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In solid tumors, angiogenesis occurs in the setting of a defective vasculature and impaired lymphatic drainage that is associated with increased vascular permeability and enhanced tumor permeability. These universal aspects of the tumor microenvironment can have a marked influence on intratumoral drug delivery that may often be underappreciated. In this study, we investigated the effect of blood vessel normalization in tumors by the antiangiogenic drug bevacizumab on antibody uptake by tumors. In mouse xenograft models of human ovarian and esophageal cancer (SKOV-3 and OE19), we evaluated antibody uptake in tumors by positron emission tomographic imaging 24 and 144 hours after injection of 89Zr-trastuzumab (SKOV-3 and OE19), 89Zr-bevacizumab (SKOV-3), or 89Zr-IgG (SKOV-3) before or after treatment with bevacizumab. Intratumor distribution was assessed by fluorescence microscopy along with mean vessel density (MVD) and vessel normalization. Notably, bevacizumab treatment decreased tumor uptake and intratumoral accumulation compared with baseline in the tumor models relative to controls. Bevacizumab treatment also reduced MVD in tumors and increased vessel pericyte coverage. These findings are clinically important, suggesting caution in designing combinatorial trials with therapeutic antibodies due to a possible reduction in tumoral accumulation that may be caused by bevacizumab cotreatment. Cancer Res; 73(11); 3347–55. ©2013 AACR.

PDF file - 53K, Biodistribution of 89Zr-Trastuzumab in the OE19 model and correlation of the in vivo PET tumor uptake with the ex vivo biodistriution tumor uptake of 89Zr-trastuzumab in the OE19 model.

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Background Adjuvant endocrine therapy (ET) in patients with breast cancer (BC) increases the risk of becoming less physically active. Physical inactivity is associated with a higher risk of treatment-related side effects and mortality. This study investigated whether supervised exercise increased the proportion of patients adhering to the national physical activity (PA) guideline during adjuvant ET in overweight or obese BC patients. Methods This multicentre single-arm clinical trial included patients with BC participating in a 12-week supervised exercise intervention. An accelerometer measured moderate to vigorous PA (MVPA) at baseline (T0), after 12 (T1) and 26 weeks (T2). The primary endpoint was change in the proportion of patients with weekly ≥150 min of MVPA at T1 compared to T0. Secondary endpoints were adherence to PA guideline at T2, metabolic syndrome (MetS), body composition, health-related quality of life (HRQoL) and BC-specific functioning and symptoms, self-reported PA, self-efficacy, exercise motivation and satisfaction with life. Results 141 patients with a median age of 61 years and a mean BMI of 31.3 participated. Adherence to the PA guideline increased from 38.3% at T0, to 40.4% at T1 (p = .112) and 44.7% at T2 (p = .003). MetS, body composition, HRQoL, BC-specific functioning and symptoms (i.e. fatigue, dyspnoea), self-reported PA, self-efficacy, exercise motivation and satisfaction with life improved significantly over time. Conclusions Supervised exercise increased the proportion of BC patients adhering to the PA guideline over time. Furthermore, MetS, body composition, HRQoL and symptoms improved. Our findings highlight the clinical relevance of supervised exercise during ET in overweight BC patients. Clinical trial information (NCT02424292)., Highlights • Overweight breast cancer patients on endocrine therapy can be motivated for supervised exercise. • Supervised exercise increases adherence to the physical activity guideline. • Metabolic syndrome and quality of life improved due to our intervention. • Participation in supervised exercise induces persistent clinical improvements.

Background: Breast cancer (BC) is a heterogeneous disease, in which estrogen receptor (ER) expression plays an important role in the majority of breast tumors. A clinical dilemma may arise when a metastasis biopsy to determine the ER status cannot be performed safely or when ER heterogeneity is suspected between tumor lesions. Whole-body ER imaging, such as 16α-18F-fluoro-17β-estradiol (18F-FES) positron emission tomography (PET), may have added value in these situations. However, the role of this imaging technique in routine clinical practice remains to be further determined. Therefore, we assessed the value of 18F-FES-PET by evaluating if the physician's clinical dilemma that remained after standard workup was solved by the 18F-FES-PET scan. Methods: In this retrospective study, 18F-FES-PET scans, performed in patients with (suspected) ER+ metastatic BC with remaining clinical dilemma after standard workup, at the University Medical Center of Groningen between November 2009 and January 2019, were included. We investigated whether the physician's clinical dilemma was solved, defined as 1) 18F-FES-PET provided a solution for the clinical dilemma, and/or 2) a treatment decision was based directly on the 18F-FES-PET. In addition, category of clinical dilemma, and rate of 18F-FES positive or negative PET scans were reported, and related to frequency of solved dilemmas. Results: One hundred 18F-FES-PET scans were performed in 83 patients. Clinical dilemma categories were: 1) inability to determine extent of (suspected) metastatic disease with standard workup (n = 52), 2) unclear ER status of the tumor (n = 31), and 3) inability to determine which primary tumor caused metastases (n = 17). Dilemmas were solved by 18F-FES-PET in 87/100 cases (87%). In 81/87 cases a treatment decision was made based directly on the 18F-FES-PET (treatment change: n = 51 cases; continuance: n = 30 cases). The frequency of solved dilemmas was not related to the clinical dilemma category (P = 0.334). However, the frequency of solved dilemmas was related to whether scans were 18F-FES positive (n = 63) or negative (n = 37; p

Background:Eftilagimod alpha (efti, IMP321),a soluble LAG-3 protein (LAG-3Ig) that binds to a subset of MHC class II molecules, mediates antigen-presenting cell (APC) activation followed by CD8 T-cell activation. AIPAC (Active Immunotherapy PAClitaxel; NCT02614833) investigatedcombinations of the APC activator efti + weekly paclitaxel compared to paclitaxel + placebo in metastatic breast carcinoma (MBC) patients (pts). Methods:The placebo-controlled, double-blinded, 1:1 randomized, multinational, phase IIb trial enrolled pts with measurable disease, hormone receptor-positive (HR+) MBC with indications for first line weekly paclitaxel without indication for HER2/neu targeted therapy. Pts received paclitaxel (80 mg/m² IV at D1, D8, D15 plus efti (30 mg) or placebo at D2, D16 (injected SC every 2 weeks) for 6 cycles (1 cycle = 4 weeks). Maintenance phase followed in which pts benefitting from treatment received efti or placebo for an additional 52 weeks. Primary endpoint was progression-free survival (PFS) (RECIST1.1) determined by blinded independent central read (BICR). Secondary endpoints included local read PFS, RECIST 1.1 tumor response, pharmacodynamic effects, quality of life and overall survival. The study was powered (80 %) to detect 0.667 hazard ratio (HR) based on 5% one-sided alpha and planned to enroll 226 pts. Results: From Jan 2017-Jul 2019, 227 pts were randomized. All except 1 received ≥ 1 treatment and were included in the full analysis set [(efti (n = 114); placebo (n = 112)]. Data cut-off was 10th Jan 2020 with 12 month median follow-up. HR for PFS assessed by BICR was 0.93 [95 % CI 0.67-1.30], p =0.341. In the efti group 63 % (95% CI 52- 71%) were progression free at 6 months compared to 54 % (95% CI 43-63%) in the placebo group. ORR was 48.3 % in the efti group compared to 38.4 % in the placebo group as assessed by BICR (p=0.118). Efti increased numbers of PBMCs and T cells (CD4 and CD8) significantly compared to placebo. In predefined subgroups such as pts with low monocytes at baseline, luminal B subtype or age Conclusion: Efti did not prolong overall median PFS in women with HR+ MBC receiving first line chemotherapy. Tested in a randomized setting against placebo in pts receiving paclitaxel, efti is well tolerated and did not add clinically significant toxicity while inducing a sustained, significant increase in CD8 T cells in blood and clinical benefits in some predefined subgroups. Relevant subgroups and overall survival will be investigated. BICRInvestigatorEfti+paclitaxel(N= 114)Placebo+paclitaxel(N= 112)Efti+paclitaxel(N= 114)Placebo+paclitaxel(N= 112)Median PFS, months (95% CI)7.29 (6.64-7.46)7.29 (5.52-7.46)7.16 (5.65-7.39)6.70 (5.52-7.33)HR (95% CI)0.93 (0.67-1.30)0.92 (0.69-1.23)P value0.3410.305Mean PFS [SE], months7.12 [0.37]6.64 [0.38]6.81 [0.33]6.30 [0.31] Citation Format: Hans Wildiers, Anne Armstrong, Eveline Cuypere, Florence Dalenc, Luc Dirix, Steve Chan, Frederik Marme, Carolina Pia Schröder, Jens Huober, Jill Wagemans, Peter Vuylsteke, Jean-Philippe Jacquin, Etienne Brain, Sherko Kümmel, Zsuzsanna Pápai, Luca Perjesi, Christian Mueller, Chrystelle Brignone, Frederic Triebel. Primary efficacy results from AIPAC: A double-blinded, placebo controlled, randomized multinational phase IIb trial comparing weekly paclitaxel plus eftilagimod alpha (soluble LAG-3 protein) vs. weekly paclitaxel plus placebo in HR-positive metastatic breast cancer patients [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PD14-08.

