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Temporal characterization and in vitro comparison of cell survival following the delivery of 3Dconformal, intensity-modulated radiation therapy (IMRT) and volumetric modulated arc therapy (VMAT)
This article has been downloaded from IOPscience. Please scroll down to see the full text article. 2011 Phys. Med. Biol. 56 2445 (http://iopscience.iop.org/0031-9155/56/8/008) View the table of contents for this issue, or go to the journal homepage for more

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IOP PUBLISHING Phys. Med. Biol. 56 (2011) 24452457

PHYSICS IN MEDICINE AND BIOLOGY

doi:10.1088/0031-9155/56/8/008

Temporal characterization and in vitro comparison of cell survival following the delivery of 3D-conformal, intensity-modulated radiation therapy (IMRT) and volumetric modulated arc therapy (VMAT)
Conor K McGarry1,2 , Karl T Butterworth2 , Colman Trainor2 , Joe M OSullivan2,3 , Kevin M Prise2 and Alan R Hounsell1,2
1 Radiotherapy Physics, Northern Ireland Cancer Centre, Belfast Health and Social Care Trust, Belfast, UK 2 Centre for Cancer Research and Cell Biology, Queens University Belfast, Belfast, UK 3 Clinical Oncology, Northern Ireland Cancer Centre, Belfast Health and Social Care Trust, Belfast, UK

E-mail: conor.mcgarry@belfasttrust.hscni.net

Received 10 December 2010, in nal form 4 February 2011 Published 22 March 2011 Online at stacks.iop.org/PMB/56/2445 Abstract A phantom was designed and implemented for the delivery of treatment plans to cells in vitro. Single beam, 3D-conformal radiotherapy (3D-CRT) plans, inverse planned ve-eld intensity-modulated radiation therapy (IMRT), nineeld IMRT, single-arc volumetric modulated arc therapy (VMAT) and dual-arc VMAT plans were created on a CT scan of the phantom to deliver 3 Gy to the cell layer and veried using a Farmer chamber, 2D ionization chamber array and gafchromic lm. Each plan was delivered to a 2D ionization chamber array to assess the temporal characteristics of the plan including delivery time and cells eye view for the central ionization chamber. The effective fraction time, dened as the percentage of the fraction time where any dose is delivered to each point examined, was also assessed across 120 ionization chambers. Each plan was delivered to human prostate cancer DU-145 cells and normal primary AGO-1522b broblast cells. Uniform beams were delivered to each cell line with the delivery time varying from 0.5 to 20.54 min. Effective fraction time was found to increase with a decreasing number of beams or arcs. For a uniform beam delivery, AGO-1552b cells exhibited a statistically signicant trend towards increased survival with increased delivery time. This trend was not repeated when the different modulated clinical delivery methods were used. Less sensitive DU-145 cells did not exhibit a signicant trend towards increased survival with increased delivery time for either the uniform or clinical deliveries. These results conrm that dose rate effects are most prevalent in more radiosensitive cells. Cell survival data generated from uniform beam
0031-9155/11/082445+13$33.00 2011 Institute of Physics and Engineering in Medicine Printed in the UK 2445

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deliveries over a range of dose rates and delivery times may not always be accurate in predicting response to more complex delivery techniques, such as IMRT and VMAT. (Some gures in this article are in colour only in the electronic version)

