Objective To evaluate tumor vascularity by threedimensional
power Doppler ultrasound (3D-PDU) in
early and advanced stage primary ovarian cancers and
in metastatic tumors to the ovary.
Patients and methods This was a retrospective analysis
of clinical and sonographic data from 49 women
with primary ovarian cancers or metastatic tumors to
the ovary. All women underwent 3D-PDU prior to
surgery. Vascularization index (VI), flow index (FI) and
vascularization flow index (VFI) from solid portions or
papillary projections in the tumors were calculated using
the Virtual Organ Computer-aided AnaLysis (VOCALTM)
program. Definitive histological diagnosis was obtained
in each case.
Results Among the 49 women, 10 had stage I primary
cancers (five low-malignant potential tumors and five
invasive tumors), 26 had advanced stage primary ovarian
cancers and 13 had metastatic tumors to the ovary. Mean
VI and VFI were significantly higher in advanced stage
tumors and metastatic tumors as compared with early
stage tumors. No differences in 3D-PDU indices were
found between advanced stage and metastatic cancers.
Conclusions Vascular indices derived from 3D-PDU tend
to be higher in advanced stage and metastatic ovarian
cancers as compared with early stage ovarian tumors

Abulafia

Alcazar

Alvarez

Bourne

Cheng

Denis

Emoto

Folkman

Hata

Hollingsworth

Kurjak

Pairleitner

Paley

Raspollini

Sedelaar

Serov

Sheperd

Sonmezer

Stone

Testa

Valentin

Yang

Alcazar, J.L. (Juan Luis)

Universidad de Navarra

Tumor angiogenesis assessed by three-dimensional power Doppler ultrasound in early, advanced and metastatic ovarian cancer: a preliminary study

