- Email: [email protected]
Feasibility Study of an Intensity‐Modulated Radiation Model for the Study of Erectile Dysfunction
411
Feasibility Study of an Intensity-Modulated Radiation Model for the Study of Erectile Dysfunction jsm_2125
411..418
Bridget F. Koontz, MD,* Hui Yan, PhD,* Masaki Kimura, MD,† Zeljko Vujaskovic, MD, PhD,* Craig Donatucci, MD,† and Fang-Fang Yin, PhD* *Duke University Medical Center, Department of Radiation Oncology, Durham, NC, USA; †Duke University Medical Center, Division of Urology, Department of Surgery, Durham, NC, USA DOI: 10.1111/j.1743-6109.2010.02125.x
ABSTRACT
Introduction. Preclinical studies of radiotherapy (RT) induced erectile dysfunction (ED) have been limited by radiation toxicity when using large fields. Aim. To develop a protocol of rat prostate irradiation using techniques mimicking the current clinical standard of intensity modulated radiotherapy (IMRT). Main Outcome Measures. Quality assurance (QA) testing of plan accuracy, animal health 9 weeks after RT, and intracavernosal pressure (ICP) measurement on cavernosal nerve stimulation. Methods. Computed tomography-based planning was used to develop a stereotactic radiosurgery (SRS) treatment plan for five young adult male Sprague-Dawley rats. Two treatment planning strategies were utilized to deliver 20 Gy in a single fraction: three-dimensional dynamic conformal arc and intensity-modulated arc (RapidArc). QA testing was performed for each plan type. Treatment was delivered using a NovalisTX (Varian Medical Systems) with high-definition multi-leaf collimators using on-board imaging prior to treatment. Each animal was evaluated for ED 2 months after treatment by nerve stimulation and ICP measurement. Results. The mean prostate volume and target volume (5 mm expansion of prostate) for the five animals was 0.36 and 0.66 cm3, respectively. Both conformal and RapidArc plans provided at least 95% coverage of the target volume, with rapid dose fall-off. QA plans demonstrated strong agreement between doses of calculated and delivered plans, although the conformal arc plan was more homogenous in treatment delivery. Treatment was well tolerated by the animals with no toxicity out to 9 weeks. Compared with control animals, significant reduction in ICP/mean arterial pressure, maximum ICP, and ICP area under the curve were noted. Conclusion. Tightly conformal dynamic arc prostate irradiation is feasible and results in minimal toxicity and measurable changes in erectile function. Koontz BF, Yan H, Kimura M, Vujaskovic Z, Donatucci C, and Yin F-F. Feasibility study of an intensity-modulated radiation model for study of erectile dysfunction. J Sex Med 2011;8:411–418. Key Words. Prostate Cancer; Intensity Modulated Radiation Therapy; Animal Model; Erectile Dysfunction; Stereotactic; Radiation-Associated Erectile Dysfunction
Introduction
R
adiation therapy (RT) is an effective and common treatment for prostate cancer, with 33% of newly diagnosed patients undergoing some form of radiation [1]. A particularly devastating side effect for many radiation patients is erectile dysfunction (ED), which is progressive over time and occurs in about 50% of men 3–5
© 2010 International Society for Sexual Medicine
years after external beam radiotherapy [2,3]. Clinical studies of RT-induced ED have found decreased penile blood flow, suggesting vascular injury as a cause [4–6]. More direct understanding has been elusive, as neurovascular bundle irradiation and dose to penile bulb and associated vasculature have inconsistently been found associated with ED in RT patients [7–9]. J Sex Med 2011;8:411–418
412 Animal models of surgical or medically induced ED have been successful in investigating the mechanisms of and possible interventions for ED [10–13]. For radiation induced ED there are only a few studies in which anesthetized rats were treated using a square field with a single or multiple fractions of radiation [14–16]. The utility of the previously developed animal models are significantly limited by the inability to focus the radiation on the prostate in a manner that simulates current treatment standards. Intensitymodulated radiotherapy (IMRT) uses tight margins with significant sparing of rectum, bladder, penile bulb, and other surrounding tissues. Accurate dose delivery to a rat prostate, about 0.3 cubic centimeters in volume, while sparing surrounding structures is technically difficult. However, with stereotactic radiosurgery (SRS), which uses multiple tightly focused beams to create a highly conformal radiation treatment [17,18], this is now feasible. This article reports our methods and success in implementing a small animal prostate radiosurgery program for preclinical research into the mechanisms of RT-induced ED. Materials and Methods
This Institutional Animal Care and Use Committee (IACUC) approved protocol used five young adult male Sprague-Dawley rats, aged approximately 2 months. All animals were housed two per cage in a temperature controlled vivarium with food and water provided ad libitum. All animal procedures within the radiation oncology clinic were performed after hours.
