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The insulin-like growth factor-1 receptor (IGF-1R) plays a pivotal role in transformation, growth, and survival of malignant cells, and has emerged as a general and promising target for cancer treatment. However, no fully selective IGF-1R inhibitors have thus far been found. This is explained by the fact that IGF-1R is highly homologous to the insulin receptor, coinhibition of which may cause diabetic response. The receptors are both tyrosine kinases, and their ATP binding sites are identical, implying that ATP inhibitors cannot discriminate between them. Therefore, the current strategy has been to identify compounds interfering with receptor autophosphorylation at the substrate level. In this study we investigated the effects of cyclolignans and related molecules on IGF-1R activity. We report that certain cyclolignans are potent and selective inhibitors of tyrosine phosphorylation of the IGF-1R. Of particular interest was picropodophyllin (PPP), which is almost nontoxic (LD50 >500 mg/kg in rodents). PPP efficiently blocked IGF-1R activity, reduced pAkt and phosphorylated extracellular signal regulated kinase 1 and 2 (pErk1/2), induced apoptosis in cultured IGF-1R-positive tumor cells, and caused complete tumor regression in xenografted and allografted mice. PPP did not affect the insulin receptor or compete with ATP in an in vitro kinase assay, suggesting that it may inhibit IGF-1R autophosphorylation at the substrate level. This is also in agreement with our molecular model of how the cyclolignans may act on the IGF-1R kinase. Our results open the possibility to use PPP or related compounds with inhibitory effects on IGF-1R as lead compounds in development of anticancer agents.

Protein-coding genes comprise only 3% of the human genome, while the genes that are transcribed into RNAs but do not code for proteins occupy majority of the genome. Once considered as biological darker matter, non-coding RNAs are now being recognized as critical regulators in cancer genome. Among the many types of non-coding RNAs, microRNAs approximately 20 nucleotides in length are best characterized and their mechanisms of action are well generalized. microRNA exerts oncogenic or tumor suppressor function by regulation of protein-coding genes via sequence complementarity. The expression of microRNA is aberrantly regulated in all cancer types, and both academia and biotech companies have been keenly pursuing the potential of microRNA as cancer biomarker for early detection, prognosis, and therapeutic response. The key involvement of microRNAs in cancer also prompted interest on exploration of therapeutic values of microRNAs as anticancer drugs and drug targets. MRX34, a liposome-formulated miRNA-34 mimic, developed by Mirna Therapeutics, becomes the first microRNA therapeutic entering clinical trial for the treatment of hepatocellular carcinoma, renal cell carcinoma, and melanoma. In this review, we presented a general overview of microRNAs in cancer biology, the potential of microRNAs as cancer biomarkers and therapeutic targets, and associated challenges.

Insulin receptor substrates (IRSs) play central roles in insulin/insulin-like growth factor (IGF) signaling and mediate a variety of their bioactivities. IRSs are tyrosine-phosphorylated by activated insulin receptor/IGF-I receptor tyrosine kinase in response to insulin/IGF, and are recognized by signaling molecules possessing the SH2 domain such as phosphatidylinositol 3-kinase (PI3K), leading to the activation of downstream pathways. Recent studies have suggested that degradation of IRSs by the ubiquitin-proteasome pathway leads to impaired insulin/IGF signaling, but the precise mechanism underlying the process is still unclear. In this study, we identified deubiquitinating enzyme ubiquitin specific protease 7 (USP7) as an IRS-2-interacting protein and demonstrated that deubiquitinase activity of USP7 plays important roles in IRS-2 stabilization through the ubiquitin-proteasome pathway. In addition, insulin treatment dissociated USP7 from IRS-2, leading to degradation of IRS-2. This dissociation was prevented by treatment with LY294002, a PI3K inhibitor, indicating that insulin activation of the PI3K pathway leads to dissociation of IRS-2 from USP7 and IRS-2 degradation. We obtained similar results for IRS-1 in cells treated with insulin and for IRS-2 in cells treated with IGF-I. Taken together, this is the first report demonstrating that USP7 is an IRS-1/2 deubiquitinating enzyme forming a negative feedback loop in insulin/IGF signaling.

