tumors in platelet miRNA-depleted mice showed more sarcomatoid growth and more advanced tumor grade, indicating roles for host platelet miRNAs in tumor plasticity. We further validated increased protein expression of selected genes associated with increased cognate mRNAs in the tumors due to platelet miRNA depletion in the host animals, providing proof of principle of widespread effects of platelet miRNAs on tumor cell functional gene expression in primary tumors in vivo. Together, these data demonstrate that platelet-derived miRNAs modulate solid tumor growth in vivo by broad-spectrum restructuring of the tumor cell transcriptome.
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Plaquetas/metabolismo , MicroARNs/genética , Neoplasias Pancreáticas/genética , ARN Mensajero/genética , Animales , Plaquetas/patología , Carcinogénesis/genética , Carcinogénesis/patología , Femenino , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Ratones , Ratones Endogámicos C57BL , Neoplasias Pancreáticas/patología , TranscriptomaRESUMEN
PURPOSE: Performance status (PS) is a subjective assessment of patients' overall health. Quantification of physical activity using a wearable tracker (Fitbit Charge [FC]) may provide an objective measure of patient's overall PS and treatment tolerance. MATERIALS AND METHODS: Patients with colorectal cancer were prospectively enrolled into two cohorts (medical and surgical) and asked to wear FC for 4 days at baseline (start of new chemotherapy [± 4 weeks] or prior to curative resection) and follow-up (4 weeks [± 2 weeks] after initial assessment in medical and postoperative discharge in surgical cohort). Primary end point was feasibility, defined as 75% of patients wearing FC for at least 12 hours/d, 3 of 4 assigned days. Mean steps per day (SPD) were correlated with toxicities of interest (postoperative complication or ≥ grade 3 toxicity). A cutoff of 5,000 SPD was selected to compare outcomes. RESULTS: Eighty patients were accrued over 3 years with 55% males and a median age of 59.5 years. Feasibility end point was met with 68 patients (85%) wearing FC more than predefined duration and majority (91%) finding its use acceptable. The mean SPD count for patients with PS 0 was 6,313, and for those with PS 1, it was 2,925 (122 and 54 active minutes, respectively) (P = .0003). Occurrence of toxicity of interest was lower among patients with SPD > 5,000 (7 of 33, 21%) compared with those with SPD < 5,000 (14 of 43, 32%), although not significant (P = .31). CONCLUSION: Assessment of physical activity with FC is feasible in patients with colorectal cancer and well-accepted. SPD may serve as an adjunct to PS assessment and a possible tool to help predict toxicities, regardless of type of therapy. Future studies incorporating FC can standardize patient assessment and help identify vulnerable population.
Asunto(s)
Neoplasias Colorrectales , Monitores de Ejercicio , Neoplasias Colorrectales/cirugía , Ejercicio Físico , Estudios de Factibilidad , Femenino , Humanos , Masculino , Persona de Mediana Edad , Complicaciones PosoperatoriasRESUMEN
Frequent mutations of genes in the histone-lysine N-methyltransferase 2 (KMT2) family members were identified in gastric cancers (GCs). Understanding how gene mutations of KMT2 family affect cancer progression and tumor immune microenvironment may provide new treatment strategies. A total of 1245 GCs were analyzed using next-generation sequencing, whole transcriptome sequencing, immunohistochemistry (Caris Life Sciences, Phoenix, AZ). The overall mutation rate of genes in the KMT2 family was 10.6%. Compared to KMT2-wild-type GCs, genes involved in epigenetic modification, receptor tyrosine kinases/MAPK/PI3K, and DNA damage repair (DDR) pathways had higher mutation rates in KMT2-mutant GCs (p < 0.05). Significantly higher rates of high tumor mutational burden, microsatellite instability-high/mismatch-repair deficiency (dMMR), and PD-L1 positivity were observed in KMT2-mutant GCs (p < 0.01), compared to KMT2-wild-type GCs. The association between PD-L1 positivity and KMT2 mutations remained significant in the proficient-MMR and microsatellite stable subgroup. Based on transcriptome data from the TCGA, cell cycle, metabolism, and interferon-α/ß response pathways were significantly upregulated in KMT2-mutant GCs than in KMT2-wild-type GCs. Patients with KMT2 mutation treated with immune checkpoint inhibitors had longer median overall survival compared to KMT2-wild-type patients with metastatic solid tumors (35 vs. 16 months, HR = 0.73, 95% CI: 0.62-0.87, p = 0.0003). In conclusion, this is the largest study to investigate the distinct molecular features between KMT2-mutant and KMT2-wild-type GCs to date. Our data indicate that GC patients with KMT2 mutations may benefit from ICIs and drugs targeting DDR, MAPK/PI3K, metabolism, and cell cycle pathways.