1. Introduction Intensity-modulated radiation therapy (IMRT) can enable highly conformal doses to be delivered to a tumour volume whilst avoiding or limiting dose to critical structures. The delivery time of IMRT may be longer than conformal therapy (3D-CRT), depending on planning parameters such as the number of beams or segments (Kuperman et al 2008). Volumetric modulated arc therapy (VMAT) is a specic type of IMRT where the gantry rotates around the patient with the MLCs moving during delivery. During gantry rotation, the speed and dose rate may or may not be varied (Palma et al 2008). VMAT has shown promise for the delivery of highly conformal plans whilst reducing overall delivery time compared to conventional IMRT (Verbakel et al 2009a, Bedford 2009). Clinical outcomes predicted by historical dose volume data of cancer patients treated with radiotherapy have been typically obtained from conventional radiotherapy or 3D-CRT that may no longer be applicable for IMRT and VMAT where long term data are not yet available (Blockhuys et al 2010). Variations in the spatio-temporal dose histories of the tumour have been demonstrated between different radiotherapy delivery techniques including 3D-CRT, dynamic IMRT, static IMRT and tomotherapy (Shaikh et al 2010). Even within a single treatment fraction the dose history will vary within a tumour volume (Wang et al 2003, Sch afer et al 2005). A detailed characterization of the temporal delivery properties of advanced techniques such as VMAT has, to date, not been performed. Randomized clinical trials have yet to clearly show any adverse effects on tumour control due to IMRT delivery (Veldeman et al 2008) and therefore pre-clinical models are of importance in predicting outcomes of advanced radiotherapy techniques. A number of modelling studies have predicted increased cell survival following radiation delivery over a prolonged time associated with IMRT delivery (Fowler et al 2004, Wang et al 2003, Paganetti 2005). Recent reports have provided in vitro evidence that the temporal pattern of dose delivery may have a signicant impact on the radiobiological response of tumour cells (Sterzing et al 2005, Moiseenko et al 2007, Bewes et al 2008, Yang et al 2009, Altman et al 2009, Joiner et al 2010). A progressive increase in cell survival for increasing delivery time has been observed (Bewes et al 2008) with a potential increase in the biological effectiveness of a given dose by up to 20% for a 1 min Rapidarc R delivery, a commercial version of VMAT delivery, compared to 17 min IMRT delivery being inferred (Verbakel et al 2009b). The analysis shows that the largest temporal effects do appear for more radiosensitive cells (SF2 < 0.5) with higher / ratios although dependence of the effects on repair half times and the relative proportion of different repair processes inhibits detailed interpretation of dose rate effects based on / ratios (Paganetti 2005). Delivery times for IMRT plans can vary. Improved inverse planning systems such as those utilizing direct aperture optimization (DAO) have resulted in reduced complexity with reduced monitor units and/or reduced leaf movement resulting in reduced delivery times (Ludlum and Xia 2008). Studies investigating temporal differences between conformal delivery and IMRT delivery either assumed that the delivery was in the order of 2045 min (Joiner et al 2010) or used IMRT plans with less efcient deliveries (Sterzing et al 2005). The delivery of IMRT