Crossref

Dadun, University of Navarra

Ultrasound Obstet Gynecol 2006; 28: 325–329Published online in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/uog.3804Tumor angiogenesis assessed by three-dimensional powerDoppler ultrasound in early, advanced and metastaticovarian cancer: a preliminary studyJ. L. ALCA´ZARDepartment of Obstetrics and Gynecology, Clı´nica Universitaria de Navarra, University of Navarra, Pamplona, SpainKEYWORDS: 3D ultrasound; ovarian cancer; power DopplerABSTRACTObjective To evaluate tumor vascularity by three-dimensional power Doppler ultrasound (3D-PDU) inearly and advanced stage primary ovarian cancers andin metastatic tumors to the ovary.Patients and methods This was a retrospective analysisof clinical and sonographic data from 49 womenwith primary ovarian cancers or metastatic tumors tothe ovary. All women underwent 3D-PDU prior tosurgery. Vascularization index (VI), flow index (FI) andvascularization flow index (VFI) from solid portions orpapillary projections in the tumors were calculated usingthe Virtual Organ Computer-aided AnaLysis (VOCALTM)program. Definitive histological diagnosis was obtainedin each case.Results Among the 49 women, 10 had stage I primarycancers (five low-malignant potential tumors and fiveinvasive tumors), 26 had advanced stage primary ovariancancers and 13 had metastatic tumors to the ovary. MeanVI and VFI were significantly higher in advanced stagetumors and metastatic tumors as compared with earlystage tumors. No differences in 3D-PDU indices werefound between advanced stage and metastatic cancers.Conclusions Vascular indices derived from 3D-PDU tendto be higher in advanced stage and metastatic ovariancancers as compared with early stage ovarian tumors.Copyright  2006 ISUOG. Published by John Wiley &Sons, Ltd.INTRODUCTIONAngiogenesis plays an essential role in tumor growth andmetastasis1. Several studies using immunohistochemicaltechniques have demonstrated that, in ovarian cancer,angiogenesis may be a prognostic factor2,3. Angiogenesismay be assessed in vivo using pulsed and color Dopplerultrasonography, the conventional methods for assess-ing tumor vascularization. In ovarian tumors Dopplerultrasound has been used mainly for discriminatingbetween benign and malignant lesions. Although ini-tial studies on color and spectral pulsed Doppler wereencouraging4, further studies revealed a wide overlap inpulsed Doppler indices between malignant and benigntumors, making this approach impractical from the clin-ical point of view5. Three-dimensional power Dopplerultrasound (3D-PDU) is a relatively new technique thatallows tumor vascularization assessment, both quantita-tively by means of 3D-PDU-derived vascular indices6 andqualitatively by depicting three-dimensionally the tumorvascular network7.Some studies using immunohistochemical staining havefound quantitative differences in angiogenesis in ovariancancer according to tumor stage8 and other studies haveshown that Doppler assessment of tumor vascularizationis related to microvessel density in cervical, prostateand breast cancer9–11. The aim of the present studywas to evaluate whether differences exist in tumorvascularization as assessed by 3D-PDU in early andadvanced stage primary ovarian cancers and in tumorsthat have metastasized to the ovary.PATIENTS AND METHODSIn this retrospective study data from 49 women diagnosedas having primary or metastatic ovarian cancer, and whowere evaluated and treated at our institution betweenJanuary 2003 and October 2005, were reviewed. A groupof these patients are included in a previous study12.Correspondence to: Dr J. L. Alca´zar, Department of Obstetrics and Gynecology, Avenida Pio XII, 36 31008 Pamplona, Spain(e-mail: jlalcazar@unav.es)Accepted: 3 April 2006Copyright  2006 ISUOG. Published by John Wiley & Sons, Ltd. ORIGINAL PAPER326 Alca´zarThe patients’ mean age was 52.9 (range, 17–82) years.Twenty-eight (57.1%) women were postmenopausal and21 (42.9%) women were premenopausal. Four womenhad bilateral tumors identifiable at ultrasound but onlyone tumor per patient was used for analysis in these fourwomen.All the women had been evaluated by transvaginalultrasound using a Voluson 730 Pro (GE Healthcare, Zipf,Austria) with a 5–7.5-MHz transvaginal probe and color,power and pulsed Doppler as well as three-dimensional(3D) ultrasound capabilities. Transabdominal ultrasound(3.5–5-MHz) was also performed in the case of largetumors.The scanning technique has been described in detailelsewhere12. Following B-mode evaluation, the two-dimensional power Doppler gate was activated to assesstumor vascularization. Power Doppler settings were setto achieve maximum sensitivity to detect low velocityflow without artifacts (frequency, 5 MHz; power Dopplergain, 20 (range, 1–30); dynamic range, 20–40 dB;edge, 1; persistence, 