Tumor Control Dose Research Articles

Feasibility Study of Real-Time Tumor-Tracking Radiotherapy for Adrenal Tumors: Three-Dimensional Movement of Internal Fiducial Gold Markers Measured in Supine and Prone Patient Positions

In the management of primary adrenal tumors, similar to pituitary tumors, radiotherapy could be an alternative treatment to surgery provided that the accuracy of irradiation is improved. Rigorous accounting of organ motion may be necessary for the accurate radiotherapy of the adrenal gland in order to give it adequate dose for tumor control. However, there has been no data about the organ motion of the adrenal gland. We have developed a method of inserting a fiducial marker near the adrenal gland and have used a real-time tumor-tracking radiotherapy (RTRT) system to investigate the three-dimensional movement of markers near adrenal tumors in supine and prone patient-positions.

Radiation Therapy Plus Angiogenesis Inhibition with Bevacizumab: Rationale and Initial Experience

Angiogenesis inhibition by monoclonal antibodies against vascular endothelial growth factor receptor (VEGFR) combined with cytotoxic chemotherapy has shown encouraging potential for improvement of cancer treatment. Several rationales exist for combining VEGFR antibodies with ionizing radiation, a primary curative cancer treatment, either in bimodal or trimodal fashion, i.e. with or without additional chemotherapy. This systematic review compares the results of preclinical and clinical studies published before December 2006. The combination of VEGFR inhibitors with irradiation in animal models consistently resulted in improved tumor growth delay (at least additive effects), despite different treatment schedules. Only one study evaluated tumor control dose (TCD)(50) as a measure of tumor cure (radiation dose yielding permanent local control in 50% of the tumors). Also in this setting, anti-VEGFR antibody treatment improved the outcome. Importantly, both radiotherapy schedule and sequence of the modalities in combined treatment may impact on the outcome. Hence, further preclinical studies examining these parameters need to be conducted. While preclinical research is ongoing, phase I and II clinical trials with bevacizumab, usually combined with radio- and chemotherapy, have been designed. Early results suggest that acute toxicity is acceptable, planned surgery after such treatment is feasible, and that further evaluation of such combined modality treatment is warranted.

Do the Dietary Supplements Epigallocatechin Gallate or Vitamin E Cause a Radiomodifying Response on Tumors in Vivo? A Pilot Study with Murine Breast Carcinoma

The primary aim of this study was to evaluate a systematic and reproducible assay to examine the potential radiomodifying effects of vitamin E (VE) or epigallocatechin gallate (EGCG), antioxidants commonly consumed by cancer patients as dietary supplements, on tumor control. C3H mice were randomized to a control diet or to the control diet supplemented with VE or EGCG. A tumor control dose 50% (TCD(50)) assay was used to evaluate for a radiomodifying response in stage IV murine cancer (MCa-IV) tumors, implanted in the hindleg of mice, and allowed to grow to 8 mm before receiving a single dose of radiation. The effects of VE and EGCG on intratumoral angiogenesis and apoptosis were evaluated in a group of nonirradiated mice using immunohistochemical staining. Cell proliferation assays were conducted on MCa-IV tumors in vitro. EGCG slowed tumor growth rate by 10%. EGCG and VE slowed tumor regrowth by 24 to 25%. There were no significant differences in TCD(50) values between the groups (control = 73.9 Gy, VE = 77.2 Gy, EGCG = 76.4 Gy); however, normal tissues were protected from late radiation effects (autoamputations) in the VE group. VE and EGCG increased tumor cell apoptosis and decreased tumor cell proliferation but had no effect on microvessel density. In this pilot study, neither VE nor EGCG exerted a significant radiomodifying effect on the MCa-IV tumor. Nonetheless, the suggestion of a small degree of tumor radioprotection by these antioxidant compounds warrants further research. As supplementation with VE radioprotected normal tissue, additional studies on this putative benefit are recommended.

Response of U87 glioma xenografts treated with concurrent rapamycin and fractionated radiotherapy: Possible role for thrombosis

