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Numerous recent works highlight the limited utility of established tumor cell lines in recapitulating the heterogeneity of tumors in patients. More realistic preclinical cancer models are thought to be provided by transplantable, patient-derived xenografts (PDXs). The inter- and intratumor heterogeneity of PDXs, however, presents several challenges in developing optimal quantitative pipelines to assess response to therapy. The objective of this work was to develop and optimize image metrics for

Purpose: To investigate whether longitudinal functional PET imaging of mammary tumors using the radiopharmaceuticals [18F]FDG (to measure glucose uptake), [18F]FES [to measure estrogen receptor (ER) levels], or [18F]FFNP [to measure progesterone receptor (PgR) levels] is predictive of response to estrogen-deprivation therapy. Experimental Design: [18F]FDG, [18F]FES, and [18F]FFNP uptake in endocrine-sensitive and -resistant mammary tumors was quantified serially by PET before ovariectomy or estrogen withdrawal in mice, and on days 3 and 4 after estrogen-deprivation therapy. Specificity of [18F]FFNP uptake in ERα+ mammary tumors was determined by competition assay using unlabeled ligands for PgR or glucocorticoid receptor (GR). PgR expression was also assayed by immunohistochemistry (IHC). Results: The levels of [18F]FES and [18F]FDG tumor uptake remained unchanged in endocrine-sensitive tumors after estrogen-deprivation therapy compared with those at pretreatment. In contrast, estrogen-deprivation therapy led to a reduction in PgR expression and [18F]FFNP uptake in endocrine-sensitive tumors, but not in endocrine-resistant tumors, as early as 3 days after treatment; the changes in PgR levels were confirmed by IHC. Unlabeled PgR ligand R5020 but not GR ligand dexamethasone blocked [18F]FFNP tumor uptake, indicating that [18F]FFNP bound specifically to PgR. Therefore, a reduction in FFNP tumor to muscle ratio in mammary tumors predicts sensitivity to estrogen-deprivation therapy. Conclusions: Monitoring the acute changes in ERα activity by measuring [18F]FFNP uptake in mammary tumors predicts tumor response to estrogen-deprivation therapy. Longitudinal noninvasive PET imaging using [18F]FFNP is a robust and effective approach to predict tumor responsiveness to endocrine treatment. Clin Cancer Res; 21(5); 1063–70. ©2014 AACR.

Overall survival of patients with newly diagnosed glioblastoma (GBM) remains dismal at 16 months with state-of-the-art treatment that includes surgical resection, radiation, and chemotherapy. GBM tumors are highly heterogeneous, and mechanisms for overcoming tumor resistance have not yet fully been elucidated. An injectable chitosan hydrogel capable of releasing chemotherapy (temozolomide [TMZ]) while retaining radioactive isotopes agents (iodine, [ 131 I]) was used as a vehicle for localized radiation and chemotherapy, within the surgical cavity. Release from hydrogels loaded with TMZ or 131 I was characterized in vitro and in vivo and their efficacy on tumor progression and survival on GBM tumors was also measured. The in vitro release of 131 I was negligible over 42 days, whereas the TMZ was completely released over the first 48 h. 131 I was completely retained in the tumor bed with negligible distribution in other tissues and that when delivered locally, the chemotherapy accumulated in the tumor at 10-fold higher concentrations than when delivered systemically. We found that the tumors were significantly decreased, and survival was improved in both treatment groups compared to the control group. Novel injectable chemo-radio-hydrogel implants may potentially improve the local control and overall outcome of aggressive, poor prognosis brain tumors.

Rationale and Objectives Non-invasive longitudinal imaging of tumor vasculature could provide new insights into the development of solid tumors, facilitating efficient delivery of therapeutics. In this study, we report three-dimensional imaging and characterization of tumor vascular architecture using a nanoparticle contrast agent and high-resolution computed tomography (CT) imaging. Materials and Methods Five Balb/c mice implanted with 4T1/Luc syngeneic breast tumors cells were used for the study. The nanoparticle contrast agent was systemically administered and longitudinal CT imaging was performed pre-contrast and at serial time points post-contrast, for up to 7 days for studying the characteristics of tumor-associated blood vessels. Gene expression of tumor angiogenic biomarkers was measured using quantitative real-time polymerase chain reaction. Results Early-phase imaging demonstrated the presence of co-opted and newly developed tumor vessels. The co-opted vessels demonstrated wall-permeability and “leakiness” characteristics evident by an increase in extravascular nanoparticle-based signal enhancement visible well beyond the margins of tumor. Diameters of tumor-associated vessels were larger than the contralateral normal vessels. Delayed-phase imaging also demonstrated significant accumulation of nanoparticle contrast agent both within and in areas surrounding the tumor. A heterogeneous pattern of signal enhancement was observed both within and among individual tumors. Gene-expression profiling demonstrated significant variability in several angiogenic biomarkers both within and among individual tumors. Conclusions The nanoparticle contrast agent and high-resolution CT imaging facilitated visualization of co-opted and newly developed tumors vessels as well as imaging of nanoparticle accumulation within tumors. The use of this agent could provide novel insights into tumor vascular biology and could have implications on the monitoring of tumor status.