Rationale: Whether assessment of potential bone lesions in metastatic breast cancer (MBC) by means of 18F-fluorodeoxyglucose (FDG)-PET instead of 99mTechnetium (99mTc) bone scintigraphy (BS) supports clinically relevant changes in MBC management, is currently unknown. Therefore, we retrospectively compared the management recommendations based on bone lesion assessment by FDG-PET plus contrast enhanced computed tomography (ceCT) or BS plus ceCT, for patients with newly diagnosed MBC. Methods: Baseline ceCT, BS and FDG-PET of all patients included in the IMPACT-MBC study (NCT01957332) at location University Medical Center Groningen (UMCG), were reviewed for bone lesions. In case of bone lesions on any imaging modality, virtual MBC management recommendations per patient were made by a multidisciplinary expert panel, based on either FDG-PET plus ceCT, or BS plus ceCT. The panel had access to standard clinicopathological information and baseline imaging findings outside the skeleton. Clinically relevant management differences between the two recommendations were defined as 1) different treatment intent, (curative, non-curative or unable to determine and/or 2) different systemic or local treatment. In case of absence of bone lesions with any imaging modality, patients were included in the analyses without expert review. Results: A total of 3,473 unequivocal bone lesions was identified in 102 evaluated patients (39% by ceCT, 26% by BS, 87% by FDG-PET). Additional bone lesions on FDG-PET plus ceCT compared to BS plus ceCT led to change of MBC management recommendations in 16% of all patients (95% confidence interval (CI) 10-24%). BS also changed management compared to FDG-PET in one patient (1%; 95%CI 0-5%). In 26% (95%CI 19-36%) of patients, an additional FDG-PET was requested, because BS provided insufficient information. Conclusion: In this exploratory analysis of newly diagnosed MBC patients, FDG-PET versus BS to assess bone lesions, resulted in clinically relevant management differences in 16% of all patients. BS delivered insufficient information in over one fourth of patients, resulting in additional request for FDG-PET. Based on this data, FDG-PET should be considered as primary imaging modality for assessment of bone lesions in newly diagnosed MBC.

Background: The androgen receptor (AR) is a potential target in metastatic breast cancer (MBC), and 16 beta-[F-18]-fluoro-5 alpha-dihydrotestosterone positron emission tomography ([F-18]-FDHT-PET) can be used for noninvasive visualisation of AR. [F-18]-FDHT uptake reduction during AR-targeting therapy reflects AR occupancy and might be predictive for treatment response. We assessed the feasibility of [F-18]-FDHT-PET to detect changes in AR availability during bicalutamide treatment and correlated these changes with treatment response.Patients and methods: Patients with AR thorn MBC, regardless of oestrogen receptor status, received an [F-18]-FDHT-PET at baseline and after 4-6 weeks bicalutamide treatment. Baseline [F-18]-FDHT uptake was expressed as maximum standardised uptake value. Percentage change in tracer uptake, corrected for background activity (SUVcor), between baseline and follow-up PET scan (% reduction), was assessed per-patient and lesion. Clinical benefit was determined in accordance with Response Evaluation Criteria in Solid Tumours (RECIST) 1.1 or clinical evaluation (absence of disease progression for >= 24 weeks).Results: Baseline [F-18]-FDHT-PET in 21 patients detected 341 of 515 lesions found with standard imaging and 21 new lesions. Follow-up [F-18]-FDHT- PET was evaluable in 17 patients with 349 lesions, showing a decrease in median SUVcor from 1.3 to 0.7 per-patient and lesion (P Conclusion: In this feasibility study, a bicalutamide-induced reduction in [F-18]-FDHT uptake could be detected by follow-up [F-18]-FDHT-PET in patients with AR thorn MBC. However, this change could not predict bicalutamide response. (C) 2020 The Authors. Published by Elsevier Ltd.

Determining the estrogen receptor (ER) status is essential in metastatic breast cancer (MBC) management. Whole-body ER imaging with 16α-[In a prospective multicenter trial, 200 patients with newly diagnosed MBC underwent extensive workup including molecular imaging. For this subanalysis, ER expression in the biopsied metastasis was related to qualitative whole-body [Whole-body [In this largest prospective series so far, we established the clinical validity of [

BACKGROUND: Adding cyclin-dependent kinase (CDK) inhibitor to endocrine treatment improves outcome in œstrogen receptor (ER) positive metastatic breast cancer, but identifying the subset of patients who benefit is challenging. Response is potentially associated with ER expression heterogeneity. This is because, unlike the primary tumour in the breast that is localized to the organ, the metastatic breast cancer has spread and continues to spread to distant locations in the body such as bones, lungs, liver, axial skeleton, even to the central nervous system like the brain, wherefrom obtaining biopsies are not easy, and also, the metastasised tissues are heterogeneous. Positron emission tomography (PET) with 16α-[18F]fluoro-17β-œstradiol (FES), briefly referred to as FES-PET, allows whole-body ER assessment. We explored whether FES-PET heterogeneity and FES uptake were related to letrozole and palbociclib outcome, in patients with ER positive, metastatic breast cancer.PATIENTS AND METHODS: Patients underwent a baseline FES-PET and 18F-fluorodeoxyglucose (FDG) PET, the FDG-PET served to help identify active sites of breast cancer with contrast-enhanced computed tomography (CT). FES-PET heterogeneity score (% FES positive lesions divided by all lesions on FDG-PET and/or CT) and FES uptake were related to outcome and 8-week FDG-PET response. Circulating tumour DNA (CtDNA) samples for ESR1 mutation analysis were collected at baseline.RESULTS: In 30 patients with 864 metastatic lesions, baseline FES-PET heterogeneity was assessed. In 27 patients with 688 lesions, response was evaluated. Median time to progression (TTP) was 73 weeks (95% confidence interval [CI] 21 to ∞) in 7 patients with 100% FES positive disease, 27 weeks (14-49) in heterogeneous FES positive disease (20 patients), and 15 weeks (9 to ∞) without FES positivity (three patients; log-rank P = 0.30). Geometric mean FES uptake was 2.3 for metabolic progressive patients, 2.5 (Pvs progression = 0.82) for metabolic stable disease, and 3.3 (Pvs progression = 0.40) for metabolic response (Ptrend = 0.21). ESR1 mutations, found in 13/23 patients, were unrelated to FES uptake.CONCLUSION: This exploratory study suggests that FES-PET heterogeneity may potentially identify the subset of ER positive, metastatic breast cancer patients who benefit from letrozole combined with CDK inhibition.CLINICAL TRIAL INFORMATION: NCT02806050.

