Cerebral blood volume , genotype and chemosensitivity in oligodendroglial tumours Introduction The biological factors responsible for differential chemoresponsiveness in oligodendroglial tumours with or without the − 1p / − 19q genotype are unknown , but tumour vascularity may contribute . We aimed to determine whether dynamic susceptibility contrast -LRB- DSC -RRB- magnetic resonance imaging -LRB- MRI -RRB- could distinguish molecular subtypes of oligodendroglial tumour , and examined the relationship between relative cerebral blood volume -LRB- rCBV -RRB- and outcome following procarbazine , lomustine and vincristine -LRB- PCV -RRB- chemotherapy . Introduction Oligodendroglial tumours with combined loss of chromosomes 1p and 19q have a more indolent clinical evolution , respond more favourably to chemotherapy and show prolonged survival compared to those with intact 1p/19q , but the basis of these clinical differences is not well understood -LSB- 1 -- 5 -RSB- . In addition , up to 30 % of oligodendroglial neoplasms with intact 1p/19q may respond to chemotherapy -LSB- 3 , 6 , 7 -RSB- , but the genetic and biological factors associated with response are unknown and not all patients with oligodendroglial tumours derive durable benefit from therapy . Assessment of the clinical utility of biological markers and non-invasive imaging techniques is essential to advance the clinical management of these patients . Recently dynamic susceptibility contrast magnetic resonance imaging -LRB- DSC-MRI -RRB- has been used to calculate perfusion parameters in gliomas -LSB- 8 , 9 -RSB- , guide biopsy -LSB- 10 -RSB- , provide prognostic information -LSB- 11 -RSB- and demonstrate differences in the vascularity of low-grade astrocytomas compared with oligodendrogliomas -LSB- 8 -RSB- . It has been suggested that increased vascularity and concomitant improved drug delivery may contribute to the chemoresponsiveness of oligodendroglial tumours , especially as lipophilic drugs -LRB- procarbazine , lomustine and temozolomide -RRB- , which easily cross the blood -- brain barrier , are used in the treatment of oligodendrogliomas -LSB- 12 -- 14 -RSB- . Currently DSC-MRI is used in the radiological evaluation of gliomas , but its role in oligodendroglial neoplasms classified by genotype and the relationship with response and outcome following chemotherapy has not so far been investigated . The aim of this study was to determine whether DSC-MRI obtained through routine pre-therapy MR diagnostic examinations may be used to distinguish molecular subtypes of oligodendroglial tumours , and to investigate the influence of tumour blood volume on outcome following procarbazine , lomustine and vincristine -LRB- PCV -RRB- chemotherapy compared to molecular genetics . Methods Patient selection Patients for investigation were selected from those participating in a prospective study of adult oligodendroglial tumours eligible for chemotherapy at the Walton Centre for Neurology and Neurosurgery/Clatterbridge Centre for Oncology between May 2000 and June 2003 -LSB- 7 , 15 -- 18 -RSB- . The study had full ethical approval . For inclusion in this study , patients had to have -LRB- 1 -RRB- given research consent , -LRB- 2 -RRB- a histological diagnosis of oligodendroglioma or oligoastrocytoma based on the current World Health Organisation -LRB- WHO -RRB- classification -LSB- 19 -RSB- , -LRB- 3 -RRB- a known 1p/19q status , and -LRB- 4 -RRB- undergone DSC-MRI before or after biopsy -LRB- 2 image-guided ; 35 serial stereotactic -RRB- , but prior to commencing therapy . All patients -LRB- with primary and recurrent disease -RRB- underwent biopsy prior to PCV . Patients with post-resection imaging were excluded . DCS-MRI Imaging studies were obtained using a 1.5-T Signa MR scanner -LRB- General Electric Medical Systems -RRB- . The MR imaging protocol consisted of T1-weighted spin-echo -LRB- SE -RRB- sagittal sections -LRB- 400/14 TR/TE -RRB- , axial fluid attenuated inversion recovery -LRB- FLAIR 8802/142/2200 TR/TE/TI -RRB- , axial fast spin-echo -LRB- FSE -RRB- T2-weighted imaging -LRB- 4360/102 TR/TE -RRB- , 2D dynamic susceptibility-weighted , SE , echoplanar imaging -LRB- EPI 1900/80 ms , bandwidth 62.5 -RRB- and postcontrast T1-weighted SE -LRB- 600/14 TR/TE -RRB- or FLAIR -LRB- 2280/11/750 TR/TE/TI -RRB- axial imaging . The T2-weighted FLAIR , FSE and postcontrast T1-weighted images were used to define regions of T2 hyperintensity and T1 enhancement . For the dynamic EPI series , the tumour size and location , and the position of the margins were determined from the T2-weighted FLAIR or FSE images to equally define low-grade and high-grade tumours . Nine slices were selected to cover the majority of the tumour volume . Slice thickness was 7 mm , with no gap . A standard dose of 0.1 mmol/kg body weight of dimeglumine gadopentetate -LRB- Gd DTPA -RRB- was injected intravenously manually at an approximate rate of 3 ml/s . A series of T2 * - weighted multislice image sets -LRB- field of view 30 × 30 , matrix 192 × 128 -RRB- were acquired every 2 s during the first pass of the contrast agent . Image postprocessing was performed using commercial software -LRB- Functool2 , version 6.0 ; Sun Microsystems -RRB- . Negative enhancement integral -LRB- NEI -RRB- maps were generated for each patient relative to the signal intensity in the superior sagittal sinus -LRB- Fig. 1 -RRB- . The axial slice with the greatest cerebral blood volume and tumour area was chosen for analysis . Fig. 1Calculation of rCBV using DSC-MR . CBV is derived from the drop in the T2 * signal after injection of a bolus of paramagnetic compound -LSB- 8 , 9 -RSB- . Using Functool2 , the radiological tumour margin was delineated on the T2 * sequence axial image and a single circular ROI placed over normal brain mirrored in the contralateral hemisphere -LRB- a , b -RRB- . Negative enhancement integral -LRB- NEI -RRB- colour maps were generated -LRB- c , d -RRB- relative to the signal intensity in the superior sagittal sinus using image data between dynamic scans immediately before and after the contrast transient . CBV is proportional to the area under the contrast agent concentration-time curve