AMR is being recognized with increasing efficiency, but continues to present a significant threat to renal allograft survival. Traditional therapies for AMR (IVIG, plasmapheresis, rituximab, and antilymphocyte preparations) in general have provided inconsistent results and do not deplete the source of antibody production, viz., the mature plasma cell. Recently, the first plasma cell-targeted therapy in humans has been developed using bortezomib (a first in class PI) for AMR treatment in kidney transplant recipients. Initial experience with bortezomib involved treatment of refractory AMR. Subsequently, the efficacy of bortezomib in primary therapy for AMR was demonstrated. In a multicenter collaborative effort, the initial results with bortezomib in AMR have been confirmed and expanded to pediatric and adult heart transplant recipients. More recently, results from a prospective, staged desensitization trial has shown that bortezomib alone can substantially reduce anti-HLA antibody levels. These results demonstrate the significant potential of proteasome inhibition in addressing humoral barriers. However, the major advantage of proteasome inhibition lies in the numerous potential strategies for achieving synergy.

Purpose of review Bortezomib is a first-in-class proteasome inhibitor that was originally Food and Drug Administration approved for the treatment of multiple myeloma. In the past few years, off-label use in solid organ transplant recipients has demonstrated its ability to provide plasma cell-targeted therapy in humans. The purpose of this review is to provide an update of recent basic science and clinical results with bortezomib in treating antibody-mediated rejection (AMR) that occurs in solid organ transplant recipients. Recent findings Proteasome inhibitor therapy for AMR in kidney transplant recipients is effective both as primary and as rescue therapy. Optimal responses with proteasome inhibitor therapy are obtained when AMR is diagnosed promptly and early in the posttransplant period. However, proteasome inhibitor therapy for late AMR (i.e., occurring 6 months or later posttransplant) provides less predictable results, likely due to the existence of a substantial bone marrow niche-resident long-lived plasma cell population. Proteasome inhibitor therapy has also recently been shown to provide effective therapy for AMR in heart, and also, transplant recipients. Summary Proteasome inhibitor therapy with bortezomib provides effective treatment for AMR in solid organ transplant recipients. As the first plasma cell-targeted therapy, proteasome inhibitor therapy provides the additional advantage of opening new possibilities for biologically defined plasma cell-targeted therapies.

Background Rejection with hemodynamic compromise (RHC) is associated with high mortality in heart recipients. This study investigates the association between genetic polymorphisms and RHC in pediatric heart recipients. Methods Data from 532 pediatric heart recipients from six centers in the Pediatric Heart Transplant Study were analyzed for time to RHC by recipient race, age at transplantation, and genotype at 13 genetic polymorphisms (TNF-α A-308G, IL-6 G-174C, INF-γ T+874A, IL-10 G-1082A, C-819T, and C-592A; FAS A-670G, FASL C-843T, and ACE I/D; and VEGF A-2578C, C-1451T, C+405G, and -2549 I/D). Results RHC occurred in 126 (23.7%) patients during the study period. Adjusting for age and race, IL-10 G-1082A, FAS A-670G, and ACE I/D genotypes were associated with RHC. IL-10 G-1082A GG genotype was associated with decreased risk of RHC with an adjusted hazard ratio (HR) of 0.49 (95% confidence interval [CI], 0.27-0.90; P=0.020). FAS A-670G AA genotype was associated with increased risk of RHC with an adjusted HR of 1.84 (95% CI, 1.25-2.69; P=0.002). ACE II genotype was associated with decreased risk of RHC with an adjusted HR of 0.58 (95% CI, 0.36-0.95; P=0.031). Conclusions Recipients with a high anti-inflammatory and immune-regulatory genetic profile (high interleukin-10) were protected from RHC. Conversely, recipients with a pro-apoptotic genetic profile (high Fas) or high angiotensin-1-converting enzyme producing genotype were at increased risk of RHC. This represents progress toward understanding the genetic risk factors of posttransplantation outcomes in pediatric heart recipients.

