Prognostic factors for local control of stage I non-small cell lung cancer in stereotactic radiotherapy: a retrospective analysis
© Shirata et al.; licensee BioMed Central Ltd. 2012
Received: 13 July 2012
Accepted: 26 October 2012
Published: 31 October 2012
The purpose of this study is to investigate the prognostic factors of stereotactic radiotherapy for stage I NSCLC to improve outcomes.
Stage I non-small cell lung cancer patients who were treated with stereotactic radiotherapy between 2005 and 2009 at our hospital were enrolled in this study. The primary endpoint was local control rate. Survival estimates were calculated from the completion date of radiotherapy using the Kaplan-Meier method. The prognostic factors including patients’ characteristics and dose-volume histogram parameters were evaluated using Cox’s proportional hazard regression model.
Eighty patients (81 lesions) treated with 3 dose levels, 48 Gy/4 fractions, 60 Gy/8 fractions and 60 Gy/15 fractions, were enrolled in this study. Median follow-up was 30.4 months (range, 0.3 – 78.5 months). A Cox regression model showed T factor (p = 0.013), biological effective dose calculated from prescribed dose (BED10) (p = 0.048), and minimum dose for PTV (p = 0.013) to be prognostic factors for local control. Three-year overall survival rate and local control rate were 89.9% (T1: 86.8%, T2: 100%) and 89.0% (T1: 97.9%; T2: 64.8%), respectively. When the 3-year local control rates were examined by prescribed doses, they were 100% for the dose per fraction of 48 Gy /4 fractions (105.6 Gy BED10), 82.1% for 60 Gy/8 fractions (105 Gy BED10), and 57.1% for 60 Gy/15 fractions (84 Gy BED10). The median value of the minimum dose for PTV (%) was 89.88 (%), and the 3-year local control rates were 100% in those with the minimum dose for PTV (%) ≥ 89.88% and 79.2% in those with the minimum dose for PTV (%) < 89.88%.
Our results suggest that T factor, BED10, and minimum dose for PTV influence the local control rate. Local control rate can be improved by securing the minimum dose for PTV.
KeywordsStereotactic radiotherapy SBRT Non-small cell lung cancer NSCLC Prognostic factor Minimum dose PTV margin
In stereotactic radiotherapy for lung tumors, the dose at the lesion has been successfully increased through advancement of irradiation devices, improvement of set-up accuracy, introduction of image-guidance technology and measures for respiratory tumor movement [1, 2] while ensuring a high level of safety. For stage I non-small cell lung cancer (NSCLC), in particular, some reports suggest that short-term outcomes of stereotactic radiotherapy are comparable to those of surgeries [3, 4]. Furthermore, patients diagnosed with lung cancer in the early stage have increased recently due to the use of computed tomography (CT) scans and educational campaigns for cancer screening [5, 6].
However, lung cancer is still the main cause of cancer death worldwide, and local recurrence after stereotactic radiotherapy is not rare [3, 7–9]. The purpose of this study is to investigate the prognostic factors of stereotactic radiotherapy for stage I NSCLC to improve outcomes.
We reviewed data for all of the 80 patients (81 lesions) with stage I NSCLC who had undergone stereotactic radiotherapy in our hospital between March 2005 and July 2009. All patients included in this study had histological or cytological diagnosis of NSCLC and were staged as Union Internationale Contre le Cancer (UICC)-6 stage IA or IB by the use of CT. If available, 18 F-fluorodeoxyglucose positron emission tomography (FDG-PET) was used for staging. They either had a medical contraindication to surgery or refused surgery. Patient eligibility was not restricted on the basis of tumor location, unless a part of the esophagus, heart, main bronchi, hilus, spinal cord, or skin would be exposed to high-dose radiation. If the treatment plan included these organs in the high-dose areas, the patients were treated with conventional radiotherapy or modified stereotactic radiotherapy with a moderate irradiation dose; those patients were excluded from this study.
The primary endpoint was local recurrence. The secondary endpoint was overall survival. Patients’ background factors, various clinical parameters, and clinical course after irradiation were surveyed using information sources including medical records, data saved at the practice support terminal of our hospital, case follow-up cards of our department, and irradiation records. Local recurrence was defined as local progression that was 1.5 times or more the dimensions of original tumor . Tumors were observed on CT and/or 18 F-fluorodeoxyglucose positron emission tomography (FDG-PET) in order to assess the primary tumor’s stage and the presence or absence of recurrence. If FDG-PET was available, the maximum standardized uptake value (SUVmax) greater than 5.0 was considered as recurrence . The physicians and radiation oncologists finally decided to be local recurrence.