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plans in more recent studies has been between 5 and 10 min (Shaikh et al 2010, Yang et al 2009, Bedford 2009, Moiseenko et al 2007). In a modelling study, Shaikh et al (2010) hypothesized that any reduced biological effect would be less for helical tomotherapy (HT) than xed eld IMRT. For HT approximately 90% of the dose was delivered over a short period of time, termed the rapid dose accumulation time (RDAT), in the middle of treatment time. With the use of this RDAT as the treatment time, ignoring the small scatter dose, it would appear that tomotherapy times are equivalent to VMAT times. However, Yang et al 2009 showed in-vitro that cell survival following tomotherapy may be closer to IMRT delivery over 7 min than 2 min continuous delivery with no MLC modulation. Therefore a simple comparison between tomotherapy and VMAT does not appear to apply. Delivery time has been cited as the major factor in differences in cell survival (Wang et al 2003, Bewes et al 2008); however, the most up to date delivery methods are yet to be studied in vitro and compared with uniform delivery over the same time. The aim of this work was to assess the variation in tumour dose history for clinically relevant inverse planned IMRT and VMAT deliveries and compare these to a conformal plan. The effect of varying the number of beams in IMRT (ve and nine beams) and the number of arcs in VMAT (single and dual arc) was also investigated. In this paper prostate cancer cell (DU-145) and normal primary AGO-1522b broblast cell survival are presented for each 3D-CRT, IMRT and VMAT treatment technique following the delivery of a clinically relevant dose (3 Gy) to a cell monolayer. To assess if the leaf sequencing has an effect on clinical delivery compared to uniform exposure, a series of single uniform beams, again for a 3 Gy dose, were delivered over a longer time range (0.520.54 min) to each cell line. 2. Methods and materials 2.1. Phantom design and construction A cell phantom was constructed of PMMA using a computer controlled (CNC) milling machine (Hass, US) with the aim of minimizing air gaps to maintain electronic equilibrium within a T80 ask (Nunc, UK) lled with culture medium. Figure 1(a) shows that the phantom has dimensions 30 30 6 cm3 with cells located 1.2 cm from the bottom of the phantom. It is modular in design to facilitate insertion and removal of culture asks. 2.2. Experimental setup and treatment planning Figure 1(b) shows the experimental setup including 10 cm thick 30 30 cm2 and 5 cm thick 30 30 cm2 Wte solid water phantom (Barts and The London NHS Trust, London, UK) below and above the phantom respectively to provide a similar thickness to a prostate patient. A CT scan was acquired of the setup using a Siemens Emotion 6 CT scanner (Siemens Medical Systems, Forcheim, Germany). Oncentra R v3.3sp1 treatment planning system (Nucletron, Veenendaal, the Netherlands) was used to outline the planning target volume (PTV), ensuring a margin of at least 1 cm around the cell layer within the ask. Two pseudo-organs-at-risk (OARs) were also outlined as avoidance structures in the planning process. All plans were created using 6 MV photon beam data from a Varian 600CD linear accelerator (Varian Medical Systems, Palo Alto, CA, USA). Treatment plans were created to deliver 3 Gy per fraction for 20 fractions ensuring that the 95% isodose encapsulated the PTV and the dose to the pseudo-OARs was minimized. Conformal plans were created using the class solution employed at our centre. This includes an anterior beam and two lateral wedged elds. IMRT and VMAT deliveries were inverse

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(b)

(c)

Figure 1. Photograph of (a) the PMMA phantom, (b) phantom setup for clinical irradiations and (c) phantom setup for uniform irradiations.

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planned using the optimization module within Oncentra R . The local class solution for a ve-eld IMRT plan includes beam angles of 35 , 100 , 180 , 260 and 325 with a maximum number of segments of 50 and a minimum eld size of 4 cm. The nine-eld IMRT plan used nine equi-spaced beams 40 apart with a starting angle of 0 (maximum number of segments = 90 and minimum eld size = 4 cm). A single-arc VMAT plan used the default gantry spacing of 4 with a start angle of 184 and arc length of 356 . For the dual arc, all properties were the same as the rst arc but the second arc started at 180 and rotated in the opposite direction to the rst arc. A constant dose rate and gantry speed was assumed and the couch accounted for (McGarry et al 2010) during the VMAT planning process. A single uniform 20 20 cm2 beam was also planned and delivered. All plans were delivered using a 6 MV photon Varian 600CD linear accelerator (Varian Medical Systems, Palo Alto, CA, USA). The aim of this work was to replicate the clinical delivery of each modality as closely as possible. A local audit of ten clinical ve-eld IMRT plans and ten clinical conformal plans showed that they would take approximately 5.0 0.8 min and 2.4 0.3 min respectively between the rst beam on time and the last beam off time on days that the patient was not imaged. The aim of the VMAT planning was to ensure that delivery time was close to the delivery of a prostate plan as reported in the literature (Bedford 2009, Boylan et al 2010). 2.3. Temporal effect on cell survival Figure 1(c) shows the experimental setup with the gantry and couch placed at 90 including 10 cm thick 30 30 cm2 and 5 cm thick 30 30 cm2 Wte solid water behind and in front of the phantom respectively to provide buildup and backscatter. The total dose was kept at 3 Gy but the average dose rate was varied by changing source-to-surface distances (SSDs) in the range from 0.75 through 3.42 m whilst ensuring that a 20 20 cm2 eld size was maintained at the surface of the solid water for each SSD studied. The extreme distances represent the minimum and maximum possible SSD in the treatment room. Similar to Bewes et al (2008), the instantaneous dose rate varies approximately according to the inverse square of the distance, while a pulse repetition frequency (PRF) of either 200 or 400 MU min1 was utilized. The time to deliver the 3 Gy dose varied from 0.55 to 20.54 min. All plans were delivered using 6 MV photons on a Varian 600CD linear accelerator (Varian Medical Systems, Palo Alto, CA, USA). 2.4. Dosimetry Each of the six clinical plans was re-calculated onto a CT scan dataset of the 30 30 19 cm3 solid water phantom with a Farmer ionization chamber (volume 0.6 cc) located at the isocentre. The active area of the chamber was outlined and the mean volume from each plan recorded. This was to avoid possible average volume effects associated with taking a point measurement using the detector. Each plan was delivered to the Farmer chamber on a Varian 600CD linear accelerator (Varian Medical Systems, Palo Alto, CA, USA). Farmer chamber readings were converted to average dose and these doses were compared to the doses calculated. Each of the six plans was re-calculated onto a CT scan dataset of a 2D ionization chamber array (IBA MatriXX Evolution) positioned between 6 cm slabs of 30 30 cm2 solid water. Irradiations were measured in movie mode as a sequence of 0.5 s frames with the frames summed using Omnipro ImRT software (IBA Dosimetry, Schwarzenbruck, Germany) to produce a cumulative dose image. A 2D ionization chamber array was used for measurements

