Improved Post-Transplant Survival in the United States for Patients with Cholangiocarcinoma After 2000

Background

The incidence of cholangiocarcinoma (CCA) continues to rise. Orthotopic liver transplantation (OLT) can be used for selected patients with localized but unresectable hilar CCA. Although initial post-OLT survival rates were poor, outcomes after introduction of the Mayo Clinic protocol have been more promising and there has been increased interest in OLT for CCA nationally.

Aims

The aim of this study is to determine post-transplant survival and prognostic factors for patients undergoing OLT for CCA.

Methods

A retrospective analysis of all patients with CCA listed nationwide for OLT between October 1987 and May 2008 was performed using the Scientific Registry of Transplant Recipients database. Survival curves were generated using the Kaplan–Meier method and compared using log-rank test.

Results

Of 595 patients with CCA listed for OLT, 359 (60.3 %) underwent OLT. Median age at OLT was 49 years, 66 % were male and 91 % were Caucasian. The median follow-up time was 2 years. There has been an increasing number of liver transplants performed for CCA since 2000. The 1- and 5-year probability of survival was 85.8 and 51.4 %, respectively. On multivariate analysis, significant prognostic factors for decreased post-OLT survival included transplant before 2000 (HR 11.25, 95 % CI 1.28–98.7) and acute cellular rejection (HR 5.64, 95 % CI 1.14–27.8).

Conclusions

Survival after transplant for CCA has improved over time, and OLT is being used more frequently in the treatment of CCA. Significant predictors of post-OLT survival include a history of acute rejection and date of transplant in relation to the publication of Mayo protocol results.     

Engineered kinase activation reveals unique morphodynamic phenotypes and associated trafficking for Src family isoforms

The Src kinase family comprises nine homologous members whose distinct expression patterns and cellular distributions indicate that they have unique roles. These roles have not been determined because genetic manipulation has not produced clearly distinct phenotypes, and the kinases’ homology complicates generation of specific inhibitors. Through insertion of a modified FK506 binding protein (insertable FKBP12, iFKBP) into the protein kinase isoforms Fyn, Src, Lyn, and Yes, we engineered kinase analogs that can be activated within minutes in living cells (RapR analogs). Combining our RapR analogs with computational tools for quantifying and characterizing cellular dynamics, we demonstrate that Src family isoforms produce very different phenotypes, encompassing cell spreading, polarized motility, and production of long, thin cell extensions. Activation of Src and Fyn led to patterns of kinase translocation that correlated with morphological changes in temporally distinct stages. Phenotypes were dependent on N-terminal acylation, not on Src homology 3 (SH3) and Src homology 2 (SH2) domains, and correlated with movement between a perinuclear compartment, adhesions, and the plasma membrane.Since its discovery, c-Src (1) has been subject to intensive research into its cellular functions and regulation. Whereas c-Src is the best-studied protooncogene, less is known about the other, closely related Src family kinase (SFK) members. Their high degree of similarity in structure and regulation suggests that SFKs can partially compensate for each other in vivo. Indeed, knockout studies have shown that only mice deficient in all three genes (src, yes, and fyn) show embryonic lethality (2). Early studies demonstrated that disruption of Src or Fyn genes individually resulted only in subtle changes in function of a few cell types (e.g., osteoclasts for src^−/−, and T cells for fyn^−/−) (3, 4). Roche et al. provided strong evidence that Src, Yes, and Fyn substitute for each other during cell cycle progression (5). These studies suggested that there is a high degree of functional redundancy among Src family kinases.Nonetheless, emerging evidence indicates that Src and Fyn regulate distinct processes in the same cell. Down-regulation of Fyn expression enhances VEGF-stimulated migration of endothelial cells, whereas down-regulation of Src does not (6). Differences in the transforming capacity of SFKs are thought to depend on their affinity for cholesterol-enriched membrane microdomains, which is determined in part by their N-terminal lipid modifications (7, 8). Src has higher tumorigenic potential than Fyn in prostate epithelium, and this is differently affected by alterations in N-terminal palmitoylation (9). Previous studies have shown that Src localizes to perinuclear endosomal compartments and translocates to the plasma membrane upon activation (10–12), whereas Fyn localizes to the plasma membrane regardless of its activity (13, 14). Although these studies suggest that localization is important in differentiating the actions of the two kinases, they do not identify specific roles associated with particular subcellular locations.Various techniques have been applied to elucidate the differences in signaling specificity of SFKs. Kinase–substrate interactions have been examined using purified substrates (15). Mutated kinases with selectivity for radiolabeled ATP analogs have identified directly phosphorylated substrates of Src (16). These methods were restricted to cell lysates or purified proteins, and so were unable to address the role of cellular localization in substrate specificity.To dissect the unique role of different SFK isoforms (2–4, 17, 18) in living cells, we engineered regulatable analogs of Fyn, Yes, and LynA kinases using our rapamycin-regulated activation (RapR) strategy, which has been developed using Src as a prototype (19, 20). Insertable FKBP12 (iFKBP, a truncated form of FKBP) was inserted into the catalytic domain of each SFK, which abolished their kinase activity. Activity was rescued by treating cells with rapamycin in the presence of the FKBP12-rapamycin binding domain (FRB) (Fig. 1A). Molecular dynamics studies have indicated that heterodimerization of the inserted iFKBP with FRB likely reduces the conformational mobility of the kinase G loop, restoring ATP binding (3, 21).Open in a separate windowFig. 1.Design of RapR kinases. (A) Schematic representation of the approach used to regulate catalytic activity of SFKs. The insertion of iFKBP at a highly conserved site in the catalytic domain of each kinase resulted in loss of kinase activity. Catalytic activity was restored by rapamycin, which induced binding of iFKBP and coexpressed FRB. (B) Sequence alignment of SFKs shows that there is a well-defined loop where iFKBP is inserted (blue). It is linked to the G loop (red) through a β-sheet in each SFK.These analogs enabled activation of each isoform specifically, within minutes, resulting in clear phenotypic differences. Unlike genetic modifications of cell populations, there was little time for the cell to compensate for kinase activation before observation. The induced cell behaviors occurred in a succession of stages, associated with changes in the subcellular distribution of each kinase. We focused on Src and Fyn, developing quantitative tools to carefully characterize the kinetics of induced behaviors and associated localization dynamics. Our results indicated that Src’s unique ability to induce polarized movement shortly after kinase activation results from its localization in a perinuclear compartment, where it phosphorylates substrates that traffic on microtubules to the cell perimeter. Both the localization dynamics and phenotype differences between Src and Fyn were dependent on N-terminal lipid modifications, and not on SH2 and SH3 domain interactions.     

