modeling-effects-hemodynamic-stress-on-circulating-tumor-cells-using

Modeling the Effects of Hemodynamic Stress on Circulating Tumor Cells using a Syringe and Needle

During metastasis, cancer cells from solid tissues, including epithelia, gain access to the lymphatic and hematogenous circulation where they are exposed ...

University of Iowa

Circadian clocks regulate membrane excitability in master pacemaker neurons to control daily rhythms of sleep and wake. Here, we find that two distinctly timed electrical drives collaborate to impose rhythmicity on Drosophila clock neurons. In the morning, a voltage-independent sodium conductance via the NA/NALCN ion channel depolarizes these neurons. This current is driven by the rhythmic expression of NCA localization factor-1, linking the molecular clock to ion channel function. In the evening, basal potassium currents peak to silence clock neurons. Remarkably, daily antiphase cycles of sodium and potassium currents also drive mouse clock neuron rhythms. Thus, we reveal an evolutionarily ancient strategy for the neural mechanisms that govern daily sleep and wake.

A Conserved Bicycle Model for Circadian Clock Control of Membrane Excitability.

The sodium leak channel NARROW ABDOMEN (NA)/ NALCN is an important component of circadian pacemaker neuronal output. In , rhythmic expression of the NA channel regulator  in a subset of adult pacemaker neurons has been proposed to contribute to circadian regulation of channel localization or activity. Here we have restricted expression of  NA channel subunits or the  regulator to either development or adulthood using the temperature-inducible -GAL80 system. Surprisingly, we find that developmental expression of endogenous channel subunits and  is sufficient to promote robust rhythmic behavior in adults. Moreover, we find that channel complex proteins produced during development persist in the  head with little decay for at least 5-7 days in adults. In contrast, restricting either endogenous or transgenic gene expression to adult stages produces only limited amounts of the functional channel complex. These data indicate that much of the NA channel complex that functions in adult circadian neurons is normally produced during development, and that the channel complex is very stable in most neurons in the  brain. Based on these findings, we propose that circadian regulation of NA channel function in adult pacemaker neurons is mediated primarily by post-translational mechanisms that are independent of 

The Narrow Abdomen Ion Channel Complex Is Highly Stable and Persists from Development into Adult Stages to Promote Behavioral Rhythmicity.

Genetic mouse models of soft tissue sarcoma provide critical insights into disease pathophysiology, which are oftentimes unable to be extracted from human tumor samples or xenograft models. In this study we describe a mouse model of soft tissue sarcoma mediated by adenoviral-Cre recombinase injection into Trp53fl/fl/Ptenfl/fl lox-stop-lox luciferase mice. Injection of adenovirus expressing Cre recombinase, either subcutaneously or intramuscularly in two experimental groups, results in viral infection and gene recombination with 100% penetrance within the first 24 hours following injection. Luciferase expression measured by real-time bioluminescence imaging increases over time, with an initial robust increase following viral injection, followed by a steady rise over the next several weeks as primary tumors develop and grow. Intramuscular injections were more commonly associated with evidence of systemic viral distribution than subcutaneous injections. All mice developed soft tissue sarcomas at the primary injection site, with histological examination identifying 93% of tumors as invasive pleomorphic sarcomas based on microscopic morphology and immunohistochemical expression of sarcoma markers. A lymphocytic infiltrate was present in 64% of the sarcomas in this immunocompetent model and 71% of tumors expressed PD-L1. This is the first report of a viral-Cre mediated Trp53/Pten mouse model of undifferentiated pleomorphic sarcoma. The bioluminescence imaging feature, along with high penetrance of the model and its immunological characteristics, makes it suited for pre-clinical studies of soft tissue sarcoma.

A Trp53fl/flPtenfl/fl mouse model of undifferentiated pleomorphic sarcoma mediated by adeno-Cre injection and in vivo bioluminescence imaging.

