RESUMO
BACKGROUND & AIMS: Gastric cancer is often accompanied by a loss of mucin 6 (MUC6), but its pathogenic role in gastric carcinogenesis remains unclear. METHODS: Muc6 knockout (Muc6-/-) mice and Muc6-dsRED mice were newly generated. Tff1Cre, Golph3-/-, R26-Golgi-mCherry, Hes1flox/flox, Cosmcflox/flox, and A4gnt-/- mice were also used. Histology, DNA and RNA, proteins, and sugar chains were analyzed by whole-exon DNA sequence, RNA sequence, immunohistochemistry, lectin-binding assays, and liquid chromatography-mass spectrometry analysis. Gastric organoids and cell lines were used for in vitro assays and xenograft experiments. RESULTS: Deletion of Muc6 in mice spontaneously causes pan-gastritis and invasive gastric cancers. Muc6-deficient tumor growth was dependent on mitogen-activated protein kinase activation, mediated by Golgi stress-induced up-regulation of Golgi phosphoprotein 3. Glycomic profiling revealed aberrant expression of mannose-rich N-linked glycans in gastric tumors, detected with banana lectin in association with lack of MUC6 expression. We identified a precursor of clusterin as a binding partner of mannose glycans. Mitogen-activated protein kinase activation, Golgi stress responses, and aberrant mannose expression are found in separate Cosmc- and A4gnt-deficient mouse models that lack normal O-glycosylation. Banana lectin-drug conjugates proved an effective treatment for mannose-rich murine and human gastric cancer. CONCLUSIONS: We propose that Golgi stress responses and aberrant glycans are important drivers of and promising new therapeutic targets for gastric cancer.
Assuntos
Camundongos Knockout , Mucina-6 , Neoplasias Gástricas , Animais , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Neoplasias Gástricas/genética , Glicosilação , Humanos , Mucina-6/metabolismo , Mucina-6/genética , Camundongos , Linhagem Celular Tumoral , Carcinogênese/metabolismo , Carcinogênese/genética , Mucosa Gástrica/metabolismo , Mucosa Gástrica/patologia , Fator Trefoil-1/metabolismo , Fator Trefoil-1/genética , Organoides/metabolismo , Complexo de Golgi/metabolismo , Mucinas Gástricas/metabolismo , Modelos Animais de DoençasRESUMO
Colorectal cancer (CRC) is the second leading cause of cancer-related death worldwide. Cytokine-induced killer (CIK) cells are an adoptive immunotherapy reported to have strong anti-tumour activity across a range of cancers. They are a heterogeneous mix of lymphoid cells generated by culturing human peripheral blood mononuclear cells with cytokines and monoclonal antibodies in vitro. In this study, we investigated the yield and function of CIK cells generated from patients with CRC liver metastases. We first showed that CIK cells generated in serum free medium X-VIVO 15 were comparable to those from RPMI medium with 10% FBS in terms of the number and percentages of the main subsets of cells in the CIK culture, and the intracellular levels of granzyme B and perforin, and the pro-inflammatory cytokines IL-2, IFN-γ and TNF-α. The CIK cells were cytotoxic to CRC cell lines grown in 2D cultures or as spheroids, and against autologous patient-derived tumour organoids. Donor attributes such as age, sex, or prior chemotherapy exposure had no significant impact on CIK cell numbers or function. These results suggest that functional CIK cells can be generated from patients with CRC liver metastatic disease, and support further investigations into the therapeutic application of autologous CIK cells in the management of patients with CRC liver metastases.
Assuntos
Neoplasias Colorretais , Células Matadoras Induzidas por Citocinas , Neoplasias Hepáticas , Humanos , Neoplasias Hepáticas/terapia , Anticorpos Monoclonais , Citocinas , Neoplasias Colorretais/terapiaRESUMO
Bone morphogenetic protein (BMP) signaling is required for early forebrain development and cortical formation. How the endogenous modulators of BMP signaling regulate the structural and functional maturation of the developing brain remains unclear. Here, we show that expression of the BMP antagonist Grem1 marks committed layer V and VI glutamatergic neurons in the embryonic mouse brain. Lineage tracing of Grem1-expressing cells in the embryonic brain was examined by administration of tamoxifen to pregnant Grem1creERT; Rosa26LSLTdtomato mice at 13.5â days post coitum (dpc), followed by collection of embryos later in gestation. In addition, at 14.5â dpc, bulk mRNA-seq analysis of differentially expressed transcripts between FACS-sorted Grem1-positive and -negative cells was performed. We also generated Emx1-cre-mediated Grem1 conditional knockout mice (Emx1-Cre;Grem1flox/flox) in which the Grem1 gene was deleted specifically in the dorsal telencephalon. Grem1Emx1cKO animals had reduced cortical thickness, especially layers V and VI, and impaired motor balance and fear sensitivity compared with littermate controls. This study has revealed new roles for Grem1 in the structural and functional maturation of the developing cortex.
