Real-Time Visualization and Quantitation of Vascular Permeability In Vivo: Implications for Drug Delivery

The leaky, heterogeneous vasculature of human tumors prevents the even distribution of systemic drugs within cancer tissues. However, techniques for studying vascular delivery systems in vivo often require complex mammalian models and time-consuming, surgical protocols. The developing chicken embryo is a well-established model for human cancer that is easily accessible for tumor imaging. To assess this model for the in vivo analysis of tumor permeability, human tumors were grown on the chorioallantoic membrane (CAM), a thin vascular membrane which overlays the growing chick embryo. The real-time movement of small fluorescent dextrans through the tumor vasculature and surrounding tissues were used to measure vascular leak within tumor xenografts. Dextran extravasation within tumor sites was selectively enhanced an interleukin-2 (IL-2) peptide fragment or vascular endothelial growth factor (VEGF). VEGF treatment increased vascular leak in the tumor core relative to surrounding normal tissue and increased doxorubicin uptake in human tumor xenografts. This new system easily visualizes vascular permeability changes in vivo and suggests that vascular permeability may be manipulated to improve chemotherapeutic targeting to tumors

Leuprorelin Acetate Long-Lasting Effects on GnRH Receptors of Prostate Cancer Cells: An Atomic Force Microscopy Study of Agonist/Receptor Interaction

High cell-surface GnRH receptor (GnRH-R) levels have been shown to have a major influence on the extent of GnRH agonist-mediated tumor growth inhibition. The ability of the GnRH agonist leuprorelin acetate (LA) to induce a post-transcriptional upregulation of GnRH-R at the plasma membrane of androgen-sensitive (LNCaP) and -insensitive (PC-3) prostate cancer (PCa) cells has been previously demonstrated by Western blotting. Here we performed single molecule force spectroscopy by using Atomic Force Microscopy (AFM), which has proven to be a powerful tool allowing for investigation of living cell surface biological features, such as the so far unclear GnRH agonist/receptor interaction. Thus, in the hormone-insensitive PC-3 cells, we characterized the strength of the LA-receptor binding, and the amount and distribution of the functional receptor molecules on the cell surface. The effect of a long and continuous treatment (up to 30 days) with the agonist (10-11 and 10-6 M) on the same parameters was also investigated. A GnRH-R increase was observed, reaching the maximum (~80%) after 30 days of treatment with the highest dose of LA (10-6 M). The analogue-induced increase in GnRH-R was also demonstrated by Western blotting. In addition, two different receptor bound strengths were detected by AFM, which suggests the existence of two GnRH-R classes. A homogeneous distribution of the unbinding events has been found on untreated and treated PC-3 cell surfaces. The persistence of high receptor levels at the membrane of these living cells may warrant the maintenance of the response to LA also in androgen-unresponsive PCa. Moreover, the determination of ligand/receptor bond strength could shed light on the poorly understood event of LA/GnRH-R interaction and/or address structural/chemical agonist optimizations. \ua9 2013 Lama et al

Luteinising hormone-releasing hormone analogue reverses the cell adhesion profile of EGFR overexpressing DU-145 human prostate carcinoma subline

Cetrorelix, a luteinising hormone-releasing hormone (LHRH) analogue, has been shown to limit growth of the human androgen-independent prostate cell line DU-145, although other inhibitory actions may also be affected. Both growth and invasion of DU-145 cells are linked to autocrine epidermal growth factor receptor (EGFR) signalling. Invasiveness requires not only cells to migrate to conduits, but also reduced adhesiveness between tumour cells to enable separation from the tumour mass. Thus, we investigated whether Cetrorelix alters the DU-145 cell–cell adhesion and if this occurs via altered EGFR signalling. Pharmacologic levels of Cetrorelix limited the invasiveness of a highly invasive DU-145 subline overexpressing full-length EGFR (DU-145 WT). Extended exposure of the cells to Cetrorelix resulted in increased levels of the cell–cell adhesion complex molecules E-cadherin, α- and β-catenin, and p120. Puromycin blocked the increases in E-cadherin and β-catenin levels, suggesting that de novo protein synthesis is required. The Cetrorelix effect appears to occur via transmodulation of EGFR by a protein kinase C (PKC)-dependent mechanism, as there were no changes in DU-145 cells expressing EGFR engineered to negate the PKC transattenuation site (DU-145 A654); downregulation of EGFR signalling produced a similar upregulation in adhesion complex proteins, further suggesting a role for autocrine signalling. Cetrorelix increased the cell–cell adhesiveness of DU-145 WT cells to an extent similar to that seen when autocrine EGFR signalling is blocked; as expected, DU-145 A654 cell–cell adhesion also was unaffected by Cetrorelix. The increased adhesiveness is expected as the adhesion complex molecules moved to the cells' periphery. These data offer direct insight into the possible crosstalk pathways between the LHRH and EGFR receptor signalling. The ability of Cetrorelix to downregulate EGFR signalling and subsequently reverse the antiadhesiveness found in metastatic prostate cancer highlights a novel potential target for therapeutic strategies

