Joshua Leach

CXCR2 Inhibition Profoundly Suppresses Metastases and Augments Immunotherapy in Pancreatic Ductal Adenocarcinoma.

Summary CXCR2 has been suggested to have both tumor-promoting and tumor-suppressive properties. Here we show that CXCR2 signaling is upregulated in human pancreatic cancer, predominantly in neutrophil/myeloid-derived suppressor cells, but rarely in tumor cells. Genetic ablation or inhibition of CXCR2 abrogated metastasis, but only inhibition slowed tumorigenesis. Depletion of neutrophils/myeloid-derived suppressor cells also suppressed metastasis suggesting a key role for CXCR2 in establishing and maintaining the metastatic niche. Importantly, loss or inhibition of CXCR2 improved T cell entry, and combined inhibition of CXCR2 and PD1 in mice with established disease significantly extended survival. We show that CXCR2 signaling in the myeloid compartment can promote pancreatic tumorigenesis and is required for pancreatic cancer metastasis, making it an excellent therapeutic target.

Targeting the LOX/hypoxia axis reverses many of the features that make pancreatic cancer deadly: inhibition of LOX abrogates metastasis and enhances drug efficacy

Pancreatic ductal adenocarcinoma (PDAC) is one of the leading causes of cancer‐related mortality. Despite significant advances made in the treatment of other cancers, current chemotherapies offer little survival benefit in this disease. Pancreaticoduodenectomy offers patients the possibility of a cure, but most will die of recurrent or metastatic disease. Hence, preventing metastatic disease in these patients would be of significant benefit. Using principal component analysis (PCA), we identified a LOX/hypoxia signature associated with poor patient survival in resectable patients. We found that LOX expression is upregulated in metastatic tumors from Pdx1‐Cre KrasG12D/+ Trp53R172H/+ (KPC) mice and that inhibition of LOX in these mice suppressed metastasis. Mechanistically, LOX inhibition suppressed both migration and invasion of KPC cells. LOX inhibition also synergized with gemcitabine to kill tumors and significantly prolonged tumor‐free survival in KPC mice with early‐stage tumors. This was associated with stromal alterations, including increased vasculature and decreased fibrillar collagen, and increased infiltration of macrophages and neutrophils into tumors. Therefore, LOX inhibition is able to reverse many of the features that make PDAC inherently refractory to conventional therapies and targeting LOX could improve outcome in surgically resectable disease.

Fumarate induces redox-dependent senescence by modifying glutathione metabolism

Mutations in the tricarboxylic acid (TCA) cycle enzyme fumarate hydratase (FH) are associated with a highly malignant form of renal cancer. We combined analytical chemistry and metabolic computational modelling to investigate the metabolic implications of FH loss in immortalized and primary mouse kidney cells. Here, we show that the accumulation of fumarate caused by the inactivation of FH leads to oxidative stress that is mediated by the formation of succinicGSH, a covalent adduct between fumarate and glutathione. Chronic succination of GSH, caused by the loss of FH, or by exogenous fumarate, leads to persistent oxidative stress and cellular senescence in vitro and in vivo. Importantly, the ablation of p21, a key mediator of senescence, in Fh1-deficient mice resulted in the transformation of benign renal cysts into a hyperplastic lesion, suggesting that fumarate-induced senescence needs to be bypassed for the initiation of renal cancers.

Mitotic Stress Is an Integral Part of the Oncogene-Induced Senescence Program that Promotes Multinucleation and Cell Cycle Arrest.

Summary Oncogene-induced senescence (OIS) is a tumor suppression mechanism that blocks cell proliferation in response to oncogenic signaling. OIS is frequently accompanied by multinucleation; however, the origin of this is unknown. Here, we show that multinucleate OIS cells originate mostly from failed mitosis. Prior to senescence, mutant H-RasV12 activation in primary human fibroblasts compromised mitosis, concordant with abnormal expression of mitotic genes functionally linked to the observed mitotic spindle and chromatin defects. Simultaneously, H-RasV12 activation enhanced survival of cells with damaged mitoses, culminating in extended mitotic arrest and aberrant exit from mitosis via mitotic slippage. ERK-dependent transcriptional upregulation of Mcl1 was, at least in part, responsible for enhanced survival and slippage of cells with mitotic defects. Importantly, mitotic slippage and oncogene signaling cooperatively induced senescence and key senescence effectors p21 and p16. In summary, activated Ras coordinately triggers mitotic disruption and enhanced cell survival to promote formation of multinucleate senescent cells.