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at intervals of 0.5 s as this has previously been shown to be accurate to within 1.5% (Evans et al 2010). The cumulative image for each delivery was compared with the plane calculated in terms of the percentage of pixels passing a gamma value of 3%/3 mm. The six clinical plans were delivered to the phantom with calibrated gafchromic EBT lm (International Specialty Products, NJ, USA) placed at the bottom of the ask. Film area measurements (to 0.5 cm inside the perimeter of the ask) were compared with those calculated using Oncentra R . The gafchromic lm was placed at the bottom of the asks at each SSD to ensure that, although the delivery time changed, the dose delivered was equivalent to 3 Gy. Measurements showed that doses were accurate to within 0.8%. 2.5. Temporal delivery characteristics of clinical plans The cells-eye-view (Goitein 2005) was used to describe the approximate cumulative dose received by the tumour cells over time. The cumulative central axis dose measurements for each plan were recorded using the 2D array data by integrating the dose over time. Effective fraction time is dened as the percentage of the treatment fraction time where any dose is delivered to the point examined (Sch afer et al 2005). This was calculated for 120 ionization chambers which approximately coincided with the cells inside the ask. 2.6. Cell culture and clonogenic assays Experiments were conducted using two cell lines, the human prostate cancer cell line, DU-145, and the human broblast cell line, AGO-1522b. Cell lines were obtained from Cancer Research UK and selected as malignant and transformed models with different radiosensitivity. DU145 cells were grown in RPMI-1640 with L- glutamine (Lonza, UK) supplemented with 10% fetal bovine serum, 1% penicillin/streptomycin (Gibco, UK). AGO-1522b cells were grown in Eagles minimum essential medium with deoxyribonucleosides and deoxyribonucleotides (Lonza, UK) supplemented with 20% fetal bovine serum and 1% penicillin/streptomycin. All cell lines were maintained at 37 C in a humidied atmosphere of 5% CO2. Cell survival was determined by clonogenic assay as previously reported (Butterworth et al 2010). Cells were plated and allowed to adhere overnight. Culture asks were lled with serum-free medium and sealed immediately prior to irradiation. Cells were irradiated at room temperature (25 2 C). Following irradiation, serum-free medium was removed and replaced with complete culture medium. Cultures were incubated for 1014 days before staining with 0.5% crystal violet in 50% methanol. DU-145 colonies were scored using a Colcount (Oxford Optronix, UK) automated counter which optimized for the cell line. AGO-1522b colonies were scored manually applying a 50 cell exclusion rule. For each experiment unexposed controls were prepared and treated as sham exposures. All exposures were performed in duplicate or triplicate and each clinical treatment plan group had 1219 samples with each uniform point derived from at least nine replicates. On each occasion unexposed controls were prepared and treated as sham exposures. For presentation purposes, and to be consistent with Moiseenko et al (2007), cell survival was normalized to the standard conformal treatment (delivered over 2.45 min). For uniform beams, the cell survival was normalized to cells irradiated over a time close to conformal treatment (2.79 min). 2.7. Statistical analysis Statistical signicance was assessed using regression analysis based on linear, quadratic, exponential and logarithmic models with the model accounting for most statistical variance used for statistical analysis. Data uncertainties were calculated as the standard error of the