Background and purpose Rapamycin, a highly specific mTOR inhibitor, has shown anti-proliferative and anti-angiogenic properties, as well as an enhancement in tumour growth delay when used in combination with radiation in mouse xenograft models. Our goal was to determine if rapamycin can also have a positive effect on the local tumour control achieved by radiotherapy. Materials and methods Nude mice bearing U87 glioblastoma xenografts were treated with concomitant rapamycin and radiotherapy over a 5 day fractionation schedule. Animals received graded total doses ranging from 24 to 100 Gy. Experimental endpoints were tumour growth delay and local tumour control. In addition, histological evaluation of tumour sections was performed to examine changes occurring within the tumour microenvironment as a result of treatment. Analysis of proliferation, mTOR signalling, hypoxia, and vessel thrombosis was conducted. Results As a single agent, rapamycin reduced the in vitro growth of U87 cells by 70% and caused a 4 day growth delay of tumour xenografts. In combination with radiation, no further increase in tumour growth delay was observed when compared to radiation alone. The tumour control dose 50% (TCD 50) was 46.8 Gy (95% CI 41; 53 Gy) in tumours treated with radiation alone and was slightly but not significantly lower at 42.8 Gy (95% CI 36; 49 Gy) after simultaneous treatment with rapamycin. Histological evaluation revealed evidence of elevated hypoxia following rapamycin treatment that may be due to vessel thrombosis. Conclusions The influence of rapamycin on thrombosis and tumour hypoxia may be a confounding factor limiting its effectiveness in combination with radiotherapy.

Preclinical safety and antitumor efficacy of insulin combined with irradiation

Background and purpose We have previously reported that insulin significantly enhances tumor oxygenation ( pO 2) and increases radiation-induced tumor regrowth delay in experimental models. Considering the large radiosensitizing effect, clinical trials might be envisioned. The aim of the present pre-clinical study was to obtain a more complete set of safety and efficacy data which would further justify the commencement of such clinical trials. Material and methods Toxicity on normal (early and late-responding) tissues was measured by the intestinal crypt regeneration assay and the late leg contracture assay. Efficacy in terms of enhancement of pO 2 (measured by in vivo EPR oximetry) and increase in radiation-induced tumor regrowth delay was evaluated with a dose–response study on mice bearing FSaII fibrosarcoma. Results The effect on regrowth delay was directly correlated with the effect on the tumor pO 2, with a maximal effect using 400 mU kg −1 insulin. Importantly, there was no increase in the radiation toxicity for normal tissues. Finally, we found that the hypoglycaemia induced by insulin can be corrected by simultaneous glucose infusion without modification of efficacy. Conclusion Insulin here demonstrated a therapeutic gain and a lack of toxicity to normal tissues. The results of this study fully justify further larger preclinical assays such as the use of fractionated irradiation and a tumor control dose assay, before determining the utility of insulin as a radiosensitizer for human patients in the clinic.

SU‐FF‐T‐105: Assessing Influence of Motion in Non‐Gated and Gated DMLC‐IMRT Delivery Using Measured Fluence Fields for Dose Reconstruction

Purpose: Recalculating dose distributions using measured IMRT fluence fields imported into the treatment planning system (TPS) to evaluate breathing synchronized irradiation. Method and Materials: DMLC‐IMRT fluence patterns acquired on radiographic film, generated in non‐gated and gated mode, have been imported into the TPS. The effect of dose blurring and the efficacy of a breathing synchronized irradiation technique have been evaluated using radiographic film mounted to a phantom simulating a breathing pattern of 16 cycles/min and 4cm amplitude. Two situations have been investigated: (a) A spherical lesion located close to the diaphragm assessing the influence of motion on the dose to the target volume and the gastro‐intestinal tract. (b) A mediastinal lesion requiring complicated fluence patterns. Results: Disturbed dose reconstructions have been observed in case of the non‐gated delivery with the phantom in motion (both orthogonal and parallel to the leaf direction), whereas the measurements from the static and gated deliveries showed good agreement with the theoretical dose distribution. These findings were confirmed by dose‐volume histograms, tumor control probabilities (comparable for the original, static and gated measurements; reduced with a factor 2 for the in‐motion‐non‐gated delivery), conformity index, and dose heterogeneity values (increased with a factor 3 to 6 when motion was induced; comparable values between the theoretical, static and gated situations). The normal tissue complication probability was affected to a lesser degree. The breathing synchronization technique introduced an increased treatment time by a factor 3 to 4. Conclusion: The use of measured fluence fields, delivered in non‐gated and gated mode, in the TPS is an interesting QA‐tool to assess the clinical impact of dose blurring, as well as the potential of breathing synchronization to resolve this issue. Conflict of Interest: Part of this work has been supported by BrainLAB AG.