The study protocol was approved by the Ethical Committee of our institution, and informed consent was obtained from all patients.
- Long time scan
- 2.5 mm slice
Radiotherapy planning system
-Algorithm: Pencil beam convolution
- Heterogeneity correction: Modified Batho Power Law
- Observation of the tumor by fluoroscopy in advance
- GTV + 0–5 mm = CTV, CTV + 5–10 mm = PTV
- non-coplanar multi-dynamic arcs and/or multi-static beams
- 48 Gy/4 fractions, 60 Gy/8 fractions, 60 Gy/15 fractions
- Prescription for the iso-center
- 6 MV - X ray
Radiation therapy equipment
- Clinac 23EX
Patients were immobilized in the supine position with an individually fashioned half-body vacuum cast. Both the upper extremities were immobilized in the raised position unless the tumor was located at the apex of the lung, in which case both the upper extremities were immobilized beside the body.
Gross tumor volume (GTV) was defined as the visible extent of the tumor on the CT image at the lung window. Clinical target volume (CTV) was defined as GTV plus 0–5 mm margin for microscopic invasion. Internal target volume (ITV) was set equal to CTV because CT scanning was performed with an acquisition time of 4 seconds, and we consider that long-time (4 seconds) scan CT depicted virtually the entire tumor trajectory . Planning target volume (PTV) was determined by allowing for a set-up margin of 5 - 10mm beyond the ITV.
Treatment planning was performed with non-coplanar multi-dynamic arcs and/or multi-static beams by using a three-dimensional radiotherapy treatment planning system (Eclipse, Varian Medical Systems, Palo Alto, CA). The algorithm to calculate the dose was pencil beam convolution (PBC). Modified Batho Power law correction was used as the tissue heterogeneity correction algorithm. The target reference point was defined as the center of the PTV, and the dose was prescribed for its point. PTV was encompassed by the minimum 90% dose line of the reference point dose as possible. X-rays of 6 MV were used in all treatments.
The treatment took place using Clinac 23EX, Varian Medical Systems, Palo Alto, CA. In each irradiation, the position of the tumor was confirmed with fluoroscopy, and set-up and/or inter-fractional errors were corrected.
The patients were treated with a radiation schedule of 12 Gy /fraction ×4 fractions (48 Gy/4 fractions), 7.5 Gy/fraction ×8 fractions (60 Gy/8 fractions), and 4 Gy/fraction ×15 fractions (60 Gy/15 fractions). When the tumor was close to a risk organ, 7.5 Gy/fraction ×8 fractions or 4 Gy/fraction ×15 fractions was used to reduce the risk of serious toxicity due to set-up error or internal motion.
The first examination, including a clinical examination and CT scanning, was performed 4–6 weeks after treatment to assess the pulmonary reaction. Thereafter, the patients underwent follow-up examinations every 3–6 months for 2 years following treatment. After 2 years, follow-up examinations were performed every 6 months.
The prognostic factors for local control, including age, sex, T factor, histology, planning target volume (PTV), minimum dose for PTV (%), biological effective dose (BED) calculated from prescribed dose (BED10), and biological effective dose calculated from minimum dose (BEDmin), were investigated by stepwise Cox’s proportional hazard regression model for multivariate analysis. Hazard ratio for continuous data was evaluated with Wald χ2 test statistics. Sex and T factor were considered as categorical data. To investigate for the presence of multicollinearity, correlation coefficients were calculated for all variables. The hazard ratio was observed graphically to check its constancy.
BED10 and BEDmin were calculated using the linear quadratic formula in order to compare the effects of treatments with different fraction sizes and total doses. BED was given by: BED= nd[1 + d/(α/β)], where n is the number of fractions, d is the dose/fraction, and α/β ratio is 10 Gy. BED was not corrected with values for overall radiation time or tumor doubling time.
