👤 Catherine A Reardon

FGFR alterations are reported across various malignancies and might act as oncogenic drivers in multiple histologies. Erdafitinib is an oral, selective pan-FGFR tyrosine kinase inhibitor with activity in FGFR-altered advanced urothelial carcinoma. We aimed to evaluate the safety and activity of erdafitinib in previously treated patients with FGFR-altered advanced solid tumours. The single-arm, phase 2 RAGNAR study was conducted at 156 investigative centres (hospitals or oncology practices that are qualified oncology study centres) across 15 countries. The study consisted of four cohorts based on tumour histology and patient age; the results reported in this Article are for the primary cohort of the study, defined as the Broad Panel Cohort, which was histology-agnostic. We recruited patients aged 12 years or older with advanced or metastatic tumours of any histology (except urothelial cancer) with predefined FGFR1-4 alterations (mutations or fusions according to local or central testing). Eligible patients had disease progression on at least one previous line of systemic therapy and no alternative standard therapy available to them, and an Eastern Cooperative Oncology Group performance status of 0-1 (or equivalent for adolescents aged 12-17 years). Patients received once-daily oral erdafitinib (8 mg/day with provision for pharmacodynamically guided up-titration to 9 mg/day) on a continuous 21-day cycle until disease progression or intolerable toxicity. The primary endpoint was objective response rate by independent review committee according to Response Evaluation Criteria In Solid Tumors (RECIST), version 1.1, or Response Assessment In Neuro-Oncology (RANO). The primary analysis was conducted on the treated population of the Broad Panel Cohort. This ongoing study is registered with ClinicalTrials.gov, number NCT04083976. Patients were recruited between Dec 5, 2019, and Feb 15, 2022. Of 217 patients treated with erdafitinib, 97 (45%) patients were female and 120 (55%) were male. The data cutoff was Aug 15, 2022. At a median follow-up of 17·9 months (IQR 13·6-23·9), an objective response was observed in 64 (30% [95% CI 24-36]) of 217 patients across 16 distinct tumour types. The most common grade 3 or higher treatment-emergent adverse events related to erdafitinib were stomatitis (25 [12%]), palmar-plantar erythrodysaesthesia syndrome (12 [6%]), and hyperphosphataemia (11 [5%]). The most commonly occurring serious treatment-related adverse events (grade 3 or higher) were stomatitis in four (2%) patients and diarrhoea in two (1%). There were no treatment-related deaths. RAGNAR results show clinical benefit for erdafitinib in the tumour-agnostic setting in patients with advanced solid tumours with susceptible FGFR alterations who have exhausted other treatment options. These results support the continued development of FGFR inhibitors in patients with advanced solid tumours. Janssen Research & Development. Show less

Using genetic and biochemical approaches, we investigated proteins that regulate macrophage cholesterol efflux capacity (CEC) and ABCA1-specific CEC (ABCA1 CEC), 2 functional assays that predict cardiovascular disease (CVD). Macrophage CEC and the concentration of HDL particles were markedly reduced in mice deficient in apolipoprotein A-I (APOA1) or apolipoprotein E (APOE) but not apolipoprotein A-IV (APOA4). ABCA1 CEC was markedly reduced in APOA1-deficient mice but was barely affected in mice deficient in APOE or APOA4. High-resolution size-exclusion chromatography of plasma produced 2 major peaks of ABCA1 CEC activity. The early-eluting peak, which coeluted with HDL, was markedly reduced in APOA1- or APOE-deficient mice. The late-eluting peak was modestly reduced in APOA1-deficient mice but little affected in APOE- or APOA4-deficient mice. Ion-exchange chromatography and shotgun proteomics suggested that plasminogen (PLG) accounted for a substantial fraction of the ABCA1 CEC activity in the peak not associated with HDL. Human PLG promoted cholesterol efflux by the ABCA1 pathway, and PLG-dependent efflux was inhibited by lipoprotein(a) [Lp(a)]. Our observations identify APOA1, APOE, and PLG as key determinants of CEC. Because PLG and Lp(a) associate with human CVD risk, interplay among the proteins might affect atherosclerosis by regulating cholesterol efflux from macrophages. Show less