and was calculated for each ROI by Functool2 using algorithms to integrate ΔR2i values -LRB- where ΔR2 is the change in the reciprocal of T2 * -RRB- . rCBV for each tumour was calculated from signal intensity time curves for individual ROIs placed within the delineated tumour region or in the large ROI in the contralateral hemisphere -LRB- as illustrated in e and f , arrows scans adjacent to the contrast transient -RRB- as described in the Materials and methods . a , c , e Low rCBV -LRB- 0.73 ± 0.17 , mean ± SD -RRB- in a grade II oligoastrocytoma with intact chromosomes 1p and 19q and ; b , d , f high rCBV -LRB- 4.15 ± 0.87 -RRB- in a grade II oligoastrocytoma with the − 1p / − 19q genotype The radiological tumour margin was delineated on the T2 * sequence axial image using a nongeometric region of interest -LRB- ROI -RRB- , and a single circular region was placed over normal brain mirrored in the contralateral hemisphere . Depending on tumour location , the mirrored area could include both white and grey matter . ROIs were placed according to published methods giving the lowest inter - and intraobserver variability -LSB- 20 -RSB- within the delineated tumour region . Briefly , five ROIs -LRB- area range 12 -- 27 mm2 -RRB- were placed over the tumour areas with the highest blood volume . Cystic and hemorrhagic regions were avoided and care was taken not to place ROIs over normal blood vessels or a biopsy tract where seen . Three independent analyses were performed by a neuroradiologist -LRB- T.S.S. -RRB- and a neurosurgeon -LRB- M.D.J. -RRB- blinded to histology and genotype , to assess inter - and intraobserver variability . For each observation , data were expressed relative to the internal reference -LRB- four ROIs placed within the delineated mirrored region of normal brain without deliberate bias towards either grey or white matter -RRB- to generate relative cerebral blood volume -LRB- rCBV -RRB- values -LRB- mean ROItumour/mean ROInormal brain -RRB- and accommodate physiological differences between patients -LSB- 21 -RSB- . For comparison with clinical data , the mean rCBV from these three data sets was used . Molecular genetics For each patient regions of tumour histology in pretherapy biopsy specimens -LRB- formalin-fixed , paraffin-embedded tissues or intraoperative diagnostic smear preparations -RRB- representative of the most aggressive tissue available and the overall pathology diagnosis were selected for laser capture microdissection to enrich the tumour component in the samples for analysis . Allelic imbalance was investigated using paired normal -LRB- microdissected uninvolved brain or DNA from blood -RRB- and tumour tissue samples and multiple simultaneous PCR amplification of a panel of microsatellite markers , capillary electrophoresis and data analysis as described previously -LSB- 16 -- 18 -RSB- . The microsatellite panel included : chromosome 1 -- D1S2667 , D1S508 , D1S214 -LRB- 1p36 -RRB- ; chromosome 19 -- D19S412 , D19S112 , D19S596 -LRB- 19q13 -RRB- ; chromosome 17 -- D17S796 , D17S1176 , D17S1353 -LRB- 17p13 -RRB- ; chromosome 10p -- D10S89 , D10S179 -LRB- 10p12-10p15 -RRB- ; chromosome 10q -- D10S1687 , D10s2491 , D10S583 , D10S587 , D10S212 -LRB- 10q22-10q26 -RRB- . Additional markers , D1S468 , D1S2795 -LRB- 1p36 -RRB- and D19S217 , D19S572 -LRB- 19q13 -RRB- were used in cases that lacked informativity for given loci . p53 mutations -LRB- exons 5 -- 8 -RRB- were detected by direct sequence analysis of PCR products . PCV chemotherapy , response and clinical outcome All patients receiving oncology therapy had evidence of advancing disease -LRB- either radiographic or clinical progression -RRB- prior to therapy . Patients with mass effect or evidence of malignancy were treated at presentation ; others were followed clinically until progression . Oncology therapy was based on individual patient 's clinical status/choice . Patients fit for PCV were offered PCV chemotherapy , or radiotherapy if fertility was an issue . PCV chemotherapy was administered according to standard clinical protocols -LSB- 22 -RSB- : lomustine -LRB- CCNU -RRB- -LRB- 110 mg/m2 -RRB- , procarbazine -LRB- 60 mg/m2 -RRB- and vincristine -LRB- 1.4 mg/m2 , maximum 2 mg -RRB- were given on days 1 , 8 -- 21 , and 8 and 29 . respectively . Cycles were repeated every 6 weeks for a maximum of six cycles . As described previously -LSB- 18 -RSB- , a consultant neuroradiologist reviewed all available MR or CT images taken before , during and after chemotherapy , and at follow-up . The largest perpendicular diameters of the tumour were measured on axial sections of T2-weighted MR images or contrast-enhanced regions on T1-weighted MR or CT images . Response was assessed using the criteria of Macdonald et al. -LSB- 23 -RSB- and postcontrast T1-weighted MRI in 25enhancing tumours . In tumours showing no enhancement -LRB- n = 6 -RRB- or in those in which contrast enhancement was not assessable -LRB- n = 1 -RRB- , response was based on T2-weighted images . Information regarding current management , follow-up and outcome was collected prospectively . Progression-free survival -LRB- PFS -RRB- and overall survival -LRB- OS -RRB- were calculated from the start of PCV to progression , last seen , or death as appropriate . Statistical analysis Inter - and intraobserver variability was determined using Bland and Altman plots -LSB- 24 -RSB- . Differences in rCBV between genetic and clinical factors were assessed using Student 's t-test for numerical data and the Chi-squared or Fisher 's exact test for categorical data . Receiver operator characteristics -LRB- ROC -RRB- analysis was performed to define the optimal rCBV cut-off for predicting 1p/19q status . Kaplan-Meier survival curves were obtained and differences in OS or PFS were tested for statistical significance using the Log Rank test . Cox regression multivariate analysis of factors shown to be significant in univariate analysis was by forward stepwise entry of parameters at a significance of 0.05 for entry and 0.01 for removal . Probability -LRB- P -RRB- values less than 0.05 were considered significant . Results The study included 30 patients with primary tumours without previous