Bert van der Vegt

Pathological and biological differences between screen-detected and interval ductal carcinoma in situ of the breast.

BackgroundThe incidence of ductal carcinoma in situ (DCIS) has risen dramatically with the introduction of screening mammography. The aim was to evaluate differences in pathological and biological characteristics between patients with screen-detected and interval DCIS.MethodsFrom January 1992 to December 2001, 128 consecutive patients had been treated for pure DCIS at our institute. From these, 102 had been attending the Dutch breast cancer screening program. Sufficient paraffin-embedded tissue was available in 74 out of the 102 cases to evaluate biological marker expression (Her2/neu, ER, PR, p53 and cyclin D1) on tissue microarrays (TMA group). Differences in clinicopathological characteristics and marker expression between screen-detected and interval patients were evaluated. Screen-detected DCIS was classified as DCIS detected by screening mammography, when the two-year earlier examination failed to reveal an abnormality. Interval patients were classified as patients with DCIS detected within the two-year interval between two subsequent screening rounds.ResultsScreen-detected DCIS was related with linear branching and coarse granular microcalcifications on mammography (p < .001) and with high-grade DCIS according to the Van Nuys classification (p = .025). In univariate analysis, screen-detected DCIS was related with Her2/neu overexpression (odds ratio [OR] = 6.5; 95%CI 1.3–31.0; p = .020), and interval DCIS was associated with low-grade (Van Nuys, OR = 7.3; 95% CI 1.6–33.3; p = .010) and PR positivity (OR = 0.3; 95%CI 0.1–1.0; p = .042). The multivariate analysis displayed an independent relation of Her2/neu overexpression with screen-detected DCIS (OR = 12.8; 95%CI 1.6–104.0; p = .018).ConclusionsThese findings suggest that screen-detected DCIS is biologically more aggressive than interval DCIS and should not be regarded as overdiagnosis.

Towards personalised treatment in primary Sjögren's syndrome: baseline parotid histopathology predicts responsiveness to rituximab treatment.

Objectives The aims of this study were (1) to assess the effect of rituximab (RTX; anti-CD20) treatment in patients with primary Sjögrens syndrome (pSS) based on sequential parotid biopsies obtained in a placebo-controlled, randomised clinical trial, and (2) to assess the prognostic value of the histological characteristics of parotid gland tissue with regard to responsiveness to RTX treatment. Methods In a double-blinded, placebo-controlled trial, sequential parotid gland biopsies were taken from 20 RTX-treated and 10 placebo-treated patients with pSS, at baseline and 12 weeks after treatment. The relative amount of lymphocytic infiltrate (stained for CD45), absolute number of T cells and B cells per mm2 parenchyma (stained for CD3 and CD20, respectively), focus score, number of germinal centres and of lymphoepithelial lesions per mm2 in parotid gland parenchyma were assessed. Histopathological data were compared between clinical responders (decrease in European League Against Rheumatism Sjögrens Syndrome Disease Activity Index (ESSDAI) score of ≥3 at 12 weeks compared with baseline) and non-responders (change in ESSDAI<3) to RTX treatment. Results In RTX-treated patients, a significant reduction in the number of CD20+ B cells/mm2 parenchyma was observed, while no such reduction was observed in placebo-treated patients. The number of CD3+ T cells/mm2 in parenchyma did not change in either group. Furthermore, the number and the severity of lymphoepithelial lesions/mm2 and number of germinal centres/mm2 was significantly reduced in RTX-treated patients, but did not change in placebo-treated patients. When comparing the pretreatment characteristics of clinical responders with non-responders, the median number of CD20+ B cells/mm2 parenchyma at baseline was significantly higher in responders (1871 vs 353 cells/mm2, p<0.05). Other histopathological baseline characteristics were not predictive for response to RTX treatment. Conclusions RTX treatment in pSS leads to a major reduction of lymphocytic infiltration and to fewer B cells, germinal centres and lymphoepithelial lesions in parotid gland parenchyma. A high pretreatment number of CD20+ B cells/mm2 parotid gland parenchyma predicts better responsiveness of patients with pSS to RTX treatment. Pretreatment parotid gland histopathological characteristics could therefore contribute to a more personalised treatment approach to pSS.

