Marian Vanhoeij

Phase I Study of 68Ga-HER2-Nanobody for PET/CT Assessment of HER2 Expression in Breast Carcinoma

Human epidermal growth factor receptor 2 (HER2) status is one of the major tumor characteristics in breast cancer to guide therapy. Anti-HER2 treatment has clear survival advantages in HER2-positive breast carcinoma patients. Heterogeneity in HER2 expression between primary tumor and metastasis has repeatedly been described, resulting in the need to reassess HER2 status during the disease course. To avoid repeated biopsy with potential bias due to tumor heterogeneity, Nanobodies directed against HER2 have been developed as probes for molecular imaging. Nanobodies, which are derived from unique heavy-chain-only antibodies, are the smallest antigen-binding antibody fragments and have ideal characteristics for PET imaging. The primary aims were assessment of safety, biodistribution, and dosimetry. The secondary aim was to investigate tumor-targeting potential. Methods: In total, 20 women with primary or metastatic breast carcinoma (score of 2+ or 3+ on HER2 immunohistochemical assessment) were included. Anti-HER2-Nanobody was labeled with 68Ga via a NOTA derivative. Administered activities were 53–174 MBq (average, 107 MBq). PET/CT scans for dosimetry assessment were obtained at 10, 60, and 90 min after administration. Physical evaluation and blood analysis were performed for safety evaluation. Biodistribution was analyzed for 11 organs using MIM software; dosimetry was assessed using OLINDA/EXM. Tumor-targeting potential was assessed in primary and metastatic lesions. Results: No adverse reactions occurred. A fast blood clearance was observed, with only 10% of injected activity remaining in the blood at 1 h after injection. Uptake was seen mainly in the kidneys, liver, and intestines. The effective dose was 0.043 mSv/MBq, resulting in an average of 4.6 mSv per patient. The critical organ was the urinary bladder wall, with a dose of 0.406 mGy/MBq. In patients with metastatic disease, tracer accumulation well above the background level was demonstrated in most identified sites of disease. Primary lesions were more variable in tracer accumulation. Conclusion: 68Ga-HER2-Nanobody PET/CT is a safe procedure with a radiation dose comparable to other routinely used PET tracers. Its biodistribution is favorable, with the highest uptake in the kidneys, liver, and intestines but very low background levels in all other organs that typically house primary breast carcinoma or tumor metastasis. Tracer accumulation in HER2-positive metastases is high, compared with normal surrounding tissues, and warrants further assessment in a phase II trial.

Scapula alata in early breast cancer patients enrolled in a randomized clinical trial of post-surgery short-course image-guided radiotherapy

BackgroundScapula alata (SA) is a known complication of breast surgery associated with palsy of the serratus anterior, but it is seldom mentioned. We evaluated the risk factors associated with SA and the relationship of SA with ipsilateral shoulder/arm morbidity in a series of patients enrolled in a trial of post-surgery radiotherapy (RT).MethodsThe trial randomized women with completely resected stage I-II breast cancer to short-course image-guided RT, versus conventional RT. SA, arm volume and shoulder-arm mobility were measured prior to RT and at one to three months post-RT. Shoulder/arm morbidities were computed as a post-RT percentage change relative to pre-RT measurements.ResultsOf 119 evaluable patients, 13 (= 10.9%) had pre-RT SA. Age younger than 50 years old, a body mass index less than 25 kg/m2, and axillary lymph node dissection were significant risk factors, with odds ratios of 4.8 (P = 0.009), 6.1 (P = 0.016), and 6.1 (P = 0.005), respectively. Randomization group was not significant. At one to three months’ post-RT, mean arm volume increased by 4.1% (P = 0.036) and abduction decreased by 8.6% (P = 0.046) among SA patients, but not among non-SA patients. SA resolved in eight, persisted in five, and appeared in one patient.ConclusionThe relationship of SA with lower body mass index suggests that SA might have been underestimated in overweight patients. Despite apparent resolution of SA in most patients, pre-RT SA portended an increased risk of shoulder/arm morbidity. We argue that SA warrants further investigation. Incidentally, the observation of SA occurring after RT in one patient represents the second case of post-RT SA reported in the literature.