Computed Tomography (CT) Scanning Each animal underwent sedation using intraperitoneal ketamine/xylagine (60 mg/kg, 5.3 mg/kg) and was immobilized in the prone position on a styrofoam block. Using a GE Advantage Sim, a diagnostic quality CT scan was performed to include the entire pelvis from iliac crest to tail base in high resolution mode (0.625-mm slice thickness, 0.196-mm pixel size, 80 kV 300 mA). Approximately 150 CT slices were generated for each scan. The animal was then removed and recovered. Treatment Planning CT images were imported into an Eclipse treatment planning workstation (Varian Medical System, Palo Alto, CA, USA). Critical structures J Sex Med 2011;8:411–418
Koontz et al. including prostate, bladder, rectum, penile bulb and penile shaft were contoured (M.K.) and approved by another investigator (B.F.K.). A planning target volume (PTV) was generated by expanding prostate volume by 1 mm then subtracting out a “rectum + 1 mm” volume. Plans were then designed to treat the PTV with a single fraction of 20 Gy. Treatment was planned for an advanced treatment linear accelerator installed in our clinic, the Novalis TX (Varian, CA, USA and BrainLAB, Heimstetten, Germany), dedicated to stereotactic radiation therapy and radiosurgery [19–21]. It is a modified Varian Trilogy machine with high-definition multi-leaf collimators with leaf width of 2.5 mm at isocenter and equipped with an on-board imager with cone beam CT (CBCT) capability. The Novalis Tx system can treat with 6 MV energy in a high dose rate mode specifically designed for SRS. Two techniques were employed for treatment planning. The first technique, using a single dynamic conformal arc with 360-degree gantry rotation (18 fields at 20° angle intervals), generated a uniform sphere of dose with very sharp edges and dose fall-off. The MLCs of all beams were initially shaped to the PTV with 5 mm margins. Then, the margin of the lateral beams was modified to 3 mm manually in order to reduce the rectal volume receiving high dose. The second technique utilized volumetric modulated arc therapy (RapidArc) consisting of 177 fields with 2-degree angle intervals optimized to further improve rectal sparing while maintaining acceptable dose coverage of the prostate. This technique allows the dose constraints of interested structures to be considered in the optimization of beam intensities. All plans were normalized to require 95% of the PTV receive 100% of the prescription dose. Both conformal arc and RapidArc plans were developed for each rat and one plan was chosen for final treatment based on the quality of treatment plan. As an example, the dose distributions of both plans for rat 1 are displayed in Figure 1. The conformal plan (Figure 1a) provides better dose coverage of PTV but more rectal dose, while the RapidArc plan provides less rectal dose but poor dose coverage of PTV (Figure 1b). The comparison of dose volume histograms (DVHs) shown in Figure 1c further demonstrate the differences between the two plans. As in our study dose coverage of PTV is critical, the conformal plan was chosen for treatment. In another example, the dose distributions of both plans developed for rat 3 are shown in
IMRT Animal Model for Erectile Dysfunction
413
Figure 1 Axial and sagittal dose distributions of the conformal arc plan (a) and RapidArc plan (b) for rat 1. The dose volume histogram (DVH) is shown in (c), with structure volume along the y-axis and planned dose along the x-axis. The plan is indicated by square (conformal arc) or triangle (RapidArc) marked lines, with colors indicating structure (orange = prostate, yellow = bladder, brown = rectum, green = penile bulb).
Figure 2. The RapidArc plan shown in Figure 2b presents less rectal dose and comparable dose coverage of PTV as the conformal plan shown in (Figure 2a). The comparison of DVHs of both plans (Figure 2c) shows the same result. As more protection of rectum is provided without compromising dose coverage of PTV, the RapidArc plan was chosen for treatment.
Target Localization and Treatment Delivery On treatment day, animals were again sedated and immobilized on the styrofoam block. Initial set-up was based on pelvic bony anatomy using two-dimensional orthogonal kV images registered to digitally reconstructed radiographs (DRRs). Additional shifts to accurately set up the prostate within the treatment field were per-
formed using a three-dimensional CBCT (360° gantry rotation, 70 kV, 80 mA, 100 MV in fullfan mode with immediate reconstruction). Treatment was then delivered, with entire set-up and treatment taking approximately 30 minutes per animal.
Quality Assurance (QA) for Treatment Plans Treatment QA according to current patientspecific QA procedures designed for dynamic arc delivery was performed. Two assessments were made in order to investigate the accuracy of dose delivered: measurement of point dose at isocenter and two-dimensional dose distribution. QA plans were generated based on the existing treatment plans and delivered to a phantom in which doses were measured. In the first test, the point dose at J Sex Med 2011;8:411–418
414
Koontz et al.
Figure 2 Axial and sagittal dose distributions of the conformal arc plan (a) and RapidArc plan (b) for rat 3. The dose volume histogram (DVH) is shown in (c), with structure volume along the y-axis and planned dose along the x-axis. The plan is indicated by square (conformal arc) or triangle (RapidArc) marked lines, with colors indicating structure (orange = prostate, yellow = bladder, brown = rectum, green = penile bulb).
isocenter was measured and compared with the planned dose. Clinical guidelines require the difference to be within 3%. In the second test, the two-dimensional dose distribution within the isocenter plane was measured using EDR2 films (Carestream Health Inc., Toronto, Canada). Dose was delivered to film inside a square solid water phantom, developed using a Kodak film processor, and digitized using a Vidar scanner (VIDAR Systems Corporation, Herndon, VA, USA). A freshly generated H&D curve was applied to the treated film to obtain a two-dimensional dose distribution. The deviation between the measured two-dimensional dose and the planned twodimensional dose exported from treatment planning system was quantified by gamma index. The J Sex Med 2011;8:411–418
gamma index is set to evaluate 4mm diameter pixels. For clinical treatment purpose, it is required that at least 95% pixels fall within +/-4% of planned dose [22,23].
Intracavernosal Pressure (ICP) Measurement Animals were observed for 9 weeks after RT treatment. At completion of this waiting period, each animal was anesthetized using pentobarbital (45 mg/kg) and secured in the supine position. Continuous monitoring of mean arterial pressure (MAP) was performed by cannulating the right carotid artery with polyethylene (PE)-50 tubing containing heparinized saline and connected to a pressure transducer (World Precision Instruments, Sarasota, FL, USA) and amplifier (Biopac Systems
415
IMRT Animal Model for Erectile Dysfunction Inc., Goleta, CA, USA). For monitoring ICP, the shaft of the penis was exposed from skin and fascia, and the crus punctured with a 23 gauge needle also connected via PE-60 tubing. Microsurgery was then performed, dissecting to and isolating the cavernosal nerve as it runs alongside the prostate. The cavernosal nerve was stimulated using a bipolar electrode connected to a Grass Instruments S48 stimulator (Grass Technologies, West Warwick, RI, USA), using 2–8 V of 1.5 mA 16 Hz 5 ms wide pulses for 1 minute with a minimum interval between stimulation of 1 minute. All data was recorded by the MP100 data acquisition system and analyzed using the Acqknowledge® software (Biopac Systems Inc.). This procedure was performed on five irradiated animals and five age-matched control animals.
pixels were within range for the conformal arc and RapidArc plans, respectively (Figure 3). It is noted that there were two areas (1.5 mm ¥ 1.5 mm) with >4% deviation in the RapidArc plan, whereas no such area was observed in the conformal arc plan. Animals showed no signs of hair loss, distress, weight loss, anorexia, or other gastrointestinal symptoms over the 9-week observation period. At sacrifice, macroscopic atrophic changes were seen in the prostates of irradiated animals compared with the control group. The mean maximum ICP was 20.5 mm Hg vs. 65.8 mm Hg in the irradiated and control groups, respectively (P = 0.013). AUC was 932.6 vs. 2,914.6 mm Hg * second (P = 0.007), and ICP/MAP ration 0.24 vs. 0.62 (P = 0.002). Data are shown in Figure 4.
