Biology contribution
Differential oxygen dynamics in two diverse Dunning prostate R3327 rat tumor sublines (MAT-Lu and HI) with respect to growth and respiratory challenge

Presented in part at the Forty-seventh Annual Meeting of the Radiation Research Society, Albuquerque, NM, April 2000.
https://doi.org/10.1016/S0360-3016(02)02822-5Get rights and content

Abstract

Purpose: Since hypoxia may influence tumor response to therapy and prognosis, we have compared oxygenation of tumors known to exhibit differential growth rate and tissue differentiation.

Methods and Materials: Regional tumor oxygen tension was measured using 19F nuclear magnetic resonance echo planar imaging relaxometry of hexafluorobenzene, which provided dynamic maps with respect to respiratory intervention. Investigations used two Dunning prostate R3327 rat tumor sublines: the fast growing, highly metastatic MAT-Lu and the moderately well-differentiated, slower growing HI.

Results: Both sublines showed significantly higher oxygen tension in smaller tumors (<2 cm3) than in larger tumors (>3.5 cm3). Pooled data showed that MAT-Lu tumors exhibited greater hypoxia compared with the size-matched HI tumors (p < 0.0001). Respiratory challenge (oxygen or carbogen) produced significant increases in mean pO2 for tumors of both sublines (p < 0.0001). However, initially hypoxic regions displayed very different behavior in each subline: those in the HI tumors responded rapidly with significant elevation in pO2, while those in the MAT-Lu tumors showed little response to respiratory intervention.

Conclusions: These results concur with hypotheses that hypoxia is related to tumor growth rate and degree of differentiation. Under baseline conditions, the differences were subtle. However, response to respiratory intervention revealed highly significant differences, which, if held valid in the clinic, could have prognostic value.

Introduction

Hypoxia in solid tumors has been widely recognized as a potent factor, which leads to resistance to radiotherapy 1, 2, photodynamic therapy (3), and some anticancer drugs (1). Further, recent studies suggest that tumor hypoxia might also be associated with malignant progression in solid tumors 4, 5. Therefore, accurate measurement of tumor oxygenation, assessment of levels of hypoxia in individual tumors, and the development of effective methods to reduce the hypoxic fraction may well contribute to therapeutic outcome. Given the importance of oxygen, many techniques for monitoring oxygen tension (pO2) have been developed (6). While each method has specific attributes, many are highly invasive and impractical for longitudinal studies of specific regions of interest. Nuclear magnetic resonance (NMR) is entirely noninvasive: 31P NMR provides an indirect estimate of hypoxia based on phosphorylation potential (7), but the measured metabolic hypoxia occurs at a higher pO2 than radiobiological hypoxia, and some studies have shown a lack of correlation between high-energy phosphate metabolites and pO2 (8). Blood Oxygen Level Dependent (BOLD) contrast proton magnetic resonance imaging (MRI) provides an indication of tumor vascular oxygenation, and heterogeneity, in response to intervention, but the method does not provide pO2 values and interpretation may be complicated by flow, hence, the concept FLOOD (FLOw and Oxygenation Dependent contrast) (9).

We recently demonstrated the feasibility of measuring tumor oxygenation based on 19F NMR echo planar imaging (EPI) after direct intratumoral injection (i.t.) of hexafluorobenzene (HFB) 10, 11, for which we have chosen the acronym FREDOM (Fluorocarbon Relaxometry using Echo planar imaging for Dynamic Oxygen Mapping). This technique allows us to assess baseline pO2 at multiple locations within a tumor, and to follow dynamic changes in response to interventions. Hexafluorobenzene has many strengths as a reporter molecule; it is readily available, cheap, and nontoxic. In terms of NMR, the sixfold symmetry provides a single 19F signal offering maximum signal to noise, and the long relaxation times (T1 and T2) facilitate echo planar imaging. The spin lattice relaxation rate R1 is very sensitive to changes in pO2, but shows minimal response to variations in temperature. HFB is readily administered through a fine needle and remains at the site of administration for several hours (t1/2 typically 600 min) (12).

We have now applied the technique to investigate oxygen distribution and dynamics in two rat prostate tumor sublines exhibiting diverse characteristics. Although the baseline oxygenation of the moderately well-differentiated subline (HI) has been investigated previously using electrodes (13), we are unaware of previous investigations of oxygenation in the highly metastatic and poorly differentiated MAT-Lu subline. Furthermore, comparison of response to interventions, here respiratory challenge with oxygen and carbogen, is now established using a single technique for comparison of both sublines.

Section snippets

Methods and materials

Experiments were approved by the Institutional Animal Care and Research Advisory Committee.

Results

Histology shows distinctly different characteristics for the two sublines: the HI appears moderately well-differentiated with uniform sized tumor cells, pseudoglandular structures, and large vesicles (Fig. 1A). By comparison, the MAT-Lu appears poorly differentiated with cellular and nuclear variations in size and shape and no glandular structure (Fig. 1B). PCNA immunostaining also shows a higher proliferation rate in the MAT-Lu than the HI tumors (Fig. 1).

Hexafluorobenzene was readily

Discussion

The oxygen tension dynamics observed here demonstrate that response to gaseous intervention can be very different for sublines of a single parental tumor type. The relatively hypoxic regions of the well-differentiated HI subline responded to elevated inhaled oxygen, whereas those of the undifferentiated MAT-Lu subline did not. Tumors of a given subline behaved consistently.

In common with our previous investigations of the undifferentiated anaplastic Dunning prostate R3327-AT1 subline (VDT ∼5

Acknowledgements

We are grateful to Drs. Peter Antich and Peter Peschke for collegial support and Drs. Sophia Ran and Mark Jeffrey for technical assistance.