Early Contralateral Shoulder-Arm Morbidity in Breast Cancer Patients Enrolled in a Randomized Trial of Post-Surgery Radiation Therapy:

Introduction Shoulder/arm morbidity is a common complication of breast cancer surgery and radiotherapy (RT), but little is known about acute contralateral morbidity. Methods Patients were 118 women enrolled in a RT trial. Arm volume and shoulder mobility were assessed before and 1–3 months after RT. Correlations and linear regression were used to analyze changes affecting ipsilateral and contralateral arms, and changes affecting relative interlimb differences (RID). Results Changes affecting one limb correlated with changes affecting the other limb. Arm volume between the two limbs correlated (R = 0.57). Risk factors were weight increase and axillary dissection. Contralateral and ipsilateral loss of abduction strongly correlated (R = 0.78). Changes of combined RID exceeding 10% affected the ipsilateral limb in 25% of patients, and the contralateral limb in 18%. Aromatase inhibitor therapy was significantly associated with contralateral loss of abduction. Conclusions High incidence of early contralateral arm morbidity warrants further investigations.

The influence of multidisciplinary rehabilitation on physical well-being and quality of life of breast cancer survivors.

143 Background: Breast cancer treatment has adverse effects. The aim of this study was to examine the effects of a multidisciplinary oncologic rehabilitation program on health related quality of life (HRQoL), cancer related fatigue (CRF), muscle strength, physical fitness and anthropometrics in breast cancer survivors. METHODS This quasi-experimental study included 30 early breast cancer patients in the first year following treatment. Patients completed a 12-week exercise program for 4 hours a week combined with lifestyle guidance for 2 hours a week. The supervised training sessions consisted of aerobic exercises combined with muscular strengthening exercises. Measurements were carried out at baseline (T0), at the end of the intervention (T1) and at 12-weeks follow-up (T2). HRQoL (EORTC QLQ-C30 questionnaire) and CRF (FACIT-Fatigue questionnaire), were measured at T0, T1 and T2. Muscle strength (handgrip dynamometer) was measured at T0 and T1. Physical fitness and anthropometrics were assessed at T0 and T1 using spiro ergometrics, bioimpedance and waist- and hip circumference. RESULTS Significant positive changes in HRQoL were found, especially for physical functioning (p = 0.004) and dyspnea (p = 0,003) at T1, but HRQoL decreased at T2. Weight, BMI, waist - and hip circumference and fat free mass decreased significantly (respectively p = 0,030; p = 0,047; p = 0,020; p = 0,041 and p = 0,003). Body impedance increased significantly over time (p = 0,034). There was a significant improvement in CRF at T1 (p = 0.03), that was no longer significant at T2. No significant improvements were found in muscle strength at the affected side (p = 0.16) and the non-affected side (p = 0.95). Physical fitness increased significantly for VO2max at the maximal progressive cycle test (p = 0.005). CONCLUSIONS This study reports significant improvements in HRQoL, anthropometric characteristics, CRF and physical fitness after a 12-week rehabilitation program. The declines between T1 and T2 may be explained by discontinuation of physical activity. Further research should use randomized clinical trials to examine the effectiveness of rehabilitation programs with different contents, duration and initiation.

Impact of Management Changes on Loco-Regional Control of Breast Carcinoma: A 30-year Single Institution Experience Breast Cancer: Basic and Clinical Research

Purpose: This institutional-based study aims to reflect changes in diagnosis, surgery, radiotherapy, systemic therapy by retrospectively analysing treatment modalities and outcome during the past 30 years of breast cancer. We hypothesized these changes result in better outcome. Material and methods: 2990 women are included, aged 18–95, no previous cancer, unilateral stage I-III primary breast tumors, breast-conserving surgery (BCS) or mastectomy (ME), postoperative radiotherapy (RT) and where indicated systemic treatment. Patients were divided in 3 cohorts stratified by year of diagnosis: 1984-1991, 1992- 1999 and 2000-2008. The interval of cohorts was based on institutional changes in systemic regimens. Results: Over time, median age at diagnosis was similar, patients >70 year increased (19.5 to 25.7%). Over the 3 cohorts: stage migration is observed, determination of tumor grading became routine, proportions of known ER/PR status increased. Over time an obvious shift to less mutilating surgery is observed. Systemic treatment increased significantly during the observed period. In stage I disease, overall (OS), local control (LC) and disease free survival (DFS) didn’t change. In stage II, a significant increase in 10 years OS and DFS (p= 0.02 and 0.001) is observed. In stage III we noticed a significant increase in 10 years DFS (p=0.04) and trend in increase of 10 years OS (p= 0.06). Local Recurrence free survival (RFS) didn’t change significantly for all stages. Conclusion: This study demonstrated an improved outcome for stage II and III over time in our population with the same local control. This is multifactorial, reflecting changes in diagnostic imaging, surgery and increased use of systemic therapy.