therapy and 7 with recurrent or persistent disease following radiotherapy . Median time between previous radiotherapy and MR imaging was 6.5 years -LRB- range 1.1 -- 14.2 years -RRB- . Median age was 44 years -LRB- range 28 -- 71 years -RRB- . Interobserver and intraobserver variability Tumour rCBV was obtained from DSC-MRI -LRB- Fig. 1 -RRB- by two independent observers . Bland and Altman plots revealed low inter - and intraobserver variation -LRB- Fig. 2 -RRB- . The closest observed limits of agreement were between the first observations of the neuroradiologist and neurosurgeon -LRB- mean − 0.17 , range − 1.27 to 0.93 ; Fig. 2a -RRB- . The widest observed limits of agreement were between the repeat observations of the neurosurgeon -LRB- mean 0.08 , range − 1.59 to 1.95 ; Fig. 2c -RRB- . The greatest variation was observed in tumours with higher rCBV values . Fig. 2Bland and Altman plots illustrating interobserver -LRB- a , b -RRB- and intraobserver -LRB- c -RRB- variation of rCBV measurements . The greatest variation is seen at the right of the plot for tumours with the higher rCBV values Histopathology , genotype and rCBV Oligodendroglial tumours of different histopathology subtypes and grade did not have significantly different rCBV measurements -LRB- Figure 3a , b -RRB- . Loss of both 1p36 and 19q13 was seen in 6/9 grade II oligodendrogliomas , 6/14 grade II oligoastrocytomas , 4/4 grade III oligodendrogliomas , and 3/10 grade III oligoastrocytomas . Tumours with the − 1p / − 19q genotype had significantly higher rCBV than those with intact 1p and 19q -LRB- Fig. 3c -RRB- . Similar findings were observed when primary tumours only were analysed -LRB- data not shown -RRB- . ROC analysis was used to determine the specificity and sensitivity of rCBV as a potential non-invasive tool to distinguish oligodendroglial neoplasms with or without the − 1p / − 19q genotype . The area under the ROC curve was 0.853 -LRB- 95 % confidence interval : 0.778 -- 0.927 -RRB- . A cut-off value of 1.59 for rCBV -LRB- sensitivity 92 % , specificity 76 % -RRB- best discriminated oligodendroglial tumours with and without the − 1p / − 19q genotype -LRB- Fig. 3c -RRB- . Fig. 3Box plots of rCBV against -LRB- a -RRB- histopathology subtype -LRB- P = 0.279 * -RRB- , -LRB- b -RRB- histopathology grade -LRB- P = 0.442 * -RRB- , and -LRB- c -RRB- genotype -LRB- P = 0.001 * -RRB- in oligodendroglial tumours . The optimal rCBV cut-off that distinguishes tumour genotype is shown by the horizontal line . * Student 's t-test To facilitate comparison of rCBV with genotype and clinicopathological characteristics -LRB- Table 1 -RRB- , tumours were considered to have high rCBV if this was greater than 1.59 as determined by the ROC analysis -LRB- Fig. 4 -RRB- . High rCBV values were significantly more common in tumours with loss of 1p36 and 19q13 and less common in tumours with 17p13 loss and p53 mutation . Three tumours in the series had loss of chromosome 10q22 -- 26 , of which two also had loss of 10p11 -- 15 ; all had high rCBV . One tumour with high rCBV had loss of 1p36 without loss of 19q13 . As for numerical data , there was no significant association between high rCBV and histology subtype or grade . Contrast enhancement was weakly associated with high rCBV in the series but not in primary tumours . No associations were seen between tumour rCBV and patient age < 50 years versus age ≥ 50 years , gender or temporal versus non-temporal location . Similar associations between rCBV , genotype and clinicopathological characteristics were obtained if only primary tumours were analysed -LRB- Table 1 -RRB- . Table 1rCBV , molecular genetics and clinical characteristics in the series and primary tumours All tumoursPrimary tumoursnNumber of tumours with high rCBV P valueanNumber of tumours with high rCBV P valueaLoss of 1p36 and 19q13Yes1917 -LRB- 89 % -RRB- 0.000 * 1614 -LRB- 88 % -RRB- 0.000 No184 -LRB- 22 % -RRB- 142 -LRB- 14 % -RRB- Loss of 17p13Yes112 -LRB- 18 % -RRB- 0.003 * 80 -LRB- 0 % -RRB- 0.001 No2619 -LRB- 73 % -RRB- 2216 -LRB- 73 % -RRB- Loss of 10q 22 -- 26 ± 10p11 -- 15Yes33 -LRB- 100 % -RRB- 0.23811 -LRB- 100 % -RRB- 1.0 no3317 -LRB- 52 % -RRB- 2915 -LRB- 52 % -RRB- p53 mutation -LRB- exon 5 -- 8 -RRB- Yes112 -LRB- 18 % -RRB- 0.004 * 80 -LRB- 0 % -RRB- 0.001 No2518 -LRB- 72 % -RRB- 2115 -LRB- 71 % -RRB- Patient age -LRB- years -RRB- < 502513 -LRB- 52 % -RRB- 0.491209 -LRB- 45 % -RRB- 0.260 > 50128 -LRB- 67 % -RRB- 107 -LRB- 70 % -RRB- Histology subtypeOligodendroglioma1310 -LRB- 77 % -RRB- 0.091118 -LRB- 73 % -RRB- 0.142 Oligoastrocytoma2411 -LRB- 46 % -RRB- 198 -LRB- 42 % -RRB- WHO histology gradeII2311 -LRB- 48 % -RRB- 0.19199 -LRB- 43 % -RRB- 0.118 III1410 -LRB- 71 % -RRB- 217 -LRB- 78 % -RRB- Contrast enhancementAbsent82 -LRB- 25 % -RRB- 0.04682 -LRB- 25 % -RRB- 0.092 Present2819 -LRB- 68 % -RRB- 2114 -LRB- 68 % -RRB- Tumour locationTemporal147 -LRB- 50 % -RRB- 0.733136 -LRB- 46 % -RRB- 0.713 Nontemporal2314 -LRB- 61 % -RRB- 1710 -LRB- 59 % -RRB- Tumour statusPrimary3016 -LRB- 53 % -RRB- 0.674 Recurrent75 -LRB- 71 % -RRB- Response to PCV All tumoursResponders2114 -LRB- 67 % -RRB- 0.1421711 -LRB- 65 % -RRB- 0.097 Nonresponders114 -LRB- 36 % -RRB- 82 -LRB- 25 % -RRB- Tumours with loss of 1p/19qResponders1614 -LRB- 88 % -RRB- 1311 -LRB- 85 % -RRB- Nonresponders00 Tumours with intact 1p/19qResponders50 -LRB- 0 % -RRB- 0.24540 -LRB- 0 % -RRB- 0.515 Nonresponders114 -LRB- 36 % -RRB- 82 -LRB- 25 % -RRB- aProbability calculated by Fisher 's Exact test . Responders showed complete , partial or minor response and non-responders had stable or progressive disease.Fig . 