Week 5: Regan

This week I spent my time purely doing research. Until now I have had little research to do because I have had to wait for my primary human umbilical vein endothelial cells (pHUVECs) to proliferate so that I had enough cells for experiments. In my previous experiments, I have had some inconsistent cell phenotypes and they seemed to be related to varying cell densities within the organoid. Because of that, this week I tried to discern a better cell density for encapsulating these cells into our PEG-MAL integrin presenting, degradable hydrogels. The current standards for culturing pHUVECs in 3D are cultures in matrigel or collagen. The problem with these systems are that they are biomolecularly complex and so it is impossible to discern which biomolecule(s) causes cell phenotypic changes as a result of culture. To dissect the role of matrix connections with specific integrins, we strategically present peptides corresponding to integrin ligands to fool the cell into thinking it has made contact with an large matrix polymer in our engineered organoids. In complex, three dimensional cultures, pHUVECs self-organize into vessel-like tubes that branch together to resemble vasculature. I am hoping to achieve this reaction in my gels, and right now I have a hunch that cell density within the gel will play a crucial role. As you can see in the images that I have included, I discovered that cell density indeed does affect cellular orientation in three dimensional cultures as seen by the branching and spreading of cells in the denser (first) image when compared to the less dense culture (second image). I am pleased by these results, and will continue to monitor the development of vasculogenic structures in these cultures over the course of the next few days. 

In other research news, through a collaboration with Shahin Rafi's lab here at WCMC, I have now received remarkably fresh pHUVECs straight from the source - umbilical cords from recent births in Obstetrics and Gynecology. These cells are supposed to be increasingly more proliferative than the commercial cells I have been using, and should last longer in culture. I plan to put them in organoids soon to assess if there are any noticeable differences between these cells and the commercial cells I have been using in the context of 3D culture.


On a separate note, I had the chance to attend a case study meeting given by the Precision Medicine Tumor Board through the Caryl and Israel Englander Institute for Precision Medicine. This was an open meeting discussing case studies of elderly women with Chronic Lymphocytic Leukemia (CLL). These cases were interesting because special next-generation sequencing techniques were used to further characterize the complexity of their diseases, and those sequencing results in one case informed prognostically how the patient would respond to a treatment. From this talk, I learned the CLL is in most cases treated with ibrutinib, a Bruton's Tyrosine Kinase (BTK) inhibitor that irreversibly inhibits its target in the B cell receptor signaling pathway. In some cases, resistance to this drug can develop that alters the binding of the inhibitor from irreversible to reversible, therefore, reducing the effectiveness of the drug. For this reason, a second generation BTK inhibitor is sometimes used. One of the patients that was discussed was put on one, Acalabrutinib, through a clinical trial here at WCM. One thing I found surprising about this talk was the vast numbers of genes that were recognized in these patients as abnormal through sequencing, that researchers and doctors don't know enough about to decide if they were relevant. This showed me how even with the incredible advances in deep sequencing technologies, there will always be a need for molecular biology and genetics research in the realm of oncology.