In this study, we have applied non-invasive NIRS and DW MRI to evaluate tumor blood oxygenation and necrosis induced by CA4P. NIRS showed rapid decrease in vascular oxygenation and perfusion during the first 2 h post CA4P. Induction of central necrosis was evident by increased ADC 24 h later and confirmed by histology. The result of tumor hypoxiation induced by CA4P indicates that tumors, if treated with radiation shortly after VDA will probably be more radioresistant. Indeed, this has been confirmed in a number of preclinical studies comparing the sequence and interval between radiation and VDA (5
). However, 24 h later increased Δ[HbO2]
(NIRS) and reduced pimonidazole staining with oxygen breathing indicated improved vascular perfusion and tumor oxygenation, consistent with our previous study using DCE MRI and 19
F NMR oximetry (7
). Exploiting these observations, we now demonstrate that tumor oxygen dynamics and modulation of tumor hypoxia was manifested in the therapeutic response to the combination of CA4P and radiation.
DCE MRI has been widely applied to many tumor types with respect to VDAs previously, both in animal models and the clinic (15
). NIRS utilizing endogenous hemoglobin provides a cheap, rapid and real-time monitoring of vascular oxygen dynamics in response to interventions. Recent clinical studies have demonstrated its usefulness in assessing breast cancer response to chemotherapy (16
). Few previous studies have applied NIRS to detect tumor blood oxygenation following VDA, though Sunar et al.
) showed that blood oxygen saturation decreased by 38% in mouse K1735 melanomas 1 h after CA4P treatment. However, Kragh et al.
reported that CA4P caused no change in tumor blood volume of C3H mouse mammary tumors in foot pad even at a higher dose of 250 mg/kg (19
), suggesting that differential responses to VDA may be tumor type dependent. Indeed, several reports indicate that tumor growing in mice require higher doses of CA4P to achieve vascular shutdown (20
In contrast to the robust increase in Δ[HbO2]
with oxygen inhalation before CA4P, follow-up NIRS revealed little response at 2 h and small response 24 h after CA4P. This observation coincides with our previous DCE MRI studies of 13762NF tumors that showed CA4P-induced global reduction in vascular perfusion at 2 h (~80%), which recovered partially only in tumor periphery 24 h later, while the tumor center remained low (7
). Our histological analysis of functional tumor vasculature by co-staining of vascular endothelium marker, CD31, and perfusion marker, Hoechst dye, confirmed the in vivo
MRI observations (7
). Thus, the observed hemoglobin dynamics in the current study reflected changes in vascular function induced by CA4P. Vascular shutdown at 2 h prevents oxygen delivery to the tumor, while smaller increase in Δ[HbO2]
at 24 h indicates partial recovery of vascular perfusion in the surviving rim.
Diffusion-weighted MRI, measuring the microscopic mobility of water can provide information about cellular structure, i.e.
, cell density (12
). ADC determined by DW MRI has been reported to correlate with cellular density, in which lower ADC values indicate higher cellularity, whereas necrotic tissue shows high ADC (13
). Vascular perfusion also affects ADC. Significant decrease in ADC 2 h after CA4P reflected the decreased vascular perfusion, while increase in ADC 24 h later indicated the enlargement of the central necrotic core. These observations coincide with several other studies of VDA by DW MRI. Indeed, application of various ranges of b values, as reported in Thoeny's study (21
), may facilitate differentiation of perfusion-sensitive ADC (low b values) from tissue-sensitive ADC (higher b values).
Hypoxia in solid tumors leads to resistance to radiotherapy and some anticancer drugs(6
F MRI provides quantitative measurement and has great potential for preclinical studies (8
). Our previous 19
F NMR oximetry studies have shown that the 13762NF tumors are relatively well oxygenated when small, typically having a hypoxic fraction (HF10
) less than 20% (7
). Consistent with the low hypoxic fraction, a single dose of 5 Gy radiation caused a significant growth delay, but breathing oxygen produced no additional benefits when comparing IR alone with IR + O2
. Measurement of tumor pO2
dynamics following CA4P was also conducted in our previous 19
F MRI study. Significant tumor hypoxiation was observed in 13762NF tumors with HF10 increasing from the baseline 30% to 80% 2 h post CA4P. However, oxygen breathing eliminated most hypoxic regions in the tumor periphery even though the pO2
was significantly lower than pretreatment. Pimonidazole data in the current study also indicated significantly reduced hypoxia by inhaling oxygen 24 h after CA4P (). This was further demonstrated in the radiation results showing that the most significant tumor growth delay was found in the tumors treated with CA4P followed by radiation plus oxygen 24 h later.
We believe imaging can provide an important role in developing new drugs and evaluating novel therapeutic efficacy for pre-clinical investigation and ultimately to achieve personalized medicine. The present results further demonstrate that rational combination of CA4P + IR, while breathing oxygen generated enhanced tumor response. This suggests a role for oxygen sensitive measurements in developing combined therapy and potentially optimizing treatment of patients.