Abstract
Exosomes mediate intercellular communication and regulation in many processes, including cancer-immune cell interactions. Exosomes are small (~100 nm) membrane vesicles that arise from the ...late endosomal pathway and contain bioactive molecules that are transferred to and affect the function of target cells. Tumor-derived exosomes are important regulators of anti-tumor immunity and have been shown to exert stimulatory and inhibitory effects on immune cells, depending on the experimental setting. Using a proteomic approach, we found that exosomes derived from tumors that undergo immune rejection (“regressors”) contain different proteins than exosomes from progressively growing tumors (“progressors”). Specifically, the signaling molecule MAP2K1 (“MEK1”) is highly enriched in regressor compared to progressor exosomes, leading us to hypothesize that exosomal MEK1 stimulates anti-tumor immunity and inhibits tumor progression. We found that forced expression of MEK1 in progressor exosomes delays tumor growth and increases anti-tumor immune cell infiltration, and that MEK1-hi exosomes are sufficient to delay tumor growth, but MEK1-low exosomes are not. Moreover, we found that MEK1 protein can be physically transferred from tumor cells to macrophages via exosomes, and that MEK1-hi exosomes induce pro-inflammatory cytokine production in macrophages, but MEK1-low exosomes do not. We propose that tumor-derived MEK1 is transferred to macrophages via exosomes, which enhances their anti-tumoricidal effector function and delays tumor growth. This cell extrinsic, anti-tumor activity of MEK1 contrasts with the known oncogenic properties of MEK1 and could limit the efficacy of MEK1 inhibitors currently in clinical trials.
Abstract
Among pediatric cancers, sarcomas, especially those with large tumor burdens and metastatic disease, often result in poor outcome. Thus, new treatments are urgently needed to inhibit tumor ...progression, prevent metastasis, and improve overall survival.
To understand the mechanisms driving sarcoma progression, we employed two mouse fibrosarcoma cell lines that display different growth phenotypes when transplanted into syngeneic immune competent mice. Progressor fibrosarcomas evade detection by the immune system and develop large tumor burdens, while regressor fibrosarcomas regress shortly after a period of limited tumor growth. This difference in the growth phenotype is mediated in part by immune cells, but the mechanisms by which progressor and regressor cells influence immune cell activity are not fully elucidated.
Our research has focused on exosomes, 50-100 nm secreted nanovesicles, that contain bioactive cargoes and have emerged as mediators of intercellular communication between various cells. Here, we hypothesize that fibrosarcoma exosomes determine the aggressiveness of the disease by either educating tumor cells themselves or altering interactions with host cells.
To investigate the role of fibrosarcoma exosomes, we first educated regressor tumor cells with progressor exosomes for 3 weeks in vitro, and vice versa. Upon in vivo inoculation, progressor cells conditioned with regressor cell-derived exosomes grew significantly slower compared to untreated progressor cells. In contrast, regressor cells educated with progressor exosomes did not regress and overgrew significantly compared to untreated regressor cells. These results indicate that exosomes have an ability to reprogram a tumor phenotype.
To determine if fibrosarcoma exosomes specifically impact the host, we evaluated whether conditioning naïve mice with progressor exosomes alters in vivo growth of regressor tumor cells injected after three weeks of education. Twenty percent of regressor tumors continued to grow following progressor exosome education, while all tumors regressed in the control PBS group. These data suggest that exosomes also play an important role in the interaction with the host cells influencing tumor growth and disease progression.
Next, to determine which exosomal cargo may mediate these effects, we sought to characterize the proteome of exosomes from progressor and regressor cells using mass spectrometry. We found basigin, also known as CD147, a transmembrane embryonic glycoprotein reported to regulate cell proliferation, tumor migration and metastasis as one of the candidate proteins highly expressed in progressor-derived exosomes.
We conclude that exosomes derived from fibrosarcoma progresssors and regressors can alter the tumor phenotype both directly and indirectly. Exosomal cargo, such as basigin, may mediate the aggressive behavior of tumor cells, making it a potential therapeutic target to inhibit fibrosarcoma progression.