Importing measured field fluences into the treatment planning system to validate a breathing synchronized DMLC–IMRT irradiation technique

Background and purpose Recalculating dose distributions using measured IMRT fluence fields imported into the treatment planning system (TPS) to evaluate the technical feasibility of a prototype developed for breathing synchronized irradiation. Patients and methods DMLC–IMRT fluence patterns acquired on radiographic film, generated by the linac in non-gated and gated mode, have been imported into the TPS. The effect of dose blurring and possible interplay between organ motion and leaf motion, and the efficacy of a breathing synchronized irradiation technique (an adapted version of a commercially available image-guidance system: NOVALIS BODY/ExacTrac4.0, BrainLAB AG) have been evaluated using radiographic film mounted to a simple phantom simulating a breathing pattern of 16 cycles per minute and covering a distance of 4 cm to obtain the resulting fluence maps. Two situations have been investigated to illustrate this principle: (a) a tumor located close to the diaphragm to assess the influence of organ motion on the dose to the target volume as well as to the gastro-intestinal tract that presents a high risk at intersecting with the beam during the breathing cycle. (b) A mediastinal lesion requiring complicated fluence patterns. Results Importing measured fluence maps yielded highly disturbed reconstructed dose distributions in case of the non-gated delivery with the phantom in motion (both orthogonal and parallel to the leaf direction), whereas the measurements from the static (film fixed in space) and the gated delivery showed good agreement with the original theoretical dose distribution. These findings have been confirmed by the dose–volume histograms, corresponding tumor control probabilities, conformity index and dose heterogeneity values. The normal tissue complication probabilities investigated in this study seem to be affected to a lesser degree, which concurs with the observation that the motion effects result in a dose spread in the direction of motion. The applied breathing synchronization technique introduced an increased treatment time with a factor 3–4. Conclusions The use of measured fluence fields, delivered by the linac in non-gated and gated mode, as imported fluence maps for the treatment planning system is an interesting quality assurance tool and revealed the dramatic impact of dose blurring and interplay between DMLC–IMRT dose delivery and organ motion, as well as the potential of breathing synchronization to resolve this issue. The possible advantage of breathing synchronized irradiation is compromised with an increased treatment time.

1359 ORAL Radiation response of parotid glands examined by dynamic 11C-methionine PET

To correlate non-invasively acquired radiobiologically relevant magnetic resonance (MR) parameters with functional histology and tumor control doses (TCD50).The MR parameters relative perfusion, re-oxygenation and lactate (Lac) concentration from eight human xenograft squamous tumor lines were compared with the histologically acquired pimonidazole hypoxic fraction, the perfused vessel area and TCD50.Good spatial correlation in the parameter maps could be observed between the pimonidazole staining and tumor regions, which can be reoxygenated when breathing carbogen. A strong positive correlation (R = 0.74) was found between whole tumor pimonidazole hypoxic fraction and re-oxygenation, as one would expect. A good correlation was also observed between Lac concentration and re-oxygenation (R = 0.71) and between TCD50 and re-oxygenation (R = 0.64), whereas Lac and TCD50 showed a moderate relation (R = 0.44). The in vivo measurement of relative perfusion could be validated to reflect the perfused vessel area (R = 0.63). No correlation was detected between perfusion and re-oxygenation or TCD50.Lac and re-oxygenation were shown to be pretreatment predictive markers independent from the pathophysiological changes induced during a fractionated course of radiotherapy. These parameters hold promise to be acquired non-invasively with results just a few minutes after measurement and to tailor radiotherapy to individual patterns of a tumor microenvironment.

Decreased repopulation as well as increased reoxygenation contribute to the improvement in local control after targeting of the EGFR by C225 during fractionated irradiation

Background and purpose Inhibition of repopulation and enhanced reoxygenation has been suggested to contribute to improvement of local tumour control after fractionated irradiation combined with inhibitors of the epidermal growth factor receptor (EGFR). The present study addresses this hypothesis in FaDu human squamous cell carcinoma. For this tumour model marked repopulation and incomplete reoxygenation during fractionated irradiation has previously been demonstrated. Furthermore, the anti-EGFR monoclonal antibody C225 has been shown to significantly improve the results of fractionated irradiation in this tumour. Materials and methods FaDu tumours in nude mice were irradiated with 18 fractions in 18 days (18f/18d) or 18 fractions in 36 days (18f/36d). Three Gy fractions were given either under ambient or under clamp hypoxic conditions. C225 or carrier was applied four times during the course of treatment. Fractionated irradiations were followed by graded top-up doses to obtain complete dose–response curves for local tumour control. Tumour control dose 50% (TCD 50) was determined at day 120 after end of treatment. Results Significant repopulation and reoxygenation occurred during fractionated irradiation of FaDu tumours ( P-values between 0.028 and <0.001). Application of C225 significantly decreased TCD 50 for 18f/36d under ambient conditions ( P=0.04). Bootstrap analysis revealed decreased repopulation and increased reoxygenation after application of C225 ( P=0.06 for the combined effect). This was further corroborated by a significant effect of C225 on the ‘repopulated’ dose under ambient conditions which is influenced by both, reoxygenation and repopulation ( P=0.012). Conclusions Our study provides evidence that both decreased repopulation as well as increased reoxygenation contribute to the improvement of local control after targeting of EGFR by C225 during fractionated irradiation of FaDu tumours.