Overall survival, cause specific, and local control rates were calculated using the Kaplan-Meier method and statistical differences were evaluated by the log-rank test. When a continuous data was used as a variable for the Kaplan-Meier method, the data was divided by the median value into two groups. Statistical significance was defined as a value of p<0.05 in the present study. All analyses were performed using the SPSS 17.0 software package (SPSS Inc, Chicago, IL).
median, 77 years (range: 54–90 years)
Male: 64 lesions, 64 patients
Female: 17 lesions, 16 patients
Adenocarcinoma: 33 patients
Squamous cell carcinoma: 22 patients
Large cell carcinoma: 5 patients
Unclassified: 20 patients
T1 (Stage IA): 63 lesions
T2 (Stage IB): 18 lesions
Fifteen (18.8%) of the 80 patients showed evidence of recurrence. Local, regional (nodal) and distant recurrences were observed in 6 patients (7.5%), 3 patients (3.8%) and 12 patients (15%), respectively. Time to local failure varied between 12.2 and 33.7 months (median, 18.1 months).
Four patients died of NSCLC treated with stereotactic radiotherapy and 6 patients died of intercurrent causes. The 4 patients who died of NSCLC treated with stereotactic radiotherapy included 1 patient with local disease and regional lymph node metastasis and 3 patients with distant metastases. Intercurrent causes were colorectal cancer, aspiration pneumonia, advanced esophageal cancer, renal failure, chronic obstructive pulmonary disease (COPD), and multiple liver metastases considered to be from another site of NSCLC.
We treated 45 patients with a radiation schedule of 12 Gy /fraction ×4 fractions, 29 patients with a schedule of 7.5 Gy/fraction ×8 fractions and 7 patients with a schedule of 4 Gy/fraction ×15 fractions. We treated the tumor in consecutive weekdays.
Multivariate analysis with variables selected by stepwise method
95% confidence interval of hazard ratio
Significance probability (p)
T factor [Group: T2]
0.002 – 0.470
Minimum dose for PTV (%) [Unit: 1%]
0.571 – 0.935
BED 10 (Gy) [Unit: 10 Gy]
0.148 – 0.993
The 3-year overall survival rate for all patients was 89.9% (95% CI, 81.9% – 97.9%). The 3-year overall survival rates were 86.8% (95% CI, 76.6% – 97.0%) in patients with T1 tumors and 100% in those with T2 tumors. No significant difference was observed between these two groups (log-rank test, p = 0.29). The 3-year cause specific survival rate was 97.0% (95% confidence interval, 92.9% - 101.1%).
Multivariate analysis indicated that T factor, BED10, and minimum dose for PTV (%) influence local control. Among these, BED and minimum dose for PTV (%) can be changed by artificial intervention. BED10 would depend on the dose per fraction and total dose at a dose prescription, and they are generally determined by patient’s factor including tumor location, size, and patient’s general condition. The minimum dose for PTV (%) is decided by the radiotherapy plan including a targeting, margin factor, respiratory gating technique, and tumor tracking method. The results of this study suggest that the local control rate after stereotactic radiotherapy for stage I NSCLC can be improved by securing the minimum dose for PTV (%) when radiation oncologists produce a radiotherapy plan.
Onimaru et al. analyzed 41 patients with stage I NSCLC treated by stereotactic radiotherapy and reported that T factor and prescribed dose were significant factors for local control in multivariate analysis. The margin status was not a significant factor in spite of the fact that it was used as a variable . Probable reasons were the heterogeneity of the patients and the confounding factors analyzed in their study. Patients in whom a narrow margin was used included 7 patients with a T1 tumor and 3 patients with a T2 tumor.
In contrast, another study showed that tumor diameter and gender were the most significant factors affecting outcomes after stereotactic radiotherapy in recursive partitioning analysis; for comparison, tumor diameter was the only significant factor for local progression in a Cox proportional hazards model, and no margin-related variable was used . This was possibly because of the female pathological inclination to have adenocarcinoma. Some reports suggest that adenocarcinoma patients with stage I NSCLC tend to have a better survival . Since only 16 (20%) of the patients in our study were females, it was difficult to determine whether female gender is a significant prognostic factor.
Small sample size and retrospective protocol limit further interpretation of such findings, while tumor size is regarded as an independent prognostic factor for stage I NSCLC patients in general . Multi-institutional phase II trials are currently be conducted by the Radiation Therapy Oncology Group (Protocol 0236) and Japan Clinical Oncology Group (Protocol 0403). Results of detailed statistical analysis of data obtained in these trials are awaited to clarify various issues including issues discussed below.