4rCBV , molecular genetics and clinical characteristics . rCBV for each tumour is given as the mean ± standard deviation of three independent observations . The arrow indicates tumours with rCBV above -LRB- high rCBV -RRB- and below the cut-off value of 1.59 -LRB- low rCBV -RRB- . CR complete response -LRB- disappearance of all tumour , off steroids and neurologically stable or improved -RRB- , PR partial response -LRB- 50 % or greater reduction in cross-sectional area , steroids stable or reduced , and neurologically stable or improved -RRB- ; MR minor response -LRB- > 25 to < 50 % reduction in cross-sectional area , steroids stable or reduced , and neurologically stable or improved -RRB- ; PD progressive disease -LRB- 25 % or greater increase in cross-sectional area or any new tumour on CT/MR images and/or neurologically worse with steroids stable or increased -RRB- ; SD stable disease -LRB- all other situations -RRB- ; P primary tumour , R recurrent tumour rCBV and outcome following PCV chemotherapy Of the 37 patients , 33 had PCV chemotherapy following DSC-MRI -LRB- median 0.7 months , range 0.1 -- 15.2 months -RRB- , of whom 26 with primary tumours received PCV chemotherapy as the first oncology therapy , 6 with recurrent or persistent disease treated initially with radiotherapy received PCV as second oncology therapy , and 1 had PCV following radiotherapy and temozolomide . One patient with primary tumour had therapy switched to radiotherapy at the end of the first cycle because of reaction to procarbazine . The remainder received a median of four cycles of PCV -LRB- range one to six -RRB- . Response to therapy was assessed in 32 patients -LRB- Fig. 4 -RRB- . Measured radiological changes and clinical factors permitted response assessment using standard criteria of Macdonald et al. -LSB- 23 -RSB- in all patients showing enhancement . However , in tumours showing no enhancement assessed using T2-weighted MR images , three showed radiological change in cross-sectional area of > 25 % to < 50 % , accompanied by clinical benefit ; these were included in the minor response -LRB- MR -RRB- category . None of those showing enhancement had measured radiological change in this range . Response -LRB- i.e. complete , partial or minor ; see Fig. 4 legend for definition -RRB- was seen in tumours with and without high rCBV . Similarly , non-responders -LRB- i.e. patients with stable or progressive disease -RRB- had either high or low tumour rCBV -LRB- Table 1 , Fig. 4 -RRB- . Response was strongly associated with genotype ; all 16 tumours with loss of 1p36 and 19q13 responded compared with 5 of 17 tumours with intact 1p36/19q13 -LRB- Chi2 P < 0.001 -RRB- . Of the 16 responders in the 1p36/19q13 loss group , 14 had high rCBV . The five responders in the intact 1p36/19q13 group all had low rCBV and the four tumours with high rCBV failed to respond . The relationship between rCBV , genotype , histopathology and survival following PCV in the series and in those with primary tumour is given in Table 2 . The − 1p / − 19q genotype and low-grade pathology were associated with longer PFS and OS . Histopathology subtype was not significantly associated with outcome . Increased rCBV was only associated with short PFS and OS when adjusted for the − 1p / − 19q genotype . Tumours with intact 1p/19q and high rCBV -LRB- two grade III primary oligoastrocytoma , one grade II recurrent oligodendroglioma , one grade III recurrent oligoastrocytoma -RRB- were associated with shorter PFS and OS than tumours with intact 1p/19q and low rCBV or those with lp/19q loss and high rCBV -LRB- Fig. 5a , b -RRB- . Similar findings were observed if primary tumour only were analysed -LRB- Fig. 5c , d -RRB- . Compared with significant clinicopathological parameters , high rCBV was an independent prognostic factor for PFS when adjusted for 1p/19q status -LRB- Cox regression : PFS : hazard ratio 13.4 , 95 % CI 1.3 -- 133 ; P = 0.008 -RRB- and for PFS and OS in patients with intact 1p/19q -LRB- Cox regression : PFS : hazard ratio 16.5 , 95 % CI 1.8 -- 152.2 ; P = 0.004 ; OS : hazard ratio 7.4 , 95 % CI 1.6 -- 33.3 ; P = 0.013 -RRB- . Table 2rCBV and survival in the series and in those with primary tumours . PFS and OS were calculated from start date of PCV -LRB- n number of cases , nc not calculated , nr median survival not reached -RRB- . Contrast enhancement -LRB- present vs absent -RRB- , age -LRB- < 50 vs ≥ 50 years -RRB- , ECOG performance status -LRB- 0 -- 1 vs ≥ 2 -RRB- and gender -LRB- male vs female -RRB- were not significantly related to PFS or OS nProgression-free survival -LRB- months -RRB- Overall survival -LRB- months -RRB- Median -LRB- 95 % CI -RRB- P valueaP valuebMedian -LRB- 95 % CI -RRB- P valueaP valuebAll tumoursLoss of 1p36 and 19q13Yes16 > 56 -LRB- nr -RRB- 0.005 > 56 -LRB- nr -RRB- 0.020 No178 -LRB- 0 -- 21 -RRB- > 49 -LRB- nr -RRB- Histopathology subtypeOligodendroglioma11 > 56 -LRB- nr -RRB- 0.0550.187 > 56 -LRB- nr -RRB- 0.1130.288 Oligoastrocytoma218 -LRB- 9 -- 41 -RRB- 50 -LRB- 9 -- 91 -RRB- Histopathology gradeII19 > 56 -LRB- nr -RRB- 0.0040.002 > 56 -LRB- nr -RRB- 0.0070.012 III144 -LRB- 8 -- 26 -RRB- 21 -LRB- 8 -- 34 -RRB- rCBVLow14 > 46 -LRB- nr -RRB- 0.7660.003 > 49 -LRB- nr -RRB- 0.5850.002 High1946 -LRB- nc -RRB- > 56 -LRB- nr -RRB- Primary tumoursLoss of 1p36 and 19q13Yes14 > 56 -LRB- nr -RRB- 0.037 > 56 -LRB- nr -RRB- 0.056 No12 > 39 -LRB- nr -RRB- 49Histopathology subtypeOligodendroglioma10 > 56 -LRB- nr -RRB- 0.0550.137 All censored0 .0920.209 Oligoastrocytoma1646 -LRB- 6 -- 86 -RRB- 50 -LRB- 6 -- 101 -RRB- Histopathology gradeII17 > 56 -LRB- nr -RRB- 0.0470.009 > 56 -LRB- nr -RRB- 0.0480.041 III96 -LRB- 13 -- 37 -RRB- 50 -LRB- nc -RRB- rCBVLow12 > 46 -LRB- nr -RRB- 0.6810.007 > 49 -LRB- nr -RRB- 0.6110.003 High14 > 56 -LRB- nr -RRB- > 56 -LRB- nr -RRB- aProbability calculated by the Log-Rank test.bProbability calculated by the Log-Rank test after adjustment for 1p/19q genotype.Fig . 