Proof of concept and feasibility studies examining the influence of combination ribose, adenine and allopurinol treatment on stroke outcome in the rat

Background: Cerebral ischaemia results in a rapid and profound depletion of adenosine triphosphate (ATP), the energy currency of the cell. This depletion leads to disruption of cellular homeostasis and cell death. Early replenishment of ATP levels might therefore have a neuroprotective effect in the injured brain. We have previously shown that the ATP precursors, D-ribose and adenine (RibAde), restored the reduced ATP levels in rat brain slices to values similar to those measured in the intact rodent brain. The aim of this study was to assess whether RibAde, either alone or in combination with the xanthine oxidase inhibitor allopurinol (RibAdeAll; to further increase the availability of ATP precursors), could improve outcome in an in vivo rodent model of transient cerebral ischaemia. Methods: After 60 min occlusion of the middle cerebral artery, and upon reperfusion, rats were administered saline, RibAde, or RibAdeAll for 6 h. Baseline lesion volume was determined by diffusion-weighted MRI prior to reperfusion and final infarct volume determined by T2-weighted MRI at Day 7. Neurological function was assessed at Days 1, 3 and 7. Results: Ischaemic lesion volume decreased between Days 1 and 7: a 50% reduction was observed for the RibAdeAll group, 38% for the RibAde group and 18% in the animals that received saline. Reductions in lesion size in treatment groups were accompanied by a trend for faster functional recovery. Conclusion: These data support the potential use of ribose, adenine and allopurinol in the treatment of cerebral ischaemic injury, especially since all compounds have been used in man.

Diagnostic Exercise: Circling and Behavioral Changes in a Cat.

A 4-year old spayed male domestic shorthair cat was presented with a history of circling and behavioral changes. Neurologic examination showed mild proprioceptive deficits. The lesion was localized in the forebrain, and magnetic resonance imaging revealed the presence of a large midline intracranial mass extending from the frontal lobe to the tentorial region of the brain. Euthanasia was elected due to poor prognosis. Histopathologic evaluation confirmed the presence of a mass composed by sheets and aggregates of large round/polygonal cells and multinucleate cells associated with deposits of cholesterol clefts, scattered hemorrhages and hemosiderin-laden macrophages. Immunohistochemistry showed that the round/polygonal cells and multinucleate cells were strongly positive for major histocompatibility complex class II antigen, variably positive for CD18, and occasionally positive for S100. Subsets of spindle cells showing variable expression of vimentin, S100, and neuron-specific enolase were also present. The final diagnosis was cholesterol granuloma. Differential diagnosis with meningioma is discussed.

Closantel toxicity in a pregnant ewe at mid gestation: the pathological evaluation of the ewe and lamb nine months later

In February 2013, a flock of 20 pastured ewes was treated with oral closantel. Within 10 days of dosing, one 2-year-old Texel-cross ewe that was approximately three months in lamb was noticed to be spending a prolonged time in recumbency. On the 6th of March, the animal was referred to Scottish Centre for Production Animal Health and Food Safety (SCPAHFS). At admission, the ewe appeared blind, pupils were dilated with a negative menace response and pupillary light reflex bilaterally. Closantel toxicity was suspected. Following admission, this ewe was housed and was stable despite remaining blind. On April 21, a single male lamb was born. This lamb was found to have normal clinical and neurological parameters over five months of observation following birth. On postmortem examination closantel toxicity was confirmed in the ewe while following detailed gross and histological examination of all tissues of the lamb there was no indication of the toxicity suffered by its mother.

Neutrophils: homing in on the myeloid mechanisms of metastasis

Highlights • Neutrophils play important roles throughout the metastatic process.• Tumour progression and invasion are influenced by infiltrating neutrophils.• Signalling from neutrophils can enhance survival of circulating tumour cells.• Neutrophils play a key role in establishing the metastatic niche.