We offer an illustrative case about estrogen receptor (ER) imaging (also known as 16 alpha-[F-18]fluoro-17 beta-estradiol ([F-18]-FES) PET) and the detection of metastatic lesions in the dural region. We present a case of a woman with ER-positive metastatic breast cancer and high [F-18]-FES uptake in the dural region on PET imaging, without associated clinical symptoms. These lesions were missed on [F-18]-FDG PET because of physiological [F-18]-FDG uptake in the brain. This case highlighted some difficulties in the interpretation of imaging of brain metastases and demonstrated the added value of [F-18]-FES PET imaging. [F-18]-FES PET could be used to prove the presence of ER-positive metastases in the brain.

Background: Positron emission tomography (PET) with 16α-[18F]-fluoro-17β-estradiol ([18F]-FES) can visualize estrogen receptor (ER) expression, but it is challenging to determine the ER status of liver metastases, due to high physiological [18F]-FES uptake. We evaluated whether [18F]-FES-PET can be used to determine the ER status of liver metastases, using corresponding liver biopsies as the gold standard. Methods: Patients with metastatic breast cancer (n = 23) were included if they had undergone a [18F]-FES-PET, liver metastasis biopsy, CT-scan, and [18F]-FDG-PET. [18F]-FES-PET scans were assessed by visual and quantitative analysis, tracer uptake was correlated with ER expression measured by immunohistochemical staining and the effects of region-of-interest size and background correction were determined. Results: Visual analysis allowed ER assessment of liver metastases with 100% specificity and 18% sensitivity. Quantitative analysis improved the sensitivity. Reduction of the region-of-interest size did not further improve the results, but background correction improved ER assessment, resulting in 83% specificity and 77% sensitivity. Using separate thresholds for ER+ and ER− metastases, positive and negative predictive values of 100% and 75%, respectively, could be obtained, although 30% of metastases remained inconclusive. Conclusion: In the majority of liver metastases, ER status can be determined with [18F]-FES-PET if background correction and separate thresholds are applied.

Introduction: Breast cancer (BC) is a heterogeneous disease, in which estrogen receptor (ER) expression plays an important role in the majority of breast tumors. A clinical dilemma may arise when a metastasis biopsy to determine the ER status cannot be performed safely or when ER heterogeneity is suspected between tumor lesions. Whole-body ER imaging, such as 16α-[18F]-fluoro-17β-estradiol ([18F]-FES) positron emission tomography (PET), may have additional value in these situations. However, the role of this imaging technique in routine clinical practice remains to be further determined. Therefore, we assessed the value of [18F]-FES-PET in a large retrospective set of patients, by evaluating if the physicians’ clinical dilemma that remained after standard workup was solved by the [18F]-FES-PET scan. Methods: In this single center study, [18F]-FES-PET scans, performed in patients with (suspected) ER+ metastatic BC with remaining clinical dilemma after standard workup (such as computed tomography, [18F]-fluorodeoxyglucose ([18F]-FDG)-PET, bone scintigraphy, magnetic resonance imaging, or biopsy), performed at the UMCG between November 2009 and January 2019, were included. A whole-body [18F]-FES-PET scan was performed 60 min after ~200 MBq of [18F]-FES was injected intravenously. ER antagonists had to be discontinued at least 5 weeks before [18F]-FES-PET. Primary endpoint was the percentage of cases in which the referring physician’s clinical dilemma was solved based on the [18F]-FES-PET results. The dilemma was considered solved if 1) the [18F]-FES-PET provided a solution for the clinical dilemma (for example an extra metastatic site to biopsy), or 2) a treatment decision (to change or continue) was made based on the [18F]-FES-PET result. If the physician had doubts about the diagnosis after the [18F]-FES-PET examination, and additional workup was necessary for treatment decision-making, the dilemma was considered not solved. Secondary endpoints were type of clinical dilemma, and rate of [18F]-FES positive or negative PET scans (visual interpretation), related to frequency of solved dilemmas. Results: One hundred [18F]-FES-PET scans were performed in 83 patients. Clinical dilemma types were: 1) inability to determine extent of (suspected) metastatic disease with standard workup (n=52), 2) unclear disease ER status (n=31), and 3) inability to determine which primary tumor caused metastases (n=17). Dilemmas were solved by [18F]-FES-PET in 87/100 cases (87%). In these 87 cases, treatment was changed in 52 cases, and continued in 35 cases. The frequency of solved dilemmas was not related to the type of clinical dilemma. In contrast, the frequency of solved dilemmas was related to whether scans were [18F]-FES positive or negative. Out of the 63 [18F]-FES positive scans, the clinical dilemma was solved in 61 cases (97%); in 26 out of the 37 [18F]-FES negative scans (70%) the dilemma was solved (p Citation Format: Jorianne Boers, Naila Loudini, Celina L. Brunsch, Sylvia A. Koza, Erik F.J. de Vries, Andor W.J.M. Glaudemans, Geke A.P. Hospers, Carolina P. Schröder. Value of [18F]-FES-PET to solve clinical dilemmas in breast cancer patients: A retrospective study [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS3-05.

The androgen receptor (AR) is a drug target in breast cancer, and AR-targeted therapies have induced tumor responses in breast cancer patients. In this review, we summarized the role of AR in breast cancer based on preclinical and clinical data. Response to AR-targeted therapies in unselected breast cancer populations is relatively low. Pre-clinical and clinical data show that AR antagonists might have a role in estrogen receptor (ER)-negative/AR-positive tumors. The prognostic value of AR for patients remains uncertain due to the use of various antibodies and cut-off values for immunohistochemical assessment. To get more insight into the role of AR in breast cancer, we additionally performed a retrospective pooled analysis to determine the prognostic value of the AR using mRNA profiles of 7270 primary breast tumors. Our analysis shows that a higher AR mRNA level is associated with improved disease outcome in patients with ER-positive/human epidermal growth factor receptor 2 (HER2)-negative tumors, but with worse disease outcome in HER2-positive subgroups. In conclusion, next to AR expression, incorporation of additional tumor characteristics will potentially make AR targeting a more valuable therapeutic strategy in breast cancer. (C) 2019 The Authors. Published by Elsevier Inc.

Programmed cell death protein-1/ligand-1 (PD-1/PD-L1) blockade is effective in a subset of patients with several tumor types, but predicting patient benefit using approved diagnostics is inexact, as some patients with PD-L1-negative tumors also show clinical benefit1,2. Moreover, all biopsy-based tests are subject to the errors and limitations of invasive tissue collection3-11. Preclinical studies of positron-emission tomography (PET) imaging with antibodies to PD-L1 suggested that this imaging method might be an approach to selecting patients12,13. Such a technique, however, requires substantial clinical development and validation. Here we present the initial results from a first-in-human study to assess the feasibility of imaging with zirconium-89-labeled atezolizumab (anti-PD-L1), including biodistribution, and secondly test its potential to predict response to PD-L1 blockade (ClinicalTrials.gov identifiers NCT02453984 and NCT02478099). We imaged 22 patients across three tumor types before the start of atezolizumab therapy. The PET signal, a function of tracer exposure and target expression, was high in lymphoid tissues and at sites of inflammation. In tumors, uptake was generally high but heterogeneous, varying within and among lesions, patients, and tumor types. Intriguingly, clinical responses in our patients were better correlated with pretreatment PET signal than with immunohistochemistry- or RNA-sequencing-based predictive biomarkers, encouraging further development of molecular PET imaging for assessment of PD-L1 status and clinical response prediction.