Statistical Analysis Treatment plans were evaluated by conformity index (CI) and median rectal dose. The CI was calculated by dividing the volume receiving 100% prescription dose by the PTV volume. A CI of 1 indicates that no nontarget tissue was given full dose. For ICP measurement, the Student’s t-test was used to analyze differences between treated and control animals using endpoints of maximum ICP, area under the curve (AUC), and the ICP/MAP ratio.
Discussion
Results
Five animals were planned and treated as described earlier. Mean prostate and PTV volumes were 0.36 and 0.66 cm3, respectively. Mean 100% dose cloud for conformal arc and RapidArc plans were 1.11 and 0.87 cm3, respectively, and the mean equivalent sphere diameter for all plans was 1.2 cm. The mean CI for all plans was 1.6, showing minimal inclusion of nontarget structures within the highdose region. Median rectal dose for conformal arc plans was 41.6% of prescription, for RapidArc 19% of prescription. Median penile bulbs doses were 0.5% and 3% of prescription and median penile shaft doses were 3.1 and 4.5% of prescription, respectively, for conformal and RapidArc plans. The two treatment plans developed and delivered for rat 1 (using conformal arc) and rat 3 (using RapidArc) were chosen for QA testing. Dose error at isocenter was 2.6% for the conformal arc plan and 1.6% for the RapidArc plan. On twodimensional dose matching, 98.4% and 97.3% of
We demonstrate here the technical feasibility to plan and deliver highly conformal radiation to the prostate of a rat and that such treatment does result in measurable ED 2 months after treatment. To our knowledge this is the first reported description of a procedure using clinical SRS technology for small animal radiation injury research. This strategy will be further tested but may allow further preclinical research into the etiology of RT-induced ED. The quality of the treatment plans was excellent overall. In comparing the RapidArc and conformal plans, the potential for under-dose is higher with RapidArc. The under-dose regions (volume outside the 95% isodose line) of PTV are most significant superiorly and inferiorly. There was also more heterogeneity in the RapidArc plans shown in the two-dimensional QA test. While both plans met clinical requirements, increased heterogeneity within the high dose region was noted in the RapidArc plan. As all animals tolerated the treatment extremely well, for future animals the conformal plan techniques was adopted. Further refinement of our technique has included use of bolus underneath the animals, which allows improved dose-build-up and tighter PTV coverage. Initial animals were treated within 2 weeks of planning, but in further experiments we have limited time to 2–3 days to minimize animal growth between planning and treatment. In this model we chose to treat animals with a single fraction of 20 Gy, which is a departure from standard fractionation currently used in humans. Significant differences in both tumor and normal J Sex Med 2011;8:411–418
416
Koontz et al.
Figure 3 Measurement of two-dimensional dose distribution of quality assurance (QA) plans. The overlapped isodose curves for conformal arc plan (a) and RapidArc plan (b). The solid lines and dash lines represent measured and planned dose distributions. The two-dimensional distributions of gamma index for conformal arc plan (c) and RapidArc plan (d). The central white circles in (d) encompass regions of more than 4% deviation (fail).
tissue responses to single vs. fractionated therapy do exist. In the normal tissues likely to be related to ED development (nerve, vascular structures) both the total dose and fraction size play a role in development of radiation effect; therefore a single dose of 20 Gy would be expected to have the same effect on these tissues as a fractionated dose of 86–92 Gy. We chose 20 Gy in one fraction for simplicity of the model as well as its comparability with dose-escalated fractionated doses in terms of late normal tissue effects. Previous models of rat prostate RT have resulted in significant toxicity [16] or used large fields, confounding normal tissue studies [14,15]. Carrier [14] found a dose-response by bio-assay and ICP approach 5 months after RT using a 16–20 cm2 square 250 kVp anterior field. Merlin [15], using a similar treatment design, found a dose-response with increased cavernosal endothelin-1 levels, a vasoconstrictor associated with ED [24,25], and reduction in maximum ICP J Sex Med 2011;8:411–418
1 month after RT. Both used lead shielding over testes, but included proximal penile and vascular structures. A more recent study by van der Wielen [16] gave 37 Gy in five fractions to 3 ¥ 4 cm square field using a single 200 kV anterior field. Investigators found histologic evidence of vascular damage within cavernosal arteries outside of the treatment field but rectal toxicity required the animals be sacrificed by 9 weeks. Compared with these earlier studies, our treatment plan is designed to mimic current IMRT radiation delivery in humans. The focal nature of this image-guided SRS technique prevents significant radiation dose to downstream erectile structures, such as penile bulb, which are considered avoidance structures during IMRT planning. Testicular shielding was not specifically performed because of the small focal field and large relative distance from treatment field to testes. However, there has been recent clinical speculation regarding internal radiation scatter and decreased testosterone levels; thus
417
IMRT Animal Model for Erectile Dysfunction Conclusion
Our lab has demonstrated the ability to use current stereotactic techniques to focus radiation to rat prostate only. Tightly conformal radiation plans to treat rat prostate with avoidance of critical structures is possible with CBCT image-guidance, allowing modeling of prostate IMRT for normal tissue effects. All five animals received a single fraction dose of 20 Gy and tolerated the therapy extremely well without demonstrable toxicities by 9 weeks post-treatment. Preliminary data shows this technique does result in ED as measured by intracavernosal pressure measurement. Acknowledgments
We sincerely appreciate Ms. Chen Qing, Dr. James Bowsher, and Dr. Sua Yoo for their clinical support and guidance. Corresponding Author: Bridget F. Koontz, MD, Department of Radiation Oncology, Duke University Medical Center, Box 3295 Medical Center, Durham, NC 27710, USA. Tel: (919) 668-5213; Fax: (919) 6687345; E-mail: [email protected] Conflicts of Interest: No actual or potential conflicts of interest exist for this work for any of the authors. This work was supported by departmental funding. Statement of Authorship
Category 1 (a) Conception and Design Bridget F. Koontz; Fang-Fang Yin; Zeljko Vujaskovic; Craig Donatucci; Hui Yan; Masaki Kimura (b) Acquisition of Data Bridget F. Koontz; Fang-Fang Yin; Masaki Kimura (c) Analysis and Interpretation of Data Bridget F. Koontz; Hui Yan; Masaki Kimura
Category 2 Figure 4 Results of intracvaernosal pressure (ICP) and mean arterial pressure (MAP) measurements at time of cavernosal nerve stimulation. (a) Maximum ICP (mm Hg) in control vs. irradiated animals. (b) Area under the curve (AUC) for ICP during stimulation. (c) ICP/MAP ratio for the same groups.