Assuntos
Proteína Morfogenética Óssea 1/antagonistas & inibidores , Encéfalo/fisiologia , Medo/fisiologia , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Neurônios Motores/metabolismo , Transdução de Sinais , Animais , Comportamento Animal , Proteína Morfogenética Óssea 1/genética , Encéfalo/embriologia , Diferenciação Celular , Proliferação de Células , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neurônios/fisiologia , Células-Tronco , TranscriptomaRESUMO
Colorectal cancer (CRC) is a common and deadly disease. Unfortunately, immune checkpoint inhibitors (ICIs) fail to elicit effective anti-tumour responses in the vast majority of CRC patients. Patients that are most likely to respond are those with DNA mismatch repair deficient (dMMR) and microsatellite instability (MSI) disease. However, reliable predictors of ICI response are lacking, even within the dMMR/MSI subtype. This, together with identification of novel mechanisms to increase response rates and prevent resistance, are ongoing and vitally important unmet needs. To address the current challenges with translation of early research findings into effective therapeutic strategies, this review summarises the present state of preclinical testing used to inform the development of immuno-regulatory treatment strategies for CRC. The shortfalls and advantages of commonly utilised mouse models of CRC, including chemically induced, transplant and transgenic approaches are highlighted. Appropriate use of existing models, incorporation of patient-derived data and development of cutting-edge models that recapitulate important features of human disease will be key to accelerating clinically relevant research in this area.
Assuntos
Neoplasias Encefálicas , Neoplasias Colorretais , Animais , Camundongos , Humanos , Pesquisa Translacional Biomédica , Oncologia , Instabilidade de Microssatélites , Reparo de Erro de Pareamento de DNARESUMO
BACKGROUND & AIMS: Cancer-associated fibroblasts (CAFs) play an important role in colorectal cancer (CRC) progression and predict poor prognosis in CRC patients. However, the cellular origins of CAFs remain unknown, making it challenging to therapeutically target these cells. Here, we aimed to identify the origins and contribution of colorectal CAFs associated with poor prognosis. METHODS: To elucidate CAF origins, we used a colitis-associated CRC mouse model in 5 different fate-mapping mouse lines with 5-bromodeoxyuridine dosing. RNA sequencing of fluorescence-activated cell sorting-purified CRC CAFs was performed to identify a potential therapeutic target in CAFs. To examine the prognostic significance of the stromal target, CRC patient RNA sequencing data and tissue microarray were used. CRC organoids were injected into the colons of knockout mice to assess the mechanism by which the stromal gene contributes to colorectal tumorigenesis. RESULTS: Our lineage-tracing studies revealed that in CRC, many ACTA2+ CAFs emerge through proliferation from intestinal pericryptal leptin receptor (Lepr)+ cells. These Lepr-lineage CAFs, in turn, express melanoma cell adhesion molecule (MCAM), a CRC stroma-specific marker that we identified with the use of RNA sequencing. High MCAM expression induced by transforming growth factor ß was inversely associated with patient survival in human CRC. In mice, stromal Mcam knockout attenuated orthotopically injected colorectal tumoroid growth and improved survival through decreased tumor-associated macrophage recruitment. Mechanistically, fibroblast MCAM interacted with interleukin-1 receptor 1 to augment nuclear factor κB-IL34/CCL8 signaling that promotes macrophage chemotaxis. CONCLUSIONS: In colorectal carcinogenesis, pericryptal Lepr-lineage cells proliferate to generate MCAM+ CAFs that shape the tumor-promoting immune microenvironment. Preventing the expansion/differentiation of Lepr-lineage CAFs or inhibiting MCAM activity could be effective therapeutic approaches for CRC.