A multi-institutional comparison of mitoxantrone,etoposide, and cytarabine vs high-dose cytarabine and mitoxantrone therapy for patients with relapsed or refractory acute myeloid leukemia

Relapsed or refractory acute myeloid leukemia (R/R AML) has a poor prognosis and is best treated with salvage chemotherapy as a bridge to allogeneic stem cell transplant (alloSCT). However, the optimal salvage therapy remains unknown. Here we compared two salvage regimens; mitoxantrone, etoposide, and cytarabine (MEC) and mitoxantrone and high-dose Ara-C (Ara-C couplets). We analyzed 155 patients treated at three academic institutions between 1998 and 2017; 87 patients received MEC and 68 received Ara-C couplets. The primary endpoint was overall response (OR). Secondary endpoints included progression-free survival (PFS), overall survival (OS), duration of hospitalization, hematologic and nonhematologic toxicities, and success in proceeding to alloSCT. Baseline characteristics of the cohorts were well matched, though patients receiving Ara-C couplets had more co-morbidities (48.5% vs 33%; P = .07). OR was achieved in 43.7% of MEC and 54.4% of Ara-C couplets patients (P = .10). Ara-C couplets patients also trended towards a longer OS and PFS, more frequently proceeded to alloSCT (31% vs 54.4%; P = .003), and experienced less febrile neutropenia (94% vs 72%; P < .001) and grade 3/4 gastrointestinal toxicities (17.2% vs 2.94%; P = .005). No significant differences in other toxicities or median duration of hospitalization were noted. This is the first multi-institutional study directly comparing these regimens in a racially diverse population of R/R AML patients. Although these regimens have equivalent efficacy in terms of achieving OR, Ara-C couplets use is associated with significant reductions in toxicities, suggesting it should be used more frequently in these patients.     

Implications of the New Centers for Disease Control and Prevention Blood Lead Reference Value

The Centers for Disease Control and Prevention recently established a new reference value (≥ 5 μg/dL) as the standard for identifying children with elevated blood lead levels (EBLs). At present, 535 000 US children aged 1 to 5 years (2.6%) are estimated to have EBLs according to the new standard, versus 0.8% according to the previous standard (≥ 10 μg/dL). Because EBLs signify the threshold for public health intervention, this new definition increases demands on lead poisoning prevention efforts. Primary prevention has been proven to reduce lead poisoning cases and is also cost effective; however, federal budget cuts threaten the existence of such programs. Protection for the highest-risk children necessitates a reinstatement of federal funding to previous levels.In May 2012, officials of the Centers for Disease Control and Prevention (CDC) announced that they had accepted the recommendations set forth by the Advisory Committee on Childhood Lead Poisoning Prevention (ACCLPP) for (1) the discontinuation of the term blood lead “level of concern,” to acknowledge that there is no safe level of lead exposure, and (2) the use of a new reference value for the identification of children with elevated blood lead levels (EBLs).1 The level of concern, previously defined as 10 micrograms of lead per deciliter of blood, was established by the CDC as the EBL that should initiate a public health response and had been previously unchanged since 1991.2 By contrast, the new reference value is 5 micrograms per deciliter.1 Figure 1 depicts the decline in CDC-recommended blood lead level (BLL) action levels, a drop of more than 90% over the past several decades.3,4 Regardless of the action level of the time, childhood EBLs have long been targeted for complete elimination.5–9Open in a separate windowFIGURE 1—Trends in Centers for Disease Control and Prevention’s childhood blood lead level of concern: United States, 1960–2014.     