Background. The efficacy of plasma cell targeted therapies for antibody-mediated rejection (AMR) has not been defined in detail. The purpose of this study was to compare early and late acute AMR in terms of immunologic characteristics and responses with proteasome inhibitor (PI) therapy. Methods. Renal transplant recipients with acute AMR were treated with PI-based regimens. Early acute AMR was defined as occurring within 6 months posttransplant. Immunodominant donor-specific antibody (iDSA) was defined as the DSA with the highest level. Results. Results are expressed as early or late acute AMR. Thirty AMR episodes (13 early, 17 late) were treated in 12 and 16 patients. Early but not late AMR was associated with presensitization. Late AMR iDSA levels were higher, and specificities were primarily class II (DQ being most frequent). Early AMR patients demonstrated greater reduction in iDSA at 7, 14, and 30 days and at the posttreatment nadir (81.5%+21.2% vs. 51.4%+27.6%; P

Aim Heart transplant (HTX) candidates with pre-formed anti-HLA antibody (HLA-Ab) have lower transplantation rates and increased waiting time due to positive crossmatches. Anti-angiotensin II type 1 receptor antibody (AT1R-Ab) has recently been associated with allograft loss and antibody-mediated rejection. The aim of the present study was to evaluate: 1) the prevalence of anti-angiotensin receptor 1 antibody (AT1R-Ab) in HTX candidates and 2) the impact of AT1R-Ab on flow cytometry crossmatch (FXM). Methods 115 HTX candidates were tested by ELISA for the presence of AT1R-Ab (U/mL after 1:50 dilution, positive cut-off = 10 U/mL, strong-positive cut-off = 20 U/mL. 43/115 cases were identified with either positive (N = 15) or negative (N = 28) flow crossmatches in the absence of any donor-specific anti-HLA alloantibody (virtual negative crossmatches for HLA antibody). Mean channel shift (MCS) for flow cytometry crossmatches was measured on a 1024 scale. Finally, two index HTX cases with pre-formed AT1R-Ab and positive flow crossmatches are discussed. Results A high proportion of HTX candidates with HLA-Ab exhibited AT1R-Ab (Strongly reactive AT1R-Ab 46/115, 40%; weakly reactive AT1R-Ab 25/115, 21.7%; no AT1R-Ab 44/115, 38.2%). Furthermore, AT1R-Ab levels were significantly higher in patients with a positive versus a negative flow crossmatch (54.81 ± 54.92 versus 9.04 ± 7.36, p = 0.00001, Fig. 1). In addition, MCS was significantly higher in patients with AT1R-Ab versus patients without antibodies (73.95 ± 94.21 versus −51.45 ± 52.07, p = 000003, Fig. 2). In the 15 cases with positive flow crossmatch without HLA-specific antibody, all 15 patients exhibited AT1R-Ab, 11/15 had strong AT1R-Ab, while only 5/25 patients with negative FXM exhibited AT1R-Ab, Chi-square = 24, p = 0.00001). Conclusions More than half of HTX candidates exhibit AT1R-Ab, which can interfere with flow crossmatch interpretation even in the absence of HLA-specific antibodies.

Anti-HLA DSAs are associated with ABMR and graft loss in KT recipients, yet the influence of DSA IgG subclass on outcomes in pediatric KT recipients is not completely understood. We performed a single-center retrospective chart review of pediatric KT recipients with anti-HLA DSAs, aiming to study the association between specific DSA IgG subclasses and graft outcomes, including ABMR and significant graft dysfunction (graft loss or 50% decrease in eGFR). Thirty-six patients (mean age 15.4y) with DSAs initially detected 1 month-14.3 years post-transplantation were followed for a median of 2.8 years. Rates of IgG1, 2, 3, and 4 subclass detection were 92%, 33%, 58%, and 25%, respectively. Twenty-two patients (61%) had clinical ABMR, whereas 19% had subclinical ABMR, and 13 (36%) experienced significant graft dysfunction. Patients with IgG3+ DSAs had a higher risk of graft dysfunction compared with IgG3- patients (52% vs 13%, P = .03). In a multiple Cox proportional regression analysis, the presence of IgG3+ DSA was independently associated with significant graft dysfunction (HR 10.45, 95% CI 1.97-55.55, P = .006). In conclusion, IgG3 subclass DSAs are associated with graft dysfunction and may be useful for risk stratification and treatment decisions in DSA-positive pediatric KT recipients.

