A technique is presented for determining the time of metastasis during fractionated radiation therapy of the rat rhabdomyosarcoma R1H by flow cytmetric DNA index measurements. The DNA index is the relative DNA content of the tumour cells compared with normal cells. The DNA index of the highly hyperploid R1H tumour decreases linearly during radiation therapy dependent on the total dose applied, with a rate of 0.22% per Gy. Since metastatic cells escape further irradiation, their DNA index should correspond to the DNA index at time of spread. If so, cells disseminated late during a course of fractionated irradiation are expected to produce symptoms later and to have a lower DNA index compared with cells that metastasized at the start of treatment. The results obtained fitted these expectations, showing that timing of metastases by DNA flow cytometry, in principle, is possible. A first application of this method indicates that the risk per clonogenic R1H tumour cell to matastasize increases during fractionated irradiation.
