# Probing Spatiotemporal Fractionation on the Preclinical Level Master Thesis

Telarovic, Irma. Probing Spatiotemporal Fractionation on the Preclinical Level Master Thesis. 2018, University of Zurich, Faculty of Medicine.

## Abstract

Objective
In contrast to conventional radiotherapy, spatiotemporally fractionated (STF) treatments deliver a distinct dose distribution in each fraction. The aim is to increase the therapeutic window by simultaneously achieving partial hypofractionation in the tumor along with near uniform fractionation in normal tissues. This approach has previously been studied in silico based on the working hypothesis that different parts of the tumor can be treated in different fractions, as long as the cumulative biological dose that each part receives corresponds to the prescribed dose. In this thesis, an initial step is taken towards verifying this assumption in an experimental animal model.
Methods
To perform partial tumor irradiation in a small animal model, first a pipeline was developed combining a dedicated small animal image-guided research platform (X-RAD SmART/SmART-Plan, Precision X-Ray Inc.) with the clinical software MIM (MIM Software Inc.). The tumor growth delay of conventionally treated xenografts was then compared to a reductionistic preclinical model of STF. Upon reaching the tumor volume of 300 mm3 $\pm$ 10%, animals were successively assigned to four treatment groups. The "No IR" group was sham irradiated. The "Half IR" group was irradiated with a dose of 12 Gy covering only one half of the tumor. The "STF IR" group received two partial irradiations separated by 24 hours, delivering 12 Gy each. The "Full IR" group received two partial irradiations delivering 12 Gy each as in the "STF IR" group, but applied successively.
Tumor volumes were determined by daily caliper measurements and CT-based volumetry. Additionally, an immunohistological study was undertaken to investigate the DNA damage- and tumor microenvironment-related endpoints in the broader context of partial tumor irradiation.
Results
Tumors irradiated to the same dose, either immediately or with a 24 hour delay between two partial irradiations, exhibited no differences in the growth delay study. A reduction in the irradiated volume resulted in an intermediate response. CT-based volumetry measured overall significantly smaller starting volumes with an increased dispersion compared to the caliper-based volumetry, while the relative tumor growth curves (each point normalized to the corresponding initial volume) did not differ between the two methods. On the histological level, in tumors that received the first partial irradiation 24 hours prior to the second partial irradiation, there was a decrease in the incidence of double-strand DNA breaks in cells covered by the second irradiation. An increase in the microvessel density was found in partially irradiated tumors compared to other treatment
groups.
Conclusion
The assumption of STF that the tumor response can be predicted by locally adding up biological doses from each fraction is supported in a reductionistic preclinical model whereby xenografts irradiated either immediately or with a 24 hour delay between two partial irradiations exhibited no difference in the growth delay study. Furthermore, the study suggests caliper- and CT-based volumetry to be interchangeable when considering relative, but not absolute volumes in tumor growth delay studies. Finally, the investigation of the effects of STF on the histological level suggests possible differences in the response of tumors to partial irradiation.

## Abstract

Objective
In contrast to conventional radiotherapy, spatiotemporally fractionated (STF) treatments deliver a distinct dose distribution in each fraction. The aim is to increase the therapeutic window by simultaneously achieving partial hypofractionation in the tumor along with near uniform fractionation in normal tissues. This approach has previously been studied in silico based on the working hypothesis that different parts of the tumor can be treated in different fractions, as long as the cumulative biological dose that each part receives corresponds to the prescribed dose. In this thesis, an initial step is taken towards verifying this assumption in an experimental animal model.
Methods
To perform partial tumor irradiation in a small animal model, first a pipeline was developed combining a dedicated small animal image-guided research platform (X-RAD SmART/SmART-Plan, Precision X-Ray Inc.) with the clinical software MIM (MIM Software Inc.). The tumor growth delay of conventionally treated xenografts was then compared to a reductionistic preclinical model of STF. Upon reaching the tumor volume of 300 mm3 $\pm$ 10%, animals were successively assigned to four treatment groups. The "No IR" group was sham irradiated. The "Half IR" group was irradiated with a dose of 12 Gy covering only one half of the tumor. The "STF IR" group received two partial irradiations separated by 24 hours, delivering 12 Gy each. The "Full IR" group received two partial irradiations delivering 12 Gy each as in the "STF IR" group, but applied successively.
Tumor volumes were determined by daily caliper measurements and CT-based volumetry. Additionally, an immunohistological study was undertaken to investigate the DNA damage- and tumor microenvironment-related endpoints in the broader context of partial tumor irradiation.
Results
Tumors irradiated to the same dose, either immediately or with a 24 hour delay between two partial irradiations, exhibited no differences in the growth delay study. A reduction in the irradiated volume resulted in an intermediate response. CT-based volumetry measured overall significantly smaller starting volumes with an increased dispersion compared to the caliper-based volumetry, while the relative tumor growth curves (each point normalized to the corresponding initial volume) did not differ between the two methods. On the histological level, in tumors that received the first partial irradiation 24 hours prior to the second partial irradiation, there was a decrease in the incidence of double-strand DNA breaks in cells covered by the second irradiation. An increase in the microvessel density was found in partially irradiated tumors compared to other treatment
groups.
Conclusion
The assumption of STF that the tumor response can be predicted by locally adding up biological doses from each fraction is supported in a reductionistic preclinical model whereby xenografts irradiated either immediately or with a 24 hour delay between two partial irradiations exhibited no difference in the growth delay study. Furthermore, the study suggests caliper- and CT-based volumetry to be interchangeable when considering relative, but not absolute volumes in tumor growth delay studies. Finally, the investigation of the effects of STF on the histological level suggests possible differences in the response of tumors to partial irradiation.