Given their association with cardiovascular disease protection, there has been intense interest in understanding the biology of high density lipoproteins (HDL). HDL is actually a family of diverse particle types, each made up of discrete - but as yet undetermined - combinations of proteins drawn from up to 95 lipophilic plasma proteins. The abundant apolipoproteins (apo) of the A class (apoA-I, apoA-II and apoA-IV) have been proposed to act as organizing platforms for auxiliary proteins, but this concept has not been systematically evaluated. We assessed the impact of genetic knock down of each platform protein on the particle size distribution of auxiliary HDL proteins. Loss of apoA-I or apoA-II massively reduced HDL lipids and changed the plasma size pattern and/or abundance of several plasma proteins. Surprisingly though, many HDL proteins were not affected, suggesting they assemble on lipid particles in the absence of apoA-I or apoA-II. In contrast, apoA-IV ablation had minor effects on plasma lipids and proteins, suggesting that it forms particles that largely exclude other apolipoproteins. Overall, the data indicate that distinct HDL subpopulations exist that do not contain, nor depend on, apoA-I, apoA-II or apoA-IV and these contribute substantially to the proteomic diversity of HDL. Plasma levels of high density lipoproteins (HDL) are inversely correlated with cardiovascular disease. These particles are becoming known as highly heterogeneous entities that have diverse compositions and functions that may impact disease. Unfortunately, we know little about the forces that maintain the composition of each particle in plasma. It has been suggested that certain 'scaffold' proteins, such as apolipoprotein (apo) A-I, apoA-II and apoA-IV, may act as organizing centers for the docking of myriad accessory proteins. To test this hypothesis, we took advantage of the genetic tractability of the mouse model and ablated these three proteins individually. We then tracked the abundance and size profile of the remaining HDL proteins by gel filtration chromatography combined with mass spectrometry. The results clearly show that certain cohorts of proteins depend on each scaffold molecule to assemble normal sized HDL particles under wild-type conditions. This work forms the basis for more detailed studies that will define the specific compositions of HDL subspecies with the possibility of connecting them to specific functions or roles in disease. Show less

Cholesterol efflux capacity associates strongly and negatively with the incidence and prevalence of human CVD. We investigated the relationships of HDL's size and protein cargo with its cholesterol efflux capacity using APOB-depleted serum and HDLs isolated from five inbred mouse strains with different susceptibilities to atherosclerosis. Like humans, mouse HDL carried >70 proteins linked to lipid metabolism, the acute-phase response, proteinase inhibition, and the immune system. HDL's content of specific proteins strongly correlated with its size and cholesterol efflux capacity, suggesting that its protein cargo regulates its function. Cholesterol efflux capacity with macrophages strongly and positively correlated with retinol binding protein 4 (RBP4) and PLTP, but not APOA1. In contrast, ABCA1-specific cholesterol efflux correlated strongly with HDL's content of APOA1, APOC3, and APOD, but not RBP4 and PLTP. Unexpectedly, APOE had a strong negative correlation with ABCA1-specific cholesterol efflux capacity. Moreover, the ABCA1-specific cholesterol efflux capacity of HDL isolated from APOE-deficient mice was significantly greater than that of HDL from wild-type mice. Our observations demonstrate that the HDL-associated APOE regulates HDL's ABCA1-specific cholesterol efflux capacity. These findings may be clinically relevant because HDL's APOE content associates with CVD risk and ABCA1 deficiency promotes unregulated cholesterol accumulation in human macrophages. Show less

The fatty acid desaturase (FADS) gene family at 11q12-13.1 includes FADS1 and FADS2, both known to mediate biosynthesis of omega-3 and omega-6 long-chain polyunsaturated fatty acids (LCPUFA). FADS3 is a putative desaturase due to its sequence similarity with FADS1 and FADS2, but its function is unknown. We have previously described 7 FADS3 alternative transcripts (AT) and 1 FADS2 AT conserved across multiple species. This study examined the effect of dietary LCPUFA levels on liver FADS gene expression in vivo and in vitro, evaluated by qRT-PCR. Fourteen baboon neonates were randomized to three diet groups for their first 12 weeks of life, C: Control, no LCPUFA, L: 0.33% docosahexaenoic acid (DHA)/0.67% arachidonic acid (ARA) (w/w); and L3: 1.00% DHA/0.67% ARA (w/w). Liver FADS1 and both FADS2 transcripts were downregulated by at least 50% in the L3 group compared to controls. In contrast, FADS3 AT were upregulated (L3 > C), with four transcripts significantly upregulated by 40% or more. However, there was no evidence for a shift in liver fatty acids to coincide with increased FADS3 expression. Significant upregulation of FADS3 AT was also observed in human liver-derived HepG2 cells after DHA or ARA treatment. The PPARγ antagonist GW9662 prevented FADS3 upregulation, while downregulation of FADS1 and FADS2 was unaffected. Thus, FADS3 AT were directly upregulated by LCPUFA by a PPARγ-dependent mechanism unrelated to regulation of other desaturases. This opposing pattern and mechanism of regulation suggests a dissimilar function for FADS3 AT compared to other FADS gene products. Show less

