Modeling Clinical Responses to Targeted Therapies by Patient-Derived Organoids of Advanced Lung Adenocarcinoma.

PURPOSE: Patient-derived organoids (PDO) of lung cancer has been recently introduced, reflecting the genomic landscape of lung cancer. However, clinical relevance of advanced lung adenocarcinoma organoids remains unknown. Here, we examined the ability of PDOs to predict clinical responses to targeted therapies in individual patients and to identify effective anticancer therapies for novel molecular targets. EXPERIMENTAL DESIGN: Eighty-four organoids were established from patients with advanced lung adenocarcinoma. Formalin-fixed, paraffin-embedded tumor specimens from corresponding patients were analyzed by whole-exome sequencing (n = 12). Organoids were analyzed by whole-exome sequencing (n = 61) and RNA sequencing (n = 55). Responses to mono or combination targeted therapies were examined in organoids and organoid-derived xenografts. RESULTS: PDOs largely retained somatic alterations including driver mutations of matching patient tumors. PDOs were able to recapitulate progression-free survival and objective responses of patients with non-small cell lung cancer receiving clinically approved tyrosine kinase inhibitors. PDOs recapitulated activity of therapeutic strategies under clinical investigation. YUO-071 harboring an EGFR exon 19 deletion and a BRAF G464A mutation and the matching patient responded to dabrafenib/trametinib combination therapy. YUO-004 and YUO-050 harboring an EGFR L747P mutation was sensitive to afatinib, consistent with the response in the matching patient of YUO-050. Furthermore, we utilized organoids to identify effective therapies for novel molecular targets by demonstrating the efficacy of poziotinib against ERBB2 exon 20 insertions and pralsetinib against RET fusions. CONCLUSIONS: We demonstrated translational relevance of PDOs in advanced lung adenocarcinoma. PDOs are an important diagnostic tool, which can assist clinical decision making and accelerate development of therapeutic strategies.

PCL/ß-TCP Composite Scaffolds Exhibit Positive Osteogenic Differentiation with Mechanical Stimulation.

We investigated the use of Polycaprolactone (PCL)/ ß-tricalcium phosphate (ß-TCP) composites with applied mechanical stimulation as scaffold for bone tissue engineering. PCL-based three-dimensional (3D) structures were fabricated in a solvent-free process using a 3D-printing technique. The mass fraction of ß-TCP was varied in the range 0-30%, and the structure and compressive modulus of the specimens was characterized. The shape and interconnectivity of the pores was found to be satisfactory, and the compressive modulus of the specimens was comparable with that of human trabecular bone. Human mesenchymal stem cells were seeded on the composites, and various biological evaluations were performed over 9 days. With a mass fraction of ß-TCP of 30%, differentiation began earlier; however, the cell proliferation rate was lower. Through the use of mechanical stimulation, however, the proliferation rate recovered, and was comparable with that of the other groups. This stimulation effect was also observed in ECM generation and other biological assays. With mechanical stimulation, expression of osteogenic markers was lower on samples with a ß-TCP content of 10 wt% than without ß-TCP; however, with mechanical stimulation, the sample with a ß-TCP content of 30 wt% exhibited significantly greater expression of those markers than the other samples. We found that mechanical stimulation and the addition of ß-TCP interacted closely, and that a mass fraction of ß-TCP of 30% was particularly useful as a bone tissue scaffold when accompanied by mechanical stimulation.

Simultaneous engagement of mechanical stretching and surface pattern promotes cardiomyogenic differentiation of human mesenchymal stem cells.

It has been widely recognized and proved that biophysical factors for mimicking in vivo conditions should be also considered to have stem cells differentiated into desired cell type in vitro along with biochemical factors. Biophysical factors include substrate and biomechanical conditions. This study focused on the effect of biomimetic mechanical stretching along with changes in substrate topography to influence on cardiomyogenic differentiation of human mesenchymal stem cells (hMSCs). Elastic micropatterned substrates were made to mimic the geometric conditions surrounding cells in vivo. To mimic biomechanical conditions due to beating of the heart, mechanical stretching was applied parallel to the direction of the pattern (10% elongation, 0.5 Hz, 4 h/day). Suberoylanilide hydroxamic acid (SAHA) was used as a biochemical factor. The micropatterned substrate was found more effective in the alignment of cytoskeleton and cardiomyogenic differentiation compared with flat substrate. Significantly higher expression levels of related markers [GATA binding protein 4 (GATA4), troponin I, troponin T, natriuretic peptide A (NPPA)] were observed when mechanical stretching was engaged on micropatterned substrate. In addition, 4 days of mechanical stretching was associated with higher levels of expression than 2 days of stretching. These results indicate that simultaneous engagement of biomimetic environment such as substrate pattern and mechanical stimuli effectively promotes the cardiomyogenic differentiation of hMSCs in vitro. The suggested method which tried to mimic in vivo microenvironment would provide systematic investigation to control cardiomyogenic differentiation of hMSCs.

A three-dimensional hierarchical scaffold fabricated by a combined rapid prototyping technique and electrospinning process to expand hematopoietic stem/progenitor cells.

OBJECTIVE: To investigate the expansion of hematopoietic stem/progenitor cells (HSPCs) from umbilical cord blood using extracellular matrix (ECM) protein-coated three-dimensional hierarchical scaffolds. RESULTS: The expansion of HSPCs was evaluated through total nucleated cell (TNC) expansion, immuno-phenotypic analysis, and clonogenic ability. After 7 days of culture, three-dimensional cultures with fibronectin-coated scaffolds achieved the highest fold increase in TNCs (164 ± 6.9 fold) and the highest CD45(+)CD34(+) (35 %) and CD34(+)CD38(-) (32 %) ratios. CONCLUSION: Three-dimensional hierarchical scaffolds were coated with ECM protein to simulate a biomimetic environment or niche, and had a significant effect on the expansion potential of HSPCs without changing their phenotype.