Development of a Patient Acceptable Symptom State (PASS) Question for Children with Consistent, Long-Term Disability

Patient Acceptable Symptom State (PASS) is a single Yes/No question where a “yes” response indicates the minimal level of symptoms and function above which a person considers their current condition satisfactory. ➢ PASS has shown validity as an anchoring question for important health outcomes in adults with various diagnoses and in children with JIA (Strand 2011; Kvien 2007; Tubach 2012; Filocamo 2012; Consolaro 2012; Roos 2019). However, it has not been researched in children with consistent, long-term disability

Quantification of recombinant and platelet P2Y1 receptors utilizing a [125I]-labeled high-affinity antagonist 2-iodo-N6-methyl-(N)-methanocarba-2′-deoxyadenosine-3′,5′-bisphosphate ([125I]MRS2500)

The ADP-activated P2Y1 receptor is broadly expressed and plays a crucial role in ADP-promoted platelet aggregation. We previously synthesized 2-iodo-N6-methyl–(N)-methanocarba-2′-deoxyadenosine 3′,5′-bisphosphate (MRS2500), as a selective, high affinity, competitive antagonist of this receptor. Here we report utilization of a trimethylstannyl precursor molecule for the multistep radiochemical synthesis of a [125I]-labeled form of MRS2500. [125I]MRS2500 bound selectively to Sf9 insect cell membranes expressing the human P2Y1 receptor but did not specifically bind to membranes isolated from empty vector-infected cells. Binding of [125I]MRS2500 to P2Y1 receptors was saturable with a Kd of 1.2 nM. Known agonists and antagonists of the P2Y1 receptor inhibited [125I]MRS2500 binding to P2Y1 receptor-expressing membranes with potencies in agreement with those previously observed in functional assays of this receptor. A high-affinity binding site for [125I]MRS2500 also was observed on intact human platelets (Kd = 0.61 nM) and mouse platelets (Kd = 1.20 nM) that exhibited the pharmacological selectivity of the P2Y1 receptor. The densities of sites observed were 151 sites/platelet and 229 sites/platelet in human and mouse platelets, respectively. In contrast, specific binding was not observed in platelets isolated from P2Y1 receptor (−/−) mice. Taken together, these data illustrate the synthesis and characterization of a novel P2Y1 receptor radioligand and its utility for examining P2Y1 receptors natively expressed on human and mouse platelets

Functionalized Congeners of P2Y 1 Receptor Antagonists: 2-Alkynyl ( N )-Methanocarba 2′-Deoxyadenosine 3′,5′-Bisphosphate Analogues and Conjugation to a Polyamidoamine (PAMAM) Dendrimer Carrier

The P2Y1 receptor is a prothrombotic G protein-coupled receptor (GPCR) activated by ADP. Preference for the North (N) ring conformation of the ribose moiety of adenine nucleotide 3′,5′-bisphosphate antagonists of the P2Y1 receptor was established by using a ring-constrained methanocarba (a bicyclo[3.1.0]hexane) ring as a ribose substitute. A series of covalently linkable N6-methyl-(N)-methanocarba-2′-deoxyadenosine-3′,5′-bisphosphates containing extended 2-alkynyl chains was designed and binding affinity at the human (h) P2Y1 receptor determined. The chain of these functionalized congeners contained hydrophilic moieties, a reactive substituent, or biotin, linked via an amide. Variation of the chain length and position of an intermediate amide group revealed high affinity of carboxylic congener 8 (Ki 23 nM) and extended amine congener 15 (Ki 132 nM), both having a 2-(1-pentynoyl) group. A biotin conjugate 18 containing an extended ε-aminocaproyl spacer chain exhibited higher affinity than a shorter biotinylated analogue. Alternatively, click coupling of terminal alkynes of homologous 2-dialkynyl nucleotide derivatives to alkyl azido groups produced triazole derivatives that bound to the P2Y1 receptor following deprotection of the bisphosphate groups. The preservation of receptor affinity of the functionalized congeners was consistent with new P2Y1 receptor modeling and ligand docking. Attempted P2Y1 antagonist conjugation to PAMAM dendrimer carriers by amide formation or palladium-catalyzed reaction between an alkyne on the dendrimer and a 2-iodopurine-derivatized nucleotide was unsuccessful. A dialkynyl intermediate containing the chain length favored in receptor binding was conjugated to an azide-derivatized dendrimer, and the conjugate inhibited ADP-promoted human platelet aggregation. This is the first example of attaching a strategically functionalized P2Y receptor antagonist to a PAMAM dendrimer to produce a multivalent conjugate exhibiting a desired biological effect, i.e. antithrombotic action

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

Stakeholder-led science: engaging resource managers to identify science needs for long-term management of floodplain conservation lands

Floodplains pose challenges to managers of conservation lands because of constantly changing interactions with their rivers. Although scientific knowledge and understanding of the dynamics and drivers of river-floodplain systems can provide guidance to floodplain managers, the scientific process often occurs in isolation from management. Further, communication barriers between scientists and managers can be obstacles to appropriate application of scientific knowledge. With the coproduction of science in mind, our objectives were the following: (1) to document management priorities of floodplain conservation lands, and (2) identify science needs required to better manage the identified management priorities under nonstationary conditions, i.e., climate change, through stakeholder queries and interactions. We conducted an online survey with 80 resource managers of floodplain conservation lands along the Upper and Middle Mississippi River and Lower Missouri River, USA, to evaluate management priority, management intensity, and available scientific information for management objectives and conservation targets. Management objectives with the least information available relative to priority included controlling invasive species, maintaining respectful relationships with neighbors, and managing native, nongame species. Conservation targets with the least information available to manage relative to management priority included pollinators, marsh birds, reptiles, and shore birds. A follow-up workshop and survey focused on clarifying science needs to achieve management objectives under nonstationary conditions. Managers agreed that metrics of inundation, including depth and extent of inundation, and frequency, duration, and timing of inundation would be the most useful metrics for management of floodplain conservation lands with multiple objectives. This assessment provides guidance for developing relevant and accessible science products to inform management of highly dynamic floodplain environments. Although the problems facing managers of these lands are complex, products focused on a small suite of inundation metrics were determined to be the most useful to guide the decision making process