Reports of stereotactic radiotherapy for stage I non-small cell lung cancer
First author (Reference No.)
Number of patients
Total dose (Gy)
Single dose (Gy/fraction)
Median follow up (months)
Local control (%)**
3-year overall survival (%)
180 - 211.2
18 - 75
4.4 - 35
57.6 - 180
48 - 60
4 – 12
84 – 105.6
When two types of prescriptions, one with BED10 of 105.6 Gy and the other with BED10 of 105 Gy, were compared, local control rates differed markedly despite only a slight difference in BED10 (Figure 2).
BED calculated by the linear quadratic formula with no correction was used in this study. The factor of overall radiation time was therefore not taken into account. During prolonged radiation delivery, sublethal damage repair takes place, leading to a decreased effect of radiation . Actually, there was no local recurrence in patients with the shortest radiotherapy (105.6 Gy BED10) in this study, although 6 patients treated with 105.6 Gy BED10 had a T2 tumor.
Many clinicians often use the linear-quadratic (LQ) model and BED to estimate the effects of various radiation schedules, but it has been suggested that the LQ model is not applicable to stereotactic radiotherapy because of its high dose per fraction . By contrast, Fowler et al. reported that the LQ model fitted the radiation response of epithelial tissues < 23 Gy/fraction . The best-fit model for tumor responses to stereotactic radiotherapy warrants further research.
Furthermore, we speculate that restriction of prescribed doses due to the vicinity of central structures and/or the radiation oncologist’s discretion in consideration of factors including performance status affected the control rate, resulting in this difference (Figure 2).
Some reports have shown that local control rate with over 100 Gy BED10 was higher than that with less than 100 Gy BED10, although a meta-analysis conducted by Zhang et al. showed no significant difference between BED10 < 100 Gy and BED10 ≥ 100 Gy . Medium BED or medium to high BED were recommended by Zhang et al. In our study, 4 Gy/fraction ×15 fractions (84 Gy BED10) was suggested to be insufficient for treatment of stage I NSCLC, particular in patients with a T2 tumor.
Further studies are needed to clarify the optimal total dose and fractions and the risk factors for relapse and side effects. When lower BED has to be prescribed because of tumor size, location of the tumor, and/or complications, use with chemotherapy can be considered .
Minimum dose for PTV (%)
Wald χ2 test indicated that minimum dose for PTV (%) was the strongest prognostic factor. We carried out the calculation of the local control, grouping by the minimum dose for PTV (%) into 2 groups: the minimum dose for PTV (%) ≥ 89.88% and < 89.88%, and there was significant difference. Baumann et al. showed by univariate analysis that radiation dose calculated in equivalent doses in 2 Gy fractions (EQD2) at the periphery of the PTV had an impact on survival but not on local recurrence rate . A trend toward smaller PTV margins for recurrent patients was also observed in their phase II trial . Among multiple factors, BED at the PTV margin was found to be the only significant factor influencing local control by Wulf et al. .
BED at the PTV margin can decrease for many reasons. The internal margin is not constant in each fraction, and sufficient management of the respiration factor is therefore important for planning particularly in the lower lobe. Dosimetric problems also arise from the limited accuracy of dose calculation algorithms in treatment planning systems. Lax et al. reported that the pencil beam algorithm significantly overestimates the dose . Radiation oncologists must have knowledge of the characteristics of the algorithm used in each institution. According to some reports, irradiation for large volumes of the lung can result in high-grade radiation pneumonitis . Radiation pneumonitis is the most important adverse reaction in stereotactic radiotherapy for a lung tumor and its prevention is a crucial task because of its severity. For this reason, the leaf margin might be reduced at the discretion of the radiation oncologist when the target is large and/or the patient has respiratory complications.
Efforts to avoid unnecessary reduction of the margin should be made. Nowadays, we use the dose prescribed by 95% of the PTV (D95) for the stereotactic radiotherapy. We will conduct further research to determine an acceptable minimum dose (%).
The continuing development of technologies including respiratory gated radiotherapy and real-time tumor tracking should enable further reduction of the margin for PTV. Therefore, further dose escalation for a larger tumor such as a T2 tumor may be possible, and severe side effects may be reduced since the amount of normal tissue irradiation will also be reduced with reduction of the margin for PTV.