Purpose: We aimed to report a case of orbital solitary fibrous tumour (SFT) in a child and to review the relevant literature. Methods: We describe an SFT in a 13-year-old boy with a 1-month history of painless proptosis in the left eye. Results: Magnetic resonance imaging revealed a well circumscribed mass filling most of the left intraconal orbit. The lesion was excised and histopathological examination revealed a malignant SFT. Postoperative follow-up for 18 months was uneventful. Conclusions: Malignant SFT of the orbit should be included in the differential diagnosis of paediatric orbital tumours. Complete surgical excision remains the preferred method of management and the longterm prognosis is guarded.

Background. Neutrophilic tubulitis accompanied by intratubular neutrophil clusters in the renal allograft is a surrogate marker for urinary tract infection (UTI). Overlapping histologic findings can occur in antibody-mediated rejection, which is characterized by peritubular capillary (PTC) deposition of C4d. This study evaluated the incidence of UTI in biopsies with concurrent neutrophilic tubulitis and PTC C4d staining. Methods. Thirty-three allograft biopsies from 27 patients selected for the presence of simultaneous C4d staining and neutrophilic tubulitis were correlated with urine culture (U/C) results. Results. U/C obtained on the same day as the biopsy confirmed UTI in 13 of 33 (39%) biopsies. Among 20 patients with negative U/C; prior culture results within 10 days of the biopsy were available for nine patients, and 5 of 9 (55%) were positive. Thus, UTI was confirmed in 18 of 33 (54%) biopsies. Biopsy interpretation and clinical management was confounded by changes of concurrent acute cellular rejection and antibody-mediated rejection confirmed by demonstration of donor-specific antibodies. Combined therapy with antibiotics and antirejection medications (ART) was administered to 12 of 18 (67%) patients. Conclusions. Neutrophilic tubulitis accompanied by neutrophil clusters in the tubular lumen is a useful marker of UTI, even in the presence of PTC C4d deposition. Therapeutic response to antibiotics is limited by co-existent T-cell or antibody-mediated rejection and underlying chronic allograft nephropathy.

Background Intestinal retransplantation (Re-ITx) has historically been associated with high morbidity and mortality. Methods The outcomes of all children receiving Re-ITx between 1990 and 2007 at our center were reviewed. Results One hundred seventy-two children received primary intestinal grafts. Fourteen children (8.1%) were retransplanted with 15 grafts. Causes of graft failure were acute cellular rejection (ACR, n=4), liver failure (n=2), chronic rejection (n=3), posttransplant lymphoproliferative disorder (n=1), graft dysmotility or dysfunction (n=3), ACR with severe infection (n=1), and arterial graft aneurysm (n=1). Initial transplants were isolated bowel in nine, liver-bowel in five, and one multivisceral. The mean time of initial graft survival was 34.2 months. Re-ITx was with isolated bowel in two, liver-bowel in four, and multivisceral in nine (four with kidney). Initial immunosuppression was Tac-Pred based in nine and rabbit antithymocyte globulin-Tac based in six cases. Re-ITx was carried out under Tac-Pred in six, rabbit antithymocyte globulin-Tac in eight, and alemtuzumab monoclonal anti-CD52 antibody in one. Ten (71.4%) patients are alive with functioning grafts at a mean current follow-up time of 55.9 months. Four patients died from posttransplant lymphoproliferative disorder, severe ACR, fungal sepsis, and bleeding from pseudoaneurysm, respectively, at a mean time of 5.7 months post-Re-ITx. All surviving patients weaned-off total parenteral nutrition at a median time of 32 days and 90% are off intravenous fluids. Conclusions Improved long-term survival and outcome in pediatric Re-ITx may be attributed to improvements in initial immunosuppression protocols, technical modifications, proper timing, and improved infectious disease monitoring. Careful patient selection and posttransplant management are essential for successful long-term outcome.