5rCBV and outcome following PCV chemotherapy : a , b all patients ; c , d patients with primary tumour only ; a , c progression-free survival ; b , d overall survival . ① Patients with tumours with low rCBV -LRB- < 1.59 -RRB- and loss of 1p36 and 19q13 -LRB- all n = 2 , primary n = 2 -RRB- . ② Patients with tumours with high rCBV -LRB- > 1.59 -RRB- and loss of 1p36 and 19q13 -LRB- all n = 15 , primary n = 12 -RRB- . ③ Patients with tumours with low rCBV -LRB- < 1.59 -RRB- with intact 1p36 and 19q13 -LRB- all n = 12 , primary n = 10 -RRB- . ④ Patients with tumours with high rCBV -LRB- > 1.59 -RRB- and intact 1p36 and 19q13 -LRB- all n = 4 , primary n = 2 -RRB- Discussion To our knowledge this is the first study investigating the relationship between rCBV , genotype and outcome in oligodendroglial tumours treated with PCV chemotherapy . The cohort was drawn from a larger series in an ongoing prospective study of oligodendroglial neoplasms -LSB- 15 -- 18 -RSB- and reflects the range of patients given PCV chemotherapy in routine clinical practice . Tumours with rCBV > 1.59 were more likely to have 1p/19q loss . PCV chemosensitivity was predicted by 1p/19q loss , but high rCBV was not necessary for response . Following chemotherapy , rCBV was significantly associated with PFS and OS only after adjustment for genotype . DSC-MRI can be used to determine cerebral perfusion parameters such as rCBV , relative cerebral blood flow and permeability , but with the instrumentation available to us was restricted to rCBV . As in other studies -LSB- 8 , 20 , 25 , 26 -RSB- , placement of ROI in regions within tumour of greatest signal intensity yielded data with good inter - and intraobserver variability . The greatest variation was seen in tumours with high rCBV , reflecting heterogeneity of blood volume within these tumours . In this series , tumours with the − 1p / − 19q genotype were more likely to have high rCBV than those with intact 1p/19q . The ROC analysis determined an optimal cut-off -LRB- rCBV > 1.59 -RRB- for predicting the − 1p / − 19q genotype with 92 % sensitivity and 76 % specificity . These data suggest that in conjunction with histopathology , rCBV may be used to distinguish tumours of the oligodendroglial lineage . MRI studies of tumour vasculature in gliomas have demonstrated that rCBV may be useful in the non-invasive determination of tumour grade and vascularity -LSB- 21 , 25 , 27 , 28 -RSB- . Compared to their low-grade counterparts , high-grade tumours have elevated rCBV , associated with increased mitotic activity -LSB- 29 -RSB- . rCBV correlates with conventional angiographic assessment of tumour vascular density and histological measures of angiogenesis such as microvascular density and VEGF expression -LSB- 26 , 30 -- 32 -RSB- , which reflects the histopathological finding of increasing neovascularization with grade -LSB- 19 -RSB- . However , most studies have investigated astrocytic gliomas or included a variety of glioma pathologies and large series of oligodendroglial neoplasms are uncommon . Recent reports suggest that rCBV values in oligodendrogliomas render DSC-MRI-based tumour grading inaccurate -LSB- 28 , 33 -RSB- , and rCBV may be similar in low - and high-grade oligodendrogliomas -LSB- 33 -RSB- . Indeed low-grade oligodendrogliomas have significantly elevated rCBV compared to low-grade astrocytomas -LSB- 8 -RSB- . In our series , rCBV did not distinguish oligodendrogliomas from oligoastrocytomas , nor was rCBV significantly different between grade II and III tumours . This lack of correlation between rCBV and grade is consistent with the increased microvessel density seen in both low - and high-grade oligodendroglial tumours -LSB- 34 , 35 -RSB- . It has been postulated that the tumour vasculature in oligodendrogliomas contributes to their chemosensitivity -LSB- 12 -- 14 -RSB- . In an anaplastic oligodendroglioma treated with PCV chemotherapy , post-therapy volume changes and normalization of rCBV were more evident in regions of the pretherapy tumour that showed high rCBV -LSB- 13 -RSB- . In the present study , tumours with and without the − 1p / − 19q genotype , and high or low rCBV showed response to chemotherapy when assessed using conventional criteria -LSB- 1 , 23 -RSB- . These data indicate that rCBV measured through DSC-MRI is not a key factor in determining PCV chemosensitivity . These findings may reflect the limitations of DSC-MRI associated with contrast recirculation and disruption of the blood -- brain barrier in tumours -LSB- 8 , 9 -RSB- . Alternatively , chemosensitivity may be influenced more by blood flow than by volume . Further studies to measure additional parameters associated with vascular perfusion , including blood flow and permeability are essential to establish whether these factors contribute to chemosensitivity . The − 1p / − 19q genotype is known to be associated with longer PFS and OS -LSB- 5 -RSB- but the impact of tumour vasculature on prognosis in oligodendrogliomas is confined to a few histopathological -LSB- 34 , 36 , 37 -RSB- and imaging studies -LSB- 12 -RSB- . Although our cohort is small , we have presented preliminary data that the prognostic significance of rCBV may differ in tumours with or without the − 1p / − 19q genotype . 1p/19q loss and low-grade pathology were associated with favourable PFS and OS following PCV . In other studies of astrocytic gliomas -LSB- 28 -RSB- or mixed low-grade gliomas -LSB- 11 -RSB- , high rCBV predicted poor prognosis . In our series , rCBV alone was an unreliable indicator of outcome , showing prognostic significance only after stratification for genotype , suggesting different effects for the two genetic subgroups at the cut-off used to classify tumours with high rCBV . With this cut-off , tumours with high rCBV and 1p/19q loss were associated with longer survival than those with high rCBV and intact 1p/19q . Unlike astrocytic gliomas , high rCBV values do not necessarily indicate aggressive biology associated with poor outcome in oligodendroglial neoplasms with 1p/19q loss , suggestive of differences in the baseline biology of these two genetic subtypes . These preliminary data therefore indicate that further investigation of the prognostic significance of rCBV in molecular subtypes of glioma in a much larger series is warranted . Conclusion In conclusion , we have shown that high rCBV is more likely in oligodendroglial tumours with the − 1p / − 19q genotype and may be a useful aid to diagnosis ; however , rCBV does not predict chemosensitivity . Lack of correlation with histopathology grade and the data suggesting that the prognostic significance of rCBV may differ in patients with or without the − 1p / − 19q genotype , indicates that different interpretation of rCBV data may be necessary for oligodendroglial compared with astrocytic gliomas . Further study is essential to fully evaluate the role of DSC-MRI in the non-invasive diagnosis and prediction of prognosis in gliomas with an oligodendroglial phenotype .