Introduction Tracer kinetic modeling used in conjunction with positron emission tomography (PET) is an excellent tool for the noninvasive quantification of physiological, biological and molecular processes and their alterations due to disease. Currently, complex multi-compartment modeling approaches are being applied in a variety of clinical studies to determine myocardial perfusion, viability and glucose utilization as well as fatty acid metabolism and oxidation in the normal and diseased heart. These kinetic models require two key measurements of tracer activity over time, tracer activity in arterial blood (input function) and its corresponding activity in the organ of interest. The alteration in the time course of tracer activity as it travels from blood to the organ of interest describes the kinetics of the tracer. To be able to implement these approaches in rodent models of disease using small-animal PET (microPET), it is imperative that the input function is measured accurately. Methods The blood input functions in rodent experiments were obtained by (1) direct blood sampling, (2) direct measurement of blood activity by a beta-detecting probe that counts the activity in the blood, (3) an arterial–venous bypass (A/V shunt), (4) factor analysis of dynamic structures from dynamic PET images and (5) measurement from region-of-interest (ROI) analysis of dynamic PET images. Direct blood sampling was used as the reference standard to which the results of the other techniques were compared. Results Beta probes are difficult to operate and may not provide accurate blood input functions unless they are used intravenously, which requires complicated microsurgery. A similar limitation applies to the A/V shunt. Factor analysis successfully extracts the blood input function for mice and rats. The ROI-based method is less accurate due to limited image resolution of the PET system, which results in severe partial volume effect and spillover from myocardium. Conclusion The current reference standard, direct blood sampling, is more invasive and has limited temporal resolution. With current imaging technology, image-based extraction of blood input functions is possible by factor analysis, while forthcoming technological developments are likely to allow extraction of input function directly from the images. These techniques will reduce the level of complexity and invasiveness for animal experiments and are likely to be used more widely in the future.

Estrogen exerts diverse biological effects in multiple tissues in both animals and humans. Much of the accumulated knowledge on the role of estrogen receptor (ER) in the heart has been obtained from studies using ovariectomized mice, whole body ER gene knock-out animal models, ex vivo heart studies, or from isolated cardiac myocytes. In light of the wide systemic influence of ER signaling in regulating a host of biological functions in multiple tissues, it is difficult to infer the direct role of ER on the heart. Therefore, we developed a mouse model with a cardiomyocyte-specific deletion of the ERα allele (cs-ERα−/−). Male and female cs-ERα−/− mice with age/sex-matched wild type controls were examined for differences in cardiac structure and function by echocardiogram and differential gene expression microarray analysis. Our study revealed sex-differences in structural parameters in the hearts of cs-ERα−/− mice, with minimal functional differences. Analysis of microarray data revealed differential variations in the expression of 208 genes affecting multiple transcriptional networks. Furthermore, we report sex-specific differences in the expression of 56 genes. Overall, we developed a mouse model with cardiac-specific deletion of ERα to characterize the role of ERα in the heart independent of systemic effects. Our results suggest that ERα is involved in controlling the expression of diverse genes and networks in the cardiomyocyte in a sex-dependent manner.

