PKI-166

Human African trypanosomiasis (HAT) is caused by the protozoan Trypanosoma brucei . New drugs are needed to treat HAT because of undesirable side effects and difficulties in the administration of the antiquated drugs that are currently used. In human proliferative diseases, protein tyrosine kinase (PTK) inhibitors (PTKIs) have been developed into drugs (e.g., lapatinib and erlotinib) by optimization of a 4-anilinoquinazoline scaffold. Two sets of facts raise a possibility that drugs targeted against human PTKs could be “hits” for antitrypanosomal lead discoveries. First, trypanosome protein kinases bind some drugs, namely, lapatinib, CI-1033, and AEE788. Second, the pan-PTK inhibitor tyrphostin A47 blocks the endocytosis of transferrin and inhibits trypanosome replication. Following up on these concepts, we performed a focused screen of various PTKI drugs as possible antitrypanosomal hits. Lapatinib, CI-1033, erlotinib, axitinib, sunitinib, PKI-166, and AEE788 inhibited the replication of bloodstream T. brucei , with a 50% growth inhibitory concentration (GI 50 ) between 1.3 μM and 2.5 μM. Imatinib had no effect (i.e., GI 50 > 10 μM). To discover leads among the drugs, a mouse model of HAT was used in a proof-of-concept study. Orally administered lapatinib reduced parasitemia, extended the survival of all treated mice, and cured the trypanosomal infection in 25% of the mice. CI-1033 and AEE788 reduced parasitemia and extended the survival of the infected mice. On the strength of these data and noting their oral bioavailabilities, we propose that the 4-anilinoquinazoline and pyrrolopyrimidine scaffolds of lapatinib, CI-1033, and AEE788 are worth optimizing against T. brucei in medicinal chemistry campaigns (i.e., scaffold repurposing) to discover new drugs against HAT.

This study aimed to determine the interaction between the EGF receptor (EGFR) and peroxisome proliferator-activated receptor γ (PPARγ) and the role of EGFR in sodium and water transport in the proximal tubule.

Primary human proximal tubule cells (PTCs) were exposed to high glucose in the presence and absence of pioglitazone. Total and phospho-EGFR levels and EGFR mRNA expression were determined by western blot and real-time PCR, respectively. Sodium-hydrogen exchanger-3 (NHE3), PPARγ and aquaporin 1 (AQP1) levels were determined by western blot. The role of EGFR was elucidated using the EGFR tyrosine kinase inhibitor, PKI166. The role of PPARγ in high-glucose conditions was determined using specific PPARγ small interfering (si)RNA. P-EGFR, PPARγ, AQP1 and NHE3 production in a rat model of diabetes (streptozotocin-induced hypertensive Ren-2 transgenic [mRen2]27 rats) and controls, with or without pioglitazone treatment, was determined by immunohistochemistry. The PPARγ and EGFR interaction was determined by chromatin immunoprecipitation assay, and the effect of pioglitazone on EGFR activation by luciferase assay.

PTCs exposed to both high glucose and pioglitazone increased protein abundance of P-EGFR, NHE3, AQP1 and PPARγ. Pioglitazone-induced upregulation of NHE3 and AQP1 was abolished by PKI166. High-glucose-induced increases in P-EGFR, NHE3 and AQP1 were decreased with PPARγ siRNA. AQP1 and NHE3 but not PPARγ were increased in a diabetic rat model and further increased by pioglitazone treatment. Pioglitazone induced PPARγ binding to the EGFR promoter and subsequent downstream activation.

Our data suggest that EGFR activation mediates PPARγ-induced sodium and water reabsorption via upregulation of NHE3 and AQP1 channels in the proximal tubule. EGFR inhibition may be a therapeutic strategy in the treatment of diabetic nephropathy and in limiting salt and water retention, which currently restricts the use of PPARγ agonists.