O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O B O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O B O O O O O B O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O B O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O B O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O B O O O B O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O B O O O O O O B O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O B O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O B O O O O O O O O B O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O B O O O O O O O O O B O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O B O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O B O O O O O O O O O O O O B O O O O O O O O O B O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O B O O O B O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

oligodendroglial tumour
rcbv
chemosensitivity
outcome


Cyclosporine-A-induced nephrotoxicity in children with minimal-change nephrotic syndrome : long-term treatment up to 10 years The impact of cyclosporine A -LRB- CsA -RRB- therapy in patients with steroid-dependent nephrotic-syndrome -LRB- SDNS -RRB- on long-term renal function is controversial . Data beyond 5 years are rare . Long-term renal function was evaluated in children with SDNS with and without CsA therapy , especially beyond 5 years . Twenty children were treated with CsA -LRB- study group -RRB- for a mean of 5.4 ± 2.2 years -LRB- ten patients for 5 -- 11 years -RRB- . Glomerular filtration rate -LRB- GFR -RRB- was calculated before and after 3 and 12 months and at latest follow-up of therapy . Fifteen children with cyclophosphamide-treated SDNS without CsA served as controls . In the study group , GFR decreased within 12 months from 136 ± 19 to 120 ± 31 , to 114 ± 14 ml/min per 1.73 m2 at latest follow-up -LRB- p < 0.0001 -RRB- . Patients with CsA > 5 years had a GFR of 111 ± 14 ml/min per 1.73 m2 at latest follow-up without a GFR below 90 ml/min per 1.73 m2 . No CsA toxicity was found in biopsies . In the control group , GFR dropped within 3 months , from 137 ± 27 to 130 ± 24 , to 126 ± 19 ml/min per 1.73 m2 at latest follow-up -LRB- p = 0.1 -RRB- . Patients with and without nephrotoxic CsA therapy showed a drop in GFR . In CsA-treated patients , GFR was about 12 % lower at latest follow-up compared with patients without nephrotoxic therapy but always remained within normal range . CsA seems to be safe , even in long-term treatment for more than 5 years . Introduction Minimal-change nephrotic syndrome -LRB- MCNS -RRB- in children is characterised by steroid responsiveness , subsequent relapses and a benign prognosis concerning renal function -LSB- 1 -RSB- . Patients with steroid-dependent nephrotic syndrome -LRB- SDNS -RRB- and frequent relapsing nephrotic syndrome -LRB- FRNS -RRB- with steroid-toxic side effects are recommended for treatment with cyclophosphamide -LRB- CP -RRB- . The overall rate of cumulative sustained remission after therapy with CP , however , was only 24 % after 10 years -LSB- 2 -RSB- . The efficacy of cyclosporine A -LRB- CsA -RRB- in the treatment of steroid-sensitive nephrotic syndrome -LRB- SSNS -RRB- has been well demonstrated -LSB- 3 -- 8 -RSB- . Early withdraw of CsA leads to relapses of the SDNS . So patients remain dependent on CsA for years . CsA was identified as being nephrotoxic , inducing tubulointerstitial fibrosis -LSB- 9 -RSB- , vasoconstriction leading to reduced renal plasma flow and glomerular filtration rate -LRB- GFR -RRB- -LSB- 10 -RSB- . The potential risk of long-term CsA therapy to induce chronic renal failure in patients with MCNS has been discussed controversially . Seikaly et al. -LSB- 11 -RSB- found no drop in GFR but a histological progression of tubulointerstitial lesions in patients treated with CsA in comparison with those without CsA . Hulton et al. -LSB- 5 , 12 -RSB- demonstrated a significant reduction of GFR within the first 3 months after introduction of CsA . Thereafter , the GFR remained stable . In contrast , Inoue et al. -LSB- 6 -RSB- could not detect a significant impairment of renal function in patients with CsA therapy . The interpretation of several studies focussing on the impact of CsA therapy on long-term renal function is limited and controversial . Most authors investigate the influence of CsA on renal function in nephrotic children with different histologic entities , not considering their different risk to progress to chronic renal failure . Thus Niaudet et al. , Habib and Niaudet , and Gregory et al. -LSB- 7 , 13 , 14 -RSB- did not differentiate between MCNS and focal segmental glomerulosclerosis -LRB- FSGS -RRB- in their studies evaluating long-term renal function in nephrotic children with CsA therapy.The physiological drop in GFR after temporary hyperfiltration during a relapse often has been neglected . Relapse-associated hypoproteinemia and hypoalbuminaemia lead to oedema and increased GFR -LSB- 15 -RSB- . Hulton et al. -LSB- 12 -RSB- considered that concomitant therapy with steroids could contribute to an increase of GFR due to the mineralocorticoid effect . Consequently , steroid cessation could lead to a physiological drop in GFR.Several studies claim to address the long-term effect of CsA therapy on renal function , but actually , many of them do not extend more than 2 -- 3 years -LSB- 5 , 6 , 7 , 12 , 16 -RSB- , depicting the short-term effect of CsA on renal function.No study evaluated an additional control group without nephrotoxic therapy to demonstrate the physiological development of GFR in patients with SDNS . In this study , we focus on the long-term renal function in 20 children with biopsy-proven MCNS treated with CsA for a mean of 5.36 ± 2.2 -LRB- range 2 -- 11 -RRB- years . The development of GFR in these patients is compared with a control group with SDNS without CsA therapy . Patients and methods Between 1993 and 2004 , 20 children -LRB- eight girls and 12 boys -RRB- with biopsy-proven MCNS were treated with CsA for a mean of 5.4 ± 2.2 -LRB- median 5.0 , range 2.1 -- 11.0 -RRB- years . Patients only qualified for this retrospective analysis after a minimum CsA therapy duration of 2 years . The definitions and criteria for NS , remission , relapse and steroid dependency were those used by the International Study of Kidney Disease in Children -LRB- ISKDC -RRB- and the Arbeitsgemeinschaft Pädiatrische Nephrolgie -LRB- APN -RRB- -LSB- 17 , 18 -RSB- . All patients had SDNS and were treated with cyclophosphamide -LRB- CP -RRB- prior to CsA without achieving sustained remission . CsA treatment was started with a dosage of 100 -- 150 mg/m2 per body surface area -LRB- BSA -RRB- in two divided doses . CsA dose was adjusted to a target blood level of 80 -- 120 ng/ml . Side effects such as hypertension , hypertrichosis and gingival hyperplasia were not consistently documented and could not be considered in this study . Renal function -LRB- GFR -RRB- was calculated by the Schwartz formula -LRB- -LSB- 19 -RSB- , κ-factor 0.55 -RRB- before and after 3 and 12 months and at the