Assuntos
Fibroblastos Associados a Câncer/patologia , Fibroblastos Associados a Câncer/fisiologia , Carcinogênese/patologia , Linhagem da Célula , Neoplasias Colorretais/patologia , Células-Tronco Mesenquimais/fisiologia , Actinas/genética , Actinas/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Antígeno CD146/genética , Antígeno CD146/metabolismo , Carcinogênese/genética , Carcinogênese/metabolismo , Diferenciação Celular , Proliferação de Células , Neoplasias Colorretais/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Mucosa Intestinal/patologia , Antígeno Ki-67/metabolismo , Masculino , Camundongos , Camundongos Transgênicos , Pessoa de Meia-Idade , Organoides/patologia , Organoides/fisiologia , Prognóstico , Receptores para Leptina/genética , Receptores para Leptina/metabolismo , Análise de Sequência de RNA , Taxa de Sobrevida , Microambiente TumoralRESUMO
BACKGROUND & AIMS: Cancer-associated fibroblasts (CAFs), key constituents of the tumor microenvironment, either promote or restrain tumor growth. Attempts to therapeutically target CAFs have been hampered by our incomplete understanding of these functionally heterogeneous cells. Key growth factors in the intestinal epithelial niche, bone morphogenetic proteins (BMPs), also play a critical role in colorectal cancer (CRC) progression. However, the crucial proteins regulating stromal BMP balance and the potential application of BMP signaling to manage CRC remain largely unexplored. METHODS: Using human CRC RNA expression data, we identified CAF-specific factors involved in BMP signaling, then verified and characterized their expression in the CRC stroma by in situ hybridization. CRC tumoroids and a mouse model of CRC hepatic metastasis were used to test approaches to modify BMP signaling and treat CRC. RESULTS: We identified Grem1 and Islr as CAF-specific genes involved in BMP signaling. Functionally, GREM1 and ISLR acted to inhibit and promote BMP signaling, respectively. Grem1 and Islr marked distinct fibroblast subpopulations and were differentially regulated by transforming growth factor ß and FOXL1, providing an underlying mechanism to explain fibroblast biological dichotomy. In patients with CRC, high GREM1 and ISLR expression levels were associated with poor and favorable survival, respectively. A GREM1-neutralizing antibody or fibroblast Islr overexpression reduced CRC tumoroid growth and promoted Lgr5+ intestinal stem cell differentiation. Finally, adeno-associated virus 8 (AAV8)-mediated delivery of Islr to hepatocytes increased BMP signaling and improved survival in our mouse model of hepatic metastasis. CONCLUSIONS: Stromal BMP signaling predicts and modifies CRC progression and survival, and it can be therapeutically targeted by novel AAV-directed gene delivery to the liver.
Assuntos
Proteínas Morfogenéticas Ósseas/metabolismo , Neoplasias Colorretais/patologia , Imunoglobulinas/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Neoplasias Hepáticas/secundário , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Fibroblastos Associados a Câncer/metabolismo , Carcinogênese/patologia , Diferenciação Celular , Linhagem Celular Tumoral , Neoplasias Colorretais/mortalidade , Progressão da Doença , Feminino , Hepatócitos/metabolismo , Humanos , Imunoglobulinas/genética , Estimativa de Kaplan-Meier , Masculino , Camundongos , Pessoa de Meia-Idade , Prognóstico , Transdução de Sinais , Microambiente Tumoral , Regulação para Cima , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Rectal Cancer is a common malignancy. The current treatment approach for patients with locally advanced rectal cancer involves neoadjuvant chemoradiotherapy followed by surgical resection of the rectum. The resection can lead to complications and long-term consequences. A clinical complete response is observed in some patients after chemoradiotherapy. A number of recent studies have shown that patients can be observed safely after completing chemoradiotherapy (without surgery), provided clinical complete response has been achieved. In this approach, resection is reserved for cases of regrowth. This is called the watch and wait approach. This approach potentially avoids unnecessary surgical resection of the rectum and the resulting complications. In this study, we will prospectively investigate this approach. METHODS: Adult patients with a diagnosis of rectal cancer planned to receive neoadjuvant long course chemoradiotherapy (± subsequent combination chemotherapy) will be consented into the study prior to commencing treatment. After completing the chemoradiotherapy (± subsequent combination chemotherapy), based on the clinical response, subjects will be allocated to one of the following arms: subjects who achieved a clinical complete response will be allocated to the watch and wait arm and others to the standard management arm (which includes resection). The aim of the study is to determine the rate of local failure and other safety and efficacy outcomes in the watch and wait arm. Patient reported outcome measures and the use of biomarkers as part of the clinical monitoring will be studied in both arms of the study. DISCUSSION: This study will prospectively investigate the safety of the watch and wait approach. We will investigate predictive biomarkers (molecular biomarkers and imaging biomarkers) and patient reported outcome measures in the study population and the cost effectiveness of the watch and wait approach. This study will also help evaluate a defined monitoring schedule for patients managed with the watch and wait approach. This protocol covers the first two years of follow up, we are planning a subsequent study which covers year 3-5 follow up for the study population. TRIAL REGISTRATION: Name of the registry: Australia and New Zealand Clinical Trials Registry (ANZCTR). TRIAL REGISTRATION NUMBER: Trial ID: ACTRN12619000207112 Registered 13 February 2019, https://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id=376810.
