A Novel Long-Acting Human Growth Hormone Fusion Protein (VRS-317): Enhanced In Vivo Potency and Half-Life

ABSTRACT:A novel recombinant human growth hormone (rhGH) fusion protein (VRS-317) was designed to minimize receptor-mediated clearance through a reduction in receptor binding without mutations to rhGH by genetically fusing with XTEN amino acid sequences to the N-terminus and the C-terminus of the native hGH sequence. Although in vitro potency of VRS-317 was reduced approximately 12-fold compared with rhGH, in vivo potency was increased because of the greatly prolonged exposure to the target tissues and organs. VRS-317 was threefold more potent than daily rhGH in hypophysectomized rats and fivefold more potent than daily rhGH in juvenile monkeys. In juvenile monkeys, a monthly dose of 1.4mg/kg VRS-317 (equivalent to 0.26mg/kg rhGH) caused a sustained pharmacodynamic response for 1month equivalent to 0.05mg/kg/day rhGH (1.4mg/kg rhGH total over 28days). In monkeys, VRS-317, having a terminal elimination half-life of approximately 110h, was rapidly and near-completely absorbed, and was well tolerated with no observed adverse effects after every alternate week subcutaneous dosing for 14weeks. VRS-317 also did not cause lipoatrophy in pig and monkey studies. VRS-317 is currently being studied in GH-deficient patients to confirm the observations in these animal studies. © 2012 Wiley Periodicals, Inc. and the American Pharmacists Associatio

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

Immune systems biology: immunoprofiling of cells and molecules

White blood cells and their secreted products are key elements of immune systems biology that are important indicators of patient health and disease. We have developed the SurroScan™ microvolume laser scanning cytometer to immunoprofile hundreds of variables, including cell populations, cell surface antigens, and intracellular molecules in antibody-based assays on small samples (about 1 mL) of whole blood, processed blood, or other fluids without cell purification or washing steps. The system enables high-throughput, robust and automated data capture and analysis. We demonstrate the utility of this immunoprofiling technology platform by surveying patient samples before and after glucocorticosteroid administration and show both the expected and novel response characteristics. This system complements recent advances in genomic and proteomic approaches to disease prediction and monitoring

GLP2-2G-XTEN: a pharmaceutical protein with improved serum half-life and efficacy in a rat Crohn's. PLoS One. 2012; 7(11): e50630.

[This corrects the article on p. e50630 in vol. 7.]

Pharmacokinetics of GLP2-2G-XTEN in cynomolgous monkeys.

<p>GLP2-2G-XTEN plasma concentration is shown following 25 nmol/kg administration via intravenous (triangle) or subcutaneous (square) route. Three animals were dosed by each route of administration. Data points are the average ± s.d. of three animals per time point.</p

GLP2-2G-XTEN administration improves small intestine histopathology in the indomethacin-induced rat disease model.

<p>A) Hematoxylin and eosin staining of jejunum sections from A) diseased, vehicle-treated rats, B) diseased, GLP2-2G-XTEN high dose treated rats (125 nmol/kg total dose divided into five daily 25 nmol/kg doses), C) diseased, GLP2-2G-XTEN low dose treated rats (75 nmol/kg total dose given once on day −3) and D) diseased, GLP2-2G-XTEN low dose treated rats (divided into three 25 nmol/kg doses on day −3, −1 and 1).</p

Efficacy of low dose GLP2-2G-XTEN treatment in rat indomethacin-induced disease model.

<p>Data shown are effects on A) small intestine adhesions and B) small intestine trans-ulcerations. Treatment with GLP2-2G peptide (12.5 nmol/kg per injection; twice per day for 5 days) or GLP2-2G-XTEN (25 nmol/kg once daily or only once at the indicated doses) was initiated on Day −3 and sacrifice is on Day 2. Data are means and SE, n = 10 per group. * P<0.05 compared to diseased, vehicle-treated rats.</p

Efficacy of equal molar GLP2-2G-XTEN and GLP2-2G peptide in rat indomethacin-induced disease model.

<p>Data shown are A) change in body weight from day −3 through day 2, B) small intestine length, C) small intestine adhesion score, D) small intestine trans-ulceration score and E) small intestine TNFα concentration. Open bars are healthy rats treated with vehicle; colored bars are indomethacin-induced diseased rats treated with vehicle, GLP2-2G peptide (12.5 nmol/kg per injection; twice per day for 5 days) or GLP2-2G-XTEN (25 nmol/kg once per day for 5 days) as indicated. Compounds were dosed starting three days prior to first indomethacin injection (Day −3) and data shown are from the time of sacrifice on Day 2 (24 hours post second indomethacin injection). Data are means and SE, n = 10 per group. * P<0.05 compared to diseased, vehicle-treated rats, ^# P<0.05 compared to diseased GLP2-2G peptide treated rats.</p

Intestinotrophic effects of GLP2-2G peptide and GLP2-2G-XTEN.

<p>Small intestine weight A) and length B) in normal rats following treatment with a total dose of 300 nmol/kg GLP2-2G peptide or GLP2-2G-XTEN. Data shown are from day 12 following SC injection of GLP2-2G peptide (12.5 nmol/kg per injection; twice per day for 12 days) and GLP2-2G-XTEN (25 nmol/kg per injection; once per day for 12 days). Data are means and SE, n = 10 per group. * P<0.05 compared to vehicle-treated rats, ^# P<0.05 compared to GLP2-2G peptide treated rats.</p