Introduction Choroidal neovascularization (CNV) is a debilitating complication of age-related macular degeneration (AMD) and a leading cause of vision loss. Along with other angiogenic factors such as vascular endothelial growth factor (VEGF), insulin-like growth factor (IGF)-1 and its receptor, IGF-1R, have been implicated in CNV. Purpose A prior study has shown that the cyclolignan picropodophyllin (PPP) efficiently blocks the insulin-like growth factor-1 receptor (IGF-1R) activity and causes cell death in uveal melanoma cell lines and in an in vivo model. In this study we investigated the effect of PPP on VEGF expression, both in vitro and in vivo, and whether this effect has antiangiogenic consequences in a murine CNV model. Methods C57BL/6J mice with laser-induced CNVs were treated with PPP. Effects on CNV area were assayed by image analysis. VEGF levels in the choroid and retinal pigment epithelial cells (ARPE-19) were measured by Western blot or ELISA. Transcriptional activation of the VEGF promoter was determined by luciferase reporter gene assay. Results Mice treated with PPP, administered intraperitoneally or orally, showed a 22% to 32% (P = 0.002) decrease in CNV area. Furthermore, VEGF levels in the choroid were significantly reduced. In cultured ARPE-19 cells, IGF-1 was shown to increase VEGF secretion. This increase was completely blocked by PPP. PPP reduced the level of transcriptional activity of the VEGF promoter. Conclusions PPP reduces IGF-1-dependent VEGF expression and CNV in vivo. Accordingly, IGF-1R inhibitors may be useful tools in the treatment of conditions associated with CNV, including neovascular AMD.

The hallmark of humoral rejection is the presence of subendothelial C4d in the allograft. A simultaneous determination of vascular C4d with soluble C4d in broncho-alveolar lavage fluid (BAL) and circulating anti-human leukocyte antigen (HLA) antibodies (HLA-Ab) has not been reported in lung transplantation.Forty-two consecutive lung-transplant patients were included in this cross-sectional study. The presence and specificity of HLA-Ab was determined at the same frequency with transbronchial biopsies. Soluble C4d levels were measured by enzyme-linked immunosorbent assay in all 42 patients. In a subgroup of 32 patients with available timely matched paraffin-embedded tissue sections, the vascular C4d deposition was also assessed.The presence of HLA-Ab in 16 patients was associated with biopsy-proven acute rejection (10/16 vs. 3/16, P0.01) and increased immunosuppression (13/16 vs. 4/16, P0.005). Pulmonary function was also decreased in patients with HLA-Ab (mean forced expiratory volume in 1 second=49%) when compared with the control group (mean forced expiratory volume in 1 second=66%, P0.05). Nine patients exhibited specific vascular C4d deposition and in eight of nine (89%) cases HLA-Ab were detected, versus 8 of 23 (35%) in C4d-negative patients (P0.05). Soluble C4d in BAL was highly (0.5 microg/mL) elevated in patients with HLA-Ab and vascular C4d and was moderately (0.2 microg/mL) increased in patients with antibodies but C4d-negative. In contrast, only a slight elevation of soluble C4d (0.1 microg/mL) was detected in patients without HLA-specific antibodies.The association of HLA-specific antibodies with vascular C4d deposition and soluble C4d in BAL, in addition to the reduced pulmonary function, might constitute a diagnostic triad for antibody-mediated rejection in lung transplant patients.

The insulin-like growth factor receptor (IGF-IR) plays several pivotal roles in cancer. Although most studies on the function of the IGF-IR have been attributed to kinase-dependent signaling, recent findings by our group and others have implicated biological roles mediated by ubiquitination of the receptor. As previously reported, the E3 ligases Mdm2 and Nedd4 mediate IGF-IR ubiquitination. Here we show that c-Cbl is a novel E3 ligase for IGF-IR. On ligand stimulation, both Mdm2 and c-Cbl associate with IGF-IR and mediate receptor polyubiquitination. Whereas Mdm2 catalyzed lysine 63 (K63) chain ubiquitination, c-Cbl modified IGF-IR through K48 chains. Mdm2-mediated ubiquitination occurred when cells were stimulated with a low concentration (5 ng/mL) of IGF-I, whereas c-Cbl required high concentrations (50–100 ng/mL). Mdm2-ubiquitinated IGF-IR was internalized through the clathrin endocytic pathway whereas c-Cbl–ubiquitinated receptors were endocytosed via the caveolin route. Taken together, our results show that c-Cbl constitutes a new ligase responsible for the ubiquitination of IGF-IR and that it complements the action of Mdm2 on ubiquitin lysine residue specificity, responsiveness to IGF-I, and type of endocytic pathway used. The actions and interactions of Mdm2 and c-Cbl in the ubiquitination and endocytosis of IGF-IR may have implications in cancer. In addition, identification and functional characterization of new E3 ligases are important in itself because therapeutic targeting of substrate-specific E3 ligases is likely to represent a critical strategy in future cancer treatment. [Cancer Res 2008;68(14):5669–77]