Background Data on the long-term effects of prenatal exposure to maternal cancer and its treatment on child development are scarce. Methods In a multicenter cohort study, the neurologic and cardiac outcomes of 6-year-old children born to women diagnosed with cancer during pregnancy were compared with the outcome of children born after an uncomplicated pregnancy. Assessment included clinical evaluation, comprehensive neuropsychological testing, electrocardiography and echocardiography. Results In total, 132 study children and 132 controls were included. In the study group, 97 children (73.5%) were prenatally exposed to chemotherapy (alone or in combination with other treatments), 14 (10.6%) to radiotherapy (alone or in combination), 1 (0.8%) to trastuzumab, 12 (9.1%) to surgery alone and 16 (12.1%) to no treatment. Although within normal ranges, statistically significant differences were found in mean verbal IQ and visuospatial long-term memory, with lower scores in the study versus control group (98.1, 95% confidence interval [CI]: 94.5–101.8, versus 104.4, 95% CI: 100.4–108.4, P = 0.001, Q, Highlights • Cancer treatment including chemotherapy is possible during pregnancy. • Children are at risk for lower verbal IQ and visuospatial long-term memory scores. • Other cognitive functions and cardiac outcomes were normal at the age of 6 years. • We documented ototoxicity in three children exposed to cisplatin. • Follow-up until adulthood is recommended.

Breast cancer (BC) is a heterogeneous disease in which estrogen receptor (ER) expression plays an important role in most tumors. A clinical dilemma may arise when a metastasis biopsy to determine the ER status cannot be performed safely or when ER heterogeneity is suspected between tumor lesions. Whole-body ER imaging, such as 16α

The androgen receptor (AR) is a potential target in metastatic breast cancer (MBC), and 16β-[Patients with AR + MBC, regardless of oestrogen receptor status, received an [Baseline [In this feasibility study, a bicalutamide-induced reduction in [NCT02697032.

Purpose of Review Molecular imaging with positron emission tomography (PET) is a powerful tool to visualize breast cancer characteristics. Nonetheless, implementation of PET imaging into cancer care is challenging, and essential steps have been outlined in the international “imaging biomarker roadmap.” In this review, we identify hurdles and provide recommendations for implementation of PET biomarkers in breast cancer care, focusing on the PET tracers 2-[18F]-fluoro-2-deoxyglucose ([18F]-FDG), sodium [18F]-fluoride ([18F]-NaF), 16α-[18F]-fluoroestradiol ([18F]-FES), and [89Zr]-trastuzumab. Recent Findings Technical validity of [18F]-FDG, [18F]-NaF, and [18F]-FES is established and supported by international guidelines. However, support for clinical validity and utility is still pending for these PET tracers in breast cancer, due to variable endpoints and procedures in clinical studies. Summary Assessment of clinical validity and utility is essential towards implementation; however, these steps are still lacking for PET biomarkers in breast cancer. This could be solved by adding PET biomarkers to randomized trials, development of imaging data warehouses, and harmonization of endpoints and procedures.

Purpose Correct identification of tumour receptor status is important for treatment decisions in breast cancer. [18F]FES PET and [18F]FDHT PET allow non-invasive assessment of the oestrogen (ER) and androgen receptor (AR) status of individual lesions within a patient. Despite standardised analysis techniques, interobserver variability can significantly affect the interpretation of PET results and thus clinical applicability. The purpose of this study was to determine visual and quantitative interobserver variability of [18F]FES PET and [18F]FDHT PET interpretation in patients with metastatic breast cancer. Methods In this prospective, two-centre study, patients with ER-positive metastatic breast cancer underwent both [18F]FES and [18F]FDHT PET/CT. In total, 120 lesions were identified in 10 patients with either conventional imaging (bone scan or lesions > 1 cm on high-resolution CT, n = 69) or only with [18F]FES and [18F]FDHT PET (n = 51). All lesions were scored visually and quantitatively by two independent observers. A visually PET-positive lesion was defined as uptake above background. For quantification, we used standardised uptake values (SUV): SUVmax, SUVpeak and SUVmean. Results Visual analysis showed an absolute positive and negative interobserver agreement for [18F]FES PET of 84% and 83%, respectively (kappa = 0.67, 95% CI 0.48–0.87), and 49% and 74% for [18F]FDHT PET, respectively (kappa = 0.23, 95% CI − 0.04–0.49). Intraclass correlation coefficients (ICC) for quantification of SUVmax, SUVpeak and SUVmean were 0.98 (95% CI 0.96–0.98), 0.97 (95% CI 0.96–0.98) and 0.89 (95% CI 0.83–0.92) for [18F]FES, and 0.78 (95% CI 0.66–0.85), 0.76 (95% CI 0.63–0.84) and 0.75 (95% CI 0.62–0.84) for [18F]FDHT, respectively. Conclusion Visual and quantitative evaluation of [18F]FES PET showed high interobserver agreement. These results support the use of [18F]FES PET in clinical practice. In contrast, visual agreement for [18F]FDHT PET was relatively low due to low tumour-background ratios, but quantitative agreement was good. This underscores the relevance of quantitative analysis of [18F]FDHT PET in breast cancer. Trial registration ClinicalTrials.gov, NCT01988324. Registered 20 November 2013, https://clinicaltrials.gov/ct2/show/NCT01988324?term=FDHT+PET&draw=1&rank=2.

It is unknown whether assessment of potential bone lesions in metastatic breast cancer (MBC) by

BackgroundBispecific antibodies redirecting T cells to the tumor obtain increasing interest as potential cancer immunotherapy. ERY974, a full-length bispecific antibody targeting CD3ε on T cells and glypican 3 (GPC3) on tumors, has been in clinical development However, information on the influence of T cells on biodistribution of bispecific antibodies, like ERY974, is scarce. Here, we report the biodistribution and tumor targeting of zirconium-89 (89Zr) labeled ERY974 in mouse models using immuno-positron emission tomography (PET) imaging.MethodsTo study both the role of GPC3 and CD3 on the biodistribution of [89Zr]Zr-N-suc-Df-ERY974,89Zr-labeled control antibodies targeting CD3 and non-mammalian protein keyhole limpet hemocyanin (KLH) or KLH only were used. GPC3 dependent tumor targeting of [89Zr]Zr-N-suc-Df-ERY974 was tested in xenograft models with different levels of GPC3 expression. In addition, CD3 influence on biodistribution of [89Zr]Zr-N-suc-Df-ERY974 was evaluated by comparing biodistribution between tumor-bearing immunodeficient mice and mice reconstituted with human immune cells using microPET imaging and ex vivo biodistribution. Ex vivo autoradiography was used to study deep tissue distribution.ResultsIn tumor-bearing immunodeficient mice, [89Zr]Zr-N-suc-Df-ERY974 tumor uptake was GPC3 dependent and specific over [89Zr]Zr-N-suc-Df-KLH/CD3 and [89Zr]Zr-N-suc-Df-KLH/KLH. In mice engrafted with human immune cells, [89Zr]Zr-N-suc-Df-ERY974 specific tumor uptake was higher than in immunodeficient mice. Ex vivo autoradiography demonstrated a preferential distribution of [89Zr]Zr-N-suc-Df-ERY974 to T cell rich tumor tissue. Next to tumor, highest specific [89Zr]Zr-N-suc-Df-ERY974 uptake was observed in spleen and lymph nodes.Conclusion[89Zr]Zr-N-suc-Df-ERY974 can potentially be used to study ERY974 biodistribution in patients to support drug development.

