ASSESSING THE EFFECT OF TREATMENT ON SOLITARY DORMANT METASTATIC CELLS BY MRI AND OPTICAL IMAGING

Metastatic disease is responsible for the majority of cancer related deaths. However, most anti-cancer drugs are not designed or tested for efficacy against this heterogeneous cancer cell population. This is in part due to the technical challenges involved in imaging and quantifying individual cells, or even large metastases, in common non-superficial sites of metastasis including lung, brain, bone marrow and liver. The purpose of the studies presented here was to develop and utilize imaging techniques capable of assessing metastatic cell progression and quantifying the effect of treatment on the majority of metastatic cell population, inclusive of solitary cells. Quantification of the solitary metastatic cells in whole mouse liver was achieved using an MRI technique in which iron labeled cells were visible as areas of signal void. Signal void area was found to be highly correlated with the number of MPIO labeled cell injected into the liver. The MR scanning protocol used here resulted in images in which both signal void (due to iron labeled cells) / and hyperintensity due to metastases (\u3e200 |j,m) were apparent and could be quantified. This technique was subsequently utilized to assess the effect of doxorubicin and the synthetic triterpenoid CDDO-lm on experimental liver metastases in mice. It was found that despite significantly inhibiting the growth of large metastases, neither treatment decreased the number of solitary cells present in the same liver. In order to better understand the effect of treatment on the metastatic cell population at a cellular level, cells that express a cell cycle reporter system that changes colour as cell cycle progresses (fudci) were used in 2D and 3D cell culture. It was found that initial cell density and CDDO-lm treatment altered cell growth, and that cell cycle progression could be monitored longitudinally in individual and groups of cells within the same enclosed culture system. It is expected that the techniques presented here will enable the screening of therapeutic strategies for their efficacy against not just a subset of the metastatic cell population, but the population as a whol

Silica bioreplication preserves three-dimensional spheroid structures of human pluripotent stem cells and HepG2 cells

Three-dimensional (3D) cell cultures produce more in vivo-like multicellular structures such as spheroids that cannot be obtained in two-dimensional (2D) cell cultures. Thus, they are increasingly employed as models for cancer and drug research, as well as tissue engineering. It has proven challenging to stabilize spheroid architectures for detailed morphological examination. Here we overcome this issue using a silica bioreplication (SBR) process employed on spheroids formed from human pluripotent stem cells (hPSCs) and hepatocellular carcinoma HepG2 cells cultured in the nanofibrillar cellulose (NFC) hydrogel. The cells in the spheroids are more round and tightly interacting with each other than those in 2D cultures, and they develop microvilli-like structures on the cell membranes as seen in 2D cultures. Furthermore, SBR preserves extracellular matrix-like materials and cellular proteins. These findings provide the first evidence of intact hPSC spheroid architectures and similar fine structures to 2D-cultured cells, providing a pathway to enable our understanding of morphogenesis in 3D cultures.Peer reviewe

Tumour dormancy in breast cancer: an update

Delayed recurrences, common in breast cancer, are well explained by the concept of tumour dormancy. Numerous publications describe clinical times to disease recurrence or death, using mathematical approaches to infer mechanisms responsible for delayed recurrences. However, most of the clinical literature discussing tumour dormancy uses data from over a half century ago and much has since changed. This review explores how current breast cancer treatment could change our understanding of the biology of breast cancer tumour dormancy, and summarizes relevant experimental models to date. Current knowledge gaps are highlighted and potential areas of future research are identified

Sentinel lymph node mapping using ICG fluorescence and cone beam CT – a feasibility study in a rabbit model of oral cancer

Abstract Background Current sentinel lymph node biopsy (SLNB) techniques, including use of radioisotopes, have disadvantages including the use of a radioactive tracer. Indocyanine green (ICG) based near-infrared (NIR) fluorescence imaging and cone beam CT (CBCT) have advantages for intraoperative use. However, limited literature exists regarding their use in head and neck cancer SLNB. Methods This was a prospective, non-randomized study using a rabbit oral cavity VX2 squamous cell carcinoma model (n = 10) which develops lymph node metastasis. Pre-operatively, images were acquired by MicroCT. During surgery, CBCT and NIR fluorescence imaging of ICG was used to map and guide the SLNB resection. Results Intraoperative use of ICG to guide fluorescence resection resulted in identification of all lymph nodes identified by pre-operative CT. CBCT was useful for near real time intraoperative imaging and 3D reconstruction. Conclusions This pre-clinical study further demonstrates the technical feasibility, limitations and advantages of intraoperative NIR-guided ICG imaging for SLN identification as a complementary method during head and neck surgery

