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Objective Familial hypercholesterolemia (FH), a prevalent genomic disorder that increases risk of coronary heart disease, remains significantly underdiagnosed. Clinical decision support (CDS) tools have the potential to increase FH detection. We describe our experience in the development and implementation of a genomic CDS for FH at a large academic medical center.Methods CDS development and implementation were conducted in four phases: (1) development and validation of an algorithm to identify “possible FH”; (2) obtaining approvals from institutional committees to develop the CDS; (3) development of the initial prototype; and (4) use of an implementation science framework to evaluate the CDS.Results The timeline for this work was approximately 4 years; algorithm development and validation occurred from August 2018 to February 2020. During this 4-year period, we engaged with 15 stakeholder groups to build and integrate the CDS, including health care providers who gave feedback at each stage of development. During CDS implementation six main challenges were identified: (1) need for multiple institutional committee approvals; (2) need to align the CDS with institutional knowledge resources; (3) need to adapt the CDS to differing workflows; (4) lack of institutional guidelines for CDS implementation; (5) transition to a new institutional electronic health record (EHR) system; and (6) limitations of the EHR related to genomic medicine.Conclusion We identified multiple challenges in different domains while developing CDS for FH and integrating it with the EHR. The lessons learned herein may be helpful in streamlining the development and deployment of CDS to facilitate genomic medicine implementation.

Restoration of kidney tubular epithelium following sublethal injury sequentially involves partial epithelial–mesenchymal transition (pEMT), proliferation, and further redifferentiation into specialized tubule epithelial cells (TECs). Because the immunosuppressant cyclosporine-A produces pEMT in TECs and inhibits the peptidyl-prolyl isomerase (PPIase) activity of cyclophilin (Cyp) proteins, we hypothesized that cyclophilins could regulate TEC phenotype. Here we demonstrate that in cultured TECs, CypA silencing triggers loss of epithelial features and enhances transforming growth factor β (TGFβ)-induced EMT in association with upregulation of epithelial repressors Slug and Snail. This pro-epithelial action of CypA relies on its PPIase activity. By contrast, CypB emerges as an epithelial repressor, because CypB silencing promotes epithelial differentiation, prevents TGFβ-induced EMT, and induces tubular structures in 3D cultures. In addition, in the kidneys of CypB knockout mice subjected to unilateral ureteral obstruction, inflammatory and pro-fibrotic events were attenuated. CypB silencing/knockout leads to Slug, but not Snail, downregulation. CypB support of Slug expression depends on its endoplasmic reticulum location, where it interacts with calreticulin, a calcium-buffering chaperone related to Slug expression. As CypB silencing reduces ionomycin-induced calcium release and Slug upregulation, we suggest that Slug expression may rely on CypB modulation of calreticulin-dependent calcium signaling. In conclusion, this work uncovers new roles for CypA and CypB in modulating TEC plasticity and identifies CypB as a druggable target potentially relevant in promoting kidney repair., Graphical abstract

Cyclophilins (Cyp) are peptidil-prolyl-isomerases and the intracellular receptors for the immunosuppressant Cyclosporine-A (CsA), which produces epithelial-mesenchymal-transition (EMT) and renal tubule-interstitial fibrosis. Since CsA inhibits Cyp enzymatic activity, we hypothesized that Cyp could be involved in EMT and fibrosis. Here, we demonstrate that CypB is a critical regulator of tubule epithelial cell plasticity on the basis that: i) CypB silencing caused epithelial differentiation in proximal tubule-derived HK-2 cells, ii) CypB silencing prevented TGFβ-induced EMT in HK-2, and iii) CypB knockdown mice exhibited reduced UUO-induced inflammation and kidney fibrosis. By contrast, silencing of CypA induces a more undifferentiated phenotype and favors TGFβ effects. EMT mediators Slug and Snail were up-regulated in CypA-silenced cells, while in CypB silencing, Slug, but not Snail, was down-regulated; thus, reinforcing the role of Slug in kidney fibrosis. CypA regulates Slug through its PPIase activity whereas CypB depends on its ER location, where interacts with calreticulin, a calcium modulator which is involved in TGFβ signaling. In conclusion, this work uncovers new roles for CypA and CypB in modulating proximal tubular cell plasticity.

Treatment of cancer is frequently unsuccessful related to the loss of apoptotic signaling in malignant cells. This is a particular problem for high-grade gliomas, such as Glioblastoma Multiforme (GBM), which are almost universally fatal within a year or so of diagnosis. Novel therapies that capitalize on non-apoptotic cell death pathways may yield more effective outcomes, if their underlying mechanisms can be more completely deciphered. In a recent publication (ref 10), the mechanisms by which cellular cyclophilins support GBM cell survival have been identified. Inhibition of cyclophilins activated paraptosis, which relied on a combination of endoplasmic reticulum (ER) stress and transient activation of autophagy. An important aspect of this effect was the relative rates of cap-dependent versus cap-independent protein synthesis, which were differentially modulated by protein synthesis inhibitors or mTOR inhibition. Although cycloheximide has previously been characterized as an inhibitor of paraptosis, in the case of cyclophilin inhibition, it appears to significantly enhance stress-related paraptosis and cell death. This work reveals an important role for cap-independent protein translation and autophagy in the ability of GBM cells to resist non-apoptotic death, and adds to our understanding of the events that underlie paraptosis.

Cancer is the second leading cause of death worldwide. Current treatment strategies based on multi-agent chemotherapy and/or radiation regimens have improved overall survival in some cases. However, resistance to apoptosis often develops in cancer cells, and its occurrence is thought to contribute to treatment failure. Non-apoptotic cell death mechanisms have become of great interest, therefore, in hopes that they would bypass tumor cell resistance. Glioblastoma multiforme (GBM), a grade IV astrocytic tumor is the most frequent brain tumor in adults, and has a high rate of mortality. We report that NIM811, a small molecule cyclophilin-binding inhibitor, induces catastrophic vacuolization and cell death in GBM cells. These unique features are distinct from many known cell death pathways, and are associated with an incompletely defined cell death mechanism known as paraptosis. We found that NIM811-induced paraptosis is due to unresolved ER stress. The abnormal upregulation of protein translation was responsible for the build-up of misfolded or unfolded proteins in ER, whereas pro-survival autophagy and UPR signals were shutdown during prolonged treatment with NIM811. Although cycloheximide has been claimed to suppress paraptosis, instead we find that it only temporarily delayed vacuole formation, but actually enhanced paraptotic cell death in the long term. On the other hand, mTOR inhibitors rescued cells from NIM811-induced paraptosis by sustaining autophagy and the UPR, while specifically restraining cap-dependent translation. These findings not only provide new insights into the mechanisms underlying paraptosis, but also shed light on a potential approach to enhance GBM treatment.