Developing new oncology drugs has increased since the improved understanding of cancer's complex biology. The oncology field has become the top therapeutic research area for new drugs. However, only a limited number of drugs entering clinical trials will be approved for use as standard of care for cancer patients. Molecular imaging is increasingly perceived as a tool to support go/no-go decisions early during drug development. It encompasses a wide range of techniques including radiolabeling a compound of interest followed by visualization with single photon emission computed tomography or positron emission tomography. Radiolabeling can be performed using a variety of radionuclides that are preferably matched to the compound based on size and half-life. Imaging can provide information on drug behavior in vivo, whole body drug target visualization, and heterogeneity in drug target expression. This review focuses on current applications of molecular imaging in the development of small molecules, antibodies, and anti-hormonal anticancer drugs.

Rationale: Knowledge on monoclonal antibody biodistribution in healthy tissues in humans can support clinical drug development. Molecular imaging with positron emission tomography (PET) can yield information in this setting. However, recent imaging studies have analyzed the behavior of single antibodies only, neglecting comparison across different antibodies. Methods: We compared the distribution of four 89Zr-labeled antibodies in healthy tissue in a retrospective analysis based on the recently published harmonization protocol for 89Zr-tracers and our delineation protocol. Results: The biodistribution patterns of 89Zr-lumretuzumab, 89Zr-MMOT0530A, 89Zr-bevacizumab and 89Zr-trastuzumab on day 4 after tracer injection were largely similar. The highest tracer concentration was seen in healthy liver, spleen, kidney and intestines. About one-third of the injected tracer dose was found in the circulation, up to 15% in the liver and only 4% in the spleen and kidney. Lower tracer concentration was seen in bone marrow, lung, compact bone, muscle, fat and the brain. Despite low tracer accumulation per gram of tissue, large-volume tissues, especially fat, can influence overall distribution: On average, 5-7% of the injected tracer dose accumulated in fat, with a peak of 19% in a patient with morbid obesity. Conclusion: The similar biodistribution of the four antibodies is probably based on their similar molecular structure, binding characteristics and similar metabolic pathways. These data provide a basis for a prospectively growing, online accessible warehouse of molecular imaging data, which enables researchers to increase and exchange knowledge on whole body drug distribution and potentially supports drug development decisions.

BackgroundEftilagimod alpha (efti; IMP321) is a soluble LAG-3 protein (LAG-3Ig) that binds to a subset of MHC class II molecules and mediates activation of antigen-presenting cells followed by CD8 T-cells. Weekly paclitaxel is a standard of care chemo-regimen after failure of endocrine-based therapy for metastatic breast carcinoma (MBC). AIPAC (Active Immunotherapy PAClitaxel) investigated the addition of efti to weekly paclitaxel in these patients (pts).MethodsThis placebo-controlled, double-blinded, 1:1 randomized phase IIb trial enrolled pts with measurable disease, HR+ HER2- MBC after endocrine-based therapy. Pts received paclitaxel (80 mg/m² IV on D1, D8, D15) + efti (30 mg) or placebo on D2, D16 (every 2 weeks) for up to 24 weeks following efti/placebo for up to 52 weeks. The primary endpoint (EP) was progression-free survival (RECIST1.1) by BICR. Secondary EPs included overall survival (OS), PFS (local read), overall response rate (ORR), biomarker, quality of life. Exploratory EPs included univariate/multivariate analyses.Results227 pts were randomized (Jan2017-Jul2019). All except 1 received ≥1 treatment and were included in the full analysis set [efti (n=114); placebo (n=112)]. Data cut-off was 14May2021 (min. follow-up= 22 months). Median age was 60 yrs with ECOG 0 in 61.5%. 91.6% had visceral disease. Pts were mostly endocrine resistant (84%) and partially pre-treated with CDK4/6 inhibitors (44.2%). Post-study treatment was similar. Median OS was 20.4 (95% CI: 14.3-25.1) months in the efti group vs. 17.5 (95% CI: 12.9-21.9) in the placebo group. HR was 0.88 (95%CI: 0.64-1.19; p=0.197). In predefined univariate analyses, younger pts, low baseline monocytes and luminal B showed significant/clinically meaningful improvement in OS (table 1).Efti increased PBMC/T cell (CD4/CD8) count vs. placebo, correlating with improved OS (Spearman Rho=0.6, p=0.02 for CD8 T cells). In a whole population multivariate cox regression model, increasing BMI and prior treatment with CDK4/6 were independent significant poor prognostic markers for PFS and OS.TEAEs leading to discontinuation were similar at 5.3%(efti) & 6.3%(placebo). PFS (Primary EP) and safety were reported at SABCS 2020 (Abstract#132).Abstract 948 Table 1Overall survival by subgroups at final analysisConclusionsEfti added to paclitaxel led to a non-significant 2.9 months median OS increase in HR+ HER2- MBC pts after endocrine-based therapy. Effects were significant in pts Trial RegistrationThe trial identifiers are IMP321-P011 (code for sponsor), 2015-002541-63 (EudraCT) and NCT02614833 (ClinicalTrials.gov).Ethics ApprovalThe study was approved by relevant ethic committees and institutional review boards.

T cell enhancing immune checkpoint inhibitors (ICI) are effective across several tumor types in a subset of patients. Insights into systemic localization of cytotoxic CD8+ T cells might support early treatment decisions. To address this, we performed a PET imaging study with a zirconium-89 (89Zr) labeled one-armed CD8-specific antibody 89ZED88082A to assess tracer performance, safety, and pharmacokinetics (PK) before and during treatment. Here we report preliminary data on uptake in tumor lesions before ICI. Methods: Patients with locally advanced or metastatic solid tumors that may benefit from ICI are eligible. In part A (imaging before treatment) and part B (imaging before and during treatment), 37 MBq (1 mCi) 89ZED88082A is administered with unlabeled one-armed antibody CED88004S to vary total protein dose. PET images are acquired at up to 4 time points: 1 h, and days (d) 2, 4, 7 post-injection followed by a tumor biopsy for CD8 immunohistochemistry and autoradiography (NCT04029181). Subsequently, patients receive atezolizumab (NCT02478099) or standard of care nivolumab ± ipilimumab. Tumor and lymph node 89ZED88082A uptake are assessed as (geometric mean) maximum standard uptake value (SUVmax), in other organs as SUVmean. Serum 89ZED88082A/CED88004S levels are measured for PK. Tumor response is according to (i)RECIST1.1. Results: For pretreatment imaging results, 32 patients (9 part A, 23 part B) were evaluable; 3 received 4 mg total tracer protein dose, 29 received 10 mg. No tracer infusion-related reactions occurred. Here we show results on d2 PET imaging with 10 mg protein dose, which was considered optimal based on superior 89Zr blood pool activity, clinical feasibility and serum antibody PK with a half-life of 28.6 h. 89ZED88082A uptake was observed within 1 h in spleen, and strong d2 imaging signal was seen across lymphoid organs including spleen (\bar{x}$ SUVmean 47.2), lymph nodes (SUVmax 4.2), bone marrow (\bar{x}$ SUVmean 5.0), small bowel and Waldeyer's ring. 89ZED88082A tumor uptake was seen at all main metastatic organ sites (overall lesion SUVmax 5.5, range 0.6-30.9) and varied across patients (\bar{x}$ per patient SUVmax 5.4, IQR 3.8-7.4). Higher tumor uptake showed a trend with better response (p=0.059) and longer PFS (p=0.033). Tumor uptake was higher in patients with mismatch-repair deficient (dMMR) than MMR proficient tumors (SUVmax 9.3 vs 4.9, p Citation Format: Laura Kist de Ruijter, Pim P. van de Donk, Jahlisa S. Hooiveld-Noeken, Danique Giesen, Alexander Ungewickell, Bernard M. Fine, Simon P. Williams, Sandra M. Sanabria Bohorquez, Mahesh Yadav, Hartmut Koeppen, Jing Jing, Sebastian Guelman, Mark T. Lin, Michael J. Mamounas, Jeffrey Eastham, Patrick K. Kimes, Andor W. Glaudemans, Adrienne H. Brouwers, Marjolijn N. Lub-de Hooge, Jourik A. Gietema, Carolina P. Schröder, Wim Timens, Mathilde Jalving, Sjoerd Elias, Sjoukje F. Oosting, Derk J. de Groot, Elisabeth G. de Vries. 89ZED88082A PET imaging to visualize CD8+ T cells in patients with cancer treated with immune checkpoint inhibitor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB037.