Impact of adjuvant inhibition of vascular endothelial growth factor receptor tyrosine kinases on tumor growth delay and local tumor control after fractionated irradiation in human squamous cell carcinomas in nude mice

Previous experiments have shown that adjuvant inhibition of the vascular endothelial growth factor receptor after fractionated irradiation prolonged tumor growth delay and may also improve local tumor control. To test the latter hypothesis, local tumor control experiments were performed. Human FaDu and UT-SCC-14 squamous cell carcinomas were studied in nude mice. The vascular endothelial growth factor receptor tyrosine kinase inhibitor PTK787/ZK222584 (50 mg/kg body weight b.i.d.) was administered for 75 days after irradiation with 30 fractions within 6 weeks. Tumor growth time and tumor control dose 50% (TCD(50)) were determined and compared to controls (carrier without PTK787/ZK222584). Adjuvant administration of PTK787/ZK222584 significantly prolonged tumor growth time to reach 5 times the volume at start of drug treatment by an average of 11 days (95% confidence interval 0.06;22) in FaDu tumors and 29 days (0.6;58) in UT-SCC-14 tumors. In both tumor models, TCD(50) values were not statistically significantly different between the groups treated with PTK787/ZK222584 compared to controls. Long-term inhibition of angiogenesis after radiotherapy significantly reduced the growth rate of local recurrences but did not improve local tumor control. This indicates that recurrences after irradiation depend on vascular endothelial growth factor-driven angiogenesis, but surviving tumor cells retain their clonogenic potential during adjuvant antiangiogenic treatment with PTK787/ZK222584.

Different classes of EGFR inhibitors may have different potential to improve local tumour control after fractionated irradiation: a study on C225 in FaDu hSCC

Background and purposePrevious experiments reported from this laboratory have shown that simultaneous application of the selective epidermal growth factor receptor tyrosine kinase (EGFR-TK) inhibitor BIBX1382BS during fractionated irradiation significantly prolonged growth delay of FaDu human squamous cell carcinoma but did not improve local tumour control. The present study investigates the effect of the EGFR monoclonal antibody (mAb) C225 on local tumour control of FaDu tumours after combined treatment with single dose and fractionated irradiation to address whether different classes of EGFR inhibitors have different potential to improve the outcome of radiotherapy in the same tumour model. Material and methodsIn unirradiated tumours, C225 was given either once or 4 times i.p. to the nude mice. Irradiation experiments were performed with graded single doses under clamp hypoxic conditions or with 30 fractions in 6 weeks with graded total doses under ambient blood flow. C225 was given 6h before or 6h before and 2, 5 and 7 days after single dose irradiation. During fractionated irradiation C225 was given once per week. Experimental endpoints were tumour growth delay and local tumour control 120 after end of irradiation. ResultsC225 treatment resulted in prolongation of tumour growth delay after drug treatment alone as well as after single dose and fractionated irradiation. TCD50 values were reduced from 56.3Gy [95% CI 50; 62Gy] after single dose irradiation alone to 46.0Gy [41;51] (enhancement ratio [ER]=1.22, P<0.01) after 1 C225 injection and 47.7Gy [44; 51] after 4 injections of the drug (ER=1.18, P=0.06). After fractionated irradiation, tumour control dose 50% (TCD50) was 73.0Gy [64; 82] in control tumours and 63.1Gy [57; 69] after simultaneous C225 treatment, corresponding to an ER of 1.2 (P=0.01). ConclusionTreatment of FaDu hSCC with the anti-EGFR mAb C225 resulted in a significant prolongation of tumour growth delay after single dose and fractionated irradiation. In contrast to previous results on the EGFR-TK inhibitor BIBX1382BS, this prolongation of growth delay translated into a slight but significant improvement of local tumour control. The data indicate that different classes of EGFR inhibitors may have different potential to improve the outcome of radiotherapy in the same tumour model.