2; color map, 1; gate, 2; filter,3; pulse repetition frequency, 0.6 kHz). Central vesselvascularization was defined as the presence of colorspots within papillary projections, solid areas or thecentral part of solid tumors. Power Doppler imaging wasused to identify vessels for subsequent pulsed Dopplerinterrogation to obtain a flow velocity waveform and toconfirm the arterial nature of the vessel. Pulsatility index(PI), resistance index (RI) and peak systolic velocity (PSV,cm/s) were recorded automatically. The lowest PI and RIand the highest PSV found in a given tumor, independentlyfrom the vessel in which they were obtained, were usedfor analysis. In cases in which no arterial flow was found(n = 2) pulsed Doppler data were not calculated.For 3D imaging the volume was activated to obtain a3D box from either papillary projections, solid areas or,in the case of mostly solid tumors, the whole tumor. Oncea 3D volume had been obtained, it was stored on a harddisk in the ultrasound machine. The stored volumes werefurther analyzed using the Virtual Organ Computer-aidedAnaLysis (VOCAL) program (SonoviewTM, Kretztechnik,Zipf, Austria). Volume acquisition time lasted from 15 to20 s depending on the size of the volume box. Using theVOCAL program, three vascular indices were calculatedfrom every solid area or papillary projection with colorsignals within it, excluding cystic (fluid-filled) areas inwhich blood vessels would not be found (Figures 1 and 2).The three-dimensional volume consists of ‘voxels’(smallest unit of volume). Voxels contain informationabout gray-scale and color according to an intensity scaleranging from 0 to 100. The vascular indices calculatedwere the vascularization index (VI), which is expressedas a percentage and represents the relative proportion ofpower Doppler data within the defined volume; the flowindex (FI), which is the mean signal intensity of this powerDoppler information; and the vascular flow index (VFI),which is a combination of both6.Measurements were undertaken using the manualmode, in plane ‘A’ and a 9◦-rotation step. If there wasFigure 1 (a) Transvaginal ultrasound scan showing a solid cysticvascularized adnexal mass. (b) Tumor volume estimation using theVirtual Organ Computer-aided AnaLysis (VOCAL) program. Inthis case only the solid portion of the tumor is included in theanalysis. (c) Three-dimensional power Doppler-derived indicesobtained using the VOCAL program. This case was a stage IIIcserous–papillary adenocarcinoma of the ovary.more than one non-contiguous solid area, each area wasevaluated separately and the highest VI, FI and VFIvalues (which could have been obtained from differentareas) were used for analysis. If the tumor showed areasCopyright  2006 ISUOG. Published by John Wiley & Sons, Ltd. Ultrasound Obstet Gynecol 2006; 28: 325–329.Tumor angiogenesis in early, advanced and metastatic ovarian cancer 327Figure 2 (a) Transvaginal ultrasound scan showing a multilocularcystic lesion with a solid area arising from internal cyst wall.(b) Transvaginal two-dimensional power Doppler ultrasound scanshowing color signals within solid area. (c) Solid area volumeestimation using the Virtual Organ Computer-aided AnaLysis(VOCAL) program.of low echogenicity, this area was not included in theVOCAL calculation. All calculations were performed onthe ultrasound machine.All ultrasound examinations and 3D analyses wereperformed by a single operator. Patients were selectedaccording to availability of the examiner and ultrasoundmachine. As this was a retrospective study no InstitutionalReview Board approval was sought.All patients underwent surgery, and a definitivehistological diagnosis was obtained in every case.Tumors were classified according to the World HealthOrganization (WHO) criteria13 and ovarian cancerswere staged according to International Federation ofGynecology and Obstetrics (FIGO) criteria14.The Kolmogorov–Smirnov test was used to assessnormal distribution of continuous data. As all continuousdata showed a skewed distribution they were comparedusing the Kruskal–Wallis test and the Mann–WhitneyU-test with Bonferroni correction. Categorical variableswere compared using the Chi-square test. P ≤ 0.05 wasconsidered to be statistically significant. All statisticalanalyses were performed using the SPSS 11.0 statisticalpackage (SPSS Inc, Chicago, IL, USA).RESULTSOf the 49 patients, five (10.2%) had tumors of lowmalignant potential, 31 (63.3%) had primary invasivecarcinomas and 13 (26.5%) had metastatic tumors to theovary (source: colon, six cases; endometrium, two cases;breast, two cases and liver, mesothelioma and lung onecase each.). Histological diagnoses in primary ovariancancers, including low-malignant potential tumors, wereas follows: epithelial cancer 32 cases, sarcoma onecase, immature teratoma one