Assuntos
Terapia Neoadjuvante , Neoplasias Retais/terapia , Conduta Expectante/métodos , Adulto , Biomarcadores Tumorais/análise , Feminino , Humanos , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Neoplasias Retais/mortalidade , Taxa de Sobrevida , Resultado do TratamentoRESUMO
The stem/progenitor cells of the developing intestine are biologically distinct from their adult counterparts. Here, we examine the microenvironmental cues that regulate the embryonic stem/progenitor population, focusing on the role of Notch pathway factor delta-like protein-1 (DLK1). mRNA-seq analyses of intestinal mesenchymal cells (IMCs) collected from embryonic day 14.5 (E14.5) or adult IMCs and a novel coculture system with E14.5 intestinal epithelial organoids were used. Following addition of recombinant DLK1 (rDLK) or Dlk1 siRNA (siDlk1), epithelial characteristics were compared using imaging, replating efficiency assays, qPCR, and immunocytochemistry. The intestinal phenotypes of littermate Dlk1+/+ and Dlk1-/- mice were compared using immunohistochemistry. Using transcriptomic analyses, we identified morphogens derived from the embryonic mesenchyme that potentially regulate the developing epithelial cells, to focus on Notch family candidate DLK1. Immunohistochemistry indicated that DLK1 was expressed exclusively in the intestinal stroma at E14.5 at the top of emerging villi, decreased after birth, and shifted to the intestinal epithelium in adulthood. In coculture experiments, addition of rDLK1 to adult IMCs inhibited organoid differentiation, whereas Dlk1 knockdown in embryonic IMCs increased epithelial differentiation to secretory lineage cells. Dlk1-/- mice had restricted Ki67+ cells in the villi base and increased secretory lineage cells compared with Dlk1+/+ embryos. Mesenchyme-derived DLK1 plays an important role in the promotion of epithelial stem/precursor expansion and prevention of differentiation to secretory lineages in the developing intestine.NEW & NOTEWORTHY Using a novel coculture system, transcriptomics, and transgenic mice, we investigated differential molecular signaling between the intestinal epithelium and mesenchyme during development and in the adult. We show that the Notch pathway factor delta-like protein-1 (DLK1) is stromally produced during development and uncover a new role for DLK1 in the regulation of intestinal epithelial stem/precursor expansion and differentiation to secretory lineages.
Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Comunicação Celular , Diferenciação Celular , Proliferação de Células , Células-Tronco Embrionárias/enzimologia , Células Epiteliais/enzimologia , Mucosa Intestinal/enzimologia , Células Estromais/enzimologia , Animais , Proteínas de Ligação ao Cálcio/deficiência , Proteínas de Ligação ao Cálcio/genética , Linhagem da Célula , Células Cultivadas , Técnicas de Cocultura , Regulação da Expressão Gênica no Desenvolvimento , Mucosa Intestinal/embriologia , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Organoides , Via Secretória , Transdução de Sinais , Nicho de Células-Tronco , TranscriptomaRESUMO
Osteoarthritis (OA) is a disease of an entire synovial joint characterized by clinical symptoms and distortion of joint tissues, including cartilage, muscles, ligaments, and bone. Although OA is a disease of all joint tissues, it is a defined accessible compartment and is thus amenable to topical surgical and regenerative therapies, including stem cells. All tissues arise from stem progenitor cells, and the relative capacity of different cellular compartments, and different individuals, to renew tissues into adulthood may be important in the onset of many different degenerative diseases. OA is driven by both mechanical and inflammatory factors, but how these factors affect the proliferation and differentiation of cells into cartilage in vivo is largely unknown. Indeed, our very basic understanding of the physiological cellular kinetics and biology of the stem-progenitor cell unit of the articular cartilage, and how this is influenced by mechano-inflammatory injury, is largely unknown. OA seems, rather deceptively, to be the low-hanging fruit for stem cell therapy. Without the basic understanding of the stem cell and progenitor unit that generate and maintain articular cartilage in vivo, we will continue to waste opportunities to both prevent and manage this disease. In this review, we discuss the biology of chondrogenesis, the stem cell populations that support articular cartilage in health and disease, and future opportunities afforded through the translation of basic articular chondrocyte stem cell biology into new clinical therapies.
Assuntos
Transplante de Células-Tronco Mesenquimais/métodos , Células-Tronco Mesenquimais/metabolismo , Osteoartrite/terapia , Medicina Regenerativa/métodos , HumanosRESUMO
BACKGROUND: In an effort to foster patient engagement, some healthcare systems provide their patients with open notes, enabling them to access their clinical notes online. In January 2013, the Veterans Health Administration (VA) implemented online access to clinical notes ("VA Notes") through the Blue Button feature of its patient portal. OBJECTIVE: To measure the association of online patient access to clinical notes with changes in healthcare utilization and clinician documentation behaviors. DESIGN: A retrospective cohort study. PATIENTS: Patients accessing My HealtheVet (MHV), the VA's online patient portal, between July 2011 and January 2015. MAIN MEASURES: Use of healthcare services (primary care clinic visits and online electronic secure messaging), and characteristics of physician clinical documentation (readability of notes). KEY RESULTS: Among 882,575 unique portal users, those who accessed clinical notes (16.2%; N = 122,972) were younger, more racially homogenous (white), and less likely to be financially vulnerable. Compared with non-users, Notes users more frequently used the secure messaging feature on the portal (mean of 2.6 messages (SD 7.0) v. 0.87 messages (SD 3.3) in January-July 2013), but their higher use of secure messaging began prior to VA Notes implementation, and thus was not temporally related to the implementation. When comparing clinic visit rates pre- and post-implementation, Notes users had a small but significant increase in rate of 0.36 primary care clinic visits (2012 v. 2013) compared to portal users who did not view their Notes (p = 0.01). At baseline, the mean reading ease of primary care clinical notes was 53.8 (SD 10.1) and did not improve after implementation of VA Notes. CONCLUSIONS: VA Notes users were different than patients with portal access who did not view their notes online, and they had higher rates of healthcare service use prior to and after VA Notes implementation. Opportunities exist to improve clinical note access and readability.