Recovery from sublethal damage during fractionated irradiation of human FaDu SCC

Background and purpose The present study addresses whether recovery of sublethal damage in tumours may change during fractionated irradiation in FaDu human squamous cell carcinoma and whether such an effect might contribute to the pronounced time factor of fractionated irradiation previously found in this tumour. Patients and methods FaDu tumours were transplanted s.c. into the right hind leg of NMRI nu/nu mice. Single doses or 2, 4, and 8 equal fractions in 3.5 days were applied in previously unirradiated tumours and after priming with 18 fractions of 3 Gy in 18 or 36 days. All irradiations were given under clamp hypoxic conditions. Experimental endpoints were tumour control dose 50% (TCD 50) and α/ β values without and after priming. Results Without priming TCD 50 increased with increasing number of fractions from 38.8 Gy (95% CI 35;45) after single dose irradiation to 54.0 Gy (42;57) after 8 fractions. No increase in TCD 50 when given in 1, 2, 4, or 8 fractions in 3.5 days was found after priming with 18 3-Gy fractions in 18 and 36 days. After priming with 18 fractions in 18 days TCD 50 remained constant at 25 Gy and after priming with 18 fractions in 36 days at 42 Gy. The α/ β ratio without priming was 68 Gy (42;127). After fractionated irradiation with 18 3-Gy fractions in 18 and 36 days the α/ β ratio increased to 317 Gy (38;∞) and to infinite, respectively. Conclusions Our results indicate that clonogenic cells in FaDu tumours lose entirely their capacity to recover from sublethal radiation damage during fractionated irradiation. Therefore, an increased repair capacity as an explanation for the pronounced time factor of fractionated irradiation in this tumour can be ruled out.

Radiomodifying responses of vitamin E and epigallocatechin gallate supplementation during irradiation of implanted tumors

9601 Background: Dietary antioxidants may play an antagonistic role during radiation treatment (RT). Antioxidants such as vitamins C and E show radioprotective effects on normal cells in vitro but limited evidence in this regard is available in healthy tissue or tumor cells in vivo. Methods: Syngeneic, murine breast carcinoma (MCa-4) cells were implanted in hind legs of C3H mice. Vitamin E (VE) or epigallocatechin gallate (EGCG, derived from green tea extract) were fed at 500 and 658 mg/kg diet, respectively, for 4 wk until tumors reached 8 mm. Three tumor control dose 50 (TCD50) assays (N=48 mice/assay) were performed to quantify the radiomodifying effect of the antioxidants. Tumor growth data were collected prior to and after RT. Tissue concentrations of VE and EGCG were analyzed by HPLC. Tissue reactions were recorded for 90 d following RT. Angiogenesis specific gene arrays were used to analyze expression in tumor and cultured tumor cells. Results: A 4% increase in TCD50 was observed in mice fed either VE or EGCG supplemented diets relative to the control diet, though these changes were not statistically significant (P=0.30 and 0.20, respectively). The gamma-50 (slope) was 6.2 for control and 3.5 for both experimental arms (P=0.20). Prior to RT, tumor growth was 10% slower in the EGCG group compared to controls (P=0.002). VE concentration in the tumor was twice that of surrounding muscle (382 vs. 189 pmole/mg) but only slightly (4%) slowed tumor growth (P=0.24) prior to RT. There were fewer RT-related soft-tissue reactions (loss of irradiated limb below the tumor) in the EGCG (9.8%) and VE (7.1%) groups compared to control (23.8%) at P=0.09 and 0.04, respectively. EGCG increased VEGF and MMP gene expression in stromal tissue but decreased it in the tumor. Conclusions: TCD50 and gamma-50 were modestly increased in mice fed diets supplemented with VE or EGCG, but these changes were not statistically significant. EGCG significantly inhibited tumor growth rate. Adverse RT-related soft tissue reactions occurred less frequently with antioxidant supplementation, particularly with VE. Author Disclosure Employment or Leadership Consultant or Advisory Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration Harvard Medical School/Osher Institute; Goldman Philanthropic Partnerships

Radiomodifying responses of vitamin E and epigallocatechin gallate supplementation during irradiation of implanted tumors

9601 Background: Dietary antioxidants may play an antagonistic role during radiation treatment (RT). Antioxidants such as vitamins C and E show radioprotective effects on normal cells in vitro but limited evidence in this regard is available in healthy tissue or tumor cells in vivo. Methods: Syngeneic, murine breast carcinoma (MCa-4) cells were implanted in hind legs of C3H mice. Vitamin E (VE) or epigallocatechin gallate (EGCG, derived from green tea extract) were fed at 500 and 658 mg/kg diet, respectively, for 4 wk until tumors reached 8 mm. Three tumor control dose 50 (TCD50) assays (N=48 mice/assay) were performed to quantify the radiomodifying effect of the antioxidants. Tumor growth data were collected prior to and after RT. Tissue concentrations of VE and EGCG were analyzed by HPLC. Tissue reactions were recorded for 90 d following RT. Angiogenesis specific gene arrays were used to analyze expression in tumor and cultured tumor cells. Results: A 4% increase in TCD50 was observed in mice fed either...

