Lyne Gagnon

2,4,6-Trisubstituted Triazines as Protein A Mimetics for the Treatment of Autoimmune Diseases

A first-in-class series of low molecular weight trisubstituted triazines were synthesized and evaluated for their ability to mimic protein A binding to human IgG antibody. The structure-activity relationship (SAR) demonstrates that the 1,3-phenylenediamine component was essential for robust activity. Twenty-two compounds, represented by lead molecule 34, displayed significant activity compared to protein A. These compounds may prove useful for the treatment of autoimmune disease.

D-LysAsnProTyr tetrapeptide : a novel B-cell stimulant and stabilized bursin mimetic

The tetrapeptide I (D-lysine-L-asparaginyl-L-prolyl-L-tyrosine or D-LysAsnProTyr), and analogue sequences, were synthesized and evaluated for the ability to stimulate immune cell subsets. These sequences were selected based on their perceived ability to readily adopt a beta-turn structure. In vitro immunological assays revealed a robust stimulation of mitogen activated B-cell proliferation and a modest to significant stimulation of cytotoxic T lymphocytes (CTLs). Further, this in vitro stimulation of B-cells was accompanied by an in vivo expansion of B-cells in C57BL/6 mice, as demonstrated by immunophenotyping experiments. Interestingly, a conformational analysis of the low energy conformers of I and the endogenous B-cell stimulant bursin (LysHisGlyNH2) shows that these molecules can be superimposed. However, I displayed significantly enhanced physiological stability. For a number of reasons, I may be a particularly useful vaccine adjuvant.

Fatty acid receptor modulator PBI-4050 inhibits kidney fibrosis and improves glycemic control

Extensive kidney fibrosis occurs in several types of chronic kidney diseases. PBI-4050, a potentially novel first-in-class orally active low-molecular weight compound, has antifibrotic and antiinflammatory properties. We examined whether PBI-4050 affected the progression of diabetic nephropathy (DN) in a mouse model of accelerated type 2 diabetes and in a model of selective tubulointerstitial fibrosis. eNOS-/- db/db mice were treated with PBI-4050 from 8-20 weeks of age (early treatment) or from 16-24 weeks of age (late treatment). PBI-4050 treatment ameliorated the fasting hyperglycemia and abnormal glucose tolerance tests seen in vehicle-treated mice. In addition, PBI-4050 preserved (early treatment) or restored (late treatment) blood insulin levels and increased autophagy in islets. PBI-4050 treatment led to significant improvements in lifespan in the diabetic mice. Both early and late PBI-4050 treatment protected against progression of DN, as indicated by reduced histological glomerular injury and albuminuria, slow decline of glomerular filtration rate, and loss of podocytes. PBI-4050 inhibited kidney macrophage infiltration, oxidative stress, and TGF-β-mediated fibrotic signaling pathways, and it also protected against the development of tubulointerstitial fibrosis. To confirm a direct antiinflammatory/antifibrotic effect in the kidney, further studies with a nondiabetic model of EGFR-mediated proximal tubule activation confirmed that PBI-4050 dramatically decreased the development of the associated tubulointerstitial injury and macrophage infiltration. These studies suggest that PBI-4050 attenuates development of DN in type 2 diabetes through improvement of glycemic control and inhibition of renal TGF-β-mediated fibrotic pathways, in association with decreases in macrophage infiltration and oxidative stress.

PBI-4050 Reduces Stellate Cell Activation and Liver Fibrosis through Modulation of Intracellular ATP Levels and the Liver Kinase B1/AMP-Activated Protein Kinase/Mammalian Target of Rapamycin Pathway