Assuntos
Registros Eletrônicos de Saúde , Portais do Paciente , Documentação , Humanos , Aceitação pelo Paciente de Cuidados de Saúde , Estudos RetrospectivosRESUMO
OBJECTIVE: Pathological Wnt pathway activation is a conserved hallmark of colorectal cancer. Wnt-activating mutations can be divided into: i) ligand-independent (LI) alterations in intracellular signal transduction proteins (Adenomatous polyposis coli, ß-catenin), causing constitutive pathway activation and ii) ligand-dependent (LD) mutations affecting the synergistic R-Spondin axis (RNF43, RSPO-fusions) acting through amplification of endogenous Wnt signal transmembrane transduction. Our aim was to exploit differential Wnt target gene expression to generate a mutation-agnostic biomarker for LD tumours. DESIGN: We undertook harmonised multi-omic analysis of discovery (n=684) and validation cohorts (n=578) of colorectal tumours collated from publicly available data and the Stratification in Colorectal Cancer Consortium. We used mutation data to establish molecular ground truth and subdivide lesions into LI/LD tumour subsets. We contrasted transcriptional, methylation, morphological and clinical characteristics between groups. RESULTS: Wnt disrupting mutations were mutually exclusive. Desmoplastic stromal upregulation of RSPO may compensate for absence of epithelial mutation in a subset of stromal-rich tumours. Key Wnt negative regulator genes were differentially expressed between LD/LI tumours, with targeted hypermethylation of some genes (AXIN2, NKD1) occurring even in CIMP-negative LD cancers. AXIN2 mRNA expression was used as a discriminatory molecular biomarker to distinguish LD/LI tumours (area under the curve >0.93). CONCLUSIONS: Epigenetic suppression of appropriate Wnt negative feedback loops is selectively advantageous in LD tumours and differential AXIN2 expression in LD/LI lesions can be exploited as a molecular biomarker. Distinguishing between LD/LI tumour types is important; patients with LD tumours retain sensitivity to Wnt ligand inhibition and may be stratified at diagnosis to clinical trials of Porcupine inhibitors.
Assuntos
Neoplasias Colorretais/diagnóstico , Transdução de Sinais/genética , Proteína Wnt1/metabolismo , Idoso , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Marcadores Genéticos/genética , Humanos , Masculino , Pessoa de Meia-Idade , Proteína Wnt1/genéticaRESUMO
SARS-CoV-2, the virus that causes coronavirus disease 2019 (COVID-19), has spread rapidly around the world since it was first recognized in late 2019. Most early reports of person-to-person SARS-CoV-2 transmission have been among household contacts, where the secondary attack rate has been estimated to exceed 10% (1), in health care facilities (2), and in congregate settings (3). However, widespread community transmission, as is currently being observed in the United States, requires more expansive transmission events between nonhousehold contacts. In February and March 2020, the Chicago Department of Public Health (CDPH) investigated a large, multifamily cluster of COVID-19. Patients with confirmed COVID-19 and their close contacts were interviewed to better understand nonhousehold, community transmission of SARS-CoV-2. This report describes the cluster of 16 cases of confirmed or probable COVID-19, including three deaths, likely resulting from transmission of SARS-CoV-2 at two family gatherings (a funeral and a birthday party). These data support current CDC social distancing recommendations intended to reduce SARS-CoV-2 transmission. U.S residents should follow stay-at-home orders when required by state or local authorities.