Repopulation of moderately well-differentiated and keratinizing GL human squamous cell carcinomas growing in nude mice

Purpose It has been suggested that, reminiscent of the regulated proliferative response of normal squamous epithelium, squamous cell carcinomas that have preserved characteristics of differentiation have a greater repopulation capacity during fractionated irradiation than undifferentiated tumors. The aim of the present study was to investigate repopulation in moderately well-differentiated and keratinizing GL human squamous cell carcinomas in nude mice. Methods and materials GL human squamous cell carcinomas were transplanted s.c. into the right hind leg of NMRI nu/nu mice. Irradiation was performed with 5.4 Gy fractions under clamp hypoxia or with 2 Gy fractions under ambient conditions. Six, 12, or 18 fractions were given daily, every second day, or every third day. Graded top-up doses were applied under clamp hypoxia to determine the tumor control dose 50% ( TCD 50). A total of 20 TCD 50 assays were performed and analyzed using maximum-likelihood techniques. Results With an increasing number of daily 5.4 Gy fractions under clamp hypoxia, the top-up TCD 50 values decreased significantly from 50.9 Gy (95% CI: 47, 54) after single doses to 0 Gy after 18 fractions. For the same number of fractions, the top-up TCD 50 increased with increasing overall treatment time. The results are consistent with a constant repopulation rate with a clonogenic doubling time ( T clon ) of 12.7 days (8.6, 16.8). Under ambient blood flow, the top-up TCD 50s for daily 2 Gy fractions decreased significantly, but were less pronounced than for 5.4 Gy fractions under clamp hypoxia. For a given number of fractions under ambient conditions, the top-up TCD 50s did not increase significantly with overall treatment time, except for irradiation with 12 fractions in 36 days compared to 12 and 24 days. The T clon value from these data was 24.0 days (11.6, 36.4). Conclusion Our data demonstrate a slow but significant rate of repopulation of clonogenic tumor cells during fractionated irradiation of GL human squamous cell carcinomas under clamp hypoxia without indication of a change of the repopulation rate during treatment. Less pronounced repopulation was observed for irradiation under ambient conditions, which might be explained by preferential survival of hypoxic and therefore nonproliferating cells. Taken together with our previous studies on poorly differentiated FaDu tumors (Petersen et al., IJROBP 2001;51:483–493), the results support important heterogeneity of kinetics and mechanisms of repopulation, in particular of the influence of the oxygenation status of surviving clonogenic cells on the repopulation rate during fractionated irradiation, in different experimental squamous cell carcinoma.

502 Repopulation of the moderately well differentiated GL human squamous cell carcinoma growing in nude mice

Purpose: To evaluate the antitumor activity of recombinant human tumor necrosis factor-alpha (rHuTNF-α) on a human glioblastoma multiforme (U87) xenograft in nude mice, and to study the effect of combining rHuTNF-α with local radiation on the tumor control probability of this tumor model.Methods and Materials: U87 xenograft was transplanted SC into the right hindleg of NCr/Sed nude mice (7–8 weeks old, male). When tumors reached a volume of about 110 mm3, mice were randomly assigned to treatment: rHuTNF-α alone compared with normal saline control; or local radiation plus rHuTNF-α vs. local radiation plus normal saline. Parameters of growth delay, volume doubling time, percentage of necrosis, and cell loss factor were used to assess teh antitumor effects of rHuTNF-α on this tumor. The TCD50 (tumor control dose 50%) was used as an endoint to determine the effect of combining rHuTNF-α with local radiation.Results: Tumor growth in mice treated with a dose of 150 μg/kg body weight rHuTNF-α, IP injection daily for 7 consecutive days, was delayed about 8 days comapred to that in controls. Tumors in the treatment group had a significantly longer volume doubling time, and were smaller in volume and more necrotic than matched tumors in control group. rHuTNF-α also induced a 2.3 times increase of cell loss factor. The administration ofthe above-mentioned dose of rHuTNF-α starting 24 h after single doses of localized irradiation under hypoxic condition, resulted in a significant reduction in TCD50 from the control value of 60.9 Gy to 5035 Gy (p < 0.01).Conclusion: rHuTNF-α exhibits an antitumor effect against U87 xnograft in nude mice, as evidence by an increased delay in tumor growth as well as cell loss factor. Also, there was an augmentation of tumor curability when given in combination with radiotherapy, resulting in a significantly lower TCD50 value in the treatment vs. the control groups.