case, limphoma one case,Sertoli–Leydig tumor one case.All low-malignant potential tumors and five primaryinvasive carcinomas were stage I, so these 10 cases weregrouped as stage I cancers. Twenty-six women withprimary invasive cancers had advanced stage tumors(three stage II, 18 stage III and five stage IV). Twowomen with stage III and two with metastatic cancers hadbilateral tumors. B-mode findings according to tumoralstage are shown in Table 1. Metastatic tumors were morefrequently solid lesions than were early and advancedstage primary ovarian cancers (P = 0.009).Median VI and VFI were significantly higher inadvanced stage tumors as compared with early stagetumors; they were also significantly higher in metastatictumors as compared with early stage tumors. Nodifferences were found in FI when comparing earlystage cancers with advanced stage and metastatic cancers.No differences were found between advanced stage andmetastatic cancers in VI, FI and VFI and no differenceswere found in RI, PI and PSV among all three groups(Table 2).DISCUSSIONUsually tumor angiogenesis is evaluated by assess-ing microvessel density following immunohistochemicalstaining for several endothelial antigens such as CD34,CD31, factor VIII related antigen and VEGF8,15. SomeCopyright  2006 ISUOG. Published by John Wiley & Sons, Ltd. Ultrasound Obstet Gynecol 2006; 28: 325–329.328 Alca´zarTable 1 B-mode findings according to tumoral stageStageUnilocular solid,n (%)Multilocularsolid, n (%) Solid, n (%) Total, n (%)Early stage, n = 10 4 (40) 5 (50) 1 (10) 10 (100)Advanced stage, n = 26 5 (19.2) 7 (26.9) 14 (53.8) 26 (100)Metastatic tumors, n = 13 1 (7.7) 3 (23.1) 9 (69.2) 13 (100)Table 2 Three-dimensional power-Doppler and pulsed Doppler indices according to stage of ovarian cancer. Data are expressed as median(interquartile range)Stage VI VFI FI RI PI PSV (cm/s)Early stagea, n = 10 9.57 (8.5) 3.06 (3.1) 31.21 (6.1) 0.40 (0.10) 0.59 (0.15) 16.8 (8.5)Advanced stageb, n = 26 18.02 (10.8) 5.91 (4.1) 33.89 (7.2) 0.45 (0.24) 0.66 (0.45) 14.2 (11.3)Metastatic tumorsc, n = 13 15.10 (11.5) 5.60 (3.2) 34.79 (6.4) 0.42 (0.15) 0.59 (0.31) 14.0 (10.9)P a vs. b 0.022 a vs. b 0.005 a vs. b 0.132 a vs. b 0.682 a vs. b 0.501 a vs. b 0.589a vs. c 0.014 a vs. c 0.032 a vs. c 0.192 a vs. c 0.740 a vs. c 0.740 a vs. c 0.976b vs. c 0.857 b vs. c 0.814 b vs. c 0.879 b vs. c 0.393 b vs. c 0.195 b vs. c 0.816FI, flow index; PI, pulsatility index; PSV, peak systolic velocity; RI, resistance index; VFI, vascularization flow index; VI, vascularizationindex.have found that angiogenesis is a prognostic factor both inearly16 and advanced stage ovarian carcinomas17. How-ever, some researchers have challenged these findings18.Nonetheless, microvessel density is higher in advancedstage tumors as compared with early stage ones3,8.Vascularization can be assessed in vivo by Dopplerultrasonography. A correlation seems to exist betweenthe pulsed Doppler indices RI and PI19 or color bloodflow mapping20 and tumor microvessel density. 3D-PDU, theoretically, provides a more comprehensivemeans of assessing tumor vascularization than does two-dimensional (2D) color or power Doppler by the use ofthe vascular indices VI, FI and VFI6 and three-dimensionaldepiction of the tumor vascular network7. Some studieshave shown that 3D-PDU may be useful for differentiatingbenign from malignant ovarian tumors21–23.Since microvessel density tends to be higher in advancedstage ovarian cancers than in early stage tumors, it islogical to assume that vascular indices derived from 3D-PDU would be higher in advanced stage ovarian cancersthan in early stage tumors. Apparently, no previousstudy has used 3D-PDU to assess tumor vascularizationaccording to tumoral stage, and only one previous studyhas been published that compared PSV (not RI and PI)according to ovarian cancer stage24. According to thatstudy PSV did not differ between early and advancedstage ovarian cancer24. In another study we found nodifferences in pulsed Doppler indices between metastaticand primary ovarian cancer25.In conclusion, vascularization, as assessed by 3D-PDU indices, is higher in advanced stage and metastaticovarian cancers than in early stage ovarian cancer. Thesepreliminary results may be of value for future research.It would be worth exploring whether vascular indicesderived from 3D-PDU in ovarian cancer could be used asa prognostic factor for this condition.REFERENCES1. Folkman J, Watson K, Ingber D, Hanahan D. Induction ofangiogenesis during the transition from hyperplasia to neoplasia.Nature 1989; 339: 58–61.2. 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Tumor angiogenesis assessed by three-dimensional power Doppler ultrasound in early, advanced and metastatic ovarian cancer: a preliminary study

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