Abstract PD7-04: Exome based germline mutation detection in a panel of 372 cancer associated genes in BRCA1/2-negative familial breast cancer patients

Background: Ten to 20% percent of all breast cancers occur in a familial context and in 20-30% of these cases a mutation in the BRCA1 or BRCA2, CHEK2 genes or, more rarely, in PALB2 can be found. The remaining cases remain routinely undiagnosed with regard to a possible genetic cause. We have examined a cohort of undiagnosed probands using exome germline sequencing in order to identify other potential breast cancer predisposition genes. Methods: In total, 63 BRCA1/2-negative high risk familial BC cases (BRCAX) were considered for pair-end whole exome germline DNA sequencing on a HiSeq1500 (Illumina). High quality reads were mapped (BWA-MEM) to the reference genome (hg19) and variants were called according to GATK best practice guideline. The variants detected within the panel of 372 cancer associated genes were annoted with ANNOVAR. Synonymous variant as well as variants with MAF>1% were discarded. In a first phase protein truncating variants were validated using another NGS method. Subsequent validation of non-synonymous missense mutations and non-frameshift indels is planned. Patients signed a multilayered informed consent also covering disclosure or not of different types of incidental findings. Results: For each exome, the mean breath of coverage was about 96 % at 10X or more and the mean depth of coverage for targeted region was about 126X. In total, 3570709 SNPs (∼56678 SNPs/sample) and 477801 INDELs (7584 INDELs/sample) were called. Of them, 20829 SNPs (331 SNPs/sample) and 16071 INDELs (255 INDELs/sample) passed quality filter. In total, 445 variants were found in the publicly available cancer genes panel. Twenty-seven stop-gain/loss, frame-shift insertion/deletion and splice site variants were considered for validation with 454 Roch Junior, among which 22 variants in genes ABCC11, AFP, BARD1, BBS10, CD96, CYP1A1, DNAH11, ESCO2, EXO1, FANCI, FLCN, FLT4, HPS6, MYH8, NME8, PALB2, PDE11A, RECQL4, TTC8 were validated as true positive. Some of these genes have been found earlier to be associated with breast cancer and/other cancer types. Functional prioritization of the remaining 416 non-synonymous and non-frameshift insertion/deletions was also done in-silico before further sequencing validation, which is ongoing. The mutations found are further clinically validated by examining other affected and non-affected family members and mining the literature. Some of the mutations in known cancer predisposing genes are considered for prudent application in clinical counseling. Genotype-phenotype correlations are being examined. Conclusion: Next-generation sequencing enabled us to detect variants with high/low penetrance in known cancer predisposing genes in > 35% of BRAX families, in addition to many novel variants in many other genes not yet tied to cancer predisposition occurring singly or in combination with known cancer gene mutations. Validated variants are further examined in the families for co-segregation with the disease and potential application in counseling. Citation Format: Shahi RB, Caljon B, De Brakeleer S, Decoster L, Fontaine C, Vanacker L, Vanhoeij M, Pauwels I, Bonduelle M, Vandooren S, Croes D, Teugels E, De Greve J. Exome based germline mutation detection in a panel of 372 cancer associated genes in BRCA1/2-negative familial breast cancer patients. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr PD7-04.

Abstract P1-12-11: Prospective study of aromatase inhibitor-induced bone loss and lipid levels in early hormone sensitive breast cancer treated with AI during 8 years

