Normalising tumour vessels aids therapy

Chemotherapy drugs often never reach the tumours they're intended to treat, and radiation therapy is not always effective, because the blood vessels feeding the tumours are abnormal-"leaky and twisty" in the words of the late Judah Folkman, MD, founder of the Vascular Biology program at Children's Hospital Boston. Now, Vascular Biology researchers have discovered an explanation for these abnormalities that could, down the road, improve chemotherapy drug delivery. Their findings were published in the August 12 issue of the Proceedings of the National Academy of Sciences.

A tumour's capillaries-small blood vessels that directly deliver oxygen and nutrients to cancer cells-are irregularly shaped, being excessively thin in some areas and forming thick, snarly clumps in others. These malformations create a turbulent, uneven blood flow, so that too much blood goes to one region of the tumour, and too little to another. In addition, the capillary endothelial cells lining the inner surface of tumour capillaries, normally a smooth, tightly-packed sheet, have gaps between them, causing vessel leakiness.

"These abnormal features of tumour vessels impair delivery of circulating chemotherapeutic drugs to the actual tumour site" says Kaustabh Ghosh, PhD, first author on the paper, and a postdoctoral fellow in the laboratory of Donald Ingber, MD, PhD, the paper's senior author and interim co-director of the Vascular Biology program.

The idea of a therapy aimed at normalizing a tumour's blood vessels, to ensure that chemotherapeutic agents reach the tumour, has already been explored, but these attempts have only targeted soluble factors, particularly vascular endothelial growth factor (VEGF). Tumours secrete VEGF in abundance; it not only promotes blood vessel growth (angiogenesis), but makes them leaky. While blocking VEGF action helps reduce leakiness and improves vessel function, the effects have been transient, Ghosh says.

Ghosh and Ingber took a different approach, focusing on the role of mechanical forces on tumour blood vessels, which had previously been ignored. Past studies by Ingber and colleagues have shown that a capillary cell's sensitivity to soluble angiogenic factors like VEGF-and subsequent blood vessel formation-are determined by the mechanical balance between the cell's internal state of tension or contraction, and that of the surrounding support structure, or matrix, to which the cell adheres. These forces guide normal vascular pattern formation. Because tumour vessels are malformed, Ghosh wondered whether tumour capillary cells have lost the normal cells' ability to sense and respond to changes in matrix stiffness and distortion.

To address this question, the researchers studied capillary cells isolated from mice prostate tumours, provided by Andrew Dudley, PhD, in the lab of Michael Klagsbrun, PhD, in the Vascular Biology Program, and exposed them to cyclic mechanical stress-mimicking the pulsatile nature of blood flow and matrix distortion resulting from rhythmic heart beats. They found that normal capillary cells aligned themselves uniformly perpendicular to the force direction, but most of the tumour capillary cells failed to reorient, says Ghosh. These cells were "all over the place," and due to this lack of alignment, gaps appeared between neighbouring cells, which may explain the increased vessel permeability.

Ghosh and colleagues also found that tumour capillary cells sense and respond to matrix rigidity differently than normal cells. When placed on a stiff surface, mimicking the tumour matrix, the cells tended to keep spreading even after normal capillary cells stopped doing so. Because of these differences in "mechanosensing," the tumour capillary cells were able to form capillaries even when cell densities were very low, while normal cells failed to do so. At higher cell densities, normal cells formed nice capillaries, whereas the tumour cells balled up into tangled clumps, creating the irregular patterns seen in many images of tumour blood vessels. "Because high cell density increases contractility across the entire cell layer, these findings suggested that tumour capillary cells are inherently hyper-contractile," says Ghosh.

The researchers went on to find that this hyper-contractility results from an increase in the levels of a protein called Rho-associated kinase (ROCK), which controls tension within the cell. When they treated tumour capillary cells with an inhibitor of ROCK, they normalised the behaviour of the tumour capillary cells, so that the treated cells exhibited near-normal mechanical responses and formed more regularly-shaped tubular vessels.

"In this study, we've uncovered a previously unrecognised role for tumour capillary cell mechanosensing and contractility in the formation of irregular tumour vessels, and have identified potential new targets for vascular normalisation therapy that might be implemented in the clinic someday," Ghosh says.

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