In a study published in the February issue of the journal Radiology, researchers describe a technique for determining the "leakiness" of tumour blood vessels using a simple digital mammography unit. The researchers designed nanometer-sized capsules containing a contrast agent that could only leak into tumours with blood vessels that were growing and therefore leaky. The digital mammography-based quantification of "leakiness" is closely correlated to the ability of a clinically approved chemotherapy agent to enter the tumour, allowing the researchers to predict the agent's therapeutic efficacy.
"We developed a quantitative way to measure the leakiness of the blood vessels, which is directly linked to the amount of drug that gets to the cancer and in turn determines effectiveness", stated Ravi Bellamkonda, a professor in the Wallace H. Coulter Department of Biomedical Engineering at Georgia Tech and Emory University. "By simply measuring how much contrast agent reaches the tumour, we can predict how much of a clinically approved chemotherapeutic will reach the tumour, allowing physicians to personalize the dose and predict effectiveness."
In some cases, one chemotherapy drug may not be effective in treating the tumour, but this new technique allows oncologists to investigate other drugs sooner since they know the drug is reaching the tumour. Studies are currently underway to determine if mammography can predict the optimal dose of a wide range of breast cancer chemotherapeutics.
Ravi Bellamkonda and Coulter Department postdoctoral fellow Efstathios Karathanasis collaborated on this study with Ioannis Sechopoulos, an assistant professor in radiology at Emory University; Andrew Karellas, a former professor in the Emory University Winship Cancer Institute currently at the University of Massachusetts Medical School; and Ananth Annapragada, an associate professor of health information sciences at the University of Texas, Houston. The project was funded by the National Science Foundation and Georgia Cancer Coalition.
For the study, a long-circulating nanometer-scale liposomal capsule filled with iodinated contrast agent was injected into rats with six-day-old breast cancer tumours. For the next three days, the researchers collected digital mammography images of the animals and compared the pre- and post-injection grayscale intensity values to study the dynamics of how the contrast agent accumulated in the tumor over time.
"During the three-day time course, some tumours exhibited a rapid and significant increase in image brightness, meaning the contrast agent was accumulating in the tumour, whereas other tumours showed a slow and low increase", stated Ravi Bellamkonda, who is also a Georgia Cancer Coalition Distinguished Scholar.
X-ray images showing two tumours with different levels of "leakiness". Tumour A exhibits higher grey levels than tumour B by 40 and 70 digital units at 72 and 120 hours post-injection, respectively. Photo: Courtesy of Karathanasis, F. et al. - Radiology 250(2).
While the brightness of the tumours in the images changed significantly, no variations were observed in non-tumour areas or in the tumours of animals that did not receive the contrast agent. Immediately after the imaging was completed and the leakiness of each individual cancer vessel was quantified, the animals were intravenously injected with a clinically approved chemotherapy drug, liposomal doxorubicin.
Results showed that the chemotherapeutic drug slowed the progress of the tumour. The variability in uptake of the contrast agent by the tumours, as measured during the three-day imaging sessions, provided an accurate prognosis of the effect of liposomal doxorubicin on tumour growth rate.
"When we plotted the post-treatment tumour growth rate versus the intensity of leakiness, there was a significant and strong correlation", noted Ravi Bellamkonda. "The tumours in which the nanocarrier leaked out and accumulated the most in the tumours during the initial three-day test were the ones that responded best to the treatment."
To verify that the intensity changes in the images were caused by the nanocarrier and not endogenous changes in the tumour tissue, liposomal probes tagged with a fluorescent dye were injected into the animals. By looking at histological tumour sections, the researchers showed that the location of the increased image brightness and the fluorescent dye were the same.
"This study showed that higher uptake of the probe by the tumour related to leakier vasculature and suggested a better therapeutic outcome of liposomal doxorubicin", stated Ravi Bellamkonda. "Imaging the integrity of the tumour vasculature like this may allow cancer treatment to be more patient-specific and potentially spare patients from chemotherapy if it is not going to be effective."
Fluorescent microscopy breast tumour images that demonstrate the intensity changes in the X-ray images were caused by the nanocarrier and not endogenous changes in the tumor tissue. (A) 48 hours after injecting liposomes tagged with a red fluorescent dye, the liposomes appear in a patchy distribution in the periphery of the tumour. (B) A green immunohistochemical microvascular stain reveals the tumour has a highly vascularized peripheral rim and a less vascularized inner core. (C) In the same histological slice, the nanocarriers containing red contrast agent appear localized within the well vascularized rim. Photo: Courtesy of Karathanasis, F. et al. - Radiology 250(2).
While the goal of the study reported in the journal was not to induce tumour regression, the researchers plan to investigate whether the liposomal probes can be used for this purpose in the future. To further develop and commercialize these multifunctional probes, Ravi Bellamkonda and Ananth Annapragada founded a start-up company called Marval Biosciences Inc.
The researchers also want to investigate whether the leakiness of tumour vasculature represents a parameter that is useful for clinical diagnosis or tumour characterization. "We want to study the molecular basis for blood vessel leakiness", stated Ravi Bellamkonda. "We want to understand why there is variation in leakiness and chemotherapy effectiveness among individuals with tumours of the same type, size and stage."