Study protocol: a multi-centre randomised study of induction chemotherapy followed by capecitabine +/- nelfinavir with high- or standard-dose radiotherapy for locally advanced pancreatic cancer (SCALOP-2)

Background Induction chemotherapy followed by chemoradiation is a treatment option for patients with locally advanced pancreatic cancer (LAPC). However, overall survival is comparable to chemotherapy alone and local progression occurs in nearly half of all patients, suggesting chemoradiation strategies should be optimised. SCALOP-2 is a randomised phase II trial testing the role of radiotherapy dose escalation and/or the addition of the radiosensitiser nelfinavir, following induction chemotherapy of gemcitabine and nab-paclitaxel (GEMABX). A safety run-in phase (stage 1) established the nelfinavir dose to administer with chemoradiation in the randomised phase (stage 2). Methods Patients with locally advanced, inoperable, non-metastatic pancreatic adenocarcinoma receive three cycles of induction GEMABX chemotherapy prior to radiological assessment. Those with stable/responding disease are eligible for further trial treatment. In Stage 1, participants received one further cycle of GEMABX followed by capecitabine-chemoradiation with escalating doses of nelfinavir in a rolling-six design. Stage 2 aims to register 262 and randomise 170 patients with responding/stable disease to one of five arms: capecitabine with high- (arms C + D) or standard-dose (arms A + B) radiotherapy with (arms A + C) or without (arms B + D) nelfinavir, or three more cycles of GEMABX (arm E). Participants allocated to the chemoradiation arms receive another cycle of GEMABX before chemoradiation begins. Co-primary outcomes are 12-month overall survival (radiotherapy dose-escalation question) and progression-free survival (nelfinavir question). Secondary outcomes include toxicity, quality of life, disease response rate, resection rate, treatment compliance, and CA19–9 response. SCALOP-2 incorporates a detailed radiotherapy quality assurance programme. Discussion SCALOP-2 aims to optimise chemoradiation in LAPC and incorporates a modern induction regimen

Clinical development of new drug-radiotherapy combinations.

In countries with the best cancer outcomes, approximately 60% of patients receive radiotherapy as part of their treatment, which is one of the most cost-effective cancer treatments. Notably, around 40% of cancer cures include the use of radiotherapy, either as a single modality or combined with other treatments. Radiotherapy can provide enormous benefit to patients with cancer. In the past decade, significant technical advances, such as image-guided radiotherapy, intensity-modulated radiotherapy, stereotactic radiotherapy, and proton therapy enable higher doses of radiotherapy to be delivered to the tumour with significantly lower doses to normal surrounding tissues. However, apart from the combination of traditional cytotoxic chemotherapy with radiotherapy, little progress has been made in identifying and defining optimal targeted therapy and radiotherapy combinations to improve the efficacy of cancer treatment. The National Cancer Research Institute Clinical and Translational Radiotherapy Research Working Group (CTRad) formed a Joint Working Group with representatives from academia, industry, patient groups and regulatory bodies to address this lack of progress and to publish recommendations for future clinical research. Herein, we highlight the Working Group's consensus recommendations to increase the number of novel drugs being successfully registered in combination with radiotherapy to improve clinical outcomes for patients with cancer.National Institute for Health ResearchThis is the final version of the article. It first appeared from Nature Publishing Group via http://dx.doi.org/10.1038/nrclinonc.2016.7

PANC Study (Pancreatitis: A National Cohort Study): national cohort study examining the first 30 days from presentation of acute pancreatitis in the UK