The fatty acid desaturase genes (FADS1 and FADS2) code for enzymes required for synthesis of omega-3 and omega-6 long-chain polyunsaturated fatty acids (LCPUFA) important in the central nervous system, inflammatory response, and cardiovascular health. SNPs in these genes are associated with numerous health outcomes, but it is unclear how genetic variation affects enzyme function. Here, lymphoblasts obtained from Japanese participants in the International HapMap Project were evaluated for association of expression microarray results with SNPs in the FADS gene cluster. Six SNPs in the first intron of the FADS2 gene were associated with FADS1 expression. A 10-SNP haplotype in FADS2 (rs2727270 to rs2851682) present in 24% of the population was associated with lower expression of FADS1. A highly conserved region coinciding with the most significant SNPs contained predicted binding sites for SREBP and PPARγ. Lymphoblasts homozygous for either the major or minor haplotype were treated with agonists for these transcription factors and expression of FADS1 and FADS2 determined. Simvastatin and the LXR agonist GW3965 both upregulated expression of FADS1 and FADS2; no response was found for PPARγ agonist rosiglitazone. The minor haplotype homozygotes had 20-40% higher induction of FADS1 and FADS2 after simvastatin or GW3965 treatment. A 22 bp polymorphic insertion-deletion (INDEL) was found 137 bp downstream from the putative sterol response element, as well as a 3 or 1 bp INDEL 81-83 bp downstream. All carriers of the minor haplotype had deletions while all carriers of the major haplotype had insertions. Individuals carrying the minor haplotype may be vulnerable to alterations in diet that reduce LCPUFA intake, and especially responsive to statin or marine oil therapy. Show less

The fatty acid desaturase (FADS) genes code for the rate-limiting enzymes required for the biosynthesis of long-chain polyunsaturated fatty acids (LCPUFA). Here we report discovery and function of a novel FADS1 splice variant. FADS1 alternative transcript 1 (FADS1AT1) enhances desaturation of FADS2, leading to increased production of eicosanoid precursors, the first case of an isoform modulating the enzymatic activity encoded by another gene. Multiple protein isoforms were detected in primate liver, thymus, and brain. In human neuronal cells, their expression patterns are modulated by differentiation and result in alteration of cellular fatty acids. FADS1, but not FADS1AT1, localizes to endoplasmic reticulum and mitochondria. Ribosomal footprinting demonstrates that all three FADS genes are translated at similar levels. The noncatalytic regulation of FADS2 desaturation by FADS1AT1 is a novel, plausible mechanism by which several phylogenetically conserved FADS isoforms may regulate LCPUFA biosynthesis in a manner specific to tissue, organelle, and developmental stage. Show less

The Δ6 desaturase, encoded by FADS2, plays a crucial role in omega-3 and omega-6 fatty acid synthesis. These fatty acids are essential components of the central nervous system, and they act as precursors for eicosanoid signaling molecules and as direct modulators of gene expression. The polypyrimidine tract binding protein (PTB or hnRNP I) is a splicing factor that regulates alternative pre-mRNA splicing. Here, PTB is shown to bind an exonic splicing silencer element and repress alternative splicing of FADS2 into FADS2 AT1. PTB and FADS2AT1 were inversely correlated in neonatal baboon tissues, implicating PTB as a major regulator of tissue-specific FADS2 splicing. In HepG2 cells, PTB knockdown modulated alternative splicing of FADS2, as well as FADS3, a putative desaturase of unknown function. Omega-3 fatty acids decreased by nearly one half relative to omega-6 fatty acids in PTB knockdown cells compared with controls, with a particularly strong decrease in eicosapentaenoic acid (EPA) concentration and its ratio to arachidonic acid (ARA). This is a rare demonstration of a mechanism specifically altering the cellular omega-3 to omega-6 fatty acid ratio without any change in diet/media. These findings reveal a novel role for PTB, regulating availability of membrane components and eicosanoid precursors for cell signaling. Show less

Fatty acid desaturase 1 and 2 (FADS1 and FADS2) code for the key desaturase enzymes involved in the biosynthesis of long chain polyunsaturated fatty acids in mammals. FADS3 shares close sequence homology to FADS1 and FADS2 but the function of its gene product remains unknown. Alternative transcripts (AT) generated by alternative splicing (AS) are increasingly recognized as an important mechanism enabling a single gene to code for multiple gene products. We report the first AT of a FADS gene, FADS3, generated by AS. Aided by ORF Finder, we identified putative coding regions of eight AT for FADS3 with 1.34 kb (classical splicing), 1.14 (AT1), 0.77 (AT2), 1.25 (AT3), 0.51 (AT4), 0.74 (AT6), and 1.11 (AT7). In addition we identified a 0.51 kb length transcript (AT5) that has a termination codon within intron 8-9. The expression of each AT was analyzed in baboon neonate tissues and in differentiated and undifferentiated human SK-N-SH neuroblastoma cells. FADS3 AT are expressed in 12 neonate baboon tissues and showed reciprocal increases and decreases in expression changes in response to human neuronal cell differentiation. FADS3 AT, conserved in primates and under metabolic control in human cells, are a putative mediator of LCPUFA biosynthesis and/or regulation. Show less