Here we describe isolation and characterization of macrophage-tumor cell fusions (MTFs) from the blood of pancreatic ductal adenocarcinoma (PDAC) patients. The MTFs were generally aneuploidy, and ...immunophenotypic characterizations showed that the MTFs express markers characteristic of PDAC and stem cells, as well as M2-polarized macrophages. Single cell RNASeq analyses showed that the MTFs express many transcripts implicated in cancer progression, LINE1 retrotransposons, and very high levels of several long non-coding transcripts involved in metastasis (such as MALAT1). When cultured MTFs were transplanted orthotopically into mouse pancreas, they grew as obvious well-differentiated islands of cells, but they also disseminated widely throughout multiple tissues in "stealth" fashion. They were found distributed throughout multiple organs at 4, 8, or 12 weeks after transplantation (including liver, spleen, lung), occurring as single cells or small groups of cells, without formation of obvious tumors or any apparent progression over the 4 to 12 week period. We suggest that MTFs form continually during PDAC development, and that they disseminate early in cancer progression, forming "niches" at distant sites for subsequent colonization by metastasis-initiating cells.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Accurate tumor identification and staging can be difficult. Aptamer-targeted indocyanine green (ICG)-nanoparticles can enhance near-infrared fluorescent imaging of pancreatic and prostate tumors and ...could improve early cancer detection. This project explored whether calcium-phosphosilicate nanoparticles, also known as NanoJackets (NJs), that were bioconjugated with a tumor-specific targeting DNA aptamer could improve the non-invasive detection of pancreatic and prostate tumors.
Using in vivo near-infrared optical imaging and ex vivo fluorescence analysis, DNA aptamer-targeted ICG-loaded NJs were compared to untargeted NJs for detection of tumors.
Nanoparticles were bioconjugated with the DNA aptamer AP1153, which binds to the CCK-B receptor (CCKBR). Aptamer bioconjugated NJs were not significantly increased in size compared with unconjugated nanoparticles. AP1153-ICG-NJ accumulation in orthotopic pancreatic tumors peaked at 18 h post-injection and the ICG signal was cleared by 36 h with no evidence on uptake by non-tumor tissues. Ex vivo tumor imaging confirmed the aptamer-targeted NJs accumulated to higher levels than untargeted NJs, were not taken up by normal pancreas, exited from the tumor vasculature, and were well-dispersed throughout pancreatic and prostate tumors despite extensive fibrosis. Specificity for AP1153-NJ binding to the CCK-B receptor on pancreatic tumor cells was confirmed by pre-treating tumor-bearing mice with the CCK receptor antagonist proglumide. Proglumide pre-treatment reduced the in vivo tumoral accumulation of AP1153-NJs to levels comparable to that of untargeted NJs.
Through specific interactions with CCK-B receptors, tumor-targeted nanoparticles containing either ICG or rhodamine WT were well distributed throughout the matrix of both pancreatic and prostate tumors. Tumor-targeted NJs carrying various imaging agents can enhance tumor detection.
Leukemia is one of the most common and aggressive adult cancers, as well as the most prevalent childhood cancer. Leukemia is a cancer of the hematological system and can be divided into a diversity ...of unique malignancies based on the onset of the disease as well as the specific cell lineages involved. Cancer stem cells, including recently identified leukemia stem cells (LSCs), are hypothesized to be responsible for cancer development, relapse, and resistance to treatment, and new therapeutics targeting these cellular populations are urgently needed. Nontoxic and nonaggregating calcium phosphosilicate nanoparticles (CPSNPs) encapsulating the near-infrared fluoroprobe indocyanine green (ICG) were recently developed for diagnostic imaging and drug delivery as well as for photodynamic therapy (PDT) of solid tumors. Prior studies revealed that specific targeting of CPSNPs allowed for enhanced accumulation within breast cancer tumors, via CD71 targeting, or pancreatic cancer tumors, via gastrin receptor targeting. In the present study, ICG-loaded CPSNPs were evaluated as photosensitizers for PDT of leukemia. Using a novel bioconjugation approach to specifically target CD117 or CD96, surface features enhanced on leukemia stem cells, in vitro ICG-CPSNP PDT of a murine leukemia cell line and human leukemia samples were dramatically improved. Furthermore, the in vivo efficacy of PDT was dramatically enhanced in a murine leukemia model by utilizing CD117-targeted ICG-CPSNPs, resulting in 29% disease-free survival. Altogether, this study demonstrates that leukemia-targeted ICG-loaded CPSNPs offer the promise to effectively treat relapsing and multidrug-resistant leukemia and to improve the life of leukemia patients.