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Table 1. Properties of each delivery modality including technique, dose rate, monitor units (MU) and delivery time. Included are local audit measurements of delivery times for ten conformal and ten IMRT (ve-eld) patients and VMAT single-arc delivery time reported (aBedford 2009, Boylan et al 2010) for prostate patients. Absorbed dose characteristics for each plan measured using 120 detectors of an ionization chamber array in the form of effective fraction time.

Plan Conformal/3D-CRT VMAT single arc VMAT dual arc IMRT 5 eld IMRT 9 eld Single eld

Technique Static Dynamic arc Dynamic arc Step and shoot Step and shoot Static

Nominal dose rate (MU min1) 400 300 200/200 400 400 400

MU 468 776 697 612 703 342

Delivery time (mm:ss) 2:27 2:35 3:42 4:30 8:17 0:51

Audit/ literature (mm:ss) 2:48 00:15 2:30a 5:00 00:50

Effective fraction time (%) 49.3 0.0 100.0 0.0 95.4 0.0 42.2 0.2 30.3 0.2 100.0 0.0

mean (SEM). Calculations were performed using Statistical Package for Social Sciences version 15.0.1.1 (SPSS, Chicago, IL, USA). 3. Results 3.1. Treatment planning and dosimetry Table 1 shows the characteristics of the six plans created. Figure 2 shows the dose distributions of the conformal, IMRT and VMAT plans. It is clearly demonstrated that the 95% isodose line fully encapsulated the PTV whilst avoiding the pseudo-OARs. Ionization chamber results were within 1.8% of the calculated values for all plans. All plans showed excellent agreement following comparison of delivery to the ionization chamber array to the calculated plan using gamma criteria of 3%/3 mm (>98.8% pixels passing). Measurements from the calibrated gafchromic EBT lm placed at the bottom of the asks compared to the same dose area calculated conrmed the accuracy of each delivery to cells in the phantom to within 1.7%. 3.2. Temporal delivery characteristics of clinical plans The cells-eye-view dose accumulation at the central axis of all delivery techniques is shown in gure 3. Conformal and single-arc VMAT plans took approximately the same time to deliver and were the fastest of all treatment modalities after single-eld delivery. The dual-arc VMAT was the next fastest to deliver with the ve- and nine-eld IMRT plans delivered over 4 min and 8 min respectively. These delivery times are also shown in table 1 and coincide with the current clinical delivery or literature values (Bedford 2009, Boylan et al 2010). Table 1 also shows that the effective fraction time is related to the number of beams or arcs with values of 30% for a nine-eld plan up to 100% for a single-arc and single-beam delivery. This is consistent with the fact that the gantry will have to move between exposures and the beam must be loaded for the next exposure which takes time, time when the tumour is not being irradiated. 3.3. Cell survival Figure 4(a) shows that AGO-1522b cell survival increases as the time to deliver a uniform radiation beam increases. Table 2 shows model parameters when the delivery time was

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(c)

(d)

(e)

(f)