References (64)

  • C.S Heacock et al.

    Induction characteristics of oxygen regulated proteins

    Int J Radiat Oncol Biol Phys

    (1986)
  • B Movsas et al.

    Hypoxic regions exist in human prostate carcinoma

    Urology

    (1999)
  • R Kallman et al.

    Tumor oxygenation and reoxygenation during radiation therapyImportance in predicting tumor response

    Int J Radiat Oncol Biol Phys

    (1986)
  • B.M Fenton

    Effects of carbogen plus fractionated irradiation on KHT tumor oxygenation

    Radiother Oncol

    (1997)
  • K Hartmann et al.

    Effects of hyperbaric oxygen and normobaric carbogen on the radiation response of the rat rhabdomyosarcoma R1H

    Int J Radiat Oncol Biol Phys

    (2001)
  • C Aquino-Parsons et al.

    Oxygen tension in primary gynaecological tumoursThe influence of carbon dioxide concentration

    Radiother Oncol

    (2000)
  • J.L Lanzen et al.

    Variability in blood flow and pO2 in tumors in response to carbogen breathing

    Int J Radiat Oncol Biol Phys

    (1998)
  • D Le et al.

    Regional tumor oxygen dynamics19F PBSR EPI of hexafluorobenzene

    Magn Reson Imaging

    (1997)
  • R.P Mason et al.

    Non-invasive determination of tumor oxygen tension and local variation with growth

    Int J Radiat Oncol Biol Phys

    (1994)
  • J.M Brown

    The hypoxic cellA target for selective cancer therapy—eighteenth Bruce F. Cain memorial award lecture

    Cancer Res

    (1999)
  • E.J Hall

    The oxygen effect and reoxygenation

  • J.D Chapman et al.

    Oxygen dependency of tumor cell killing in vitro by light activated photofrin II

    Radiat Res

    (1991)
  • H Zhong et al.

    Increased expression of hypoxia inducible factor-1 alpha in rat and human prostate cancer

    Cancer Res

    (1998)
  • E.K Rofstad et al.

    Hypoxia-induced metastasis of human melanoma cellsInvolvement of vascular endothelial growth factor-mediated angiogenesis

    Br J Cancer

    (1999)
  • H.B Stone et al.

    Oxygen in human tumorsCorrelations between methods of measurement and response to therapy

    Radiat Res

    (1993)
  • H.D Sostman et al.

    Evaluation of BA 1112 rhabdomyosarcoma oxygenation with microelectrodes, optical spectrometry, radiosensitivity, and MRS

    Magn Reson Med

    (1991)
  • P Vaupel et al.

    Stable bioenergetic status despite substantial changes in blood flow and tissue oxygenation in a rat tumour

    Br J Cancer

    (1994)
  • S.P Robinson et al.

    Tumor response to hypercapnia and hyperoxia monitored by FLOOD magnetic resonance imaging

    NMR Biomed

    (1999)
  • R.P Mason et al.

    Regional tumor oxygenation and measurement of dynamic changes

    Radiat Res

    (1999)
  • R.P Mason et al.

    HexafluorobenzeneA sensitive 19F NMR indicator of tumor oxygenation

    NMR Biomed

    (1996)
  • M.J Eble et al.

    Tissue oxygen tension distribution in two sublines of the Dunning prostate tumor R3327

  • P Peschke et al.

    Differential sensitivity of three sublines of the rat Dunning prostate tumor system R3327 to radiation and/or local tumor hyperthermia

    Radiat Res

    (1998)
  • Cited by (45)

    • New frontiers and developing applications in <sup>19</sup>F NMR

      2013, Progress in Nuclear Magnetic Resonance Spectroscopy
      Citation Excerpt :

      Comparing identical tissue locations has not been feasible, but typical behavior was highly consistent. As an example, hypoxic regions in many tumors resist modulation with hyperoxic gas breathing, while well oxygenated regions show a large response (Fig. 4): matching data have been reported for 19F NMR or electrodes [236,243,250]. Meanwhile, some tumors show a large response to hyperoxic gas breathing despite baseline hypoxia and again this was seen using 19F MRI and fiber optic probes [240].

    • Preliminary Study of Oxygen-Enhanced Longitudinal Relaxation in MRI: A Potential Novel Biomarker of Oxygenation Changes in Solid Tumors

      2009, International Journal of Radiation Oncology Biology Physics
      Citation Excerpt :

      Similarly, carbogen-induced increase in 1H MRI T1-weighted signal intensity has been measured in the rat Dunning prostate R3327 model, with largest signal intensity increases were in small, well vascularized H subline tumors (34). Significantly less signal change was detected in large, poorly vascular tumors, known to be relatively hypoxic and resistant to modulation by hyperoxic gas (37). These 1H MRI findings from previous studies concur with 19F MRI studies that report increased median pO2 in H tumors but not in large AT1 subline tumors after hyperoxic gas administration (38).

    • Non Invasive Physiology and Pharmacology Using 19F Magnetic Resonance

      2008, Fluorine and Health: Molecular Imaging, Biomedical Materials and Pharmaceuticals
    View all citing articles on Scopus

    This work was supported in part by NIH RO1 CA79515, the American Cancer Society (RPG-97-116-010CCE), and a postdoctoral fellowship from the DOD Prostate Cancer Initiative (DAMD 170110108) (DZ). NMR experiments were performed at the Mary Nell and Ralph B. Rogers MR Center, an NIH BRTP Facility P41-RR02584.

    View full text