Growth factor dependency in mammary organoids regulates ductal morphogenesis during organ regeneration

AbstractSignaling pathways play an important role in cell fate determination in stem cells and regulate a plethora of developmental programs, the dysregulation of which can lead to human diseases. Growth factors (GFs) regulating these signaling pathways therefore play a major role in the plasticity of adult stem cells and modulate cellular differentiation and tissue repair outcomes. We consider murine mammary organoid generation from self-organizing adult stem cells as a tool to understand the role of GFs in organ development and tissue regeneration. The astounding capacity of mammary organoids to regenerate a gland in vivo after transplantation makes it a convenient model to study organ regeneration. We show organoids grown in suspension with minimal concentration of Matrigel and in the presence of a cocktail of GFs regulating EGF and FGF signaling can recapitulate key epithelial layers of adult mammary gland. We establish a toolkit utilizing in vivo whole animal imaging and ultrasound imaging combined with ex vivo approaches including tissue clearing and confocal imaging to study organ regeneration and ductal morphogenesis. Although the organoid structures were severely impaired in vitro when cultured in the presence of individual GFs, ex vivo imaging revealed ductal branching after transplantation albeit with significantly reduced number of terminal end buds. We anticipate these imaging modalities will open novel avenues to study mammary gland morphogenesis in vivo and can be beneficial for monitoring mammary tumor progression in pre-clinical and clinical settings.</jats:p

Aspects of Three-Dimensional Imaging by Classical Tomography for Dual Detector Positron Emission Mammography (PEM)

Images from dual detector positron emission mammography (PEM) systems are commonly reconstructed by backprojection methods of classical tomography. Characteristics of three-dimensional (3-D) PEM images were investigated using analytic models, computer simulations, and experimental acquisitions with compact pixellated detectors, in particular depth resolution normal to the detectors. An analytic formula was developed using circular image pixels that models blurring normal to the detectors. The amount of blurring is dependent on the acceptance angle for coincidence events and may vary across the field of view due to geometric limitations on the maximum angle of lines of response normal to the detectors. For experimental acquisitions with line sources and a pixellated lutetium gadolinium oxyorthosilicate (LGSO) detector, depth resolution is broader than predicted by numerical simulations, possibly due to uncorrected randoms or scatter within the scintillator arrays. Iterative image reconstruction with the maximum likelihood expectation maximization (MLEM) algorithm of a compressed breast phantom acquisition with a pixellated gadolinium oxyorthosilicate (GSO) detector shows improved contract compared with backprojection reconstruction. Image reconstruction for dual detector PEM with static detectors represents a case of limited angle tomography with truncated projection data, and there is the opportunity to improve three-dimensional PEM imaging by the use of more sophisticated image reconstruction techniques

Abstract LB-510: Longitudinal imaging of melanoma cancer cell metastasis in a preclinical model by bioluminescence, PET/CT, and MRI

Abstract Metastasis of cancer cells from the primary tumor are a leading cause of poor prognosis and death from cancer. The lung is the most common organ for metastatic spread of cancer and leads to 20-50% of cancer-related deaths. Early detection of cancer metastasis to the lung and other organs could be beneficial in disease management. In animal studies, most secondary-site disseminated tumors are evaluated by optical imaging, and gross examination at necropsy and subsequent histopathological analysis. These techniques are limited in that they do not offer the ability to monitor animals during the course of tumor development, and optical microscopy may have inadequate field of view (3-D morphometrics) for thorough tumor evaluation. Noninvasive imaging allows for serial imaging of the same animal, with the ability to repeat measurements in real-time to enhance the observation of disease progression and therapeutic response. Here, we compared the multimodal noninvasive whole-body imaging techniques bioluminescence, 2-deoxy-2-(18F)fluoro-D-glucose (18F-FDG) positron emission tomography (PET)/ computed tomography (CT), and magnetic resonance imaging (MRI) to image lung metastasis development following intravenous injection of melanoma cells (B16-F10 melanoma cell line expressing luciferase gene) in C57BL/6. By bioluminescence imaging, tumor cell localization in the lungs was evident as early as 3 days (1 x 105 photons/sec; total flux) post cancer cell inoculation and steadily increased through the end of week four (1 x 108 photons/sec; total flux). The increase in the lung tumor burden was confirmed by lung histopathology. As it was difficult to de-convolve the effects of photon flux spreading from lungs over other organs, other metastatic sites were not evaluated by bioluminescence. CT and 18F-FDG-PET are used clinically to image lung metastasis and offer excellent resolution of anatomical and metabolic details of cancer cells. We were able to image metastatic nodules in the lung by PET/CT at week four, which showed a strong correlation with histopathology analysis. [18F]FDG-PET/CT also revealed metastasis in the kidney and mesentery and were confirmed by histopathology. T2-weighted MRI at week 4 identified several lung tumors and metastatic cancers in the abdomen. A comparison of [18F]FDG/PET at week 4 to MRI and CT displayed several highly metabolic regions within the tumors. Histopathology confirmed the metastatic spread of the melanoma cells to the lung, mediastinum, kidney, and mesentery. Funded by NCI contract No. HHSN261200800001E. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-510. doi:1538-7445.AM2012-LB-510</jats:p