Assuntos
Betacoronavirus/isolamento & purificação , Infecções Comunitárias Adquiridas/transmissão , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/transmissão , Pneumonia Viral/diagnóstico , Pneumonia Viral/transmissão , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , COVID-19 , Chicago/epidemiologia , Criança , Pré-Escolar , Análise por Conglomerados , Infecções Comunitárias Adquiridas/epidemiologia , Infecções Comunitárias Adquiridas/mortalidade , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/mortalidade , Família , Humanos , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/mortalidade , SARS-CoV-2 , Adulto JovemRESUMO
OBJECTIVE: Serrated colorectal cancer (CRC) accounts for approximately 25% of cases and includes tumours that are among the most treatment resistant and with worst outcomes. This CRC subtype is associated with activating mutations in the mitogen-activated kinase pathway gene, BRAF, and epigenetic modifications termed the CpG Island Methylator Phenotype, leading to epigenetic silencing of key tumour suppressor genes. It is still not clear which (epi-)genetic changes are most important in neoplastic progression and we begin to address this knowledge gap herein. DESIGN: We use organoid culture combined with CRISPR/Cas9 genome engineering to sequentially introduce genetic alterations associated with serrated CRC and which regulate the stem cell niche, senescence and DNA mismatch repair. RESULTS: Targeted biallelic gene alterations were verified by DNA sequencing. Organoid growth in the absence of niche factors was assessed, as well as analysis of downstream molecular pathway activity. Orthotopic engraftment of complex organoid lines, but not BrafV600E alone, quickly generated adenocarcinoma in vivo with serrated features consistent with human disease. Loss of the essential DNA mismatch repair enzyme, Mlh1, led to microsatellite instability. Sphingolipid metabolism genes are differentially regulated in both our mouse models of serrated CRC and human CRC, with key members of this pathway having prognostic significance in the human setting. CONCLUSION: We generate rapid, complex models of serrated CRC to determine the contribution of specific genetic alterations to carcinogenesis. Analysis of our models alongside patient data has led to the identification of a potential susceptibility for this tumour type.
Assuntos
Adenocarcinoma/genética , Adenocarcinoma/patologia , Colo/patologia , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Organoides/patologia , Proteínas Proto-Oncogênicas B-raf/genética , Adenocarcinoma/metabolismo , Alelos , Colo/metabolismo , Neoplasias Colorretais/metabolismo , Ilhas de CpG/genética , Reparo de Erro de Pareamento de DNA , Análise Mutacional de DNA , Progressão da Doença , Epigenômica , Regulação Neoplásica da Expressão Gênica , Genes Supressores de Tumor , Humanos , Modelos Genéticos , Mutação , Organoides/metabolismo , Fenótipo , Proteínas Proto-Oncogênicas B-raf/metabolismoRESUMO
Cancer-associated fibroblasts (CAFs) were originally presumed to represent a homogeneous population uniformly driving tumorigenesis, united by their morphology and peritumoural location. Our understanding of CAFs has since been shaped by sophisticated in vitro and in vivo experiments, pathological association and, more recently, ablation, and it is now widely appreciated that CAFs form a group of highly heterogeneous cells with no single overarching marker. Studies have demonstrated that the CAF population contains different subtypes based on the expression of marker proteins with the capacity to promote or inhibit cancer, with their biological role as accomplices or adversaries dependent on many factors, including the cancer stage. So, while CAFs have been endlessly shown to promote the growth, survival and spread of tumours via improvements in functionality and an altered secretome, they are also capable of retarding tumorigenesis via largely unknown mechanisms. It is important to reconcile these disparate results so that the functions of, or factors produced by, tumour-promoting subtypes can be specifically targeted to improve cancer patient outcomes. This review will dissect out CAF complexity and CAF-directed cancer treatment strategies in order to provide a case for future, rational therapies.
Assuntos
Fibroblastos Associados a Câncer/fisiologia , Neoplasias/tratamento farmacológico , Animais , Fibroblastos Associados a Câncer/efeitos dos fármacos , Metabolismo Energético , Matriz Extracelular/fisiologia , Humanos , Camundongos , Invasividade Neoplásica , Neoplasias/metabolismo , Neoplasias/patologia , Fenótipo , PrognósticoRESUMO
Gastric adenocarcinoma and proximal polyposis of the stomach (GAPPS) is an autosomal-dominant cancer-predisposition syndrome with a significant risk of gastric, but not colorectal, adenocarcinoma. We mapped the gene to 5q22 and found loss of the wild-type allele on 5q in fundic gland polyps from affected individuals. Whole-exome and -genome sequencing failed to find causal mutations but, through Sanger sequencing, we identified point mutations in APC promoter 1B that co-segregated with disease in all six families. The mutations reduced binding of the YY1 transcription factor and impaired activity of the APC promoter 1B in luciferase assays. Analysis of blood and saliva from carriers showed allelic imbalance of APC, suggesting that these mutations lead to decreased allele-specific expression in vivo. Similar mutations in APC promoter 1B occur in rare families with familial adenomatous polyposis (FAP). Promoter 1A is methylated in GAPPS and sporadic FGPs and in normal stomach, which suggests that 1B transcripts are more important than 1A in gastric mucosa. This might explain why all known GAPPS-affected families carry promoter 1B point mutations but only rare FAP-affected families carry similar mutations, the colonic cells usually being protected by the expression of the 1A isoform. Gastric polyposis and cancer have been previously described in some FAP-affected individuals with large deletions around promoter 1B. Our finding that GAPPS is caused by point mutations in the same promoter suggests that families with mutations affecting the promoter 1B are at risk of gastric adenocarcinoma, regardless of whether or not colorectal polyps are present.