Cell-free DNA and circulating tumor cell kinetics in a pre-clinical head and neck Cancer model undergoing radiation therapy

Abstract Background Monitoring circulating tumor DNA (ctDNA) and circulating tumor cells (CTCs), known as liquid biopsies, continue to be developed as diagnostic and prognostic markers for a wide variety of cancer indications, mainly due to their minimally invasive nature and ability to offer a wide range of phenotypic and genetic information. While liquid biopsies maintain significant promising benefits, there is still limited information regarding the kinetics of ctDNA and CTCs following radiation therapy which remains a vital treatment modality in head and neck cancers. This study aims to describe the kinetics of ctDNA and CTCs following radiation exposure in a preclinical rabbit model with VX2 induced buccal carcinoma. Methods Seven rabbits were inoculated with VX2 cells in the buccal mucosa and subjected to radiation. At selected time points, blood sampling was performed to monitor differing levels of ctDNA and CTC. Plasma ctDNA was measured with quantitative PCR for papillomavirus E6 while CTCs were quantified using an immunomagnetic nanoparticles within a microfluidic device. Comparisons of CTC detection with EpCAM compared to multiple surface markers (EGFR, HER2 and PSMA) was evaluated and correlated with the tumor size. Results Plasma ctDNA reflects the overall tumor burden within the animal model. Analysis of correlations between ctDNA with tumor and lymph node volumes showed a positive correlation (R = 0.452 and R = 0.433 [p < 0.05]), respectively. Over the course of treatment, ctDNA levels declined and quickly becomes undetectable following tumor eradication. While during the course of treatment, ctDNA levels were noted to rise particularly upon initiation of radiation following scheduled treatment breaks. Levels of CTCs were observed to increase 1 week following inoculation of tumor to the primary site. For CTC detection, the use of multiple surface markers showed a greater sensitivity when compared to detection using only EpCAM. Plasma CTC levels remained elevated following radiation therapy which may account for an increased shedding of CTCs following radiation. Conclusion This study demonstrates the utility of ctDNA and CTCs detection in response to radiation treatment in a preclinical head and neck model, allowing for better understanding of liquid biopsy applications in both clinical practice and research development

Re-examining the Size/Charge Paradigm: Differing in Vivo Characteristics of Size- and Charge-Matched Mesoporous Silica Nanoparticles

The combination of nanoparticle (NP) size, charge, and surface chemistry (e.g., extent of modification with polyethylene glycol (PEG)) is accepted as a key determinant of NP/cellular interactions. However, the influence of spatial arrangement and accessibility of the charged molecules on the NP surface <i>vis-à-vis</i> the average surface charge (zeta (ζ) potential) is incompletely understood. Here we demonstrate that two types of mesoporous silica nanoparticles (MSNP) that are matched in terms of primary and hydrodynamic particle size, shape, pore structure, colloidal stability, and ζ potential, but differ in surface chemistry, <i>viz</i>. the spatial arrangement and relative exposure of surface amines, have profoundly different interactions with cells and tissues when evaluated <i>in vitro</i> and <i>in vivo</i>. While both particles are ∼50 nm in diameter, PEGylated, and positively charged (ζ = +40 mV), PEG-PEI (MSNPs modified with exposed polyamines), but not PEG-NMe₃⁺ (MSNP modified with distributed, obstructed amines) rapidly bind serum proteins, diverse cells types <i>in vitro</i>, and endothelial and white blood cells <i>in vivo</i> (ex ovo chick embryo model). This finding helps elucidate the relative role of surface exposure of charged molecules vs ζ potential in otherwise physicochemically matched MSNP and highlights protein corona neutrality as an important design consideration when synthesizing cationic NPs for biological applications

Re-examining the Size/Charge Paradigm: Differing in Vivo Characteristics of Size- and Charge-Matched Mesoporous Silica Nanoparticles