Tumor-specific uptake of fluorescent bevacizumab-IRDye800CW microdosing in patients with primary breast cancer: a phase I feasibility study

Purpose: To provide proof of principle of safety, breast tumor–specific uptake, and positive tumor margin assessment of the systemically administered near-infrared fluorescent tracer bevacizumab–IRDye800CW targeting VEGF-A in patients with breast cancer. Experimental Design: Twenty patients with primary invasive breast cancer eligible for primary surgery received 4.5 mg bevacizumab–IRDye800CW as intravenous bolus injection. Safety aspects were assessed as well as tracer uptake and tumor delineation during surgery and ex vivo in surgical specimens using an optical imaging system. Ex vivo multiplexed histopathology analyses were performed for evaluation of biodistribution of tracer uptake and coregistration of tumor tissue and healthy tissue. Results: None of the patients experienced adverse events. Tracer levels in primary tumor tissue were higher compared with those in the tumor margin (P < 0.05) and healthy tissue (P < 0.0001). VEGF-A tumor levels also correlated with tracer levels (r = 0.63, P < 0.0002). All but one tumor showed specific tracer uptake. Two of 20 surgically excised lumps contained microscopic positive margins detected ex vivo by fluorescent macro- and microscopy and confirmed at the cellular level. Conclusions: Our study shows that systemic administration of the bevacizumab–IRDye800CW tracer is safe for breast cancer guidance and confirms tumor and tumor margin uptake as evaluated by a systematic validation methodology. The findings are a step toward a phase II dose-finding study aimed at in vivo margin assessment and point to a novel drug assessment tool that provides a detailed picture of drug distribution in the tumor tissue. Clin Cancer Res; 23(11); 2730–41. ©2016 AACR.

B Cell Depletion Therapy Normalizes Circulating Follicular Th Cells in Primary Sjögren Syndrome

Objective. To assess the effect of B cell depletion therapy on effector CD4+ T cell homeostasis and its relation to objective measures of disease activity in patients with primary Sjögren syndrome (pSS). Methods. Twenty-four patients with pSS treated with rituximab (RTX) and 24 healthy controls (HC) were included. Frequencies of circulating effector CD4+ T cell subsets were examined by flow cytometry at baseline and 16, 24, 36, and 48 weeks after the first RTX infusion. Th1, Th2, follicular Th (TFH), and Th17 cells were discerned based on surface marker expression patterns. Additionally, intracellular cytokine staining was performed for interferon-γ, interleukin (IL)-4, IL-21, and IL-17 and serum levels of these cytokines were analyzed. Results. In patients with pSS, frequencies of circulating TFH cells and Th17 cells were increased at baseline compared with HC, whereas frequencies of Th1 and Th2 cells were unchanged. B cell depletion therapy resulted in a pronounced decrease in circulating TFH cells, whereas Th17 cells were only slightly lowered. Frequencies of IL-21–producing and IL-17–producing CD4+ T cells and serum levels of IL-21 and IL-17 were also reduced. Importantly, the decrease in circulating TFH cells was associated with lower systemic disease activity over time, as measured by the European League Against Rheumatism Sjögren’s Syndrome Disease Activity Index scores and serum IgG levels. Conclusion. B cell depletion therapy in patients with pSS results in normalization of the elevated levels of circulating TFH cells. This reduction is associated with improved objective clinical disease activity measures. Our observations illustrate the pivotal role of the crosstalk between B cells and TFH cells in the pathogenesis of pSS.

Aggressiveness of 'true' interval invasive ductal carcinomas of the breast in postmenopausal women