(a) Drafting the Article Bridget F. Koontz; Hui Yan (b) Revising It for Intellectual Content Zeljko Vujaskovic; Masaki Kimura; Fang-Fang Yin; Craig Donatucci
Category 3 with future experiments serum testosterone levels will be tested at sacrifice. Current ongoing experiments are evaluating the dose and time relationship of ED and radiation treatment, as well as investigating the histologic changes of the neurovascular bundles, penile bulb, and shaft for clues to the pathogenesis of RT-induced ED.
(a) Final Approval of the Completed Article Bridget F. Koontz References 1 Meltzer D, Egleston B, Abdalla I. Patterns of prostate cancer treatment by clinical stage and age. Am J Public Health 2001;91:126–8.
J Sex Med 2011;8:411–418
418 2 Incrocci L. Sexual function after external-beam radiotherapy for prostate cancer: What do we know? Crit Rev Oncol Hematol 2006;57:165–73. 3 Teloken PE, Parker M, Mohideen N, Mulhall JP. Predictors of response to sildenafil citrate following radiation therapy for prostate cancer. J Sex Med 2009;6:1135–40. 4 Goldstein I, Feldman MI, Deckers PJ, Babayan RK, Krane RJ. Radiation-associated impotence. A clinical study of its mechanism. JAMA 1984;251:903–10. 5 Mulhall J, Ahmed A, Parker M, Mohideen N. The hemodynamics of erectile dysfunction following external beam radiation for prostate cancer. J Sex Med 2005;2:432–7. 6 Zelefsky MJ, Eid JF. Elucidating the etiology of erectile dysfunction after definitive therapy for prostatic cancer. Int J Radiat Oncol Biol Phys 1998;40:129–33. 7 Incrocci L. Radiation therapy for prostate cancer and erectile (dys)function: The role of imaging. Acta Oncol 2005;44:673–8. 8 van der Wielen GJ, Mulhall JP, Incrocci L. Erectile dysfunction after radiotherapy for prostate cancer and radiation dose to the penile structures: A critical review. Radiother Oncol 2007;84:107–13. 9 Sadovsky R, Basson R, Krychman M, Morales AM, Schover L, Wang R, Incrocci L. Cancer and sexual problems. The J Sexual Medicine 2010;7:349–73. 10 Azadzoi KM, Goldstein I. Erectile dysfunction due to atherosclerotic vascular disease: The development of an animal model. J Urol 1992;147:1675–81. 11 Canguven O, Burnett A. Cavernous nerve injury using rodent animal models. J Sex Med 2008;5:1776–85. 12 Gur S, Kadowitz PJ, Hellstrom WJ. A critical appraisal of erectile function in animal models of diabetes mellitus. Int J Androl 2008. 13 Kovanecz I, Ferrini MG, Vernet D, Nolazco G, Rajfer J, Gonzalez-Cadavid NF. Ageing-related corpora veno-occlusive dysfunction in the rat is ameliorated by pioglitazone. BJU Int 2007;100:867–74. 14 Carrier S, Hricak H, Lee SS, Baba K, Morgan DM, Nunes L, Ross GY, Phillips TL, Lue TF. Radiation-induced decrease in nitric oxide synthase—Containing nerves in the rat penis. Radiology 1995;195:95–9. 15 Merlin SL, Brock GB, Begin LR, Hiou Tim FF, Macramalla AN, Seyam RM, Shenouda G, Dion SB. New insights into the
J Sex Med 2011;8:411–418
Koontz et al.
16
17
18
19
20
21
22
23
24
25
role of endothelin-1 in radiation-associated impotence. Int J Impot Res 2001;13:104–9. van der Wielen GJ, Vermeij M, de Jong BW, Schuit M, Marijnissen J, Kok DJ, van Weerden WM, Incrocci L. Changes in the penile arteries of the rat after fractionated irradiation of the prostate: A pilot study. J Sex Med 2009;6:1908–13. Nelson JW, Yoo DS, Sampson JH, Isaacs RE, Larrier NA, Marks LB, Yin FF, Wu QJ, Wang Z, Kirkpatrick JP. Stereotactic body radiotherapy for lesions of the spine and paraspinal regions. Int J Radiat Oncol Biol Phys 2009;73: 1369–75. Timmerman RD, Kavanagh BD, Cho LC, Papiez L, Xing L. Stereotactic body radiation therapy in multiple organ sites. J Clin Oncol 2007;25:947–52. Chang Z, Wang Z, Wu QJ, Yan H, Bowsher J, Zhang J, Yin FF. Dosimetric characteristics of novalis Tx system with high definition multileaf collimator. Med Phys 2008;35:4460–3. Wu QJ, Yoo S, Kirkpatrick JP, Thongphiew D, Yin FF. Volumetric arc intensity-modulated therapy for spine body radiotherapy: Comparison with static intensity-modulated treatment. Int J Radiat Oncol Biol Phys 2009;75:1596–604. Yin FF, Ryu S, Ajlouni M, Yan H, Jin JY, Lee SW, Kim J, Rock J, Rosenblum M, Kim JH. Image-guided procedures for intensity-modulated spinal radiosurgery. Technical note. J Neurosurg 2004;101(3 suppl):419–24. Ju SG, Han Y, Kum O, Cheong KH, Shin EH, Shin JS, Kim JS, Ahn YC. Comparison of film dosimetry techniques used for quality assurance of intensity modulated radiation therapy. Med Phys 2010;37:2925–33. Low DA, Harms WB, Mutic S, Purdy JA. A technique for the quantitative evaluation of dose distributions. Med Phys 1998; 25:656–61. Carneiro FS, Nunes KP, Giachini FR, Lima VV, Carneiro ZN, Nogueira EF, Leite R, Ergul A, Rainey WE, Clinton Webb R, Tostes RC. Activation of the ET-1/ETA pathway contributes to erectile dysfunction associated with mineralocorticoid hypertension. J Sex Med 2008;5:2793–807. Proietti M, Aversa A, Letizia C, Rossi C, Menghi G, Bruzziches R, Merla A, Spera G, Salsano F. Erectile dysfunction in systemic sclerosis: Effects of longterm inhibition of phosphodiesterase type-5 on erectile function and plasma endothelin-1 levels. J Rheumatol 2007;34:1712–7.