Axial Multicentric Osteosarcoma in an English Cocker Spaniel

An 8-year-old, male neutered English Cocker Spaniel presented to the Royal Veterinary College for evaluation of reluctance to exercise and spinal hyperesthesia of 1-year duration. Medical treatment by referring veterinarian with antibiotics and tramadol did not result in clinical improvement. Survey radiographs performed before referral revealed multiple lytic bone lesions involving sternum and ribs. The dog had been fed for several months on a hypoallergenic diet and administered prednisolone for presumed inflammatory bowel disease. General physical examination did not reveal abnormalities. Neurological examination revealed a kyphotic posture and a short and stilted pelvic limb gait. Spinal hyperesthesia could be elicited on thoracic, thoracolumbar, and lumbar spinal palpation. A complete blood count and free catch urine analysis did not reveal abnormalities. Serum biochemistry profile revealed hypoalbuminemia (20.3 g/L [RI (reference interval) 28– 39 g/L]) and increased inorganic phosphorus concentration (2.68 mmol/L [RI 0.8–2 mmol/L]), alanine aminotransferase activity (234 U/L [RI 13–88 U/L]) and alkaline phosphatase activity (1425 U/L [RI 19–285 U/ L]). Ionized calcium concentration was within the RI. Magnetic resonance imaging (MRI) imaging was performed using a protocol that included sagittal and transverse plane T2-weighted (repetition time, TR [ms], echo time, TE [ms], 3000/120) (Fig 1A), sagittal T2weighted short-tau inversion recovery (TR/TE, 3612/80) (Fig 1C) and sagittal and transverse plane T1-weighted spectral presaturation with inversion recovery (TR/TE, 533/8) sequences. Sagittal and transverse plane T1weighted (T1W turbo spin echo) (TR/TE, 400/8) images were acquired before and after IV injection with gadolinium contrast (Fig 1B, D). Magnetic resonance imaging demonstrated multiple poorly defined, expansile lytic lesions affecting the spinous processes of the sacrum, L5, L3, T13, T12, T11, T8, and T2, the vertebral bodies of the sacrum, L5, L4, T13, T12, T11, T9, T7, T6, and T2, and multiple sternebrae. These lesions were characterized by a heterogeneous intensity in all sequences and patchy contrast enhancement. A fracture of the vertebral body of T11 with bone remodeling was present without spinal cord compression. After MRI, thorax and abdomen computed tomography (CT) was performed using a 16-slice scanner. CT imaging (Fig 2A) confirmed the expansile lytic bone lesions and revealed additional lesions affecting multiple ribs and pelvis and accounting for more than 20 bone lytic lesions with variable sizes. CT did not reveal any abnormalities involving thoracic or abdominal visceral structures and no enhancement was seen after IV administration of contrast medium. Based on imaging and laboratory results, nonproductive multiple myeloma was considered the most likely differential diagnosis. The lack of a clear primary lesion, the numerous bones involved and absence of visceral lesions was not typical for axial osteosarcoma (OS), even considering concurrent multiple metastases. The imaging findings were more suggestive of synchronous multicentric osteosarcoma (MOS), a rare OS clinical presentation in people, with simultaneous multiple bone lesions without pulmonary involvement and extremely rare in domestic animals, which was considered as a rare differential diagnosis. Bone marrow aspirate (left humerus) and serum electrophoresis did not reveal abnormalities and evaluation of urine for Bence-Jones proteins was negative. Ultrasound guided fine needle aspirates of affected lesions (dorsal spinous process of L5 and rib) were inconclusive. Subsequently, the dog was anesthetized and a From the Department of Clinical Science and Services, (Parzefall, De Decker, Carvalho, Lara-Garcia); Department of Pathology and Pathogen Biology, Royal Veterinary College, University of London, Hatfield, (Terr, Smith); and the Veterinary Diagnostic Services, School of Veterinary Medicine, University of Glasgow, Glasgow, UK(Leach). The work was done at the Royal Veterinary College, United Kingdom. No financial support was given for this study. This case was presented in part at the 27th Annual Symposium of the European Society of Veterinary Neurology – European College of Veterinary Neurology, 18–20 September 2014, Madrid, Spain. Corresponding author: A. Lara-Garcia, DVM, MSc, PhD, DACVIM & DECVIM-CA (Oncology), Department of Clinical Science and Services, Royal Veterinary College, University of London, Hatfield, AL9 7TA, UK; e-mail: algarcia57@rvc.ac.uk. Submitted April 29, 2016; Revised July 1, 2016; Accepted July 5, 2016. Copyright

Multiple congenital cardiovascular defects including type IV persistent truncus arteriosus in a Shetland pony – Short communication

This case report describes the pathological findings of multiple congenital cardiac defects in a 2-year-old female Shetland pony with clinical signs of chronic respiratory distress. Persistent truncus arteriosus (PTA) type IV, interventricular septal defect, overriding aorta, pulmonary trunk agenesis, pulmonary arteries arising from the descending aorta, and compensatory right ventricular hypertrophy were observed.