Unsaturated malonyl esters underwent Pd-catalyzed intramolecular allylic alkylation to give 4-vinyl-substituted gamma-lactones. In contrast to the formerly studied cyclization of malonamides, this reaction could be achieved only with a substrate incorporating a suitably positioned silicon moiety, which directs the ionization toward the desired eta(3)-allylpalladium complex. The resulting 4-[dimethyl-(2-thienyl)silylvinyl]lactone could be subsequently engaged into Hiyama couplings with various iodoarenes, to give the corresponding 4-(alpha-styryl)-gamma-lactones. The use of a specifically substituted iodoarene generated an advanced tetracyclic lactone intermediate incorporating rings A-D of lignans belonging to the podophyllotoxin family. Subsequent electrophilic aromatic substitution with a variety of electron-rich arenes afforded the target picropodophyllin analogs.

The Cylex ImmuKnow Test (Cylex, Columbia, MD) measures immune cell function (ICF) and is based on the amount of adenosine triphosphate (ATP) released when T cells are stimulated by phytohemagglutinin. This preliminary study sought to determine if ICF measurements can be used to stratify kidney transplant recipients according to the risk for developing BK virus infection. ICF measurements were done in 15 samples from 8 patients with BK viremia, 38 samples from 25 patients with BK viruria, and 243 samples from 148 patients with no BK viruria or viremia. The mean+/-SD amounts of ATP released in these 3 groups were 102.9+/-58.6, 227.2+/-146.4, and 231.8+/-150.8 ng/mL, respectively (P= .002, viremia vs all other samples). Within the viruria group, lower ICF values were associated with higher urinary viral load (P= .037). These results show that a decreased ICF test result correlates with active viral replication in kidney transplant recipients.

Systemic sclerosis is a rare autoimmune disorder with a wide spectrum of clinical manifestations and a multitude of autoantibodies that are associated with it. In the past several years, advances in serologic testing have led to research indicating important prognostic and phenotypic associations with certain subsets of autoantibodies. In particular, anti-RNA polymerase III (anti-RNAP III) has been associated with diffuse cutaneous disease, scleroderma renal crisis, a temporal relationship with malignancy, myositis, synovitis, joint contractures, and gastric antral vascular ectasia. However, anti-RNAP III has not been associated with systemic sclerosis sine scleroderma. We describe a patient with an atypical presentation of anti-RNAP III positive systemic sclerosis sine scleroderma who presented without the typical features of anti-RNAP III disease. Instead, she presented with critical digital ischemia, pulmonary arterial hypertension, gastroesophageal reflux disease, interstitial lung disease, and no clinically detectable sclerodactyly.

Inosine 5'-monophosphate dehydrogenase 2 is required for purine synthesis in activated lymphocytes. Variants in the IMPDH2 gene may account for the large inter-individual variability in baseline enzyme activity, immunosuppressive efficacy and side effects in transplant recipients receiving mycophenolic acid. Therefore, the objective of this study was to identify and functionally characterize IMPDH2 variants.DNA samples from 152 solid organ transplant patients were screened at exons and exon/intron junctions of the IMPDH2 genes by PCR amplification followed by bidirectional direct DNA sequencing. Genetic variant was constructed by site-directed mutagenesis and transformed to an inosine 5'-monophosphate dehydrogenase-deficient strain of Escherichia coli h712. Proteins were purified to homogeneity and the enzymatic activity was measured by reduced nicotinamide adenine dinucleotide production.Nine genetic variants were identified in the IMPDH2 gene, with frequencies of the rarer alleles ranging from 0.5 to 10.2%. A novel nonsynonymous variant L263F was identified, and the kinetic assay demonstrated that the inosine 5'-monophosphate dehydrogenase activity of L263F variant was decreased to 10% of the wild-type. The Ki for mycophenolic acid inhibition of the L263F variant was comparable with the wild-type, and the variant Km for inosine 5'-monophosphate and nicotinamide adenine dinucleotide did not change significantly.IMPDH2 has low genetic diversity, but the nonsynonymous variant L263F has a significant impact on inosine 5'-monophosphate dehydrogenase activity. This novel functional variant may be one of the factors contributing to the inter-individual difference of baseline inosine 5'-monophosphate dehydrogenase activity as well as drug efficacy and adverse events in transplant patients.