Feasibility Study of an Intensity-Modulated Radiation Model for the Study of Erectile Dysfunction jsm_2125
411..418
Bridget F. Koontz, MD,* Hui Yan, PhD,* Masaki Kimura, MD,† Zeljko Vujaskovic, MD, PhD,* Craig Donatucci, MD,† and Fang-Fang Yin, PhD* *Duke University Medical Center, Department of Radiation Oncology, Durham, NC, USA; †Duke University Medical Center, Division of Urology, Department of Surgery, Durham, NC, USA DOI: 10.1111/j.1743-6109.2010.02125.x
ABSTRACT
Introduction. Preclinical studies of radiotherapy (RT) induced erectile dysfunction (ED) have been limited by radiation toxicity when using large fields. Aim. To develop a protocol of rat prostate irradiation using techniques mimicking the current clinical standard of intensity modulated radiotherapy (IMRT). Main Outcome Measures. Quality assurance (QA) testing of plan accuracy, animal health 9 weeks after RT, and intracavernosal pressure (ICP) measurement on cavernosal nerve stimulation. Methods. Computed tomography-based planning was used to develop a stereotactic radiosurgery (SRS) treatment plan for five young adult male Sprague-Dawley rats. Two treatment planning strategies were utilized to deliver 20 Gy in a single fraction: three-dimensional dynamic conformal arc and intensity-modulated arc (RapidArc). QA testing was performed for each plan type. Treatment was delivered using a NovalisTX (Varian Medical Systems) with high-definition multi-leaf collimators using on-board imaging prior to treatment. Each animal was evaluated for ED 2 months after treatment by nerve stimulation and ICP measurement. Results. The mean prostate volume and target volume (5 mm expansion of prostate) for the five animals was 0.36 and 0.66 cm3, respectively. Both conformal and RapidArc plans provided at least 95% coverage of the target volume, with rapid dose fall-off. QA plans demonstrated strong agreement between doses of calculated and delivered plans, although the conformal arc plan was more homogenous in treatment delivery. Treatment was well tolerated by the animals with no toxicity out to 9 weeks. Compared with control animals, significant reduction in ICP/mean arterial pressure, maximum ICP, and ICP area under the curve were noted. Conclusion. Tightly conformal dynamic arc prostate irradiation is feasible and results in minimal toxicity and measurable changes in erectile function. Koontz BF, Yan H, Kimura M, Vujaskovic Z, Donatucci C, and Yin F-F. Feasibility study of an intensity-modulated radiation model for study of erectile dysfunction. J Sex Med 2011;8:411–418. Key Words. Prostate Cancer; Intensity Modulated Radiation Therapy; Animal Model; Erectile Dysfunction; Stereotactic; Radiation-Associated Erectile Dysfunction
Introduction
R
adiation therapy (RT) is an effective and common treatment for prostate cancer, with 33% of newly diagnosed patients undergoing some form of radiation [1]. A particularly devastating side effect for many radiation patients is erectile dysfunction (ED), which is progressive over time and occurs in about 50% of men 3–5
© 2010 International Society for Sexual Medicine
years after external beam radiotherapy [2,3]. Clinical studies of RT-induced ED have found decreased penile blood flow, suggesting vascular injury as a cause [4–6]. More direct understanding has been elusive, as neurovascular bundle irradiation and dose to penile bulb and associated vasculature have inconsistently been found associated with ED in RT patients [7–9]. J Sex Med 2011;8:411–418
412 Animal models of surgical or medically induced ED have been successful in investigating the mechanisms of and possible interventions for ED [10–13]. For radiation induced ED there are only a few studies in which anesthetized rats were treated using a square field with a single or multiple fractions of radiation [14–16]. The utility of the previously developed animal models are significantly limited by the inability to focus the radiation on the prostate in a manner that simulates current treatment standards. Intensitymodulated radiotherapy (IMRT) uses tight margins with significant sparing of rectum, bladder, penile bulb, and other surrounding tissues. Accurate dose delivery to a rat prostate, about 0.3 cubic centimeters in volume, while sparing surrounding structures is technically difficult. However, with stereotactic radiosurgery (SRS), which uses multiple tightly focused beams to create a highly conformal radiation treatment [17,18], this is now feasible. This article reports our methods and success in implementing a small animal prostate radiosurgery program for preclinical research into the mechanisms of RT-induced ED. Materials and Methods
This Institutional Animal Care and Use Committee (IACUC) approved protocol used five young adult male Sprague-Dawley rats, aged approximately 2 months. All animals were housed two per cage in a temperature controlled vivarium with food and water provided ad libitum. All animal procedures within the radiation oncology clinic were performed after hours.
Computed Tomography (CT) Scanning Each animal underwent sedation using intraperitoneal ketamine/xylagine (60 mg/kg, 5.3 mg/kg) and was immobilized in the prone position on a styrofoam block. Using a GE Advantage Sim, a diagnostic quality CT scan was performed to include the entire pelvis from iliac crest to tail base in high resolution mode (0.625-mm slice thickness, 0.196-mm pixel size, 80 kV 300 mA). Approximately 150 CT slices were generated for each scan. The animal was then removed and recovered. Treatment Planning CT images were imported into an Eclipse treatment planning workstation (Varian Medical System, Palo Alto, CA, USA). Critical structures J Sex Med 2011;8:411–418
Koontz et al. including prostate, bladder, rectum, penile bulb and penile shaft were contoured (M.K.) and approved by another investigator (B.F.K.). A planning target volume (PTV) was generated by expanding prostate volume by 1 mm then subtracting out a “rectum + 1 mm” volume. Plans were then designed to treat the PTV with a single fraction of 20 Gy. Treatment was planned for an advanced treatment linear accelerator installed in our clinic, the Novalis TX (Varian, CA, USA and BrainLAB, Heimstetten, Germany), dedicated to stereotactic radiation therapy and radiosurgery [19–21]. It is a modified Varian Trilogy machine with high-definition multi-leaf collimators with leaf width of 2.5 mm at isocenter and equipped with an on-board imager with cone beam CT (CBCT) capability. The Novalis Tx system can treat with 6 MV energy in a high dose rate mode specifically designed for SRS. Two techniques were employed for treatment planning. The first technique, using a single dynamic conformal arc with 360-degree gantry rotation (18 fields at 20° angle intervals), generated a uniform sphere of dose with very sharp edges and dose fall-off. The MLCs of all beams were initially shaped to the PTV with 5 mm margins. Then, the margin of the lateral beams was modified to 3 mm manually in order to reduce the rectal volume receiving high dose. The second technique utilized volumetric modulated arc therapy (RapidArc) consisting of 177 fields with 2-degree angle intervals optimized to further improve rectal sparing while maintaining acceptable dose coverage of the prostate. This technique allows the dose constraints of interested structures to be considered in the optimization of beam intensities. All plans were normalized to require 95% of the PTV receive 100% of the prescription dose. Both conformal arc and RapidArc plans were developed for each rat and one plan was chosen for final treatment based on the quality of treatment plan. As an example, the dose distributions of both plans for rat 1 are displayed in Figure 1. The conformal plan (Figure 1a) provides better dose coverage of PTV but more rectal dose, while the RapidArc plan provides less rectal dose but poor dose coverage of PTV (Figure 1b). The comparison of dose volume histograms (DVHs) shown in Figure 1c further demonstrate the differences between the two plans. As in our study dose coverage of PTV is critical, the conformal plan was chosen for treatment. In another example, the dose distributions of both plans developed for rat 3 are shown in
IMRT Animal Model for Erectile Dysfunction
413
Figure 1 Axial and sagittal dose distributions of the conformal arc plan (a) and RapidArc plan (b) for rat 1. The dose volume histogram (DVH) is shown in (c), with structure volume along the y-axis and planned dose along the x-axis. The plan is indicated by square (conformal arc) or triangle (RapidArc) marked lines, with colors indicating structure (orange = prostate, yellow = bladder, brown = rectum, green = penile bulb).