In addition to the well-known estrogen receptor (ER) and human epidermal growth factor receptor 2, the androgen receptor (AR) is also a potential drug target in breast cancer treatment. Whole-body imaging can provide information across lesions within a patient. ER expression in tumor lesions can be visualized by F-18-fluoroestradiol (F-18-FES) PET, and AR expression has been visualized in prostate cancer patients with F-18-fluorodihydrotestosterone (F-18-FDHT) PET. Our aim was to assess the concordance between F-18-FDHT and F-18-FES PET and tumor AR and ER expression measured immunohistochemically in patients with metastatic breast cancer. Methods: Patients with ER-positive metastatic breast cancer were eligible for the study, irrespective of tumor AR status. The concordance of F-18-FDHT and F-18-FES uptake on PET with immunohistochemical expression of AR and ER in biopsies of corresponding metastases was analyzed. Patients underwent F-18-FDHT PET and F-18-FES PET. A metastasis was biopsied within 8 wk of the PET procedures. Tumor samples with more than 10% and 1% nuclear tumor cell staining were considered, respectively, AR-and ER-positive. Correlations between PET uptake and semiquantitative immunohistochemical scoring (percentage positive cells x intensity) were calculated. The optimum threshold of SUV to discriminate positive and negative lesions for both AR and ER was determined by receiver-operating-characteristic analysis. Results: In the 13 evaluable patients, correlation (R-2) between semiquantitative AR expression and F-18-FDHT uptake was 0.47 (P = 0.01) and between semiquantitative ER expression and F-18-FES uptake 0.78 (P = 0.01). The optimal cutoff for AR-positive lesions was an SUVmax of 1.94 for F-18-FDHT PET, yielding a sensitivity of 91% and a specificity of 100%; the optimal cutoff was an SUVmax of 1.54 for F-18-FES PET, resulting in a sensitivity and specificity of 100% for ER. Conclusion: F-18-FDHT and F-18-FES uptake correlate well with AR and ER expression levels in representative biopsies. These results show the potential use of whole-body imaging for receptor status assessment, particularly in view of biopsy-associated sampling errors and heterogeneous receptor expression in breast cancer metastases.

The estrogen derivative 16a-F-18-fluoro-17 beta-estradiol (FES) is a PET tracer that has been used in a variety of preclinical and clinical studies to detect estrogen receptor (ER) expression, mainly in breast cancer, but also for other oncological indications. As a result of the success of these studies and the potential applications of the tracer, FES starts to be implemented in routine clinical practice. However, the number of centers using this tracer is still limited and many nuclear medicine physicians and medical oncologists are still unaware of the possibilities FES PET imaging offers. The aim of this article is therefore to give an overview of the main indications of FES PET in oncology and to provide recommendations on correct use of this imaging technique. This includes precautions that have to be taken for patient preparation, procedures for the acquisition of the scans, the physiological distribution of the tracer, factors that might influence tracer uptake and guidance for image analysis, quantification of tracer uptake, and reporting of the scans.

Lobular breast cancer is the second most common type of invasive breast cancer. Lobular breast cancer lesions are often difficult to detect with conventional imaging because they tend to grow less cohesively than ductal cancer. These images represent 3 lobular breast cancer cases, in whom confirmation of metastatic disease would make the crucial difference between curative or noncurative treatment. Staging with conventional imaging, however, yielded equivocal results, and a biopsy was not feasible. In contrast, FES (16 alpha-[F-18]fluoro-17 beta-estradiol) PET provided a decisive contribution to clinical decision making in these patients. This indicates that FES PET may have added value in relation to conventional staging.

BACKGROUND: The androgen receptor (AR) is a potential new therapeutic target in metastatic breast cancer, but it is likely that only patients with tumors overexpressing AR can benefit. To non-invasively assess AR expression in all metastatic lesions, whole-body imaging techniques such as 16β-18F-fluoro-5α-dihydrotestosterone positron emission tomography (18F-FDHT-PET), can be used. 18F-FDHT uptake reduction during treatment with an AR antagonist may indicate effective targeting of AR, and therefore might be predictive for response to treatment. This study assessed the feasibility of 18F-FDHT-PET to detect changes in 18F-FDHT uptake during treatment with AR antagonist bicalutamide, and evaluated whether these changes were related to bicalutamide response. PATIENTS AND METHODS: Postmenopausal women with AR positive (>10% staining on primary tumor or metastasis biopsy), human epidermal growth factor receptor 2-negative metastatic breast cancer, regardless of estrogen receptor status, were eligible. At baseline, patients underwent a bone scintigraphy, and contrast-enhanced computed tomography, as part of standard staging. Metastatic lesions were identified on both imaging modalities. All patients received an 18F-FDHT-PET at baseline and after 4 to 6 weeks bicalutamide 150mg daily. Baseline 18F-FDHT uptake was expressed as maximum standardized uptake value (SUVmax). Change between baseline and repeated 18F-FDHT uptake was assessed as (difference of) SUVmax corrected for physiological background activity (SUVcor and ΔSUVcor), per patient and per lesion. Response evaluation was performed by radiological imaging according to RECIST 1.1 and/or clinical assessment. Clinical benefit was defined as absence of disease progression for at least 24 weeks. RESULTS: Twenty-one patients (mean age 65 years) with in total 536 metastatic lesions were included. Baseline 18F-FDHT-PET detected 341 of 515 lesions found with standard imaging, and 21 new lesions. Baseline 18F-FDHT uptake varied widely between patients (median SUVmax 2.62; range 1.58 - 5.58) and lesions (3.07; 0.64 - 20.24). Repeated 18F-FDHT-PET was evaluable in 17 patients with 349 metastatic lesions. Repeated 18F-FDHT uptake showed median SUVcor decrease from 1.30 to 0.72 per patient, and 1.26 to 0.69 per lesion (P < 0.001). Median ΔSUVcor per patient was −45% (range −72% to −7%), and per lesion −39% (−95% to +100%). In patients with clinical benefit on bicalutamide (n = 3), median ΔSUVcor was −49% versus −39% in patients without benefit (n = 14; P = 0.801). CONCLUSION: In this feasibility study, bicalutamide induced 18F-FDHT uptake change could be detected by repeated 18F-FDHT-PET in patients with AR positive metastatic breast cancer. However, in this small study, this change was not significantly related to bicalutamide response per patient. (NCT02697032) Funding by UMCG Healthy Ageing Pilots and de Cock-Hadders Foundation Citation Format: Jorianne Boers, Clasina M. Venema, Erik FJ de Vries, Geke AP Hospers, Hendrikus H. Boersma, Bart Rikhof, Christine Dorbritz, Andor WJM Glaudemans, Carolina P. Schröder. Visualizing bicalutamide effect on androgen receptor availability in patients with metastatic breast cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 803.