Citation Format: Miho Nakajima, Stephen C. Searles, Ayuko Hoshino, Katherine M. Offer, Candia M. Kenific, Jack D. Bui, David C. Lyden. The role of exosomes in fibrosarcoma progression abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4805. doi:10.1158/1538-7445.AM2017-4805
Increased levels of the transcription factor hypoxia inducible factor (HIF)-1 occur only in hypoxic tissue. The authors propose a therapeutic strategy that relies on HIF-1, the enhancer hypoxia ...response element (HRE), and the delivery vector adeno-associated virus-2 (AAV2) to direct ischemia specific gene therapy to skin.
An expression cassette containing the CMV promoter driving the reporter gene green fluorescent protein (GFP) was used to assess cutaneous tropism of AAV2. Transfection of dermal fibroblasts and immortalized keratinocytes (HaCat) was assessed with flow cytometry. Human embryonic kidney 293 (HEK) cells were used to produce vector stocks and test the authors' therapeutic strategy in quadruplicate. An expression cassette with nine repeats of HRE linked to beta-galactosidase (LacZ) within the AAV2 vector was constructed. HEK cells were transfected and exposed to normoxic (21% oxygen) and hypoxic (1% oxygen) conditions. LacZ activity was measured by conversion of galactoside red-beta-D-galactopyranoside.
Approximately 50 percent of dermal fibroblasts and HaCat cells were transfected when treated with 1 x 10(4) genome copies/cell of AAV2-CMV-GFP. Using the same titration of AAV2-9HRE-LacZ, transfected HEK cells demonstrated LacZ activity of 0.496 +/- 0.068 U/microg in normoxia and 2.9 +/- 0.58 U/microg in hypoxia. Transfected cells exposed to 24 hours of hypoxia show greater than an 11-fold increase in LacZ activity (p < 0.05) compared with baseline normoxic controls.
The authors' results confirm that AAV2 has in vitro tropism for skin-derived cell lines. Furthermore, HRE will drive gene expression in ischemia but not normoxia. This is the first step toward the authors' goal of HIF-1-regulated gene therapy to prevent ischemia related skin injury.
Pooled analysis of individual patient data from stroke trials can deliver more precise estimates of treatment effect, enhance power to examine prespecified subgroups, and facilitate exploration of ...treatment-modifying influences. Analysis plans should be declared, and preferably published, before trial results are known. For pooling trials that used diverse analytic approaches, an ordinal analysis is favored, with justification for considering deaths and severe disability jointly. Because trial pooling is an incremental process, analyses should follow a sequential approach, with statistical adjustment for iterations. Updated analyses should be published when revised conclusions have a clinical implication. However, caution is recommended in declaring pooled findings that may prejudice ongoing trials, unless clinical implications are compelling. All contributing trial teams should contribute to leadership, data verification, and authorship of pooled analyses. Development work is needed to enable reliable inferences to be drawn about individual drug or device effects that contribute to a pooled analysis, versus a class effect, if the treatment strategy combines ≥2 such drugs or devices. Despite the practical challenges, pooled analyses are powerful and essential tools in interpreting clinical trial findings and advancing clinical care.