Figure 2. The RapidArc plan shown in Figure 2b presents less rectal dose and comparable dose coverage of PTV as the conformal plan shown in (Figure 2a). The comparison of DVHs of both plans (Figure 2c) shows the same result. As more protection of rectum is provided without compromising dose coverage of PTV, the RapidArc plan was chosen for treatment.
Target Localization and Treatment Delivery On treatment day, animals were again sedated and immobilized on the styrofoam block. Initial set-up was based on pelvic bony anatomy using two-dimensional orthogonal kV images registered to digitally reconstructed radiographs (DRRs). Additional shifts to accurately set up the prostate within the treatment field were per-
formed using a three-dimensional CBCT (360° gantry rotation, 70 kV, 80 mA, 100 MV in fullfan mode with immediate reconstruction). Treatment was then delivered, with entire set-up and treatment taking approximately 30 minutes per animal.
Quality Assurance (QA) for Treatment Plans Treatment QA according to current patientspecific QA procedures designed for dynamic arc delivery was performed. Two assessments were made in order to investigate the accuracy of dose delivered: measurement of point dose at isocenter and two-dimensional dose distribution. QA plans were generated based on the existing treatment plans and delivered to a phantom in which doses were measured. In the first test, the point dose at J Sex Med 2011;8:411–418
414
Koontz et al.
Figure 2 Axial and sagittal dose distributions of the conformal arc plan (a) and RapidArc plan (b) for rat 3. The dose volume histogram (DVH) is shown in (c), with structure volume along the y-axis and planned dose along the x-axis. The plan is indicated by square (conformal arc) or triangle (RapidArc) marked lines, with colors indicating structure (orange = prostate, yellow = bladder, brown = rectum, green = penile bulb).
isocenter was measured and compared with the planned dose. Clinical guidelines require the difference to be within 3%. In the second test, the two-dimensional dose distribution within the isocenter plane was measured using EDR2 films (Carestream Health Inc., Toronto, Canada). Dose was delivered to film inside a square solid water phantom, developed using a Kodak film processor, and digitized using a Vidar scanner (VIDAR Systems Corporation, Herndon, VA, USA). A freshly generated H&D curve was applied to the treated film to obtain a two-dimensional dose distribution. The deviation between the measured two-dimensional dose and the planned twodimensional dose exported from treatment planning system was quantified by gamma index. The J Sex Med 2011;8:411–418
gamma index is set to evaluate 4mm diameter pixels. For clinical treatment purpose, it is required that at least 95% pixels fall within +/-4% of planned dose [22,23].
Intracavernosal Pressure (ICP) Measurement Animals were observed for 9 weeks after RT treatment. At completion of this waiting period, each animal was anesthetized using pentobarbital (45 mg/kg) and secured in the supine position. Continuous monitoring of mean arterial pressure (MAP) was performed by cannulating the right carotid artery with polyethylene (PE)-50 tubing containing heparinized saline and connected to a pressure transducer (World Precision Instruments, Sarasota, FL, USA) and amplifier (Biopac Systems
415
IMRT Animal Model for Erectile Dysfunction Inc., Goleta, CA, USA). For monitoring ICP, the shaft of the penis was exposed from skin and fascia, and the crus punctured with a 23 gauge needle also connected via PE-60 tubing. Microsurgery was then performed, dissecting to and isolating the cavernosal nerve as it runs alongside the prostate. The cavernosal nerve was stimulated using a bipolar electrode connected to a Grass Instruments S48 stimulator (Grass Technologies, West Warwick, RI, USA), using 2–8 V of 1.5 mA 16 Hz 5 ms wide pulses for 1 minute with a minimum interval between stimulation of 1 minute. All data was recorded by the MP100 data acquisition system and analyzed using the Acqknowledge® software (Biopac Systems Inc.). This procedure was performed on five irradiated animals and five age-matched control animals.
pixels were within range for the conformal arc and RapidArc plans, respectively (Figure 3). It is noted that there were two areas (1.5 mm ¥ 1.5 mm) with >4% deviation in the RapidArc plan, whereas no such area was observed in the conformal arc plan. Animals showed no signs of hair loss, distress, weight loss, anorexia, or other gastrointestinal symptoms over the 9-week observation period. At sacrifice, macroscopic atrophic changes were seen in the prostates of irradiated animals compared with the control group. The mean maximum ICP was 20.5 mm Hg vs. 65.8 mm Hg in the irradiated and control groups, respectively (P = 0.013). AUC was 932.6 vs. 2,914.6 mm Hg * second (P = 0.007), and ICP/MAP ration 0.24 vs. 0.62 (P = 0.002). Data are shown in Figure 4.