Background: Eftilagimod alpha (efti, previously IMP321) is a recombinant LAG-3Ig fusion protein that binds to MHC class II and mediates antigen-presenting cell (APC) activation followed by CD8 T-cell activation. Efti may lead to the activation of the dendritic cell network and to a strong anti-tumor CD8 T cell response when injected s.c. after chemotherapy at a time when these APC are loaded with tumor antigens. AIPAC (Active Immunotherapy PAClitaxel; NCT02614833) is a Phase IIb trial in hormone receptor (HR)-positive metastatic breast carcinoma patients receiving efti or placebo as adjunctive to weekly paclitaxel as a first-line chemotherapy. The randomized double blinded part (stage 2) has finished recruitment in June 2019. Primary analysis is expected Q1 2020. Methods: This clinical trial is a placebo-controlled, double-blind, 1:1 randomized Phase IIb study aimed to enrol 226 patients at multiple centres across 7 different European countries. Patients with metastatic, HR-positive breast adenocarcinoma receiving first line chemotherapy with weekly paclitaxel and with measurable disease according to RECIST 1.1 are eligible. Patients suitable for Her2/neu targeted therapy are excluded from the trial. In the first treatment phase, paclitaxel (80 mg/m² IV at D1, D8, D15 plus efti (30 mg s.c.) or placebo at D2, D16 will be administered for 6 cycles (1 cycle = 4 weeks). This is followed by a maintenance phase in which stable or responding patients will receive efti or placebo for up to additional 52 weeks (12 injections). The primary endpoint is progression-free survival (PFS) by blinded independent central imaging review. Secondary endpoints include PFS by local assessment, overall survival, tumor response according to RECIST 1.1., time to and duration of response, duration of stable disease and quality of life. Citation Format: Luc Dirix, Hans Wildiers, Manon T Huizing, Eveline De Cuypere, Carolina P Schröder, Anne C Armstrong, Jans Huober, Frederik Marmé, Andreas Schneeweiss, Sarah Khan, Fréderic Triebel. AIPAC (Active Immunotherapy PAClitaxel): A randomized, double blind, placebo controlled, multinational phase IIb trial evaluating the efficacy of eftilagimod alpha (a soluble LAG-3 fusion protein) in combination with paclitaxel in hormone receptor positive metastatic breast cancer [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr OT1-01-05.

Rationale: Knowledge on monoclonal antibody biodistribution in healthy tissues in humans can support clinical drug development. Molecular imaging with positron emission tomography (PET) can yield information in this setting. However, recent imaging studies have analyzed the behavior of single antibodies only, neglecting comparison across different antibodies. Methods: We compared the distribution of four 89Zr-labeled antibodies in healthy tissue in a retrospective analysis based on the recently published harmonization protocol for 89Zr-tracers and our delineation protocol. Results: The biodistribution patterns of 89Zr-lumretuzumab, 89Zr-MMOT0530A, 89Zr-bevacizumab and 89Zr-trastuzumab on day 4 after tracer injection were largely similar. The highest tracer concentration was seen in healthy liver, spleen, kidney and intestines. About one-third of the injected tracer dose was found in the circulation, up to 15% in the liver and only 4% in the spleen and kidney. Lower tracer concentration was seen in bone marrow, lung, compact bone, muscle, fat and the brain. Despite low tracer accumulation per gram of tissue, large-volume tissues, especially fat, can influence overall distribution: On average, 5-7% of the injected tracer dose accumulated in fat, with a peak of 19% in a patient with morbid obesity. Conclusion: The similar biodistribution of the four antibodies is probably based on their similar molecular structure, binding characteristics and similar metabolic pathways. These data provide a basis for a prospectively growing, online accessible warehouse of molecular imaging data, which enables researchers to increase and exchange knowledge on whole body drug distribution and potentially supports drug development decisions.

// Hilde H. Nienhuis 1,* , Marlous Arjaans 1,* , Hetty Timmer-Bosscha 1 , Elisabeth G.E. de Vries 1 and Carolina P. Schroder 1 1 Department of Medical Oncology, University of Groningen and University Medical, Center Groningen, Groningen, The Netherlands * These authors have contributed equally to this work Correspondence to: Carolina P. Schroder, email: // Keywords : breast cancer, microenvironment, zoledronic acid, TGF-β, CAM model Received : February 16, 2015 Accepted : May 30, 2015 Published : June 10, 2015 Abstract Previous studies suggested that bisphosphonate zoledronic acid exerts an anti-tumor effect by interacting with the microenvironment. In this study, we aimed to elucidate the mechanism behind the anti-breast cancer effect of zoledronic acid. Here we showed that zoledronic acid did not influence in vitro human breast cancer cell survival, but did affect human stromal cell survival. Breast cancer cell death in co-culture with stromal cells was analyzed in vitro by fluorescent microscopy and flowcytometry analysis. In co-culture, the addition of stromal cells to breast cancer cells induced tumor cell death by zoledronic acid, which was abolished by transforming growth factor (TGF)-β. In the in vivo chicken chorioallantoic membrane model, zoledronic acid reduced the breast cancer cells fraction per tumor only in the presence of human stromal cells. Zoledronic acid decreased TGF-β excretion by stromal cells and co-cultures. Moreover, supernatant of zoledronic acid treated stromal cells reduced phospho-Smad2 protein levels in breast cancer cells. Thus, zoledronic acid exerts an anti-breast cancer effect via stromal cells, accompanied by decreased stromal TGF-β excretion and reduced TGF-β signaling in cancer cells.

The human epidermal growth factor receptor (HER) family members are targeted by a growing numbers of small molecules and monoclonal antibodies. Resistance against the epidermal growth factor receptor (EGFR) and HER2-targeting agents is a clinically relevant problem forcing research on optimizing targeting of the HER family. In view of its overexpression in tumors, and compensatory role in HER signaling, HER3 has gained much interest as a potential additional target within the HER family. It is the only member of the HER family lacking intrinsic tyrosine kinase activity and therefore its role in cancer has long been underestimated. Drugs that block HER3 or interfere with HER3 dimer signaling, including fully human anti-HER3 antibodies, bispecific antibodies and tyrosine kinase inhibitors (TKIs), are currently becoming available. Several compounds have already entered clinical trial. In the meantime potential biomarkers are tested such as tumor analysis of HER3 expression, functional assays for downstream effector molecules and molecular imaging techniques. This review describes the biology and relevance of HER3 in cancer, agents targeting HER3 and potential biomarkers for effect of HER3-targeting.