Long-term infusions of p-boronophenylalanine for boron neutron capture therapy: evaluation using rat brain tumor and spinal cord models.

Rat 9L gliosarcoma cells infiltrating the normal brain have been shown previously to accumulate only approximately 30% as much boron as the intact tumor after administration of the boronated amino acid p-boronophenylalanine (BPA). Long-term i.v. infusions of BPA were shown previously to increase the boron content of these infiltrating tumor cells significantly. Experiments to determine whether this improved BPA distribution into infiltrating tumor cells after a long-term i.v. infusion improves tumor control after BNCT in this brain tumor model and whether it has any deleterious effects in the response of the rat spinal cord to BNCT are the subjects of the present report. BPA was administered in a fructose solution at a dose of 650 mg BPA/kg by single i.p. injection or by i.v. infusion for 2 h or 6 h, at 330 mg BPA/kg h(-1). At 1 h after the end of either the 2-h or the 6-h infusion, the CNS:blood (10)B partition ratio was 0.9:1. At 3 h after the single i.p. injection, the ratio was 0.6:1. After spinal cord irradiations, the ED(50) for myeloparesis was 14.7 +/- 0.4 Gy after i.p. administration of BPA and 12.9 +/- 0.3 Gy in rats irradiated after a 6-h i.v. infusion of BPA; these values were significantly different (P < 0.001). After irradiation with 100 kVp X rays, the ED(50) was 18.6 +/- 0.1 Gy. The boron compound biological effectiveness (CBE) factors calculated for the boron neutron capture dose component were 1.2 +/- 0.1 for the i.p. BPA administration protocol and 1.5 +/- 0.1 after irradiation using the 6-h i.v. BPA infusion protocol (P < 0.05). In the rat 9L gliosarcoma brain tumor model, the blood boron concentrations at 1 h after the end of the 2-h infusion (330 mg BPA/kg h(-1); n = 15) or after the 6-h infusion (190 mg BPA/kg h(-1); n = 13) were 18.9 +/- 2.2 microg 10B/g and 20.7 +/- 1.8 microg 10B/g, respectively. The irradiation times were adjusted individually, based on the preirradiation blood sample, to deliver a predicted 50% tumor control dose of 8.2 Gy ( approximately 30 photon-equivalent Gy) to all tumors. In the present study, the long-term survival was approximately 50% and was not significantly different between the 2-h and the 6-h infusion groups. The mode of BPA administration and the time between administration and irradiation influence the 10B partition ratio between the CNS and the blood, which in turn influences the measured CBE factor. These findings underline the need for clinical biodistribution studies to be carried out to establish 10B partition ratios as a key component in the evaluation of modified administration protocols involving BPA.

No evidence for a different magnitude of the time factor for continuously fractionated irradiation and protocols including gaps in two human squamous cell carcinoma in nude mice

Background and purpose: To study whether the magnitude of the time factor is different for continuously fractionated irradiation and for fractionation protocols including gaps. Materials and methods: Two human head and neck squamous cell carcinomas (SCCs), FaDu and GL, were transplanted subcutaneously into the right hindleg of NMRI (nu/nu) mice and irradiated with 30 fractions under ambient conditions within 2, 6 and 10 weeks. Irradiations within 6 and 10 weeks were given either as a continuous course or with a mid-course gap of 3 weeks. The end-point of the experiments was local tumor control at day 120 (FaDu) or day 180 (GL) after the end of treatment. Results: In FaDu tumors, two experimental cohorts (A, B) yielded significantly different results and were analyzed separately. In cohort A, the tumor control dose 50% ( TCD 50) increased from 37 to 89 Gy when the treatment time of continuous fractionated irradiation was extended from 2 to 10 weeks. The recovered dose/day ( D r) was 0.98 Gy (95% confidence interval, 0.72; 1.27). In cohort B, the TCD 50 increased from 35 to 63 Gy, and the D r was 0.51 Gy (0.24; 0.75). In GL tumors, the TCD 50 for continuously fractionated irradiation increased from 41 to 48 Gy. This increase was not significant, and the D r was 0.15 Gy (0; 0.30). None of the TCD 50 and D r values obtained in both tumor models for continuous irradiation vs. irradiation with a gap were significantly different. Conclusions: Prolongation of the overall treatment time of fractionated irradiation resulted in a pronounced decrease of local control in human FaDu SCC and little decrease of local control in human GL SCC. No evidence was found that the magnitude of the time factor in these tumors is different for continuous fractionation or fractionation protocols including gaps.