Statistical Analysis Treatment plans were evaluated by conformity index (CI) and median rectal dose. The CI was calculated by dividing the volume receiving 100% prescription dose by the PTV volume. A CI of 1 indicates that no nontarget tissue was given full dose. For ICP measurement, the Student’s t-test was used to analyze differences between treated and control animals using endpoints of maximum ICP, area under the curve (AUC), and the ICP/MAP ratio.
Discussion
Results
Five animals were planned and treated as described earlier. Mean prostate and PTV volumes were 0.36 and 0.66 cm3, respectively. Mean 100% dose cloud for conformal arc and RapidArc plans were 1.11 and 0.87 cm3, respectively, and the mean equivalent sphere diameter for all plans was 1.2 cm. The mean CI for all plans was 1.6, showing minimal inclusion of nontarget structures within the highdose region. Median rectal dose for conformal arc plans was 41.6% of prescription, for RapidArc 19% of prescription. Median penile bulbs doses were 0.5% and 3% of prescription and median penile shaft doses were 3.1 and 4.5% of prescription, respectively, for conformal and RapidArc plans. The two treatment plans developed and delivered for rat 1 (using conformal arc) and rat 3 (using RapidArc) were chosen for QA testing. Dose error at isocenter was 2.6% for the conformal arc plan and 1.6% for the RapidArc plan. On twodimensional dose matching, 98.4% and 97.3% of
We demonstrate here the technical feasibility to plan and deliver highly conformal radiation to the prostate of a rat and that such treatment does result in measurable ED 2 months after treatment. To our knowledge this is the first reported description of a procedure using clinical SRS technology for small animal radiation injury research. This strategy will be further tested but may allow further preclinical research into the etiology of RT-induced ED. The quality of the treatment plans was excellent overall. In comparing the RapidArc and conformal plans, the potential for under-dose is higher with RapidArc. The under-dose regions (volume outside the 95% isodose line) of PTV are most significant superiorly and inferiorly. There was also more heterogeneity in the RapidArc plans shown in the two-dimensional QA test. While both plans met clinical requirements, increased heterogeneity within the high dose region was noted in the RapidArc plan. As all animals tolerated the treatment extremely well, for future animals the conformal plan techniques was adopted. Further refinement of our technique has included use of bolus underneath the animals, which allows improved dose-build-up and tighter PTV coverage. Initial animals were treated within 2 weeks of planning, but in further experiments we have limited time to 2–3 days to minimize animal growth between planning and treatment. In this model we chose to treat animals with a single fraction of 20 Gy, which is a departure from standard fractionation currently used in humans. Significant differences in both tumor and normal J Sex Med 2011;8:411–418
416
Koontz et al.
Figure 3 Measurement of two-dimensional dose distribution of quality assurance (QA) plans. The overlapped isodose curves for conformal arc plan (a) and RapidArc plan (b). The solid lines and dash lines represent measured and planned dose distributions. The two-dimensional distributions of gamma index for conformal arc plan (c) and RapidArc plan (d). The central white circles in (d) encompass regions of more than 4% deviation (fail).
tissue responses to single vs. fractionated therapy do exist. In the normal tissues likely to be related to ED development (nerve, vascular structures) both the total dose and fraction size play a role in development of radiation effect; therefore a single dose of 20 Gy would be expected to have the same effect on these tissues as a fractionated dose of 86–92 Gy. We chose 20 Gy in one fraction for simplicity of the model as well as its comparability with dose-escalated fractionated doses in terms of late normal tissue effects. Previous models of rat prostate RT have resulted in significant toxicity [16] or used large fields, confounding normal tissue studies [14,15]. Carrier [14] found a dose-response by bio-assay and ICP approach 5 months after RT using a 16–20 cm2 square 250 kVp anterior field. Merlin [15], using a similar treatment design, found a dose-response with increased cavernosal endothelin-1 levels, a vasoconstrictor associated with ED [24,25], and reduction in maximum ICP J Sex Med 2011;8:411–418
1 month after RT. Both used lead shielding over testes, but included proximal penile and vascular structures. A more recent study by van der Wielen [16] gave 37 Gy in five fractions to 3 ¥ 4 cm square field using a single 200 kV anterior field. Investigators found histologic evidence of vascular damage within cavernosal arteries outside of the treatment field but rectal toxicity required the animals be sacrificed by 9 weeks. Compared with these earlier studies, our treatment plan is designed to mimic current IMRT radiation delivery in humans. The focal nature of this image-guided SRS technique prevents significant radiation dose to downstream erectile structures, such as penile bulb, which are considered avoidance structures during IMRT planning. Testicular shielding was not specifically performed because of the small focal field and large relative distance from treatment field to testes. However, there has been recent clinical speculation regarding internal radiation scatter and decreased testosterone levels; thus
417
IMRT Animal Model for Erectile Dysfunction Conclusion
Our lab has demonstrated the ability to use current stereotactic techniques to focus radiation to rat prostate only. Tightly conformal radiation plans to treat rat prostate with avoidance of critical structures is possible with CBCT image-guidance, allowing modeling of prostate IMRT for normal tissue effects. All five animals received a single fraction dose of 20 Gy and tolerated the therapy extremely well without demonstrable toxicities by 9 weeks post-treatment. Preliminary data shows this technique does result in ED as measured by intracavernosal pressure measurement. Acknowledgments
We sincerely appreciate Ms. Chen Qing, Dr. James Bowsher, and Dr. Sua Yoo for their clinical support and guidance. Corresponding Author: Bridget F. Koontz, MD, Department of Radiation Oncology, Duke University Medical Center, Box 3295 Medical Center, Durham, NC 27710, USA. Tel: (919) 668-5213; Fax: (919) 6687345; E-mail: [email protected] Conflicts of Interest: No actual or potential conflicts of interest exist for this work for any of the authors. This work was supported by departmental funding. Statement of Authorship
Category 1 (a) Conception and Design Bridget F. Koontz; Fang-Fang Yin; Zeljko Vujaskovic; Craig Donatucci; Hui Yan; Masaki Kimura (b) Acquisition of Data Bridget F. Koontz; Fang-Fang Yin; Masaki Kimura (c) Analysis and Interpretation of Data Bridget F. Koontz; Hui Yan; Masaki Kimura
Category 2 Figure 4 Results of intracvaernosal pressure (ICP) and mean arterial pressure (MAP) measurements at time of cavernosal nerve stimulation. (a) Maximum ICP (mm Hg) in control vs. irradiated animals. (b) Area under the curve (AUC) for ICP during stimulation. (c) ICP/MAP ratio for the same groups.