Assuntos
Adenocarcinoma/genética , Proteína da Polipose Adenomatosa do Colo/genética , Polipose Adenomatosa do Colo/genética , Pólipos Adenomatosos/genética , Éxons/genética , Mutação Puntual/genética , Neoplasias Gástricas/genética , Desequilíbrio Alélico/genética , Variações do Número de Cópias de DNA/genética , Exoma/genética , Feminino , Mucosa Gástrica/metabolismo , Ligação Genética/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Perda de Heterozigosidade , Masculino , Linhagem , Regiões Promotoras Genéticas/genéticaRESUMO
BACKGROUND: The OpenNotes initiative provides patients online access to their clinical notes. Mental health clinicians in the Veterans Health Administration report a need for guidance on how to provide care, write notes, and discuss them in the context of OpenNotes. AIM: To provide mental health clinicians recommendations identified by patients and clinicians that help them effectively practice in the context of OpenNotes. METHOD: Twenty-eight mental health clinicians and 28 patients in mental health care participated in semi-structured interviews about their experiences and perceptions with OpenNotes. A rapid review approach was used to analyze transcripts. RESULTS: Analysis of interviews identified three domains of advice for mental health clinicians: writing notes that maintain the therapeutic relationship, communicating with patients about their notes and utilizing clinical notes as a patient resource to enhance care. Specific recommendations are provided. CONCLUSION: Findings provide mental health clinicians with guidance from service users and clinicians on how to leverage clinical notes to maintain - and potentially enhance -therapeutic relationships in a healthcare system in which patients are able to read their mental health notes online.
Assuntos
Registros Eletrônicos de Saúde/normas , Serviços de Saúde Mental/normas , Relações Médico-Paciente , Adulto , Feminino , Comunicação em Saúde , Humanos , Masculino , Pessoa de Meia-Idade , Software , VeteranosRESUMO
So far, two genes associated with familial melanoma have been identified, accounting for a minority of genetic risk in families. Mutations in CDKN2A account for approximately 40% of familial cases, and predisposing mutations in CDK4 have been reported in a very small number of melanoma kindreds. Here we report the whole-genome sequencing of probands from several melanoma families, which we performed in order to identify other genes associated with familial melanoma. We identify one individual carrying a novel germline variant (coding DNA sequence c.G1075A; protein sequence p.E318K; rs149617956) in the melanoma-lineage-specific oncogene microphthalmia-associated transcription factor (MITF). Although the variant co-segregated with melanoma in some but not all cases in the family, linkage analysis of 31 families subsequently identified to carry the variant generated a log of odds (lod) score of 2.7 under a dominant model, indicating E318K as a possible intermediate risk variant. Consistent with this, the E318K variant was significantly associated with melanoma in a large Australian case-control sample. Likewise, it was similarly associated in an independent case-control sample from the United Kingdom. In the Australian sample, the variant allele was significantly over-represented in cases with a family history of melanoma, multiple primary melanomas, or both. The variant allele was also associated with increased naevus count and non-blue eye colour. Functional analysis of E318K showed that MITF encoded by the variant allele had impaired sumoylation and differentially regulated several MITF targets. These data indicate that MITF is a melanoma-predisposition gene and highlight the utility of whole-genome sequencing to identify novel rare variants associated with disease susceptibility.
Assuntos
Predisposição Genética para Doença , Melanoma/genética , Fator de Transcrição Associado à Microftalmia/genética , Mutação , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Sumoilação/genética , Adulto JovemRESUMO
BACKGROUND: As electronic health records and computerized workflows expand, there are unprecedented opportunities to digitally connect with patients using secure portals. To realize the value of patient portals, initial reach across populations will need to be demonstrated, as well as sustained usage over time. OBJECTIVE: The study aim was to identify patient factors associated with short-term and long-term portal usage after patients registered to access all portal functions. METHODS: We prospectively followed a cohort of patients at a large Department of Veterans Affairs (VA) health care facility who recently completed identity proofing to use the VA patient portal. Information collected at baseline encompassed patient factors potentially associated with portal usage, including: demographics, Internet access and use, health literacy, patient activation, and self-reported health conditions. The primary outcome was the frequency of portal log-ins during 6-month and 18-month time intervals after study enrollment. RESULTS: A total of 270 study participants were followed prospectively. Almost all participants (260/268, 97.0%) reported going online, typically at home (248/268, 92.5%). At 6 months, 84.1% (227/270) of participants had visited the portal, with some variation in usage across demographic and health-related subgroups. There were no significant differences in portal log-ins by age, gender, education, marital status, race/ethnicity, distance to a VA facility, or patient activation measure. Significantly higher portal usage was seen among participants using high-speed broadband at home, greater self-reported ability using the Internet, and routinely going online. By 18 months, 91% participants had logged in to the portal, and no significant associations were found between usage and demographics, health status, or patient activation. When examining portal activity between 6 and 18 months, patients who were infrequent or high portal users remained in those categories, respectively. CONCLUSIONS: Short-term and long-term portal usage was associated with having broadband at home, high self-rated ability when using the Internet, and overall online behavior. Digital inclusion, or ready access to the Internet and digital skills, appears to be a social determinant in patient exposure to portal services.