Abstract
Introduction: Pancreatic cancer (PC) is a highly lethal malignancy, largely due to its propensity for early dissemination and protracted subclinical course until it reaches advanced stages. ...Available diagnostic modalities are often unable to diagnose early events in the metastatic process, including pre-metastatic niches, micrometastases, or small macrometastases. We have previously shown that quantitative image features (QIFs) of the liver in pre-operative contrast-enhanced CT (CECT) scans can identify PC patients at risk of impending liver metastasis (PMID 35366706). Further, we found that perioperative liver biopsy in patients with untreated localized PC can capture cellular and molecular features of evolving pre-metastatic niches, which can predict timing and patterns of PC recurrence (doi.org/10.1158/1538-7445.PANCA22-PR012). In the present study, we sought to examine whether liver QIFs can capture specific biologic processes at the cellular level, which could in turn inform non-invasively on the status of the liver milieu that determine metastatic risk and potentially reveal opportunities for therapeutic intervention. Methods: Intraoperative liver biopsies were obtained from patients undergoing pancreatectomy for resectable PC (n=29) or non-cancerous pancreatic lesions (n=7) and were analyzed histologically for fibrosis, steatosis, portal and lobular inflammation. Immunostaining for immune cell populations and extracellular matrix proteins were performed. Liver biopsy histopathological features were then compared to 255 QIFs. Univariate analysis (UVA) was performed with logistic/linear regression, after stratification for biliary obstruction and presence of PC. For continuous variables, multivariate analysis (MVA) was performed using generalized linear model. For categorical variables, MVA was performed with Naïve-bayes classifier and ‘Leave-one image out’ method of cross validation after selecting the features with MRMR algorithm. Results: On UVA, multiple features correlated significantly with liver fibrosis, steatosis, and portal inflammation, as well fibronectin and collagen IV immunostaining. Further, significant correlations were found with CD11b+ cell density (cells/mm2), IBA1+ area, CD8+ staining intensity, CD3+ and CD8+ lobular infiltration, and total Ki67+ cell density. No significant associations were found with lobular inflammation, CD3+ intensity, or CD68+ cell density. On MVA, several QIFs remained significant predictors of the aforementioned 11 histopathologic variables. The best prediction was achieved for liver fibrosis (QIFs: RLM9, LBP56, LBP118, LBP3, LBP9, and LBP102; AUC 0.78) and steatosis (FD1_47; AUC 0.79). Conclusion: CECT liver texture analysis may predict biological processes affecting differential contrast perfusion, tissue distribution and parenchymal retention, enabling determination of liver histopathology non-invasively. Therefore, this approach may hold promise in identifying pre-metastatic niches or micrometastatic disease in evolution and risk-stratifying PC patients in a non-invasive fashion.
Citation Format: Constantinos P. Zambririnis, Linda Bojmar, Jonathan Hernandez, Gokce Askan, Jian Zheng, Ahmad B. Barekzai, Natally Horvat, Vinod P. Balachandran, Jeffrey A. Drebin, Peter Kingham, Michael I. D’Angelica, Olca Basturk, Mithat Gönen, Alice Wei, Robert Schwartz, David C. Lyden, William R. Jarnagin, Jayasree Chakraborty. Radiomics of pre-operative CT scans capture biologic processes within the liver in patients undergoing pancreatectomy abstract. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr C071.
The concept of the tumour microenvironment recognizes that the interplay between cancer cells and stromal cells is a crucial determinant of cancer growth. In this Perspectives article, we propose the ...novel concept that the tumour microenvironment is built through rate-limiting steps during multistage carcinogenesis. Construction of a 'precancer niche' is a necessary and early step that is required for initiated cells to survive and evolve; subsequent niche expansion and maturation accompany tumour promotion and progression, respectively. As such, cancer niches represent an emergent property of a tumour that could be a robust target for cancer prevention and therapy.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Skin is an ideal gene therapy target because it is readily accessible and is involved in many pathologic processes. Viruses are the most common gene vectors, however, few comparative studies exist ...examining their efficacy in skin. This study evaluates adenovirus serotype 5, adeno-associated virus type 2 and 5, MMLV-derived retrovirus, and human immunodeficiency virus-1 derived lentivirus for gene vector activity in human dermal fibroblasts and other skin cell lines. Human immunodeficiency virus-1-based lentiviral vector resulted in over 90% transduction in all cell lines tested. Transduced cells maintained reporter expression over several passages after a single exposure. In contrast, gene activity fell rapidly over cell divisions with adenoviral and adeno-associated vectors. Therefore, lentiviral vectors are the delivery mechanism of choice for long-term therapeutic gene expression in dermal fibroblasts and other skin cell lines, whereas adenoviral or adeno-associated vectors may be preferred for short-term therapy.
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