(a) Drafting the Article Bridget F. Koontz; Hui Yan (b) Revising It for Intellectual Content Zeljko Vujaskovic; Masaki Kimura; Fang-Fang Yin; Craig Donatucci
Category 3 with future experiments serum testosterone levels will be tested at sacrifice. Current ongoing experiments are evaluating the dose and time relationship of ED and radiation treatment, as well as investigating the histologic changes of the neurovascular bundles, penile bulb, and shaft for clues to the pathogenesis of RT-induced ED.
(a) Final Approval of the Completed Article Bridget F. Koontz References 1 Meltzer D, Egleston B, Abdalla I. Patterns of prostate cancer treatment by clinical stage and age. Am J Public Health 2001;91:126–8.
J Sex Med 2011;8:411–418
418 2 Incrocci L. Sexual function after external-beam radiotherapy for prostate cancer: What do we know? Crit Rev Oncol Hematol 2006;57:165–73. 3 Teloken PE, Parker M, Mohideen N, Mulhall JP. Predictors of response to sildenafil citrate following radiation therapy for prostate cancer. J Sex Med 2009;6:1135–40. 4 Goldstein I, Feldman MI, Deckers PJ, Babayan RK, Krane RJ. Radiation-associated impotence. A clinical study of its mechanism. JAMA 1984;251:903–10. 5 Mulhall J, Ahmed A, Parker M, Mohideen N. The hemodynamics of erectile dysfunction following external beam radiation for prostate cancer. J Sex Med 2005;2:432–7. 6 Zelefsky MJ, Eid JF. Elucidating the etiology of erectile dysfunction after definitive therapy for prostatic cancer. Int J Radiat Oncol Biol Phys 1998;40:129–33. 7 Incrocci L. Radiation therapy for prostate cancer and erectile (dys)function: The role of imaging. Acta Oncol 2005;44:673–8. 8 van der Wielen GJ, Mulhall JP, Incrocci L. Erectile dysfunction after radiotherapy for prostate cancer and radiation dose to the penile structures: A critical review. Radiother Oncol 2007;84:107–13. 9 Sadovsky R, Basson R, Krychman M, Morales AM, Schover L, Wang R, Incrocci L. Cancer and sexual problems. The J Sexual Medicine 2010;7:349–73. 10 Azadzoi KM, Goldstein I. Erectile dysfunction due to atherosclerotic vascular disease: The development of an animal model. J Urol 1992;147:1675–81. 11 Canguven O, Burnett A. Cavernous nerve injury using rodent animal models. J Sex Med 2008;5:1776–85. 12 Gur S, Kadowitz PJ, Hellstrom WJ. A critical appraisal of erectile function in animal models of diabetes mellitus. Int J Androl 2008. 13 Kovanecz I, Ferrini MG, Vernet D, Nolazco G, Rajfer J, Gonzalez-Cadavid NF. Ageing-related corpora veno-occlusive dysfunction in the rat is ameliorated by pioglitazone. BJU Int 2007;100:867–74. 14 Carrier S, Hricak H, Lee SS, Baba K, Morgan DM, Nunes L, Ross GY, Phillips TL, Lue TF. Radiation-induced decrease in nitric oxide synthase—Containing nerves in the rat penis. Radiology 1995;195:95–9. 15 Merlin SL, Brock GB, Begin LR, Hiou Tim FF, Macramalla AN, Seyam RM, Shenouda G, Dion SB. New insights into the
J Sex Med 2011;8:411–418
Koontz et al.
16
17
18
19
20
21
22
23
24
25
role of endothelin-1 in radiation-associated impotence. Int J Impot Res 2001;13:104–9. van der Wielen GJ, Vermeij M, de Jong BW, Schuit M, Marijnissen J, Kok DJ, van Weerden WM, Incrocci L. Changes in the penile arteries of the rat after fractionated irradiation of the prostate: A pilot study. J Sex Med 2009;6:1908–13. Nelson JW, Yoo DS, Sampson JH, Isaacs RE, Larrier NA, Marks LB, Yin FF, Wu QJ, Wang Z, Kirkpatrick JP. Stereotactic body radiotherapy for lesions of the spine and paraspinal regions. Int J Radiat Oncol Biol Phys 2009;73: 1369–75. Timmerman RD, Kavanagh BD, Cho LC, Papiez L, Xing L. Stereotactic body radiation therapy in multiple organ sites. J Clin Oncol 2007;25:947–52. Chang Z, Wang Z, Wu QJ, Yan H, Bowsher J, Zhang J, Yin FF. Dosimetric characteristics of novalis Tx system with high definition multileaf collimator. Med Phys 2008;35:4460–3. Wu QJ, Yoo S, Kirkpatrick JP, Thongphiew D, Yin FF. Volumetric arc intensity-modulated therapy for spine body radiotherapy: Comparison with static intensity-modulated treatment. Int J Radiat Oncol Biol Phys 2009;75:1596–604. Yin FF, Ryu S, Ajlouni M, Yan H, Jin JY, Lee SW, Kim J, Rock J, Rosenblum M, Kim JH. Image-guided procedures for intensity-modulated spinal radiosurgery. Technical note. J Neurosurg 2004;101(3 suppl):419–24. Ju SG, Han Y, Kum O, Cheong KH, Shin EH, Shin JS, Kim JS, Ahn YC. Comparison of film dosimetry techniques used for quality assurance of intensity modulated radiation therapy. Med Phys 2010;37:2925–33. Low DA, Harms WB, Mutic S, Purdy JA. A technique for the quantitative evaluation of dose distributions. Med Phys 1998; 25:656–61. Carneiro FS, Nunes KP, Giachini FR, Lima VV, Carneiro ZN, Nogueira EF, Leite R, Ergul A, Rainey WE, Clinton Webb R, Tostes RC. Activation of the ET-1/ETA pathway contributes to erectile dysfunction associated with mineralocorticoid hypertension. J Sex Med 2008;5:2793–807. Proietti M, Aversa A, Letizia C, Rossi C, Menghi G, Bruzziches R, Merla A, Spera G, Salsano F. Erectile dysfunction in systemic sclerosis: Effects of longterm inhibition of phosphodiesterase type-5 on erectile function and plasma endothelin-1 levels. J Rheumatol 2007;34:1712–7.