Background
Endogenous opioid peptides have been shown to play a role in the development and/or perpetuation of inflammation. We hypothesize that the endogenous opioid system is involved in ...inflammatory bowel disease, and antagonism of the opioid–opioid receptor will lead to reversal of inflammation.
Aims
A randomized double-blind placebo-controlled study was designed to test the efficacy and safety of an opioid antagonist for 12 weeks in adults with active Crohn’s disease.
Methods
Forty subjects with active Crohn’s disease were enrolled in the study. Randomized patients received daily oral administration of 4.5-mg naltrexone or placebo. Providers and patients were masked to treatment assignment. The primary outcome was the proportion of subjects in each arm with a 70-point decline in Crohn’s Disease Activity Index score (CDAI). The secondary outcome included mucosal healing based upon colonoscopy appearance and histology.
Results
Eighty-eight percent of those treated with naltrexone had at least a 70-point decline in CDAI scores compared to 40% of placebo-treated patients (
p
= 0.009). After 12 weeks, 78% of subjects treated with naltrexone exhibited an endoscopic response as indicated by a 5-point decline in the Crohn’s disease endoscopy index severity score (CDEIS) from baseline compared to 28% response in placebo-treated controls (
p
= 0.008), and 33% achieved remission with a CDEIS score <6, whereas only 8% of those on placebo showed the same change. Fatigue was the only side effect reported that was significantly greater in subjects receiving placebo.
Conclusions
Naltrexone improves clinical and inflammatory activity of subjects with moderate to severe Crohn’s disease compared to placebo-treated controls. Strategies to alter the endogenous opioid system provide promise for the treatment of Crohn’s disease.
Pancreatic ductal adenocarcinomas (PDACs) constitutively express the G-protein-coupled cholecystokinin B receptor (
CCKBR
). In this study, we identified DNA aptamers (APs) that bind to the
CCKBR
and ...describe their characterization and targeting efficacy. Using dual SELEX selection against “exposed”
CCKBR
peptides and
CCKBR
-expressing PDAC cells, a pool of DNA APs was identified. Further downselection was based on predicted structures and properties, and we selected eight APs for initial characterizations. The APs bound specifically to the
CCKBR
, and we showed not only that they did not stimulate proliferation of PDAC cell lines but rather inhibited their proliferation. We chose one AP, termed AP1153, for further binding and localization studies. We found that AP1153 did not activate
CCKBR
signaling pathways, and three-dimensional Confocal microscopy showed that AP1153 was internalized by PDAC cells in a receptor-mediated manner. AP1153 showed a binding affinity of 15 pM. Bioconjugation of AP1153 to the surface of fluorescent NPs greatly facilitated delivery of NPs to PDAC tumors
in vivo
. The selectivity of this AP-targeted NP delivery system holds promise for enhanced early detection of PDAC lesions as well as improved chemotherapeutic treatments for PDAC patients.
Background
Obesity and dietary fat are associated with increased risk of several malignancies including pancreatic cancer. The incidence of pancreatic cancer is increased in countries that consume ...diets high in fat.
Aim
The purpose of this study was to assess the relationship and mechanism of action between dietary fat and endogenous cholecystokinin (CCK) on pancreatic tumor growth and metastasis in an immunocompetent animal model.