T factor, BED10, and minimum dose for PTV (%) influence the local control rate. Local control rates can be improved by securing the minimum dose for PTV when radiation oncologists produce a radiotherapy plan for stage I non-small cell lung cancer in stereotactic radiotherapy.
- Buyyounouski MK, Balter P, Lewis B, et al.: Stereotactic body radiotherapy for early-stage non-small-cell lung cancer: report of the ASTRO emerging technology committee. Int J Radiat Oncol Biol Phys 2010, 78:3–10.PubMedView Article
- Onimaru R, Fujino M, Yamazaki K, et al.: Steep dose–response relationship for stage I non-small-cell lung cancer using hypofractionated high-dose irradiation by real-time tumor-tracking radiotherapy. Int J Radiat Oncol Biol Phys 2008, 70:374–381.PubMedView Article
- Onishi H, Shirato H, Nagata Y, et al.: Hypofractionated Stereotactic Radiotherapy (HypoFXSRT) for Stage I Non-small cell lung cancer: Updated results of 257 patients in a Japanese multi-institutional study. J Thorac Oncol 2007, 2:S94-S100.PubMedView Article
- Asamura H, Goya T, Koshiishi Y, et al.: A Japanese Lung Cancer Registry Study: Prognosis of 13010 Resected Lung Cancers. J Thorac Oncol 2007, 3:46–52.View Article
- Willard AF, Jerri LP, Herman RM: Ten-year survey of lung cancer treatment and survival in hospitals in the United States. A national cancer data base report. Cancer 1999,86(9):1867–1876.View Article
- Aberle DR, Adams AM, Berq CD, Black WC, Clapp JD, Faqerstrom RM, Gareen IF, Gatsonis C, Marcus PM, Sicks JD, National Lung Screening Trial Research Team: Reduced lung-cancer mortality with low-dose computed tomographic screening. N Engl J Med 2011,365(5):395–409.PubMedView Article
- Nagata Y, Takayama K, Matsuo Y, Norihisa Y, Mizowaki T, Sakamoto T, Sakamoto M, Mitsumori M, Shibuya K, Araki N, Yano S, Hiraoka M: Clinical outcomes of a phase I/II study of 48 Gy of stereotactic body radiotherapy in 4 fractions for primary lung cancer using a stereotactic body frame. Int J Radiat Oncol Biol Phys 2005,63(5):1427–1431.PubMedView Article
- Timmerman R, Paulus R, Galvin J, Michalski J, Straube W, Bradley J, Fakiris A, Bezjak A, Videtic G, Johnstone D, Fowler J, Gore E, Choy H: Stereotactic body radiation therapy for inoperable early stage lung cancer. JAMA 2010,303(11):1070–1076.PubMedView Article
- Fakiris AJ, McGarry RC, Yiannoutsos CT, Papiez L, Williams M, Henderson MA, Timmerman R: Stereotactic body radiation therapy for early-stage non-small-cell lung carcinoma: four-year results of a prospective phase II study. Int J Radiat Oncol Biol Phys 2009,75(3):677–682.PubMedView Article
- Koto M, Takai Y, Ogawa Y, Matsushita H, Takeda K, Takahashi C, Britton KR, Jingu K, Takai K, Mitsuya M, Nemoto K, Yamada S: A phase II study on stereotactic body radiotherapy for stage I non-small cell lung cancer. Raditother Oncol 2007,85(3):429–434.View Article
- Shiono S, Abiko M, Sato T: Positron emission tomography/computed tomography and lymphovascular invasion predict recurrence in stage I lung cancers. J Thorac Oncol 2010, 6:43–47.View Article
- Takeda A, Kunieda E, Shigematsu N, Hossain DM, Kawase T, Ohashi T, Fukada J, Kawaguchi O, Uematsu M, Takeda T, Takemasa K, Takahashi T, Kubo A: Small lung tumors: long-scan -time CT for planning of hypofractionated stereotactic radiation therapy-initial findings. Radiology 2005, 237:295–300.PubMedView Article
- Matsuo Y, Shibuya K, Nagata Y, Takayama K, Norihisa Y, Mizowaki T, Narabayashi M, Sakanaka K, Hiraoka M: Prognostic factors in stereotactic body radiotherapy for non-small –cell lung cancer. Int J Radiat Oncol Biol Phys 2011, 79:1104–1111.PubMedView Article