The benefits of redesigning Benin's vaccine supply chain

Introduction

New vaccine introductions have put strains on vaccine supply chains around the world. While increasing storage and transportation may be the most straightforward options, it is also important to consider what financial and operational benefits can be incurred. In 2012, suboptimal vaccine coverage and impending vaccine introductions prompted the Republic of Benin's Ministry of Health (MOH) to explore ways to improve their vaccine supply chain.

Methods

Working alongside the Beninese MOH, we utilized our computational model, HERMES, to explore the impact on cost and vaccine availability of three possible options: (1) consolidating the Commune level to a Health Zone level, (2) removing the Commune level completely, and (3) removing the Commune level and expanding to 12 Department Stores. We also analyzed the impact of adding shipping loops during delivery.

Results

At baseline, new vaccine introductions without any changes to the current system increased the logistics cost per dose ($0.23 to $0.26) and dropped the vaccine availability to 71%. While implementing the Commune level removal scenario had the same capital costs as implementing the Health Zone scenario, the Health Zone scenario had lower operating costs. This increased to an overall cost savings of $504,255 when implementing shipping loops.

Discussion

The best redesign option proved to be the synergistic approach of converting to the Health Zone design and using shipping loops (serving ten Health Posts/loop). While a transition to either redesign or only adding shipping loops was beneficial, implementing a redesign option and shipping loops can yield both lower capital expenditures and operating costs.     

Whole-Genome Analysis of Exserohilum rostratum from an Outbreak of Fungal Meningitis and Other Infections

Exserohilum rostratum was the cause of most cases of fungal meningitis and other infections associated with the injection of contaminated methylprednisolone acetate produced by the New England Compounding Center (NECC). Until this outbreak, very few human cases of Exserohilum infection had been reported, and very little was known about this dematiaceous fungus, which usually infects plants. Here, we report using whole-genome sequencing (WGS) for the detection of single nucleotide polymorphisms (SNPs) and phylogenetic analysis to investigate the molecular origin of the outbreak using 22 isolates of E. rostratum retrieved from 19 case patients with meningitis or epidural/spinal abscesses, 6 isolates from contaminated NECC vials, and 7 isolates unrelated to the outbreak. Our analysis indicates that all 28 isolates associated with the outbreak had nearly identical genomes of 33.8 Mb. A total of 8 SNPs were detected among the outbreak genomes, with no more than 2 SNPs separating any 2 of the 28 genomes. The outbreak genomes were separated from the next most closely related control strain by ∼136,000 SNPs. We also observed significant genomic variability among strains unrelated to the outbreak, which may suggest the possibility of cryptic speciation in E. rostratum.     

Detection of Chronic Wasting Disease in the Lymph Nodes of Free-Ranging Cervids by Real-Time Quaking-Induced Conversion

Chronic wasting disease (CWD), a transmissible spongiform encephalopathy of deer, elk, and moose, is the only prion disease affecting free-ranging animals. Since the disease was first identified in northern Colorado and southern Wyoming in 1967, new epidemic foci of the disease have been identified in 20 additional states, as well as two Canadian provinces and the Republic of South Korea. Identification of CWD-affected animals currently requires postmortem analysis of brain or lymphoid tissues using immunohistochemistry (IHC) or an enzyme-linked immunosorbent assay (ELISA), with no practical way to evaluate potential strain types or to investigate the epidemiology of existing or novel foci of disease. Using a standardized real-time (RT)-quaking-induced conversion (QuIC) assay, a seeded amplification assay employing recombinant prion protein as a conversion substrate and thioflavin T (ThT) as an amyloid-binding fluorophore, we analyzed, in a blinded manner, 1,243 retropharyngeal lymph node samples from white-tailed deer, mule deer, and moose, collected in the field from areas with current or historic CWD endemicity. RT-QuIC results were then compared with those obtained by conventional IHC and ELISA, and amplification metrics using ThT and thioflavin S were examined in relation to the clinical history of the sampled deer. The results indicate that RT-QuIC is useful for both identifying CWD-infected animals and facilitating epidemiological studies in areas in which CWD is endemic or not endemic.