Assuntos
Educação a Distância/métodos , Registros Eletrônicos de Saúde/estatística & dados numéricos , Participação do Paciente/métodos , Portais do Paciente/estatística & dados numéricos , Estudos de Coortes , Feminino , Humanos , Internet/estatística & dados numéricos , Masculino , Pessoa de Meia-Idade , Estudos ProspectivosRESUMO
Rubinstein-Taybi syndrome (RTS) and Cornelia de Lange syndrome (CdLS) are genetically heterogeneous multiple anomalies syndromes, each having a distinctive facial gestalt. Two genes (CREBBP and EP300) are known to cause RTS, and five (NIPBL, SMC1A, SMC3, RAD21, and HDAC8) have been associated with CdLS. A diagnosis of RTS or CdLS is molecularly confirmed in only 65% of clinically identified cases, suggesting that additional causative genes exist for both conditions. In addition, although EP300 and CREBBP encode homologous proteins and perform similar functions, only eight EP300 positive RTS patients have been reported, suggesting that patients with EP300 mutations might be escaping clinical recognition. We report on a child with multiple congenital abnormalities and intellectual disability whose facial features and complex phenotype resemble CdLS. However, no mutations in CdLS-related genes were identified. Rather, a novel EP300 mutation was found on whole exome sequencing. Possible links between EP300 and genes causing CdLS are evident in the literature. Both EP300 and HDAC8 are involved in the regulation of TP53 transcriptional activity. In addition, p300 and other chromatin associated proteins, including NIPBL, SMCA1, and SMC3, have been found at enhancer regions in different cell types. It is therefore possible that EP300 and CdLS-related genes are involved in additional shared pathways, producing overlapping phenotypes. As whole exome sequencing becomes more widely utilized, the diverse phenotypes associated with EP300 mutations should be better understood. In the meantime, testing for EP300 mutations in those with features of CdLS may be warranted.
Assuntos
Síndrome de Cornélia de Lange/diagnóstico , Síndrome de Cornélia de Lange/genética , Proteína p300 Associada a E1A/genética , Exoma , Mutação da Fase de Leitura , Fenótipo , Autopsia , Diagnóstico Diferencial , Fácies , Evolução Fatal , Heterozigoto , Humanos , Lactente , Masculino , Síndrome de Rubinstein-Taybi/diagnóstico , Síndrome de Rubinstein-Taybi/genéticaRESUMO
Medical extended reality (MXR) has emerged as a dynamic field at the intersection of health care and immersive technology, encompassing virtual, augmented, and mixed reality applications across a wide range of medical disciplines. Despite its rapid growth and recognition by regulatory bodies, the field lacks a standardized taxonomy to categorize its diverse research and applications. This American Medical Extended Reality Association guideline, authored by the editorial board of the Journal of Medical Extended Reality, introduces a comprehensive taxonomy for MXR, developed through a multidisciplinary and international collaboration of experts. The guideline seeks to standardize terminology, categorize existing work, and provide a structured framework for future research and development in MXR. An international and multidisciplinary panel of experts was convened, selected based on publication track record, contributions to MXR, and other objective measures. Through an iterative process, the panel identified primary and secondary topics in MXR. These topics were refined over several rounds of review, leading to the final taxonomy. The taxonomy comprises 13 primary topics that jointly expand into 180 secondary topics, demonstrating the field's breadth and depth. At the core of the taxonomy are five overarching domains: (1) technological integration and innovation; (2) design, development, and deployment; (3) clinical and therapeutic applications; (4) education, training, and communication; and (5) ethical, regulatory, and socioeconomic considerations. The developed taxonomy offers a framework for categorizing the diverse research and applications within MXR. It may serve as a foundational tool for researchers, clinicians, funders, academic publishers, and regulators, facilitating clearer communication and categorization in this rapidly evolving field. As MXR continues to grow, this taxonomy will be instrumental in guiding its development and ensuring a cohesive understanding of its multifaceted nature.