Estrogen receptor-α (ERα) is expressed in most human breast cancers and is an important predictive factor for directing therapy. Compared with patients with ERα-negative tumors, patients with ERα-positive (ERα+) tumors typically have longer overall survival and are more likely to respond to hormone-based therapies designed to decrease endogenous estrogens or block ER transcriptional activity, such as aromatase inhibitors, tamoxifen, or fulvestrant. However, some patients with ERα+ tumors do not respond to endocrine therapy (1). The progesterone receptor gene (PGR) is a classic estrogen-stimulated target gene, and currently both ERα and progesterone receptor (PR) are routinely measured using immunohistochemistry as part of the pathologic assessment of breast cancer (2). Although immunohistochemistry is the current gold standard for detecting steroid hormone receptor expression, noninvasive imaging offers several advantages. For instance, imaging can assess the entire lesion, which is advantageous for tumors with heterogeneous receptor expression that could lead to biopsy sample error. Furthermore, noninvasive imaging is useful for metastatic lesions that may be technically difficult to biopsy or for patients at high risk for biopsy-related complications. Repeated imaging can assess whole-body disease burden over time, as one cannot presume that recurrent or metastatic lesions retain the same steroid hormone expression profile as the primary tumor and discordance occurs in up to 30% of the patients who ultimately have a decreased response to hormonal therapy (3,4). The ability of these nuclear receptors to bind ligand provides a convenient target for the development of imaging agents (5). The estrogen-based radiopharmaceutical 18F-fluoroestradiol (18F-FES) has high specific activity, binding affinity, and selectivity for ERα over the ERβ subtype (6) and has been shown to exhibit high specific uptake by ER-rich target tissues and ERα+ mammary tumors using small-animal models (7,8). For human breast cancer patients, use of 18F-FES PET as a measure of tumoral ERα expression has been validated by the gold standards of immunohistologic and biochemical receptor assays with significant concordance (9–11). 18F-FES imaging has also been shown to be a predictive assay for endocrine therapy since patients with pretreatment 18F-FES standardized uptake values greater than 1.5–2.0 correlate with a positive response to therapy (12,13). Although not as well studied as 18F-FES, progestin-based radiopharmaceuticals such as 18F-fluoro furanyl norprogesterone (18F-FFNP) have been developed for PR imaging as an indicator of a functionally intact ERα signaling pathway (14). Preclinical studies have demonstrated high 18F-FFNP uptake in the uterus and ovaries of estrogen-primed immature female rats (14), and initial results for 18F-FFNP imaging of human breast cancer patients have recently been published (15). The goal of this study was to determine whether changes in tumoral steroid hormone receptor expression as assessed by molecular imaging after endocrine therapy are of predictive value using a preclinical animal model of human luminal breast cancer. Disruption of the gene for the signal transducer and activator of transcription-1 (STAT1) in mice results in a complete lack of responsiveness to interferons and, interestingly, spontaneous development of mammary tumors (16). These adenocarcinomas are remarkably similar to human luminal breast cancers since they express ERα and PR, require ovarian hormones for growth, and have significantly overlapping gene expression profiles (17). Furthermore, the tumors are HER2/neu-negative (17) and have no detectable epidermal growth factor receptor or ERβ protein expression (Szeman Ruby Chan and Robert D. Schreiber, unpublished data, May 29, 2012). In the current study, we performed 18F-FES and 18F-FFNP PET of primary and implanted STAT1-deficient (STAT1−/−) mammary tumors at baseline and in response to fulvestrant, a pure ERα antagonist, to determine whether changes in these imaging bio-markers can serve as an early predictive indicator of the ultimate response to therapy. An imaging approach that successfully predicts tumor response to standard endocrine therapy would be a useful tool for the further evaluation of novel breast cancer treatments.

Evidence is emerging that systemic metabolic disturbances contribute to cardiac myocyte dysfunction and clinically apparent heart failure, independent of associated coronary artery disease. To test the hypothesis that perturbation of lipid homeostasis in cardiomyocytes contributes to cardiac dysfunction, we engineered transgenic mice with cardiac-specific overexpression of fatty acid transport protein 1 (FATP1) using the α-myosin heavy chain gene promoter. Two independent transgenic lines demonstrate 4-fold increased myocardial free fatty acid (FFA) uptake that is consistent with the known function of FATP1. Increased FFA uptake in this model likely contributes to early cardiomyocyte FFA accumulation (2-fold increased) and subsequent increased cardiac FFA metabolism (2-fold). By 3 months of age, transgenic mice have echocardiographic evidence of impaired left ventricular filling and biatrial enlargement, but preserved systolic function. Doppler tissue imaging and hemodynamic studies confirm that these mice have predominantly diastolic dysfunction. Furthermore, ambulatory ECG monitoring reveals prolonged QT c intervals, reflecting reductions in the densities of repolarizing, voltage-gated K + currents in ventricular myocytes. Our results show that in the absence of systemic metabolic disturbances, such as diabetes or hyperlipidemia, perturbation of cardiomyocyte lipid homeostasis leads to cardiac dysfunction with pathophysiological findings similar to those in diabetic cardiomyopathy. Moreover, the MHC-FATP model supports a role for FATPs in FFA import into the heart in vivo.