Hepatic fibrosis is a major cause of morbidity and mortality for which there is currently no effective therapy. We previously showed that 2-(3-pentylphenyl)acetic acid (PBI-4050) is a dual G protein–coupled receptor GPR40 agonist/GPR84 antagonist that exerts antifibrotic, anti-inflammatory, and antiproliferative action. We evaluated PBI-4050 for the treatment of liver fibrosis in vivo and elucidated its mechanism of action on human hepatic stellate cells (HSCs). The antifibrotic effect of PBI-4050 was evaluated in carbon tetrachloride (CCl4)– and bile duct ligation–induced liver fibrosis rodent models. Treatment with PBI-4050 suppressed CCl4-induced serum aspartate aminotransferase levels, inflammatory marker nitric oxide synthase, epithelial to mesenchymal transition transcription factor Snail, and multiple profibrotic factors. PBI-4050 also decreased GPR84 mRNA expression in CCl4-induced injury, while restoring peroxisome proliferator–activated receptor γ (PPARγ) to the control level. Collagen deposition and α-smooth muscle actin (α-SMA) protein levels were also attenuated by PBI-4050 treatment in the bile duct ligation rat model. Transforming growth factor-β–activated primary HSCs were used to examine the effect of PBI-4050 and its mechanism of action in vitro. PBI-4050 inhibited HSC proliferation by arresting cells in the G0/G1 cycle phase. Subsequent analysis demonstrated that PBI-4050 signals through a reduction of intracellular ATP levels, activation of liver kinase B1 (LKB1) and AMP-activated protein kinase (AMPK), and blockade of mammalian target of rapamycin (mTOR), resulting in reduced protein and mRNA levels of α-SMA and connective tissue growth factor and restored PPARγ mRNA expression. Our findings suggest that PBI-4050 may exert antifibrotic activity in the liver through a novel mechanism of action involving modulation of intracellular ATP levels and the LKB1/AMPK/mTOR pathway in stellate cells, and PBI-4050 may be a promising agent for treating liver fibrosis.

Design and Synthesis of New 1,3,5-Trisubstituted Triazines for the Treatment of Cancer and Inflammation

Abstract Low‐molecular‐weight synthetic molecules 1 with the general 2‐(fluorophenylamino)‐4,6‐disubstituted 1,3,5‐triazine structure and showing anti‐inflammatory and anticancer activities were explored. Structure–activity relationship studies demonstrated the importance of the aminopentyl chain, the 3‐ or 4‐fluorophenylaniline component, and the presence of at least one substituent, such as a tyramine moiety, attached directly to the triazine ring as essential for good activity. These compounds, represented by leads 4‐{2‐[4‐(5‐Aminopentylamino)‐6‐(3‐fluorophenylamino)‐1,3,5‐triazin‐2‐ylamino]ethyl}phenol (6) and 4‐{2‐[4‐(5‐Aminopentylamino)‐6‐(4‐fluorophenylamino)‐1,3,5‐triazin‐2‐ylamino]ethyl}phenol (10), displayed moderate and significant in vitro and in vivo dual activities, respectively, and address the molecular link between inflammation and cancer. Compound 10 demonstrated significant antitumor efficacy upon administration by the oral and intravenous routes in several animal models. This class of triazine compounds is new, safe, and nontoxic and offers a novel approach to the treatment of inflammation and cancer.

P126 Oral treatment with PBI-4050 ameliorates bleomycin-induced pulmonary fibrosis by reducing serum and lung anti-inflammatory/fibrotic biomarkers.

PBI-4050 is a first-in-class orally active compound which displays antifibrotic activities in kidney, heart, liver, pancreas and lung models. PBI-4050 was found to be safe and well tolerated in healthy volunteers and in patients with CKD and Type 2 diabetes without any SAEs. Translation of pharmacological efficacy in humans …

Abstract 3534: PBI-1402, a first-in-class erythropoiesis regulating agent, possesses differentiation properties and demonstrates synergistic anticancer activity in combination with chemotherapy