Methods
C57BL/6 mice were placed on regular, low-fat, or high-fat diets for 8 weeks before establishment of Panc-02 orthotopic pancreatic tumors. Mice were then treated with a CCK-A receptor antagonist, devazepide, or vehicle for an additional 2.5 weeks. Pancreas tumors were weighed and metastases counted. Blood CCK levels were measured by radioimmunoassay (RIA). Tissues were examined histologically and studied for genes associated with metastasis by RT-PCR array. Effects of the CCK antagonist on Panc-02 cells invasiveness was assessed in a Matrigel invasion assay.
Results
Mice that received the high-fat diet had larger tumors and tenfold higher serum CCK levels by RIA compared to normal diet controls (
p
< 0.01). Pancreatic tumors in high-fat diet mice treated with the antagonist had fewer intravascular tumor emboli and metastases compared to controls. The reduction in tumor emboli correlated with decreased vascular endothelial growth factor-A (VEGF-A) expression in tumors (
p
< 6 × 10
−9
). In vitro invasiveness of Panc-02 cells also was reduced by CCK-A receptor antagonist treatment (
p
= 1.33 × 10
−6
).
Conclusion
CCK is a mediator of dietary fat-associated pancreatic cancer. CCK is also involved in the invasiveness of pancreatic tumors through a mechanism involving VEGF-A.
Drug resistant cancers like pancreatic ductal adenocarcinoma (PDAC) are difficult to treat, and nanoparticle drug delivery systems can overcome some of the limitations of conventional systemic ...chemotherapy. In this study, we demonstrate that FdUMP and dFdCMP, the bioactive, phosphorylated metabolites of the chemotherapy drugs 5-FU and gemcitabine, can be encapsulated into calcium phosphosilicate nanoparticles (CPSNPs). The non-phosphorylated drug analogs were not well encapsulated by CPSNPs, suggesting the phosphate modification is essential for effective encapsulation. In vitro proliferation assays, cell cycle analyses and/or thymidylate synthase inhibition assays verified that CPSNP-encapsulated phospho-drugs retained biological activity. Analysis of orthotopic tumors from mice treated systemically with tumor-targeted FdUMP-CPSNPs confirmed the in vivo up take of these particles by PDAC tumor cells and release of active drug cargos intracellularly. These findings demonstrate a novel methodology to efficiently encapsulate chemotherapeutic agents into the CPSNPs and to effectively deliver them to pancreatic tumor cells.
By encapsulating chemotherapeutic agents into nanocarriers, such as calcium phosphosilicate nanoparticles (CPSNPs), metabolic inactivation of pro-drugs, for instance the inactivation of 5-FU by the liver enzyme dihydropyrimidine dehydrogenase (DPD), often can be avoided. Herein we demonstrate that the bioactive metabolite of 5-FU, FdUMP, can be effectively encapsulated into CPSNPs. Once taken up by tumor cells, the mPEG-FdUMP-CPSNPs dissolve in the late endosome to release the FdUMP cargo. In the presence of folate (CH2THF), FdUMP irreversibly binds to and inhibits thymidylate synthase (TS), resulting in a cellular depletion of dTMP, increased DNA damage, and ultimately cell death. Display omitted
The poor prognosis for pancreatic ductal adenocarcinoma (PDAC) patients is due in part to the highly fibrotic nature of the tumors that impedes delivery of therapeutics, including nanoparticles ...(NPs). Our prior studies demonstrated that proglumide, a cholecystokinin receptor (CCKR) antagonist, reduced fibrosis pervading PanIN lesions in mice. Here, we further detail how the reduced fibrosis elicited by proglumide achieves the normalization of the desmoplastic tumor microenvironment (TME) and improves nanoparticle uptake. One week following the orthotopic injection of PDAC cells, mice were randomized to normal or proglumide-treated water for 3-6 weeks. Tumors were analyzed ex vivo for fibrosis, vascularity, stellate cell activation, vascular patency, and nanoparticle distribution. The histological staining and three-dimensional imaging of tumors each indicated a reduction in stromal collagen in proglumide-treated mice. Proglumide treatment increased tumor vascularity and decreased the activation of cancer-associated fibroblasts (CAFs). Additionally, PANC-1 cells with the shRNA-mediated knockdown of the CCK2 receptor showed an even greater reduction in collagen, indicating the CCK2 receptors on tumor cells contribute to the desmoplastic TME. Proglumide-mediated reduction in fibrosis also led to functional changes in the TME as evidenced by the enhanced intra-tumoral distribution of small (<12 nm) Rhodamine-loaded nanoparticles. The documented in vivo, tumor cell-intrinsic anti-fibrotic effects of CCK2R blockade in both an immunocompetent syngeneic murine PDAC model as well as a human PDAC xenograft model demonstrates that CCK2R antagonists, such as proglumide, can improve the delivery of nano-encapsulated therapeutics or imaging agents to pancreatic tumors.