There is debate whether interval carcinomas differ from screen-detected tumours biologically. In this study, clinico-pathological parameters and the expression of well-validated biological markers were compared between ‘true’ interval carcinomas and screen-detected/missed carcinomas hypothesising that ‘true’ interval carcinomas show a more aggressive biological behaviour. The study group consisted of 92 consecutive postmenopausal women attending the breast screening programme and presenting with an invasive ductal carcinoma. All screening mammograms were re-reviewed. Sixteen patients had a ‘true’ interval carcinoma. Seven carcinomas were missed at screening, but detected on re-reviewing of the screening mammogram. Radiological characteristics were assessed from diagnostic mammograms. Data on patient- and tumour characteristics and follow-up data were recorded from hospital records. Median follow-up was 61 months. Immunohistochemistry for ER, PR, Her2/neu and p53 was performed on TMA sections. Univariate and multivariate logistic regression analyses were performed. In univariate analysis, ‘true’ interval carcinomas were significantly larger (odd ratios (OR) 7.2, 95% CI 1.8–28.1) and less frequently ER (OR 0.3, 95% CI 0.1–0.9) and PR (OR 0.3, 95% CI 0.1–1.0) positive. In multivariate analysis, ‘true’ interval carcinoma was independently associated with larger tumours (OR 7.0, 95% CI 1.4–36.2). A trend toward ER negativity was found (OR 0.3, 95% CI 0.1–1.1). ‘True’ interval carcinomas showed a trend toward a decreased relapse-free survival (HR 1.7 95% CI 0.9–3.1). Although ‘true’ interval carcinomas were significantly larger than screen-detected/missed interval carcinomas, it remains challenging to observe parameters that determine this difference between ‘true’ interval carcinomas and screen-detected lesions.

Breast Cancer Incidence After Risk-Reducing Salpingo-Oophorectomy in BRCA1 and BRCA2 Mutation Carriers

Premenopausal risk-reducing salpingo-oophorectomy (RRSO) in BRCA1/2 mutation carriers effectively reduces ovarian cancer risk, but also reduces breast cancer risk. Breast cancer risk reductions up to 50% have been reported for both BRCA1 and BRCA2 mutation carriers, but recent prospective studies were not able to reproduce this finding for BRCA1 mutation carriers. Breast cancer incidence after RRSO was assessed in a consecutive series of 104 BRCA1 and 58 BRCA2 mutation carriers. On the basis of data from our own centre, and assuming a 50% risk reduction through RRSO at premenopausal age, we expected to find 8 breast cancers (range 6–10) in this population for the reported screening period (532 women-years). In 162 carriers with a median age of 41 years at RRSO, 13 incident breast cancers were diagnosed. In BRCA1 mutation carriers, 12 incident breast cancers were found compared with 5 (range 3–6) expected and in BRCA2 mutation carriers 1 breast cancer was found compared with 3 (range 2–5) expected. Breast cancer incidence after premenopausal RRSO is still high, especially in BRCA1 mutation carriers. Previously reported breast cancer risk reductions up to 50% were not confirmed. As a consequence, continued intensive screening for breast cancer is warranted in BRCA1 and BRCA2 mutation carriers after RRSO. Cancer Prev Res; 5(11); 1291–7. ©2012 AACR.

Threshold Analysis and Biodistribution of Fluorescently Labeled Bevacizumab in Human Breast Cancer.

In vivo tumor labeling with fluorescent agents may assist endoscopic and surgical guidance for cancer therapy as well as create opportunities to directly observe cancer biology in patients. However, malignant and nonmalignant tissues are usually distinguished on fluorescence images by applying empirically determined fluorescence intensity thresholds. Here, we report the development of fSTREAM, a set of analytic methods designed to streamline the analysis of surgically excised breast tissues by collecting and statistically processing hybrid multiscale fluorescence, color, and histology readouts toward precision fluorescence imaging. fSTREAM addresses core questions of how to relate fluorescence intensity to tumor tissue and how to quantitatively assign a normalized threshold that sufficiently differentiates tumor tissue from healthy tissue. Using fSTREAM we assessed human breast tumors stained in vivo with fluorescent bevacizumab at microdose levels. Showing that detection of such levels is achievable, we validated fSTREAM for high-resolution mapping of the spatial pattern of labeled antibody and its relation to the underlying cancer pathophysiology and tumor border on a per patient basis. We demonstrated a 98% sensitivity and 79% specificity when using labeled bevacizumab to outline the tumor mass. Overall, our results illustrate a quantitative approach to relate fluorescence signals to malignant tissues and improve the theranostic application of fluorescence molecular imaging. Cancer Res; 77(3); 623-31. ©2016 AACR.

