CD73 controls Myosin II-driven invasion, metastasis, and immunosuppression in amoeboid pancreatic cancer cells.

Pancreatic ductal adenocarcinoma (PDAC) has a very poor prognosis because of its high propensity to metastasize and its immunosuppressive microenvironment. Using a panel of pancreatic cancer cell lines, three-dimensional (3D) invasion systems, microarray gene signatures, microfluidic devices, mouse models, and intravital imaging, we demonstrate that ROCK-Myosin II activity in PDAC cells supports a transcriptional program conferring amoeboid invasive and immunosuppressive traits and in vivo metastatic abilities. Moreover, we find that immune checkpoint CD73 is highly expressed in amoeboid PDAC cells and drives their invasive, metastatic, and immunomodulatory traits. Mechanistically, CD73 activates RhoA-ROCK-Myosin II downstream of PI3K. Tissue microarrays of human PDAC biopsies combined with bioinformatic analysis reveal that rounded-amoeboid invasive cells with high CD73-ROCK-Myosin II activity and their immunosuppressive microenvironment confer poor prognosis to patients. We propose targeting amoeboid PDAC cells as a therapeutic strategy.

AMPK is a mechano-metabolic sensor linking cell adhesion and mitochondrial dynamics to Myosin-dependent cell migration.

Cell migration is crucial for cancer dissemination. We find that AMP-activated protein kinase (AMPK) controls cell migration by acting as an adhesion sensing molecular hub. In 3-dimensional matrices, fast-migrating amoeboid cancer cells exert low adhesion/low traction linked to low ATP/AMP, leading to AMPK activation. In turn, AMPK plays a dual role controlling mitochondrial dynamics and cytoskeletal remodelling. High AMPK activity in low adhering migratory cells, induces mitochondrial fission, resulting in lower oxidative phosphorylation and lower mitochondrial ATP. Concurrently, AMPK inactivates Myosin Phosphatase, increasing Myosin II-dependent amoeboid migration. Reducing adhesion or mitochondrial fusion or activating AMPK induces efficient rounded-amoeboid migration. AMPK inhibition suppresses metastatic potential of amoeboid cancer cells in vivo, while a mitochondrial/AMPK-driven switch is observed in regions of human tumours where amoeboid cells are disseminating. We unveil how mitochondrial dynamics control cell migration and suggest that AMPK is a mechano-metabolic sensor linking energetics and the cytoskeleton.

LAP1 supports nuclear adaptability during constrained melanoma cell migration and invasion.

Metastasis involves dissemination of cancer cells away from a primary tumour and colonization at distal sites. During this process, the mechanical properties of the nucleus must be tuned since they pose a challenge to the negotiation of physical constraints imposed by the microenvironment and tissue structure. We discovered increased expression of the inner nuclear membrane protein LAP1 in metastatic melanoma cells, at the invasive front of human primary melanoma tumours and in metastases. Human cells express two LAP1 isoforms (LAP1B and LAP1C), which differ in their amino terminus. Here, using in vitro and in vivo models that recapitulate human melanoma progression, we found that expression of the shorter isoform, LAP1C, supports nuclear envelope blebbing, constrained migration and invasion by allowing a weaker coupling between the nuclear envelope and the nuclear lamina. We propose that LAP1 renders the nucleus highly adaptable and contributes to melanoma aggressiveness.

WNT11-FZD7-DAAM1 signalling supports tumour initiating abilities and melanoma amoeboid invasion.

Melanoma is a highly aggressive tumour that can metastasize very early in disease progression. Notably, melanoma can disseminate using amoeboid invasive strategies. We show here that high Myosin II activity, high levels of ki-67 and high tumour-initiating abilities are characteristic of invasive amoeboid melanoma cells. Mechanistically, we find that WNT11-FZD7-DAAM1 activates Rho-ROCK1/2-Myosin II and plays a crucial role in regulating tumour-initiating potential, local invasion and distant metastasis formation. Importantly, amoeboid melanoma cells express both proliferative and invasive gene signatures. As such, invasive fronts of human and mouse melanomas are enriched in amoeboid cells that are also ki-67 positive. This pattern is further enhanced in metastatic lesions. We propose eradication of amoeboid melanoma cells after surgical removal as a therapeutic strategy.

Prevalence, molecular typing and risk factor analysis for Giardia duodenalis infections in dogs in a central London rescue shelter.

A cross-sectional survey to investigate canine infections with Giardia duodenalis was undertaken at a central London rescue shelter between October 2006 and March 2007. The objectives of the study were to (i) estimate the prevalence of infection in dogs admitted to a London dog shelter using a commercially available ELISA-based test kit; (ii) identify the relative importance of potential dog level risk factors for infection; and (iii) identify the occurrence of different G. duodenalis assemblages present in this population in order to identify presence of any potentially zoonotic assemblages. A faecal sample was collected from each dog entering the shelter within 1 day of arrival. Each sample was tested for the presence of parasite cyst wall protein using the Giardia SNAP test kit (Idexx Laboratories). Samples were graded for faecal consistency on a standard scale and data on age, breed, categorized breed group, sex and neutered status were collected for each dog. Associations between infection status and each dog level variable were investigated using univariable and multivariable logistic regression. Selected G. duodenalis-positive samples were genotyped using previously described primers targeting the 18S rDNA gene and the beta-giardin gene. Samples from a total of 878 dogs were collected and the true prevalence found to be 21.0% (95% CI 16.7-25.4%). In the present study, the odds of infection decreased with increasing age (adjusted odds ratio 0.66, 95% CI 0.54-0.80, p<0.0001) and was increased for Rottweilers (adjusted odds ratio 2.12, 95% CI 1.03-4.34, p=0.04). Of the 51 samples selected for genotyping, 41 samples yielded a good amplification at one or both of the targeted genes, demonstrating the occurrence of mainly dog-specific assemblages C and D. The potentially zoonotic assemblage A and a mixed template C/D were found in two individual dogs. The results of the present study illustrate the high prevalence of G. duodenalis in shelter dogs. Although predominantly infected with dog-specific assemblages, the identification of assemblage A suggests that appropriate precautions should be taken to minimize the risk of transmission to staff.