In solid tumors, angiogenesis occurs in the setting of a defective vasculature and impaired lymphatic drainage that is associated with increased vascular permeability and enhanced tumor permeability. These universal aspects of the tumor microenvironment can have a marked influence on intratumoral drug delivery that may often be underappreciated. In this study, we investigated the effect of blood vessel normalization in tumors by the antiangiogenic drug bevacizumab on antibody uptake by tumors. In mouse xenograft models of human ovarian and esophageal cancer (SKOV-3 and OE19), we evaluated antibody uptake in tumors by positron emission tomographic imaging 24 and 144 hours after injection of 89Zr-trastuzumab (SKOV-3 and OE19), 89Zr-bevacizumab (SKOV-3), or 89Zr-IgG (SKOV-3) before or after treatment with bevacizumab. Intratumor distribution was assessed by fluorescence microscopy along with mean vessel density (MVD) and vessel normalization. Notably, bevacizumab treatment decreased tumor uptake and intratumoral accumulation compared with baseline in the tumor models relative to controls. Bevacizumab treatment also reduced MVD in tumors and increased vessel pericyte coverage. These findings are clinically important, suggesting caution in designing combinatorial trials with therapeutic antibodies due to a possible reduction in tumoral accumulation that may be caused by bevacizumab cotreatment. Cancer Res; 73(11); 3347–55. ©2013 AACR.

Molecular imaging with PET is a rapidly emerging technique. In breast cancer patients, more than 45 different PET tracers have been or are presently being tested. With a good rationale, after development of the tracer and proven feasibility, it is of interest to evaluate whether there is a potential meaningful role for the tracer in the clinical setting-such as in staging, in the (early) prediction of a treatment response, or in supporting drug choices. So far, only (18)F-FDG PET has been incorporated into breast cancer guidelines. For proof of the clinical relevance of tracers, especially for analysis in a multicenter setting, standardization of the technology and access to the novel PET tracer are required. However, resources for PET implementation research are limited. Therefore, next to randomized studies, novel approaches are required for proving the clinical value of PET tracers with the smallest possible number of patients. The aim of this review is to describe the process of the development of PET tracers and the level of evidence needed for the use of these tracers in breast cancer. Several breast cancer trials have been performed with the PET tracers (18)F-FDG, 3'-deoxy-3'-(18)F-fluorothymidine ((18)F-FLT), and (18)F-fluoroestradiol ((18)F-FES). We studied them to learn lessons for the implementation of novel tracers. After defining the gap between a good rationale for a tracer and implementation in the clinical setting, we propose solutions to fill the gap to try to bring more PET tracers to daily clinical practice.

Human epidermal growth factor receptor-2 (HER2) directed therapy potentially can be improved by insight in drug effects on HER2 expression. This study evaluates the effects of the EGFR/HER2 tyrosine kinase inhibitor lapatinib, the heat shock protein-90 inhibitor 17AAG, and their combination, on HER2 expression with in vivo HER2-PET imaging. Lapatinib and 17AAG effects on EGFR and HER2 membrane expression were determined in vitro using flow cytometry of human SKBR3 tumor cells. Effect of lapatinib on HER2 internalization was studied in vitro by Zr-89-trastuzumab-F(ab')(2) internalization. For in vivo evaluation, Zr-89-trastuzurnab-F(ab')(2) mu PET imaging was performed two times with a 7 day interval. Lapatinib was administered for 6 days, starting 1 day after the baseline scan. 17AAG was given 1 day before the second Zr-89-trastuzumab-F(ab')(2) injection. Imaging data were compared with ex vivo biodistribution analysis and HER2 immunohistochemical staining. 17AAG treatment lowered EGFR expression by 41% (P = 0.016) and HER2 by 76% (P = 0.022). EGFR/HER2 downregulation by 17AAG was inhibited by lapatinib pretreatment. Lapatinib reduced internalization of Zr-89-trastuzumab-F(ab')(2) with 25% (P = 0.0022). Zr-89-trastuzumab-F(ab')(2) tumor to blood ratio was lowered 32% by lapatinib (P = 0.00004), 34% by 17AAG (P = 0.0022) and even 53% by the combination (P = 0.011). Lapatinib inhibits HER2 internalization and 17AAG lowers HER2 membrane expression. Both drugs reduce Zr-89-trastuzumab-F(ab')(2) tumor uptake. Based on our findings, supported by previous preclinical data indicating the antitumor potency of lapatinib in combination with HSP90 inhibition, combination of these drugs deserves further investigation.

Transforming growth factor-beta (TGF-beta) promotes cancer invasion and metastasis and is therefore a potential drug target for cancer treatment. Fresolimumab, which neutralizes all mammalian active isoforms of TGF-beta, was radiolabeled with Zr-89 for PET to analyze TGF-beta expression, antibody tumor uptake, and organ distribution. Methods: Zr-89 was conjugated to fresolimumab using the chelator N-succinyldesferrioxamine-B-tetrafluorphenol. Zr-89-fresolimumab was analyzed for conjugation ratio, aggregation, radiochemical purity, stability, and immunoreactivity. Zr-89-fresolimumab tumor uptake and organ distribution were assessed using 3 protein doses (10, 50, and 100 mu g) and compared with In-111-IgG in a human TGF-beta-transfected Chinese hamster ovary xenograft model, human breast cancer MDA-MB-231 xenograft, and metastatic model. Latent and active TGF-beta 1 expression was analyzed in tissue homogenates with enzyme-linked immunosorbent assay. Results: Zr-89 was labeled to fresolimumab with high specific activity (>1 GBq/mg), high yield, and high purity. In vitro validation of 89Zr-fresolimumab showed a fully preserved immunoreactivity and long (>1 wk) stability in solution and in human serum. In vivo validation showed an Zr-89-fresolimumab distribution similar to IgG in most organs, except for a higher uptake in the liver in all mice and higher kidney uptake in the 10-mu g group. Zr-89-fresolimumab induced no toxicity in mice; it accumulated in primary tumors and metastases in a manner similar to IgG. Both latent and active TGF-beta was detected in tumor homogenates, whereas only latent TGF-beta could be detected in liver homogenates. Remarkably high Zr-89-fresolimumab uptake was seen in sites of tumor ulceration and in scar tissue, processes in which TGF-beta is known to be highly active. Conclusion: Fresolimumab tumor uptake and organ distribution can be visualized and quantified with Zr-89-fresolimumab PET. This technique will be used to guide further clinical development of fresolimumab and could possibly identify patients most likely to benefit.

We would like to reply to the letter by Huang and colleagues on the impact of vascular changes induced by antiangiogenic therapy on tumor uptake of other drugs in relation to our findings of reduced antibody uptake after bevacizumab treatment ([1][1]). Their comments focus on the bevacizumab dose

The human epidermal growth factor receptor (HER) family members are targeted by a growing numbers of small molecules and monoclonal antibodies. Resistance against the epidermal growth factor receptor (EGFR) and HER2-targeting agents is a clinically relevant problem forcing research on optimizing targeting of the HER family. In view of its overexpression in tumors, and compensatory role in HER signaling, HER3 has gained much interest as a potential additional target within the HER family. It is the only member of the HER family lacking intrinsic tyrosine kinase activity and therefore its role in cancer has long been underestimated. Drugs that block HER3 or interfere with HER3 dimer signaling, including fully human anti-HER3 antibodies, bispecific antibodies and tyrosine kinase inhibitors (TKIs), are currently becoming available. Several compounds have already entered clinical trial. In the meantime potential biomarkers are tested such as tumor analysis of HER3 expression, functional assays for downstream effector molecules and molecular imaging techniques. This review describes the biology and relevance of HER3 in cancer, agents targeting HER3 and potential biomarkers for effect of HER3-targeting.

(1996). Glutathione and Drug Resistance. Cancer Investigation: Vol. 14, No. 2, pp. 158-168.