The IGF-1R pathway is essential for the initiation and progression of many cancers. In contrast to other receptor tyrosine kinases involved in cancer, it is not frequently mutated or amplified. The classical model of signaling through the IGF-1R centers on ligand initiated kinase activation, allowing binding of adaptor molecules and downstream activation of the MAPK and PI3K pathways. The signaling is terminated through receptor ubiquitination and subsequent degradation. To date, therapies targeting IGF-1R have been designed solely aiming to block phosphorylation mediated signaling by preventing receptor-ligand interaction or by limiting kinase activation. Yet, the classical model is insufficient to explain receptor behavior induced by some IGF-1R inhibitors. This review advocates an updated model of IGF-1R signaling, accommodating the "classical" kinase signaling and the IGF-1R-kinase independent signaling thus providing the theoretical background for receptor downregulation induced by IGF-1R inhibitors. This model should be considered for future design of effective therapies targeting the IGF-1R pathway.

β-Arrestins are multifunctional proteins that play central roles in G protein-coupled receptor (GPCR) trafficking and signaling. β-Arrestin1 is also recruited to the insulin-like growth factor-1 receptor (IGF-1R), a receptor tyrosine kinase (RTK), mediating receptor degradation and signaling. Because GPCR phosphorylation by GPCR-kinases (GRKs) governs interactions of the receptors with β-arrestins, we investigated the regulatory roles of the four widely expressed GRKs on IGF-1R signaling/degradation. By suppressing GRK expression with siRNA, we demonstrated that lowering GRK5/6 abolishes IGF1-mediated ERK and AKT activation, whereas GRK2 inhibition increases ERK activation and partially inhibits AKT signaling. Conversely, β-arrestin–mediated ERK signaling is enhanced by overexpression of GRK6 and diminished by GRK2. Similarly, we demonstrated opposing effects of GRK2 and -6 on IGF-1R degradation: GRK2 decreases whereas GRK6 enhances ligand-induced degradation. GRK2 and GRK6 coimmunoprecipitate with IGF-1R and increase IGF-1R serine phosphorylation, promoting β-arrestin1 association. Using immunoprecipitation, confocal microscopy, and FRET analysis, we demonstrated β-arrestin/IGF-1R association to be transient for GRK2 and stable for GRK6. Using bioinformatic studies we identified serines 1248 and 1291 as the major serine phosphorylation sites of the IGF-1R, and subsequent mutation analysis demonstrated clear effects on IGF-1R signaling and degradation, mirroring alterations by GRKs. Targeted mutation of S1248 recapitulates GRK2 modulation, whereas S1291 mutation resembles GRK6 effects on IGF-1R signaling/degradation, consistent with GRK isoform-specific serine phosphorylation. This study demonstrates distinct roles for GRK isoforms in IGF-1R signaling through β-arrestin binding with divergent functional outcomes.

Aims Renal transplant (RTX) candidates with pre-formed anti-HLA antibody (HLA-Ab) have lower transplantation rates and increased mortality. Anti-angiotensin II type 1 receptor antibody (AT1R-Ab) has recently been associated with allograft loss and antibody-mediated rejection. The aim of the present study was to evaluate: (1) the prevalence of anti-angiotensin receptor 1 antibody (AT1R-Ab) in RTX candidates with circulating HLA-Ab and (2) AT1R-Ab response to VDS. Methods Thirty-nine patients receiving desensitization therapy had HLA-Ab levels determined by single-antigen bead array (Luminex). Mean fluorescence intensity (MFI) of the strongest HLA-Ab (immunodominant antibody -iDSA) was measured before and after therapy. A successful response was considered when iDSA decreased >50% at nadir versus pre-desensitization. Serum samples were also tested by ELISA for the presence of AT1R-Ab (U/mL after 1:50 dilution), before and after desensitization. Results A high proportion of candidates with HLA-Ab exhibited AT1R-Ab (20/39, 51%). A good response of AT1R-Ab to therapy was observed in 18/20 patients [Figure 1]: 17.4 ± 12.3 pre versus 10.9 ± 13.1 U/mL post therapy, p = 0.0009). iDSA reduction with desensitization was observed in 13/20 patients – 7996 ± 2580 pre versus 4410 ± 2695 MFI post therapy, p = 0.0001). 12/20 patients had both iDSA and AT1R-Ab reduction by therapy, 6/20 responded to AT1R-Ab only, 1/20 responded to iDSA only, and 1/20 was a non-responder. AT1R-Ab levels were more elevated in Caucasians then in African-Americans both before (20.7 ± 14.9 versus 12.3 ± 3.7 U/mL, p = 0.1), and after desensitization (14.7 ± 15.9 versus 5.2 ± 2.9 U/mL, p = 0.1). Conclusions More than half of HLA-sensitized candidates exhibit AT1R-Ab. This initial desensitization experience consistently provided substantial reductions both in HLA-Ab and AT1R-Ab levels in an unselected, highly sensitized kidney transplant candidate population. Download : Download high-res image (380KB) Download : Download full-size image