The combination of nanoparticle (NP) size, charge, and surface chemistry (e.g., extent of modification with polyethylene glycol (PEG)) is accepted as a key determinant of NP/cellular interactions. However, the influence of spatial arrangement and accessibility of the charged molecules on the NP surface <i>vis-à-vis</i> the average surface charge (zeta (ζ) potential) is incompletely understood. Here we demonstrate that two types of mesoporous silica nanoparticles (MSNP) that are matched in terms of primary and hydrodynamic particle size, shape, pore structure, colloidal stability, and ζ potential, but differ in surface chemistry, <i>viz</i>. the spatial arrangement and relative exposure of surface amines, have profoundly different interactions with cells and tissues when evaluated <i>in vitro</i> and <i>in vivo</i>. While both particles are ∼50 nm in diameter, PEGylated, and positively charged (ζ = +40 mV), PEG-PEI (MSNPs modified with exposed polyamines), but not PEG-NMe₃⁺ (MSNP modified with distributed, obstructed amines) rapidly bind serum proteins, diverse cells types <i>in vitro</i>, and endothelial and white blood cells <i>in vivo</i> (ex ovo chick embryo model). This finding helps elucidate the relative role of surface exposure of charged molecules vs ζ potential in otherwise physicochemically matched MSNP and highlights protein corona neutrality as an important design consideration when synthesizing cationic NPs for biological applications

Re-examining the Size/Charge Paradigm: Differing in Vivo Characteristics of Size- and Charge-Matched Mesoporous Silica Nanoparticles

The combination of nanoparticle (NP) size, charge, and surface chemistry (e.g., extent of modification with polyethylene glycol (PEG)) is accepted as a key determinant of NP/cellular interactions. However, the influence of spatial arrangement and accessibility of the charged molecules on the NP surface <i>vis-à-vis</i> the average surface charge (zeta (ζ) potential) is incompletely understood. Here we demonstrate that two types of mesoporous silica nanoparticles (MSNP) that are matched in terms of primary and hydrodynamic particle size, shape, pore structure, colloidal stability, and ζ potential, but differ in surface chemistry, <i>viz</i>. the spatial arrangement and relative exposure of surface amines, have profoundly different interactions with cells and tissues when evaluated <i>in vitro</i> and <i>in vivo</i>. While both particles are ∼50 nm in diameter, PEGylated, and positively charged (ζ = +40 mV), PEG-PEI (MSNPs modified with exposed polyamines), but not PEG-NMe₃⁺ (MSNP modified with distributed, obstructed amines) rapidly bind serum proteins, diverse cells types <i>in vitro</i>, and endothelial and white blood cells <i>in vivo</i> (ex ovo chick embryo model). This finding helps elucidate the relative role of surface exposure of charged molecules vs ζ potential in otherwise physicochemically matched MSNP and highlights protein corona neutrality as an important design consideration when synthesizing cationic NPs for biological applications

Hybrid Core-Shell Polymer Scaffold for Bone Tissue Regeneration

A great promise for tissue engineering is represented by scaffolds that host stem cells during proliferation and differentiation and simultaneously replace damaged tissue while maintaining the main vital functions. In this paper, a novel process was adopted to develop composite scaffolds with a core-shell structure for bone tissue regeneration, in which the core has the main function of temporary mechanical support, and the shell enhances biocompatibility and provides bioactive properties. An interconnected porous core was safely obtained, avoiding solvents or other chemical issues, by blending poly(lactic acid), poly(&epsilon;-caprolactone) and leachable superabsorbent polymer particles. After particle leaching in water, the core was grafted with a gelatin/chitosan hydrogel shell to create a cell-friendly bioactive environment within its pores. The physicochemical, morphological, and mechanical characterization of the hybrid structure and of its component materials was carried out by means of infrared spectroscopy, thermogravimetric analysis, scanning electron microscopy, and mechanical testing under different loading conditions. These hybrid polymer devices were found to closely mimic both the morphology and the stiffness of bones. In addition, in vitro studies showed that the core-shell scaffolds are efficiently seeded by human mesenchymal stromal cells, which remain viable, proliferate, and are capable of differentiating towards the osteogenic phenotype if adequately stimulated