During metastasis, cancer cells are exposed to potentially destructive hemodynamic forces including fluid shear stress (FSS) while en route to distant sites. However, prior work indicates that cancer cells are more resistant to brief pulses of high-level FSS in vitro relative to non-transformed epithelial cells. Herein, we identify a mechano-adaptive mechanism of FSS resistance in cancer cells. Our findings demonstrate that cancer cells activate RhoA in response to FSS, which protects them from FSS-induced plasma membrane damage. We show that cancer cells freshly isolated from mouse and human tumors are resistant to FSS, that formin and myosin II activity protects circulating tumor cells (CTCs) from destruction, and that short-term inhibition of myosin II delays metastasis in mouse models. Collectively, our data indicate that viable CTCs actively resist destruction by hemodynamic forces and are likely to be more mechanically robust than is commonly thought.

Cancer Cells Resist Mechanical Destruction in Circulation via RhoA/Actomyosin-Dependent Mechano-Adaptation.

During metastasis, cancer cells traverse the circulation to reach distant organs. Conventionally, this journey has been regarded as mechanically destructive to circulating tumor cells from solid tissues. We have recently shown that cancer cells from diverse tissues actively resist destruction by fluid shear stress through a mechano-adaptive RhoA-actomyosin mechanism.

Survival of the resilient: Mechano-adaptation of circulating tumor cells to fluid shear stress.

Here we have improved an existing mouse model of prostate cancer based on prostate-specific deletion of Pten and Trp53 by incorporating a Cre-activatable luciferase reporter. By coupling the deletion of those genes to the activation of a luciferase reporter, we were able to monitor tumor burden non-invasively over time. We show that, consistent with previous reports, deletion of both Pten and Trp53 on a C57BL/6 background accelerates tumor growth and results in both the loss of androgen receptor expression and castrate resistant tumors as compared with loss of Pten alone. Loss of Trp53 results in the development of sarcomatoid histology and the expression of markers of epithelial-to-mesenchymal transition Zeb1 and vimentin, with kinetics and penetrance dependent on whether one or both alleles of Trp53 were deleted. Homozygous deletion of Trp53 and Pten resulted in uniformly lethal disease by 25 weeks. While we were able to detect locally invasive disease in the peritoneal cavity in aggressive tumors from the double knockout mice, we were unable to detect lymphatic or hematogenous metastatic disease in lymph nodes or at distant sites.

Locally invasive, castrate-resistant prostate cancer in a Pten/Trp53 double knockout mouse model of prostate cancer monitored with non-invasive bioluminescent imaging.

Pharmacological ascorbate (P-AscH, high-dose, intravenous vitamin C) is cytotoxic to tumor cells in doses achievable in humans. Phase I studies in pancreatic cancer (PDAC) utilizing P-AscH have demonstrated increases in progression free survival, suggesting a reduction in metastatic disease burden. The purpose of this study was to determine the effects of P-AscH on metastatic PDAC. Several in vitro and in vivo mechanisms involved in PDAC metastases were investigated following treatment with P-AscH. Serum from PDAC patients in clinical trials with P-AscH were tested for the presence and quantity of circulating tumor cell-derived nucleases. P-AscH inhibited invasion, basement membrane degradation, decreased matrix metalloproteinase expression, as well as clonogenic survival and viability during exposure to fluid shear stress. In vivo, P-AscH significantly decreased formation of ascites, tumor burden over time, circulating tumor cells, and hepatic metastases. Both in vitro and in vivo findings were reversed with the addition of catalase suggesting that the effect of P-AscH on metastatic disease is mediated by hydrogen peroxide. Finally, P-AscH decreased CTC-derived nucleases in subjects with stage IV PDAC in a phase I clinical trial. We conclude that P-AscH attenuates the metastatic potential of PDAC and may prove to be effective for treating advanced disease.

Devon L. Moose

Department of Molecular Physiology and Biophysics,
Carver College of Medicine,
Holden Comprehensive Cancer Center,
Department of Molecular Physiology and Biophysics, Carver College of Medicine

University of Iowa

Modeling the Effects of Hemodynamic Stress on Circulating Tumor Cells using a Syringe and Needle

Devon L. Moose

Department of Molecular Physiology and Biophysics,Carver College of Medicine,Holden Comprehensive Cancer Center,Department of Molecular Physiology and Biophysics, Carver College of Medicine

University of Iowa

Modeling the Effects of Hemodynamic Stress on Circulating Tumor Cells using a Syringe and Needle

Department of Molecular Physiology and Biophysics,
Carver College of Medicine,
Holden Comprehensive Cancer Center,
Department of Molecular Physiology and Biophysics, Carver College of Medicine