Introduction : AI’s are the preferred adjuvant hormonal treatment for postmenopausal(PM) estrogen receptor positive (ER+) disease because of the improved efficacy over tamoxifen in terms of disease free survival and reduction in distant recurrence. AI’s have been associated with increased bone loss by blocking peripheral tissue estrogen synthesis, arthralgia and a negative effect on lipid metabolism. In this prospective study we wanted to assess the amount of bone loss and the changes in lipid levels in early breast cancer(EBC) patients (pts) treated with 3yrs of extended letrozole as part of the SOLE study after 5 yrs of adjuvant AI. Objectives: the primary objective of the study is to calculate the mean percentage change in bone mineral density(BMD)(g/cm 2 ) after 8 yrs of letrozole and to compare between the continuous(C) and intermittent(I) intake of letrozole. The secondary objectives are to correlate the mean change in BMD with the initial value, BMI and to describe the evolution of the lipid levels in both groups. Patients and methods: BMD was measured at the lumbar spine (L2-4) and hip by dual energy X-ray absorptiometry (DEXA) in PM pts. with ER+ EBC. We also measured the fasting lipid levels baseline and after 8 yrs of an AI. Differences between the two groups were assessed by the independent samples T-test and the correlation by the linear regression analysis. Results: Fifty four pts were included in the study with a mean age of 62.5 yrs(+/-8.6 yrs). Seventeen pts are too early to be evaluable, 8 pts stopped letrozole due to adverse events and 2 pts had no baseline DEXA values. Two pts received tamoxifen 2 to 5 yrs before inclusion in the SOLE study. The remaining 25 pts showed a mean change in BMD for the lumbar spine of -1.1 (10.6SD) and for the hip -4.4 (7.04SD). Currently 13 pts in the C arm demonstrated a mean decrease in BMD for the (LS) of -2.7(10.2SD), and for the hip of -2.7(5.5SD). Twelve pts included in the I arm experienced a mean increase in BMD for the LS of 0.65(SD11.2) and a mean decrease in BMD for the hip of -4.9(9.1SD). The difference between the two treatment groups was not significant for the LS measurements after 8yrs (p=0.4) nor for the hip (p=0.23) neither was it at baseline (p=0.9; 0.1). Only three pts had fractures due to trauma. The overall fracture rate was only 0.05%.There was no correlation with the BMI, but there was a significant correlation with baseline BMD.(p=0.002)The mean fasting cholesterol levels at 8yrs in the C arm was 214mg/dl(41.3SD) and in the I arm was 218mg/dl(35.6SD)and was not significantly different(p=0.8), nor was it at baseline(p=0.5). Conclusion : This first prospective long term BMD and lipid follow-up study in PM EBC pts taking adjuvant letrozole beyond 5 yrs shows a decline in BMD. Until now no significant differences were observed between continuous and intermittent letrozole. An updated and detailed follow-up on more patients will be presented. Citation Format: Christel Fontaine, Lore Decoster, Denis Schallier, Leen Vanacker, Jacques De Greve, Marian Vanhoeij, Guy Verfaillie, Jan Lamote. Prospective study of aromatase inhibitor-induced bone loss and lipid levels in early hormone sensitive breast cancer treated with AI during 8 years [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P1-12-11.

Abstract P1-13-06: Correlation between aromatase-induced arthralgia and inflammatory cytokines in postmenopausal patients with early breast cancer treated with surgery followed by adjuvant hormone therapy: A prospective open-label single center study

Introduction: Nine randomized trials in early breast cancer(EBC) have demonstrated an advantage of aromatase inhibitors(AI’s) over tamoxifen in disease-free survival. Joint symptoms are common toxicities and lead to treatment interruption in up to 20% in case of severe aromatase-induced arthralgia(AIA). During menopause levels of markers of inflammation such as IL-1b, TNF-alpha and IL-6 increase. We hypothesized that estrogen deprivation by aromatase inhibition could be associated with similar changes in these markers and that this could play a role in AIA. In an earlier study, similar toxicities were observed in patients(pts) treated with a recombinant form of IL-6. In several cell types it has been shown that ligand activated estrogen receptor blocks nuclear factor kappa beta controlled gene transcription of IL-6. Aim of the study: We initiated a prospective open-label study to examine the role of inflammatory cytokines and other serum markers(CRP and hormones) in the pathogenesis of AIA. Methods: 29 evaluable postmenopausal pts with hormone sensitive, Her 2 negative EBC stage I-III, with baseline G0-1 arthralgia were included. Before chemotherapy, at baseline, at month 3, 6, 9, 12 and 18 after AI initiation, serum samples were taken for CRP and hormones (estradiol, androstenedion) and cytokines were analyzed with a human cytokine 25-plex panel. A detailed rheumatologic questionnaire and Visual Analog Scale(VAS) was performed at each visit. Arthralgia grading was assessed using the CTCAE criteria 4.0. The T-test and the Wilcoxon Signed Rank test were used to look for the difference in terms of IL-6, Il-1b, IL-8, TNF, CRP, estradiol and androstenedion between G0 versus G1-2-3 arthralgia. Results: The mean age was 56 yrs. (34-72yrs). All patients were treated with surgery followed by concomitant chemoradiation and letrozole (31) or anastrazole (1). In 16 pts G1 arthralgia was present before the start of the chemotherapy. Grade 2 and 3 arthralgia appeared at mth 3. The proportion of pts with grade 2 remained more or less the same, while grade 3 pts declined from month 6 onwards and disappeared at month 18. There was a significant correlation between the grade of arthralgia and the VAS score at all-time points (p Conclusion: This study indicates that both menopause and AIA have inflammatory mediators (IL6 and CRP) that correlate with the degree of lowering of estradiol levels. This is consistent with the regulation of IL-6 by the ligand activated ER through NFkB in vitro. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P1-13-06.