Background Acute pancreatitis is a common, yet complex, emergency surgical presentation. Multiple guidelines exist and management can vary significantly. The aim of this first UK, multicentre, prospective cohort study was to assess the variation in management of acute pancreatitis to guide resource planning and optimize treatment. Methods All patients aged greater than or equal to 18 years presenting with acute pancreatitis, as per the Atlanta criteria, from March to April 2021 were eligible for inclusion and followed up for 30 days. Anonymized data were uploaded to a secure electronic database in line with local governance approvals. Results A total of 113 hospitals contributed data on 2580 patients, with an equal sex distribution and a mean age of 57 years. The aetiology was gallstones in 50.6 per cent, with idiopathic the next most common (22.4 per cent). In addition to the 7.6 per cent with a diagnosis of chronic pancreatitis, 20.1 per cent of patients had a previous episode of acute pancreatitis. One in 20 patients were classed as having severe pancreatitis, as per the Atlanta criteria. The overall mortality rate was 2.3 per cent at 30 days, but rose to one in three in the severe group. Predictors of death included male sex, increased age, and frailty; previous acute pancreatitis and gallstones as aetiologies were protective. Smoking status and body mass index did not affect death. Conclusion Most patients presenting with acute pancreatitis have a mild, self-limiting disease. Rates of patients with idiopathic pancreatitis are high. Recurrent attacks of pancreatitis are common, but are likely to have reduced risk of death on subsequent admissions

PANC Study (Pancreatitis: A National Cohort Study): national cohort study examining the first 30 days from presentation of acute pancreatitis in the UK

Abstract Background Acute pancreatitis is a common, yet complex, emergency surgical presentation. Multiple guidelines exist and management can vary significantly. The aim of this first UK, multicentre, prospective cohort study was to assess the variation in management of acute pancreatitis to guide resource planning and optimize treatment. Methods All patients aged greater than or equal to 18 years presenting with acute pancreatitis, as per the Atlanta criteria, from March to April 2021 were eligible for inclusion and followed up for 30 days. Anonymized data were uploaded to a secure electronic database in line with local governance approvals. Results A total of 113 hospitals contributed data on 2580 patients, with an equal sex distribution and a mean age of 57 years. The aetiology was gallstones in 50.6 per cent, with idiopathic the next most common (22.4 per cent). In addition to the 7.6 per cent with a diagnosis of chronic pancreatitis, 20.1 per cent of patients had a previous episode of acute pancreatitis. One in 20 patients were classed as having severe pancreatitis, as per the Atlanta criteria. The overall mortality rate was 2.3 per cent at 30 days, but rose to one in three in the severe group. Predictors of death included male sex, increased age, and frailty; previous acute pancreatitis and gallstones as aetiologies were protective. Smoking status and body mass index did not affect death. Conclusion Most patients presenting with acute pancreatitis have a mild, self-limiting disease. Rates of patients with idiopathic pancreatitis are high. Recurrent attacks of pancreatitis are common, but are likely to have reduced risk of death on subsequent admissions. </jats:sec

Modulation of the tumor microvasculature by phosphoinositide-3 kinase inhibition increases doxorubicin delivery in vivo.

PURPOSE: Because effective drug delivery is often limited by inadequate vasculature within the tumor, the ability to modulate the tumor microenvironment is one strategy that may achieve better drug distribution. We have previously shown that treatment of mice bearing tumors with phosphoinositide-3 kinase (PI3K) inhibitors alters vascular structure in a manner analogous to vascular normalization and results in increased perfusion of the tumor. On the basis of that result, we asked whether inhibition of PI3K would improve chemotherapy delivery. EXPERIMENTAL DESIGN: Mice with xenografts using the cell line SQ20B bearing a hypoxia marker or MMTV-neu transgenic mice with spontaneous breast tumors were treated with the class I PI3K inhibitor GDC-0941. The tumor vasculature was evaluated by Doppler ultrasound, and histology. The delivery of doxorubicin was assessed using whole animal fluorescence, distribution on histologic sections, high-performance liquid chromatography on tumor lysates, and tumor growth delay. RESULTS: Treatment with GDC-0941 led to approximately three-fold increases in perfusion, substantially reduced hypoxia and vascular normalization by histology. Significantly increased amounts of doxorubicin were delivered to the tumors correlating with synergistic tumor growth delay. The GDC-0941 itself had no effect on tumor growth. CONCLUSION: Inhibition of PI3K led to vascular normalization and improved delivery of a chemotherapeutic agent. This study highlights the importance of the microvascular effects of some novel oncogenic signaling inhibitors and the need to take those changes into account in the design of clinical trials many of which use combinations of chemotherapeutic agents