Exogenous administration of cholecystokinin (CCK) induces hypertrophy and hyperplasia of the pancreas with an increase in DNA content. We hypothesized that endogenous CCK is involved in the malignant ...progression of pancreatic intraepithelial neoplasia (PanIN) lesions and the fibrosis associated with pancreatic cancer.
The presence of CCK receptors in early PanIN lesions was examined by immunohistochemistry in mouse and human pancreas. Pdx1-Cre/LSL-Kras transgenic mice were randomized to receive either untreated drinking water or water supplemented with a CCK receptor antagonist (proglumide, 0.1 mg/mL). Pancreas from the mice were removed and examined histologically for number and grade of PanINs after 1, 2, or 4 months of antagonist therapy.
Both CCK-A and CCK-B receptors were identified in early stage PanINs from mouse and human pancreas. The grade of PanIN lesions was reversed, and progression to advanced lesions arrested in mice treated with proglumide compared with the controls (P = 0.004). Furthermore, pancreatic fibrosis was significantly reduced in antagonist-treated animals compared with vehicle (P < 0.001).
These findings demonstrate that endogenous CCK is in part responsible for the development and progression of pancreatic cancer. The use of CCK receptor antagonists may have a role in cancer prophylaxis in high-risk subjects and may reduce fibrosis in the microenvironment.
Early deep sedation in mechanically ventilated patients during the first 48 h of intensive care unit (ICU) admission can be associated with adverse outcomes. We hypothesised that moving the ‘daily ...sedation break’ process forwards, might allow earlier titration of sedation to target levels – an ‘early sedation cessation’ (ESC).
We commenced a quality improvement project with the primary outcome being to stop sedation completely, within 4 h of ICU admission, in 95% of eligible patients. This was done by small, step-wise tests of change. No ethical approval was required.
Between 1 February 2014 and 31 January 2018, 1787 intubated patients were included. 1052 received an ‘ESC’ within 4 h (‘Yes’), 545 were excluded (‘Excluded’), and 190 were inadvertently omitted from ‘ESC’ (‘No’). The primary aim was achieved for the first time after 12 months. Compared to the ‘Yes’ group, the ‘Excluded’ group received 38% more propofol in the first 48 h of admission (IRR 1.38 (1.31–1.47), p<0.001), while the ‘No’ group received 32% more (IRR 1.32 (1.22–1.43), p<0.001). At four hours, 19·6% (12·9–27·9) of the ‘Yes’ group had attained a target RASS of -1, 0 or 1, compared to 13·6% (8·0–21·0) of those in the ‘No’ group. This proportion increased to 55·6% (46·1–64·9) at 24 h compared with 44·9% (35·6–54·4) in the ‘No’ group.
Ceasing sedative infusions as soon as possible, is safe and feasible, in both medical and surgical patients, and can be implemented into ‘real life’ with no additional staffing.
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