Fraction of radiobiologically hypoxic cells in human melanoma xenografts measured by using single-cell survival, tumour growth delay and local tumour control as end points

Abstract

Summary Four human melanoma xenograft lines (A-07, D-12, R-18, U-25) grown orthotopically in Balb/c nu/nu mice were characterzed with respect to the fraction of radiobiologically hypoxic cells. The purpose of the study was to establish a firm radiobiological basis for future use of the lines in the development and evaluation of non-invasive assays of tumour hypoxia. The hypoxic fractions were assessed using three different assays. the single cell survival assay, the tumour growth delay assay and the local tumour control assay, and the means + s.e.were found to be 6 ± 30%, 3 ± 1 0 and 5 ± 2%o respectively (A-07), 26 ± 50/0, 25 ± 60/o and 22 + 6%o respectively (D-12), 55 ± 9°o 65 ± 80/o and 48 ± 7°O respectively (R-1 8) and 52 ± 8%, 59 ± 7% and 47 ± 7% respectively (U-25). The three assays gave numerical values for the hypoxic fraction that were not significantly different for any of the lines. The hypoxic fraction differed significantly among the lines; the R-18 and U-25 lines showed higher hypoxic fractions than the D-12 line (P < 0.05). which in tum showed a higher hypoxic fraction than the A-07 line (P < 0.05). regardless of the assay. The wide range of the hypoxic fractions and the signfficant differences among the lines suggest that A-07. D-12. R-18 and U-25 tumours should be useful models in future studies attempting to develop non-invasive assays of tumour hypoxia.Keywords: treatment-response assay; hypoxic fraction; melanoma xenograft; radiation biology Manv human tumours develop regions of hxpoxic cells during gro,wth (Vaupel et al. 1989: 1991: Hockel et al. 1991. The fraction of hxpoxic cells differs substantially among individual tumours of the same histological tIpe (Nordsmark et al. 1994: Brizel et al. 1995: Wong et al. 1997. Hypoxia may cause resistance to radiation therapy (Coleman. 1988: Gatenbv et al. 1988: Hockel et al. 1993: Nordsmark et al. 1996: Brizel et al. 1997: Fx-les et al. 1997 and some forms of chemotherapx (Teicher. 1994) and mav promote the development of metastatic disease (Schwickert et al. 1995: Brizel et al. 1996: Hockel et al. 1996: Walenta et al. 1997 March 1998 Correspondence to: EK Rofstad treatment resistance and malignant progression (Kim et al. 1993: Fenton et al. 1995

Assessment of microvascular density, extracellular volume fraction, and radiobiological hypoxia in human melanoma xenografts by dynamic contrast‐enhanced MRI

“…The A‐07 and R‐18 melanomas used in the present work were inoculated intradermally in the mouse flank and were thus growing in an orthotopic site. Orthotopic A‐07 and R‐18 tumors do not develop necrotic regions during growth before they exceed a volume of 600 mm 3 (17, 26, 27), a general observation that was confirmed in the work reported here. Consequently, the pattern of contrast enhancement seen in orthotopically growing A‐07 and R‐18 tumors differs from that of many subcutaneous experimental tumors, but is similar to that seen for many types of human cancer.…”

“…The uptake of Gd‐DTPA in tumor tissue depends on several microenvironmental parameters, including blood perfusion, microvascular density, cell density, and extracellular volume fraction (8). Xenografted A‐07 and R‐18 melanomas differ substantially in these microenvironmental parameters as well as in tumor oxygenation status (16, 27). Consequently, the A‐07 and R‐18 melanomas should be excellent models for testing the hypothesis put forward in the present study.…”

Assessment of fraction of radiobiologically hypoxic cells in human melanoma xenografts by dynamic contrast‐enhanced MRI

“…Fraction of hypoxic cells was then calculated from the cell surviving fraction. An alternative approach is to irradiate tumors with a single dose of 10 Gy, measure response to treatment as growth delay, and then calculate fraction of hypoxic cells from the tumor growth delay (24, 32). In both approaches, tumors are subjected to measurement of response to single‐dose irradiation, and fraction of hypoxic cells is calculated from the radiation response.…”

“…In both approaches, tumors are subjected to measurement of response to single‐dose irradiation, and fraction of hypoxic cells is calculated from the radiation response. In the present work, we chose to measure radiation response as cell surviving fraction rather than as tumor growth delay, primarily because studies of clamped tumors have shown that the radiation response of A‐07 tumors can be measured more accurately by using the cell survival assay than by using the tumor growth delay assay (32). Nevertheless, studies in our laboratory of human melanoma xenografts have shown that fraction of hypoxic cells as calculated from tumor growth delay data are strongly correlated to fraction of hypoxic cells as calculated from cell survival data (32).…”

True

“…By comparing the fraction of metabolically hypoxic cells to data published previously (Rofstad and Måseide, 1998) (Rofstad and Måseide, 1998) and discrepancies between the assays were observed. A-07 showed a significantly lower fraction both of metabolically hypoxic cells and of radiobiologically hypoxic cells than the other three lines ( < 0.05).…”