latest follow-up of CsA therapy . Renal biopsy under CsA therapy was performed in five patients after 4.9 -- 7.0 years . All patients starting CsA therapy were still on a standard relapse treatment with prednisone -LRB- 40 mg/m2 every 48 h -RRB- that was discontinued after 4 weeks . At the reviews after 3 and 12 months and latest follow-up , all patients were in remission and had no steroid therapy . In the control group , 15 children -LRB- seven girls , eight boys -RRB- with SDNS treated with CP were evaluated for long-term renal function . GFR was calculated before and after therapy with CP and at the latest follow-up -LRB- mean 4.9 ± 3.4 years -RRB- . According to APN recommendations , these patients were treated with CP with 2 mg/kg body weight for 12 weeks -LSB- 18 , 20 -RSB- . As concomitant medication , all patients in the control group received prednisone 60 mg/m2 every 48 h tapered to 10 mg/m2 every 48 h and discontinued when CP was stopped . At latest follow-up , all patients were in remission without further steroid therapy . Statistical analysis Statistical analysis was performed employing SPSS 12.0 for windows . Data were expressed as mean ± standard deviation . Student 's t test was performed , and a p value < 0.05 was regarded as significant . Results Twenty paediatric patients with SDNS were treated with CsA due to failing CP therapy for a mean of 5.4 ± 2.2 -LRB- median 5.0 , range 2.1 -- 11.0 -RRB- years . The mean starting dosage was 130 ± 32 mg/m2 BSA per day . The mean CsA dosage after 12 months was 126 ± 26 mg/m2 BSA per day , with a trough level of 99 ± 27 ng/ml . The mean age at the beginning of CsA therapy was 8.4 ± 3.0 -LRB- median 8.6 , range 3.1 -- 14.2 -RRB- years . GFR calculated at the beginning , at months 3 and 12 and at the latest follow-up of CsA treatment was 136 ± 19 -LRB- median 136 -RRB- , 130 ± 20 -LRB- median 135 -RRB- , 120 ± 31 -LRB- median 113 -RRB- and 114 ± 14 -LRB- median 114 -RRB- ml/min per 1.73 m2 , respectively -LRB- Table 1 and Fig. 1 -RRB- . Unfortunately , data of only 12 patients were available for the calculation of GFR at month 3 . The decline in GFR from the beginning to month 12 and to the latest follow-up was significant , with p = 0.025 and p < 0.0001 , respectively -LRB- Fig. 2 -RRB- . The drop in GFR within the first 12 months was 11.8 % and during the whole follow-up 14.9 % . Three patients reestablished their GFR after cessation of CsA . No patient had a GFR below 90 ml/min per 1.73 m2 , and 17 patients had a GFR above 105 ml/min per 1.73 m2 at latest follow-up . CsA trough level at latest follow-up was 85 ± 62 ng/ml ; mean CsA dosage was 105 ± 34 mg/m2 BSA per day . Table 1Patient characteristics -LSB- mean ± standard deviation -LRB- SD -RRB- -RSB- and renal function calculated by glomerular filtration rate -LRB- GFR in ml/min per 1.73 m2 ; mean ± SD -RRB- in patients with cyclosporine A -LRB- CsA , study group -RRB- and with cyclophosphamide -LRB- CP , control group -RRB- Study group n = 20Control group n = 15Age at start of MCNS -LRB- years -RRB- 4.4 ± 2.2 4.0 ± 2.9 Age at start of CP -LRB- years -RRB- 5.7 ± 2.26.0 ± 3.2 Age at start of CsA -LRB- years -RRB- 8.4 ± 3.0 -- CsA trough level at month 12 -LRB- ng/ml -RRB- 98.8 ± 26.9 -- Follow-up time -LRB- years -RRB- 5.4 ± 2.24.9 ± 3.4 GFR at start of CP138 .7 ± 24.6 p = 0.4137.7 ± 27.6 p = 0.3 Median 132Median 131.6 Range 119 -- 181Range 95 -- 196GFR at stop of CP130 ± 30.9130.3 ± 24.7 Median 135.9 Median 131.1 Range 82 -- 164Range 93 -- 178GFR at start of CsA136 .3 ± 19.0 p < 0.0001 Median 136.5 Range 97 -- 186GFR at latest follow-up114.5 ± 14.5126.4 ± 19.8 p = 0.1 Median 114.2 Median 124.6 Range 89 -- 135Range 92 -- 156MCNS minimal-change nephrotic syndrome , GFR glomerular filtration rateFig . 1Development of the glomerular filtration rate -LRB- GFR -RRB- of all patients on cyclosporin A -LRB- CsA -RRB- therapy -LRB- n = 20 -RRB- Fig. 2Boxplot demonstrating the development of glomerular filtration rate -LRB- GFR -RRB- in patients on cyclosporin A -LRB- CsA -RRB- therapy -LRB- n = 20 -RRB- CsA was effective in 19 of 20 patients , leading to long-term remission or reduction of relapses to an infrequent relapsing NS -LRB- IRNS -RRB- . Chlorambucil was introduced in one patient due to frequent relapses after CsA cessation . In three patients , arterial hypertension was treated ; in one patient , antihypertensive therapy could be stopped after CsA cessation . Ten patients were treated with CsA for more than 5 -LRB- median 6.5 , mean 7.0 ± 1.8 , range 5.0 -- 11.0 -RRB- years . GFR at latest follow-up was 111 ± 14 -LRB- median 109 , range 90 -- 132 -RRB- ml/min per 1.73 m2 . Five patients underwent renal biopsy after 5.1 -- 7.3 years after CsA therapy had been started before they were transferred to adult nephrologists . CsA-associated nephrotoxicity was not reported by an experienced renal pathologist -LRB- Prof. U. Helmchen , Hamburg , Germany -RRB- . CsA was stopped in nine patients -LRB- mean age 13.1 ± 2.5 years -RRB- after a mean of 4.9 ± 1.9 -LRB- median 4.4 , range 2.3 -- 9.0 -RRB- years due to long-term sustained remission in all but one patient . GFR remained normal after CsA cessation , with 120 ± 25 -LRB- median 116 -RRB- and 123 ± 10 -LRB- median 125 , range 105.18 -- 134.98 -RRB- ml/min per 1.73 m2 at time of therapy cessation and at latest follow-up , respectively . The control group consisted of 15 patients with SDNS who were treated with CP at a mean age of 6.0 ± 3.2 -LRB- median 4.6 -RRB- years . At the start of CP , GFR was 138 ± 28 -LRB- median 132 -RRB- ml/min per 1.73 m2 . After 3 months , GFR dropped to 130 ± 25 ml -LRB- median 131 -RRB- ml/min per 1.73 m2 -LRB- p = 0.3 -RRB- . GFR at latest follow-up -LRB- mean follow-up 4.9 ± 3.4 years -RRB- was 126 ± 20 -LRB- median 125 -RRB- ml/min per 1.73 m2 -LRB- p = 0.1 , Figs. 3 and 4 -RRB- . This drop was in the order of 8 % but not statistically significant . Fig. 3Development of glomerular filtration rate -LRB- GFR -RRB- of patients without cyclosporin A -LRB- CsA -RRB- therapy -LRB- n = 15 -RRB- Fig. 4Boxplot demonstrating the development of glomerular filtration rate -LRB- GFR -RRB- in patients without cyclosporin A -LRB- CsA -RRB- therapy -LRB- n = 15 -RRB- Table 1 summarises the development of GFR in both patient groups , starting with the GFR before CP therapy . Patients of both groups had an initial decline in GFR during CP therapy . Discussion Histologically , the nephrotoxic effect of CsA has been defined by Mihatch et al. -LSB- 9 -RSB- : most frequent lesions attributable to CsA are tubular atrophy , interstitial fibrosis and arteriolar hyalinosis . However , a review of the literature demonstrates that it is difficult to predict the influence of CsA on long-term renal function in patients with MCNS . Some authors did not find a significant reduction in renal function under CsA therapy in nephrotic children -LSB- 4 , 8 -RSB- and only minor changes in renal histology without progression of CsA tubulointerstitial lesions -LSB- 14 , 21 -RSB- . Others emphasised reduced GFR -LSB- 5 , 12 -RSB- , reduced