Ultrasonography has become a valuable tool for the assessment of salivary gland involvement in Sjogren syndrome. Diagnostic pitfalls can be encountered in late stages of the disease, in which morphologic changes may overlap with other pathologies of the salivary glands. We present the case of a female Sjogren syndrome patient with lack of significant sicca symptoms and unilateral occurrence of parotidomegaly, which prompted the suspicion of a parotid gland tumor. Due to the atypical clinical profile, diagnosis of Sjogren syndrome was delayed, at which point, severe sialadenitis produced a cystic transformation of the parotid gland parenchyma.

When debilitating, hyperhidrosis can be seen as a disease and not just as a symptom. It is most often a primary condition but can be secondary to other diseases. Aluminum chloride products are the initial treatment modality for palmar hyperhidrosis followed by anticholinergics, iontophoresis and botulinum toxin. The Dermatology Department of the Karolinska University Hospital in Stockholm, Sweden treated 151 patients at 289 visits with botulinum toxin for palmar hyperhidrosis during a two year period (2012-2013). It was found that botulinum toxin had good effect, which lasted between two and five months in 72% of cases. Muscle weakness (pincer grip) was reported at 41% of return visits and was present for less than one to four weeks in 62% of cases. At 56% of return visits, no side effects of botulinum toxin were reported. 90% of patients surveyed thought that botulinum toxin worked well or very well for their condition and 99% valued the treatment they received at the clinic as good to excellent.

Aim Analyze the HLA and MICA antibody detection data from multiple years of screening female blood donors for TRALI mitigation. Methods Over 4000 female blood donors with history of at least 1 pregnancy were screened using One Lambda, Inc. LabScreen Mixed Product. The samples were analyzed using HLA Fusion™ software. Normalized/NBG ratios were used to assign negative or positive results. Previously determined Cutoff Ratios were used: Table 1. Results HLA antibody was detected in over 20 % of the female blood donors that presented for donation. Table 2. Conclusions Significant levels of either HLA Class I or Class II antibody were detected in over 20% of female donors (reporting 1 plus pregnancy). This information is important as blood centers assess the impact of additional testing (i.e. Zika) on recruitment strategy, despite reduced blood product usage. Download : Download high-res image (48KB) Download : Download full-size image Download : Download high-res image (102KB) Download : Download full-size image

Aim We have previously reported that more than half of HLA-sensitized renal transplant candidates also exhibit anti-angiotensin II type 1 receptor antibody (AT1R-Ab). AT1R-Ab has been associated with antibody-mediated rejection and worse allograft survival. The aim of the present study was to prospectively compare the natural evolution of AT1R-Ab levels with response to desensitization therapy. Methods Ninety-three patients receiving desensitization therapy (VDS group, N = 23) or without desensitization therapy (non-VDS group, N = 70) had AT1R-Ab levels determined ELISA (U/mL after 1:50 dilution), before and after desensitization (VDS group) or before and after transplantation (non-VDS group). Results Initial AT1R-Ab levels were higher in VDS (17.64 ± 10.62 U/mL) then in non-VDS group (13.01 ± 9.14 U/mL, p Conclusions More than half of HLA-sensitized candidates exhibit AT1R-Ab. This initial desensitization experience consistently provided substantial reductions of AT1R-Ab levels in an unselected, highly sensitized kidney transplant candidate population, reduction that is more pronounced that post-transplant natural antibody dynamics. Download : Download high-res image (104KB) Download : Download full-size image