In vivo evaluation of a novel antitumor prodrug, 1-(2′-oxopropyl)-5-fluorouracil (OFU001), which releases 5-fluorouracil upon hypoxic irradiation

Purpose: We previously proposed that a prodrug of 5-fluorouracil (5-FU), OFU001, is activated through capturing of hydrated electrons produced by hypoxic irradiation. Because hydrated electrons are readily deactivated by oxygen, the 5-FU release occurs specifically upon hypoxic irradiation. In this study, we investigated the in vivo efficacy, pharmacokinetics, and toxicity of OFU001. Methods and Materials: Female 10-week-old C3H/He mice bearing SCCVII tumors were used. To measure release of 5-FU from OFU001 in vivo, the mice were given 100 mg/kg of OFU001 intraperitoneally and irradiated. Thereafter, 5-FU levels in the tumor and serum were measured by high-performance liquid chromatography. To evaluate in vivo efficacy, OFU001 was administered 30 min before irradiation, and radiation-potentiating effects were investigated by means of a tumor growth delay assay and a 50% tumor control dose (TCD-50) assay. The lethal dose of OFU001 was evaluated in the same mice. Results: Following administration of OFU001 and irradiation at 30 Gy, the average 5-FU levels in the tumor and serum were 179 ng/g and 83 ng/mL, respectively. Administration of OFU001 (100–200 mg/kg) to the tumor-bearing mice before a single dose of 15-Gy irradiation produced a mean tumor growth delay of 1–5 days as compared to radiation alone (although the delay was not significant). However, no additional growth delay was observed when OFU001 was combined with 5 radiation fractions of 4 Gy each. The enhancement ratio of OFU001 in the TCD-50 assay was 1.2. No mice died after administration of 0.6–1.2 g/kg of OFU001. Conclusions: OFU001 appears to work in vivo via the proposed mechanism of activation. Although the in vivo effect of this compound was not strong enough for clinical efficacy, these results should encourage further research on the development of prodrugs of more potent anticancer agents activated through the same mechanism.

Impact of the tumour bed effect on microenvironment, radiobiological hypoxia and the outcome of fractionated radiotherapy of human FaDu squamous-cell carcinoma growing in the nude mouse

Purpose : To investigate the impact of the tumour bed effect (TBE) on histological parameters of the micromilieu, radiobiological hypoxic fraction and local control after fractionated irradiation in FaDu squamous-cell carcinoma in the nude mouse. This tumour has previously shown a clear-cut TBE caused by increased necrotic cell loss at a constant cell production rate in the viable tumour compartment. Materials and methods : Human FaDu tumours were studied in the NMRI nude mouse. Tumours were transplanted either into unirradiated subcutaneous (s.c.) tissues (controls) or s.c. tissues pre-irradiated with 12.5 Gy (TBE group). In both groups we measured the volume doubling time (VDT), potential doubling time (T pot) , relative necrotic area, and in the viable tumour compartment the relative vascular area (9F1 mAb), relative hypoxic area (NITP or pimonidazole), relative perfused area (Hoechst 33342), and the perfused fraction of vasculature. The tumour control dose 50% (TCD 50), radiobiological hypoxic fraction (rHF) and dose-modifying factors (DMF) for the comparison of tumours in the TBE and control groups were determined from local tumour control data after treatment with single doses under ambient conditions or under clamp hypoxia, and after irradiation with 30 fractions under ambient conditions within 6 weeks using maximum-likelihood analysis. Results : A clear-cut TBE (VDT = 4.0 days (95%CI 2.9;4.4) for the control group versus 7.2 days (6.4;8.9) for the TBE group; p <0.0001) caused by increased necrosis (mean relative necrotic area of 12% (5;20)) versus 33% (10;41); p = 0.07) at a constant cell production rate (T pot = 2.2 days (1.4;2.3) versus 2.2 days (1.7;2.6); p = 0.30) was confirmed. Histological analysis of the micromilieu within the vital subarea revealed no systematic differences between the TBE and control groups. The rHF of 2% (0.1;27) for control tumours was lower than the 15% (95% CI 2;91) for the TBE group, but this difference was nonsignificant (p = 0.12). Compared with control tumours, the TCD 50 for irradiation under clamped hypoxia was in a statistical trend lower for tumours in the TBE group (DMF 1.11 (0.98;1.28), p = 0.09). After fractionated irradiation, tumours of the TBE group were significantly more radiosensitive (TCD 50 56.6 Gy (46;70) versus 78.7 Gy (63;100); p = 0.003). Conclusions : The results on FaDu tumours growing in pre-irradiated tissues indicate that increased necrosis caused by impairment of the vascular supply may increase the radiosensitivity of tumours treated by fractioned irradiation.