Figure 2. Isodose distributions with target (blue) and pseudo-organs-at-risk (red) produced using Oncentra R . (a) Coronal slice through the CT scan with calculated isodoses for a 3D-conformal plan. Transverse slices with isodoses also shown for (b) a conformal plan, (c) a ve-eld IMRT plan, (d) a nine-eld IMRT plan, (e) a VMAT single-arc plan and (f) a VMAT dual-arc plan.

logarithmically regressed on cell survival. It was found that the dose delivery time was a statistically signicant predictor of clonogenic cell survival for AGO-1522b cells for a uniform eld (F = 10.73, n = 70, p < 0.01). However, the dose delivery time was not found to be a statistically signicant predictor of clonogenic cell survival for AGO-1522b cells when all modalities, including modulated deliveries, were analysed using linear regression (r = 0.020 0.156, t(1, 62) = 0.157, p = 0.876). Figure 4(b) shows that DU-145 cell survival does not change signicantly as the time to deliver a uniform radiation beam increases. It was found that the dose delivery time was not a statistically signicant predictor of clonogenic cell survival for DU-145 cells (F = 0.325, n = 75, p = 0.570). As with AGO-1522b cells, the dose delivery time was not found to be a statistically signicant predictor of clonogenic cell survival for DU-145 cells when all modalities were analysed using linear regression (r = 0.040 0.317, t(1, 107) = 0.410, p = 0.683).

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Figure 3. Cells-eye-view at the central axis for all six plans.

Table 2. Parameters derived from a logarithmic regression analysis showing the relationship between delivery time and cell survival for a uniform beam. Survival fraction at 3 Gy at 2.79 min shown for each cell line is included to indicate radiosensitivity.

Cell line

/ range

SF3 normalization R SE

Model summary F

Parameter estimates a 0.011 0.064

Signicant Constant 1.027 0.969

DU-145 2.332.67a 0.484 (0.034) 0.066 (0.190) 0.325 p = 0.57 AGO-1522b 10.7526.5a 0.244 (0.013) 0.365 (0.192) 10.73 p < 0.01
a

For reference, previously derived / values are included (Butterworth et al 2010).

4. Discussion In this study a phantom (gure 1) was designed and implemented to compare cell survival following the delivery of different radiation therapy techniques. Single-beam, conformal, ve-eld IMRT, nine-eld IMRT, single-arc VMAT and dual-arc VMAT plans were created and veried following a 3 Gy delivery using each modality to the cell layer within the ask in the phantom. Variation in the absorbed dose rate at a cells-eye-level was observed using a 2D ionization chamber array across all ve plans. Dose delivery time was found to be a statistically signicant predictor of clonogenic cell survival for AGO-1522b cells but not DU-145 cells for uniform deliveries over a time range 0.520 min. No dose rate effects were observed for either cell line when survival was analysed against delivery time using the clinical deliveries. All treatment plans (gure 2) ensured that the 95% isodose line encapsulated the PTV. Delivery times were in accordance with what was expected locally and from the literature (Bedford 2009, Boylan et al 2010). Delivery patterns at the central axis for IMRT and 3DCRT plans were consistent with previous studies (Sch afer et al 2005, Kuperman et al 2008, Moiseenko et al 2007, Shaikh et al 2010). This type of analysis had not yet been performed on inverse planned VMAT plans prior to this study. In addition to measurements of variation in delivery time, effective fraction time was also calculated across 120 detectors for 3D-CRT, IMRT and VMAT plans. Effective fraction times ranging from 11.6 to 37.3%, depending on the number of IMRT beams, have been previously reported by Sch afer et al (2005). Similarly, we have shown the effective fraction time to be inversely proportional to the number of beams

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Figure 4. Clonogenic cell survival fractions at 3 Gy (SF3) for (a) AGO-1522b cell line and (b) DU-145 cell line comparing dose delivery from a single uniform beam and ve treatment modalities. Error bars show the standard error of the mean. Linear trend lines are shown to guide the eye (dashed line). The normalization condition corresponds to 3D-CRT delivery. Uniform delivery of 3 Gy over 0.520.54 min range is also plotted for each cell line. Logarithmic trend lines are shown to guide the eye (solid line). The normalization condition corresponds to 2.79 min delivery.