Abstract P1-13-05: A prospective study to evaluate joint changes by ultrasound and magnetic resonance imaging induced by adjuvant chemotherapy followed by aromatase inhibitors in postmenopausal early breast cancer patients developing G3 arthralgia

Introduction: Aromatase inhibitors (AI) are a well-established component of the adjuvant hormonal therapy in postmenopausal (PM) EBC patients (pts). AI have been associated with joint symptoms in 20 to 40% of pts adversely affecting quality of life and compliance. The subjective nature of these toxicities warrants objective correlates. In this study we have compared synovial changes on MRI with ultrasound (US). Aim of the study : The primary aim was to assess joint changes in PM EBC pts by MRI and US at baseline and at occurrence of arthralgia G3 during AI therapy. The secondary aim was to investigate the relationship between clinical severity of articular symptoms and markers of bone metabolism. Methods: PM EBC pts, stages I-III, with a grade 0, 1 arthralgia (CTCAE version 4.0) at baseline were recruited for the study. At baseline and at 3, 6, 9, 12 and 18 months (mths) a rheumatologic questionnaire was performed. At the same time points serum samples for bone markers (Ca, ostase, intact PTH, alkaline phosphatase and vit. D), rheumatoid factor (RF) and urine samples for urinary N-telopeptide (uNTX) were taken. Pain was assessed by the Visual Analog Scale (VAS). Before start of AI, osteoarthritis was quantified by plain radiography (hands, wrists and knees), joint changes were assessed baseline and at occurrence of arthralgia G3 by MRI and US (hands and wrists), and bone density by DEXA. Results: 29 evaluable PM EBC pts were included in the study. At baseline, 60% of the pts had a mean VAS score of 2 (0/10-5/10), which increased to 2.5 after the adjuvant chemotherapy and to 4.5 (0-8) at mth 12 (p = 0.013) of AI therapy, and subsequently dropped to 2.5 at mth 18 (p = 0.2). The most commonly affected joints at baseline were knees (47%), shoulders (41%), and ankles/feet (29%); whereas during AI therapy it involved mainly knees (53%), shoulders (46%) and hands/wrists (32%). Fifty percent of the pts had signs of osteoarthritis of the hands (25%), knees (16%) and both (8%) on plain radiographs obtained at baseline. Grade 0-1 arthralgia was not significantly correlated with the imaging of osteoarthritis or grade of osteoporosis. At baseline intra-articular fluid was detected on MRI of the hands/wrists in 6 pts, 14 pts had synovial changes and 4 had both. Six out of 29 pts had also signs of synovitis with hyperaemia on US before the start of an AI. G3 arthralgia occurred in five pts of which only three agreed to the repeat radiological imaging. In two of these pts a worsening of imaging both by MRI and US was observed, whereas in the third pt imaging was unchanged. Markers of bone metabolism and uNTX did not differ significantly between groups with arthralgia and without (p>0.05). RF was positive in only two pts, one with arthralgia G1 at baseline and one pt at the moment of G3 arthralgia. Conclusions: This study is the first to prospectively report on synovial changes by combined MRI and US before initiation of adjuvant AI in PM women and at occurrence of arthralgia G3. Only 17% of pts developed G3 arthralgia which corresponded to worsening imaging signs. From the small sample size it seems that baseline MRI changes predicts the development of G3 arthralgia with higher sensitivity than baseline US. Sensitivity of US improved during AI therapy. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P1-13-05.