Germinal centres in diagnostic labial gland biopsies of patients with primary Sjogren's syndrome are not predictive for parotid MALT lymphoma development

Objective Patients with primary Sjögren’s syndrome (pSS) have an increased risk of developing non-Hodgkin’s lymphoma (NHL), particularly parotid gland mucosa-associated lymphoid tissue (MALT) lymphomas. Presence of germinal centres (GCs) in labial gland biopsies has been suggested as predictive factor for NHL. We assessed whether presence of GCs is increased in labial gland biopsies from patients with pSS who developed parotid MALT lymphoma, the dominant NHL-subtype in pSS, compared with patients with pSS who did not develop lymphoma. Methods Eleven labial gland biopsies from patients with pSS that were taken prior to parotid MALT lymphoma development were compared with biopsies of 22 matched pSS controls (1:2) who did not develop lymphoma. Biopsies were evaluated for GCs (H&E and Bcl6). Results Labial gland biopsies of pSS MALT lymphoma patients, revealed GCs in 2/11 (18%) H&E sections and 3/11 (27%) Bcl6 stained sections. In controls, GCs were present in 4/22 (18%) of H&E sections and 5/22 (23%) of Bcl6 stained sections. Conclusion Presence of GCs in labial gland biopsies does not differ between patients with pSS that develop parotid MALT lymphoma and patients with pSS who do not develop lymphoma. The presence of GCs in labial gland biopsies is therefore not a predictive factor for pSS-associated parotid MALT lymphomas.

In primary Sjögren's syndrome high absolute numbers and proportions of B cells in parotid glands predict responsiveness to rituximab as defined by ESSDAI, but not by SSRI

With great interest we have read the letter to the editor by Cornec et al 1 regarding our paper ‘Towards personalised treatment in primary Sjogrens syndrome (pSS): baseline parotid histopathology predicts responsiveness to rituximab treatment’.2 In essence, we showed in our paper that absolute numbers of CD20+ cells/mm2 of parenchyma of parotid gland tissue are predictive for the responsiveness of patients with pSS to rituximab (RTX) treatment. Cornec et al argue that there is a discrepancy in outcomes presented in their study and our study,1 as they observed that a high proportion of minor salivary gland B cells predict the absence of a clinical response to RTX.3 As we will show and explain here, there is no inconsistency between the two studies and most of the apparent discrepancy is likely the result of differences in how the tissues are analysed and how the disease activity is established. A major difference in the two studies is how B cells are assessed in tissue sections of salivary gland biopsies of patients with pSS before (and after) RTX treatment. We measured absolute numbers of CD20+ B cells/mm2 of parenchyma, while Cornec et al assessed the proportion of B cells.1 ,3 Obviously, even when there is a change in absolute numbers of B cells in the tissue, the B/T cell ratio still can remain the same. Thus, although higher numbers of B cells, do not need to be reflected per se in higher proportions of B cells, we also found in our study that patients with higher absolute numbers of B cells in the glandular tissue, had a higher B/B+T cell ratio. Furthermore, responders to RTX, as defined by a decrease in European League Against …

Need for consensus guidelines to standardise the assessment of germinal centres and other histopathological parameters in salivary gland tissue of patients with primary Sjögren's syndrome

We have read with great interest the letter to the editor by van Roon et al 1 commenting on our paper ‘Towards personalised treatment in primary Sjogrens syndrome: baseline parotid histopathology predicts responsiveness to rituximab treatment’.2 The authors argue that there is a need for standardisation of the histopathological characteristics of salivary gland tissue of patients with primary Sjogrens syndrome (pSS), in general, and of the presence of germinal centres (GCs), in particular. We fully agree with van Roon et al 1 and other authors about the need for consensus guidelines to standardise the histopathological evaluation of salivary gland biopsies in patients with pSS.3 A standardised scoring system may facilitate prognostication and stratification of patients with pSS and is needed for a valid evaluation of various clinical trials.3 In particular, histological definition of GCs in salivary gland tissue is warranted, since these structures can be difficult to detect in diagnostic H&E-stained tissue sections. Detection of GCs in the periductal lymphoid infiltrates of the salivary glands is clinically relevant, because the presence of these structures is associated with more severe disease.4 Importantly, the presence of GCs in minor salivary gland biopsies has been postulated to be a predictor of patients who are at risk of lymphoma development.5 …