or arcs as shown in table 1. The nine-eld IMRT plan (effective fraction time = 30.3 0.2%) was least efcient with 100 0.0% efciency being observed for VMAT single-arc plans. Less variation in the dose rates across detectors was observed for the IMRT plans compared to Sch afer et al (2005). This may be attributed to the use of a 2D array and not three-dimensional

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dose points although other factors such as the use of a more modern inverse planning system may also have contributed. Each of the six clinical plans created were delivered to give a uniform dose of 3 Gy to prostate tumour cells (DU-145) and normal primary AGO-1522b broblast cells grown as a monolayer at the bottom surface of the T80 ask within the phantom. The framework used to study the outcome following cell survival was proposed by Moiseenko et al (2007) which revealed increased cell survival following longer IMRT delivery compared to acute delivery for more radiosensitive cells. Within this framework it is important that new delivery techniques are compared to typical clinical techniques. Using a similar method, Yang et al (2009) showed signicantly increased cell kill with a 2 min arc therapy delivery compared to 7 min IMRT or HT delivery. Rapid MLC modulation had not been simulated in either arc or IMRT delivery. The data presented in this paper have focussed on cell survival following the delivery of clinically relevant inverse planned IMRT and VMAT plans compared to a conventional 3D conformal plan. Dose delivery time was not found to be a statistically signicant predictor of clonogenic cell survival for AGO-1522b or DU-145 cells when all modalities were analysed. This is consistent with our previous work comparing survival responses to modulated and non-modulated delivery (Butterworth et al 2010). When MLC modulation was not included in any plans and a wider time range used, dose delivery time was a statistically signicant predictor of clonogenic cell survival for AGO-1522b cells but not DU-145 cells. As with reports from other authors (Moiseenko et al 2007, Bewes et al 2008, Yang et al 2009, Joiner et al 2010) increased survival over time was largest for the more radiosensitive cells. This effect appears to reduce when absorbed dose properties vary between delivery techniques. The properties of plans such as effective fraction time vary widely between modalities. As the time range used for the dose-rate experiments extended well beyond the longest time for the clinical irradiations investigated here, the dose-rate data were re-tted excluding time-points beyond 10.24 min to ensure that these data points were not affecting the comparison with the clinical exposures. Even after excluding these data points there was still a statistically signicant correlation between treatment time and survival fraction following logarithmic regression (p = 0.04). Clinical trials have shown no clinical indications that IMRT has led to adverse effects on locoregional control or survival (Veldeman et al 2008). This is consistent with our ndings, although the small differences observed between different techniques may require a more sensitive method of detecting outcomes. Further work is required to extrapolate to the in vivo scenario where there will be a complex relationship between tumour cells and surrounding normal tissue.

5. Conclusion A phantom to study the delivery of radiotherapy plans in vitro was designed and validated. Single-beam, conformal, IMRT and VMAT plans were created for this phantom and the absorbed dose rates characterized using a 2D ionization chamber array. The delivery time and effective fraction time were found to vary widely between modalities. For uniform irradiations, a statistically signicant trend towards increased survival with increased delivery time was observed for AGO-1522b cells, but not for more radioresistant DU-145 cells. No trend was observed in either cell line when survival was analysed against delivery time using the modulated clinical plans with widely differing absorbed dose rate histories. Differences in absorbed dose rate histories of techniques such as 3D-CRT, IMRT and VMAT may not allow

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direct interpolation of cell survival using data generated from uniform beam delivery over a range of treatment times.

Acknowledgments We are indebted to Cyril Mitchell for his invaluable expertise in manufacturing the phantom. We also thank Dr Christina Agnew for proof reading the manuscript. CKM is supported by a Health & Social Care Research & Development Ofce of the Public Health Agency Training Fellowship Award. We wish to acknowledge nancial support from Cancer Research UK (grant number C1513/A7047 to KM Prise).

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