- Koike T, Tsuchiya R, Goya T, Sohara Y, Miyaoka E: Prognostic factors in 3315 Completely Resected Cases of Clinical Stage I Non-small Cell Lung Cancer in Japan. J Thorac Oncol 2007, 2:408–413.PubMedView Article
- Baumann P, Nyman J, Lax I, Friesland S, Hoyer M, Ericsson SR, Johansson KA, Ekberg L, Morhed E, Paludan M, Wittgren L, Blomgren H, Lewensohn R: Factors important for efficacy of stereotactic body radiotherapy of medically inoperable stage I lung cancer. A retrospective analysis of patients treated in the Nordic countries. Acta Oncol 2006, 45:787–795.PubMedView Article
- Fowler JF, Tomé WA, Fenwick JD, Mehta MP: A challenge to traditional radiation oncology. Int J Radiat Oncol Biol Phys 2004, 60:1241–1256.PubMedView Article
- Brunner TB, Kunz-Schughart LA, Grosse-Gehling P, Baumann M: Cancer Stem Cells as a Predictive Factor in Radiotherapy. Semin Radiat Oncol 2012, 22:151–174.PubMedView Article
- Matsuo Y, Shibuya K, Nagata Y, Norihisa Y, Narabayashi M, Sakanaka K, Ueki N, Mizowaki T, Hiraoka M: Preliminary Report of Late Recurrences, at 5 Years or more, after Stereotactic Body Radiation Therapy for Non-small Cell Lung Cancer. J Thorac Oncol 2012, 7:453–456.PubMedView Article
- Baba F, Shibamoto Y, Ogino H, Murata R, Sugie C, Iwata H, Otsuka S, Kosaki K, Nagai A, Murai T, Miyakawa A: Clinical outcomes of stereotactic body radiotherapy for stage I non-small cell lung cancer using different doses depending on tumor size. Radiat Oncol 2010,5(81):1–7.
- Shibamoto Y, Otsuka S, Iwata H, Sugie C, Ogino H, Tomita N: Radiobiological Evaluation of the Radiation Dose as Used in High-precision Radiotherapy: Effect of Prolonged Delivery Time and Applicability of the Linear-quadratic Model. J Radiat Res 2012, 53:1–9.PubMedView Article
- Zhang J, Yang F, Li B, Li H, Liu J, Huang W, Wang D, Yi Y, Wang J: Which is the optimal biological effective dose of stereotactic body radiotherapy for stage I non-small-cell lung cancer? A meta-analysis. Int J Radiat Oncol Biol Phys 2011, 81:e305-e316.PubMedView Article
- Chi A, Liao Z, Nguyen NP, Xu J, Stea B, Komaki R: Systemic review of the patterns of failure following stereotactic body radiation therapy in early-stage non-small-cell lung cancer: Clinical implications. Radiat Oncol 2010, 94:1–11.View Article
- Baumann P, Nyman J, Hoyer M, Wennberg B, Gagliardi G, Lax I, Drugge N, Ekberg L, Friesland S, Johansson KA, Lund JÅ, Morhed E, Nilsson K, Levin N, Paludan M, Sederholm C, Traberg A, Wittgren L, Lewensohn R: Outcome in a Prospective Phase II Trial of Medically Inoperable Stage I Non-Small Cell Lung Cancer Patients Treated With Stereotactic Body Radiotherapy. J Clin Oncol 2009, 27:3290–3296.PubMedView Article
- Wulf J, Baier K, Mueller G, Flentje MP: Dose–response in stereotactic irradiation of lung tumors. Radiat Oncol 2005, 77:83–87.View Article
- Lax I, Panettieri V, Berit W, et al.: Dose-distributions in SBRT of lung tumors: Comparison between two different treatment planning algorithms and Monte-Carlo simulation including breathing motions. Acta Oncol 2006, 45:978–988.PubMedView Article
- Yamashita H, Nakagawa K, Nakamura N, Duch MA, Näslund I, Baumann P, Gagliard G: Exceptionally high incidence of symptomatic grade 2–5 radiation pneumonitis after stereotactic radiation therapy for lung tumors. Radiat Oncol 2007, 2:1–11.View Article
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.