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Background: PBI-1402 reduces the need for transfusion and increases hemoglobin (Hb) level and red blood cell count (RBC) in chemotherapy-induced anemia (CIA) patients by a mechanism of action which is distinct from erythropoietin (EPO). The PBI-1402 receptor is also expressed on certain cancer cells such as leukemia (K562), lung (LL-2), prostate (PC-3) and pancreas (Panc-02). Aim: The objective of this study was to determine the role of the PBI-1402 receptor on tumor growth. Methods: Cell proliferation and differentiation of K562 (human erythroleukemia) was studied in presence of PBI-1402 or EPO using 2,7-diaminofluorene for hemoglobin quantification. The effect of oral administration of PBI-1402 in combination with chemotherapy agents (gemcitabine or cyclophosphamide) was studied in subcutaneous syngeneic Panc02, LL-2 and xenogeneic PC-3 models. Results: PBI-1402 inhibits proliferation of K562 cells and promotes differentiation of the remaining cells. K562 cells express both EPO and PBI-1402 receptors. EPO increases phosphorylation of ERK1/2 and Stat3 (linked to cell proliferation) while PBI-1402 decreases it in a dose dependent manner. The antitumor efficacy of oral administration of PBI-1402 was studied in combination with gemcitabine in subcutaneous LL2 and Panc02 as well as in orthotopic Panc02 cancers. In subcutaneous Panc02, gemcitabine induced a significant inhibition (p<0.05) of tumor growth from day 27 to 34 with T/C from 55% to 78%. PBI-1402 induced a significant inhibition (p<0.05) of tumor growth from day 23 to 44 with T/C from 26% to 58%. The combination therapy induced a significant (p<0.05) inhibition of tumor growth and T/C<40% from day 23 to 37 when compared to control and at day 23, 25 and 30 to 39 when compared to gemcitabine alone. In orthotopic Panc02, mice treated with gemcitabine or a combination of gemcitabine and PBI-1402 displayed prolonged survival (71 and 88 days, respectively) compared to control (48 days). In LL-2 cancer, at day 25, mice treated with PBI-1402 or gemcitabine alone had a T/C of 94% and 72%, respectively. Mice treated with a combination of gemcitabine plus PBI-1402 displayed a significant (synergistic) reduction of tumor growth (T/C: 33%). The effect of PBI-1402 alone or in combination with cyclophosphamide was also studied in a xenograft prostate PC-3 tumor. Mice treated with PBI-1402 or cyclophosphamide alone have a significant T/C<40% from day 21 to 56 and day 35 to 56, respectively. Mice treated with a combination of cyclophosphamide plus PBI-1402 displayed tumor regression from day 21 to 56. Conclusion: These results suggest that PBI-1402 has the potential to inhibit the growth of lung, prostate and pancreatic cancer by induction of cell differentiation. PBI-1402-differentiated cancer cells appear to be more susceptible to chemotherapeutic agents. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3534. doi:10.1158/1538-7445.AM2011-3534

Abstract 672: PBI-1402, a first-in-class erythropoiesis regulating agent, inhibits tumor growth and metastasis of murine mastocytoma and does not overshoot hemoglobin: Comparison with erythropoietin

BACKGROUND: PBI-1402 reduces the need for transfusion and increases hemoglobin (Hb) level and red blood cell count (RBC) in chemotherapy-induced anemia (CIA) by a mechanism of action distinct from erythropoietin (EPO). AIM: The objective was to compare the effect of PBI-1402 and EPO on modulation of tumor growth. METHODS: The effect of PBI-1402 (200 mg/kg, oral, once a day) or EPO (200 or 2000 U/kg, s.c. injection, twice a week) was studied in a subcutaneous P815 mastocytoma model. P815 cells express both PBI-1402 and EPO receptors. Tumor growth, metastasis, serum nitric oxide (NO) and hematocrit (Ht) were assessed. RESULTS: P815 tumor growth is rapid and metastasizes to the liver. Inflammation is also associated with P815 tumor growth. PBI-1402 significantly reduced tumor growth (T/C=37%) and invasion as demonstrated by a 33% reduction of mice with liver metastasis. In comparison, EPO had no effect (200 U/kg, equivalent dose used in CKD treatment). However, high dose EPO (2000 U/kg, equivalent dose used in human cancer treatment) exacerbated tumor growth (T/C: 135%). Both doses of EPO induced a significant increase in the percentage of mice with liver metastasis (2X, 200 U/kg, and 3X, 2000 U/kg) compared to control. PBI-1402 treatment had no effect on NO and did not overshoot normal hematocrit. However, treatment with high dose EPO significantly increased NO and Ht levels (Table 1). CONCLUSION: These results suggest that oral treatment with PBI-1402 may inhibit growth and metastasis of cancer cells. As an important safety feature, PBI-1402 does not induce overshoot of hemoglobin. In contrast to EPO, PBI-1402 may offer the advantage of reducing tumor growth and metastasis while preventing anemia induced by chemotherapy. In conclusion, PBI-1402 has the potential to be a safe erythropoiesis-regulating agent useful in treating anemia in cancer patients. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 672. doi:10.1158/1538-7445.AM2011-672