A spontaneously metastatic model of bladder cancer: imaging characterization

Abstract Background Spontaneously metastatic xenograft models of cancer are infrequent and the few that exist are resource intensive. In xenografts, caliper measurements can be used to determine primary tumor burden and response to therapy but in metastatic disease models determination of the presence of metastatic disease, metastatic burden, and response to therapy are difficult, often requiring serial necropsy. In this study we characterized the development of visceral metastases in a patient derived xenograft model (PDXM) using in vivo imaging. Results We identified and characterized the previously unreported development of spontaneous liver and bone metastasis in a known patient derived xenograft, bladder xenograft BL0293F, developed by Jackson Laboratories and the University of California at Davis and available from the National Cancer Institute Patient-Derived Models Repository [1]. Among FDG-PET/CT, contrast-enhanced MRI and non-contrast MRI, non-contrast T2w MRI was the most effective and efficient imaging technique. On non-contrast T2 weighted MRI, hepatic metastases were observed in over 70% of animals at 52 days post tumor implantation without resection of the xenograft and in 100% of animals at day 52 following resection of the xenograft. In a group of animals receiving one cycle of effective chemotherapy, no animals demonstrated metastasis by imaging, confirming the utility of this model for therapy evaluation. There was good agreement between pathologic grade and extent of involvement observed on MRI T2w imaging. Conclusion PDX BL0293F is a reliable visceral organ (liver) metastatic model with high penetrance in both non-aggravated and post excisional situations, providing a reliable window for therapy intervention prior to required excision of the xenograft. The imaging characteristics of this model are highly favorable for non-clinical research studies of metastatic disease when used in conjunction with non-contrast T2 weighted MRI. </jats:sec

Cyanine Masking: A Strategy to Test Functional Group Effects on Antibody Conjugate Targeting

Conjugates of small molecules and antibodies are broadly employed diagnostic and therapeutic agents. Appending a small molecule to an antibody often significantly impacts the properties of the resulting conjugate. Here, we detail a systematic study investigating the effect of various functional groups on the properties of antibody–fluorophore conjugates. This was done through the preparation and analysis of a series of masked heptamethine cyanines (CyMasks)-bearing amides with varied functional groups. These were designed to exhibit a broad range of physical properties, and include hydrophobic (−NMe2), pegylated (NH-PEG-8 or NH-PEG-24), cationic (NH-(CH2)2NMe3+), anionic (NH-(CH2)2SO3–), and zwitterionic (N-(CH2)2NMe3+)-(CH2)3SO3–) variants. The CyMask series was appended to monoclonal antibodies (mAbs) and analyzed for the effects on tumor targeting, clearance, and non-specific organ uptake. Among the series, zwitterionic and pegylated dye conjugates had the highest tumor-to-background ratio (TBR) and a low liver-to-background ratio. By contrast, the cationic and zwitterionic probes had high tumor signal and high TBR, although the latter also exhibited an elevated liver-to-background ratio (LBR). Overall, these studies provide a strategy to test the functional group effects and suggest that zwitterionic substituents possess an optimal combination of high tumor signal, TBR, and low LBR. These results suggest an appealing strategy to mask hydrophobic payloads, with the potential to improve the properties of bioconjugates in vivo

Design and Implementation of the Pre-Clinical DICOM Standard in Multi-Cohort Murine Studies

The small animal imaging Digital Imaging and Communications in Medicine (DICOM) acquisition context structured report (SR) was developed to incorporate pre-clinical data in an established DICOM format for rapid queries and comparison of clinical and non-clinical datasets. Established terminologies (i.e., anesthesia, mouse model nomenclature, veterinary definitions, NCI Metathesaurus) were utilized to assist in defining terms implemented in pre-clinical imaging and new codes were added to integrate the specific small animal procedures and handling processes, such as housing, biosafety level, and pre-imaging rodent preparation. In addition to the standard DICOM fields, the small animal SR includes fields specific to small animal imaging such as tumor graft (i.e., melanoma), tissue of origin, mouse strain, and exogenous material, including the date and site of injection. Additionally, the mapping and harmonization developed by the Mouse-Human Anatomy Project were implemented to assist co-clinical research by providing cross-reference human-to-mouse anatomies. Furthermore, since small animal imaging performs multi-mouse imaging for high throughput, and queries for co-clinical research requires a one-to-one relation, an imaging splitting routine was developed, new Unique Identifiers (UID’s) were created, and the original patient name and ID were saved for reference to the original dataset. We report the implementation of the small animal SR using MRI datasets (as an example) of patient-derived xenograft mouse models and uploaded to The Cancer Imaging Archive (TCIA) for public dissemination, and also implemented this on PET/CT datasets. The small animal SR enhancement provides researchers the ability to query any DICOM modality pre-clinical and clinical datasets using standard vocabularies and enhances co-clinical studies.</jats:p