renal plasma flow -LSB- 22 -RSB- or progression of tubulointerstitial lesions histologically -LSB- 11 , 23 -RSB- in patients with NS and CsA therapy . Histology progression could not be associated with decrease in renal function -LSB- 11 , 23 -RSB- or duration of CsA therapy -LSB- 7 -RSB- . Myers et al. -LSB- 24 -RSB- postulated that in case of striped interstitial fibrosis , some nephrons undergo atrophy while their function could be compensated for by unaffected nephrons . This may mask nephrotoxicity by measurements of normal GFR . In summary , a clear proven association between impaired renal function and long-term CsA therapy in patients with MCNS could not be found . But hints permit the question regarding the safety of long-term CsA therapy in children with a benign renal disease . Comparison of results of this study with established opinions -LRB- e.g. -LSB- 8 , 12 , 23 -RSB- -RRB- is difficult : Most studies mix different aetiological entities with different potential to progress to chronic renal failure -LRB- MCNS vs. SRNS and FSGS ; -LSB- 4 , 7 , 13 , 16 , 21 , 22 -RSB- -RRB- . The impairment of renal function in patients with FSGS and other types of SRNS can not reliably be distinguished from CsA nephrotoxicity , as those entities have a high risk for chronic renal failure on their own.In most studies , the follow-up period is restricted to 2 -- 3 years ; in single studies up to 5 years . In this study , at least ten patients underwent long-term CsA therapy > 5 years . No other study investigated patients with such a long treatment period -LSB- 5 -- 8 , 11 -- 14 , 23 -RSB- . Neuhaus et al. and Ganesan et al. . -LSB- 8 , 23 -RSB- evaluated patients with MCNS only and an extended follow-up time of 5 years . They evaluated nine patients , with one being identified with FSGS as the underlying disease in the follow-up biopsy . All patients received CsA with higher trough levels and higher CsA dosage -LRB- level : mean of 220 ng/ml , range 141 -- 270 ng/ml , CsA dosage 126 ± 29 mg/m2 per day -RRB- than accepted for our study group . The method for CsA level measurements was a polyclonal assay resulting in higher levels compared with monoclonal assays . Nevertheless , the CsA levels and dosage were still higher than in our study group . CsA toxicity was proven in 3/9 patients histologically . Ganesan et al. -LSB- 23 -RSB- found histological signs for CsA toxicity in 79 % and renal insufficiency with a GFR < 80 in 4/19 patients without correlation to histological changes . Again , that study mixed patients with steroid-resistant and steroid-responsive NS , not distinguishing between the different clinical entities . In our study , only patients with histologically proven MCNS were evaluated for long-term renal function under CsA therapy . Patients qualified for the study only after a minimum treatment duration of 2 years . The follow-up period extended over a mean of 5.4 years , with ten patients who had been treated between 5 and 11 years . At least in half of the patients with long-term CsA therapy > 5 years , CsA toxicity was excluded by biopsy . Also in our study group , the GFR of patients with CsA therapy remained within the normal range , with a drop within the first 12 months -LRB- mean drop 11.8 % -RRB- and remaining stable thereafter , with a mean GFR of 115 ± 16 ml/min per 1.73 m2 . No patient progressed to chronic renal failure , even after therapy duration of more than 10 years . Hulton et al. -LSB- 5 , 12 -RSB- showed comparable results for patients with CsA therapy , with an initial drop in GFR and a constant renal function afterwards . But the study is restricted to a short follow-up of 2 years in four patients only . In our study , a significant reincrease in GFR after therapy cessation was not found . In general , these patients showed excellent renal function in the long-term follow-up -LRB- mean 123 ml/min per 1.73 m2 -RRB- . In contrast , patients in Hulton et al. 's studies showed a worse GFR of 93 that reincreased back to a GFR > 100 ml/min per 1.73 m2 after CsA had been stopped . Interestingly , in our control group of patients without CsA therapy , an initial drop in GFR was also found . Those patients showed a mean drop in GFR of about 5 % within the 12 weeks of CP therapy , and GFR even dropped about 8 % until the latest follow-up . Probably , the observed decrease in GFR in patients with CsA therapy can not only be attributed to the hitherto described nephrotoxic effect of CsA . Hulton et al. -LSB- 12 -RSB- mentioned the potential increase of GFR caused by the mineralocorticoid effect of concomitant steroid medication . One could speculate that the cessation of steroids might lead to a physiological drop in GFR . Additionally , a possible but not proven interpretation should be discussed : Part of the GFR reduction may be attributable to normalisation of hyperfiltration in nephrotic children during the first months after initiation of immunosuppressive therapy . In patients with relapsing nephrotic syndrome , long-term immunosuppressive drugs -LRB- e.g. cyclophosphamide , CsA , chlorambucil -RRB- are initiated after urine remission has been achieved ; complete remission of serum albumin normally is not awaited . Relapse-associated hypoproteinemia and hypoalbuminemia lead to oedema and an increase in GFR . This is explained by the decrease of the colloid osmotic pressure in the glomerular capillary . Reduced total serum protein leads to decreased colloid osmotic pressure and consequently to increased GFR -LSB- 15 -RSB- . Normalisation of the increased filtration extends over weeks and leads to a decrease in GFR physiologically . This phenomenon may explain the drop in GFR , even in patients without nephrotoxic treatment . In summary , this study offers long-term follow-up of 20 patients with proven SSNS treated with CsA for a median of 5 years . Initially , the patients showed a drop in GFR but remained stable afterwards . None developed chronic renal failure . Nevertheless , the generalisation of these results are limited , as the study was retrospective , and only 5/20 patients underwent renal biopsy under long-term nephrotoxic treatment . It is questionable whether a prospective study to evaluate the outcome of long-term CsA therapy in children with MCNS is reasonable in the future . Newer , nonnephrotoxic drugs , e.g. mycophenolate mofetil -LRB- MMF -RRB- are of concern and need to be evaluated for efficacy and safety in the treatment of SDNS and FRNS . However , our analysis demonstrates that even long-term therapy with CsA for more than 5 years in children with MCNS is safe and does not impair renal function .

O B O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I I O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I I O O O O O O O O O O O O O O O O O O O O O O O O B I I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I I O O O O O O O O O O O O O O O O O O O O O O O B I I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

cyclosporine a
nephrotoxicity
glomerular filtration rate

minimal-change nephrotic syndrome
long-term outcome