Chuan-Chu Chou

SCH-C (SCH 351125), an orally bioavailable, small molecule antagonist of the chemokine receptor CCR5, is a potent inhibitor of HIV-1 infection in vitro and in vivo

We describe here the identification and properties of SCH-C (SCH 351125), a small molecule inhibitor of HIV-1 entry via the CCR5 coreceptor. SCH-C, an oxime–piperidine compound, is a specific CCR5 antagonist as determined in multiple receptor binding and signal transduction assays. This compound specifically inhibits HIV-1 infection mediated by CCR5 in U-87 astroglioma cells but has no effect on infection of CXCR4-expressing cells. SCH-C has broad and potent antiviral activity in vitro against primary HIV-1 isolates that use CCR5 as their entry coreceptor, with mean 50% inhibitory concentrations ranging between 0.4 and 9 nM. Moreover, SCH-C strongly inhibits the replication of an R5-using HIV-1 isolate in SCID-hu Thy/Liv mice. SCH-C has a favorable pharmacokinetic profile in rodents and primates with an oral bioavailability of 50–60% and a serum half-life of 5–6 h. On the basis of its novel mechanism of action, potent antiviral activity, and in vivo pharmacokinetic profile, SCH-C is a promising new candidate for therapeutic intervention of HIV infection.

Discovery and characterization of vicriviroc (SCH 417690), a CCR5 antagonist with potent activity against human immunodeficiency virus type 1.

ABSTRACT Inhibiting human immunodeficiency virus type 1 (HIV-1) infection by blocking the host cell coreceptors CCR5 and CXCR4 is an emerging strategy for antiretroviral therapy. Currently, several novel coreceptor inhibitors are being developed in the clinic, and early results have proven promising. In this report, we describe a novel CCR5 antagonist, vicriviroc (formerly SCH-D or SCH 417690), with improved antiviral activity and pharmacokinetic properties compared to those of SCH-C, a previously described CCR5 antagonist. Like SCH-C, vicriviroc binds specifically to the CCR5 receptor and prevents infection of target cells by CCR5-tropic HIV-1 isolates. In antiviral assays, vicriviroc showed potent, broad-spectrum activity against genetically diverse and drug-resistant HIV-1 isolates and was consistently more active than SCH-C in inhibiting viral replication. This compound demonstrated synergistic anti-HIV activity in combination with drugs from all other classes of approved antiretrovirals. Competition binding assays revealed that vicriviroc binds with higher affinity to CCR5 than SCH-C. Functional assays, including inhibition of calcium flux, guanosine 5′-[35S]triphosphate exchange, and chemotaxis, confirmed that vicriviroc acts as a receptor antagonist by inhibiting signaling of CCR5 by chemokines. Finally, vicriviroc demonstrated diminished affinity for the human ether a-go-go related gene transcript ion channel compared to SCH-C, suggesting a reduced potential for cardiac effects. Vicriviroc represents a promising new candidate for the treatment of HIV-1 infection.

Blocking ion channel KCNN4 alleviates the symptoms of experimental autoimmune encephalomyelitis in mice

The KCNN4 potassium‐ion channel has been reported to play an important role in regulating antigen‐induced T cell effector functions in vitro. This study presents the first evidence that a selective KCNN4 blocker, TRAM‐34, confers protection against experimental autoimmune encephalomyelitis (EAE) in the mouse model. Treatment with the KCNN4 blocker did not prevent infiltration of T cells in the spinal cord, but resulted in the reduction of both the protein and the message levels of TNF‐α and IFN‐γ as well as the message levels of several other pro‐inflammatory molecules in the spinal cord. Plasma concentrations of TRAM‐34 within a 24‐h period were between the in vitro IC50 and IC90 values for the KCNN4 channel. The effect of TRAM‐34 was reversible, as indicated by the development of clinical EAE symptoms within 48 h after withdrawal of treatment. In summary, our data support the idea that KCNN4 channels play a critical role in the immune response during the development of MOG‐induced EAE in C57BL/6 mice.

Disruption of CCL21-Induced Chemotaxis In Vitro and In Vivo by M3, a Chemokine-Binding Protein Encoded by Murine Gammaherpesvirus 68

ABSTRACT Chemokine-binding proteins represent a novel class of antichemokine agents encoded by poxviruses and herpesviruses. One such protein is encoded by the M3 gene present in the murine gammaherpesvirus 68 (MHV-68) genome. The M3 gene encodes a secreted 44-kDa protein that binds with high affinity to certain murine and human chemokines and has been shown to block chemokine signaling in vitro. However, there has been no direct evidence that M3 blocks chemokine activity in vivo, nor has the nature of M3-chemokine interaction been defined. To better understand the ability of M3 to block chemokine activity in vivo, we examined its interaction with a specific subset of chemokines expressed in lymphoid tissues, areas where gammaherpesviruses characteristically establish latency. Here we show that M3 blocks in vitro chemotaxis induced by CCL19 and CCL21, chemokines expressed constitutively in secondary lymphoid tissues. Moreover, we provide evidence that chemokine M3 binding exhibits positive cooperativity. In vivo, the expression of M3 in the pancreas of transgenic mice inhibits recruitment of lymphocytes induced by transgenic expression of CCL21 in this organ. The ability of M3 to block the biological activity of chemokines may represent an important strategy used by MHV-68 to evade immune detection and favor viral replication in the infected host.

KCa3.1: target and marker for cancer, autoimmune disorder and vascular inflammation?

KCa3.1 is a calcium-activated intermediate-conductance potassium ion channel. In humans the channel is expressed in several secretory organs and subtypes of hematopoietic cells, but not detected in excitable tissues. The mRNA level for KCa3.1 is upregulated in activated leukocytes, mitogen-induced endothelial cells and vascular smooth muscle cells, and several types of human cancers, suggesting a possible role for the channel in inflammatory and oncology diseases. Several potent and selective KCa3.1 blockers, including clotrimazole and its analogs TRAM-34 and ICA-17043, have been used to investigate the involvement of the channel in human disease. The compounds have been shown to suppress the proliferation of several cancer cells in vitro and the growth of the corresponding cancers in vivo, consistent with an oncologic indication. TRAM-34 also ameliorates symptoms in experimental autoimmune encephalomyelitis and several models of cardiovascular diseases, arguing for a role of the channel in inflammatory diseases. These results suggest several important opportunities for therapeutics based on KCa3.1. Further efforts will establish the optimal indication for these ion channel inhibitors.

Pharmacological characterization of the chemokine receptor, hCCR1 in a stable transfectant and differentiated HL-60 cells: antagonism of hCCR1 activation by MIP-1β

C‐C chemokine receptor‐1 (CCR1) has been implicated in mediating a variety of inflammatory conditions including multiple sclerosis and organ rejection. Although originally referred to as the MIP‐1α/RANTES receptor, CCR1 is quite promiscuous and can be activated by numerous chemokines. We used radioligand binding and [35S]‐GTPγS exchange assays in membranes from a cell line transfected to express CCR1 (Ba/F3‐hCCR1) to characterize a panel of chemokines (HCC‐1, MIP‐1α, MIP‐1β, MIP‐1δ, MPIF‐1, MCP‐2, MCP‐3, and RANTES) as CCR1 ligands. In this recombinant model, these chemokines displaced 125I‐MIP‐1α with a wide range of potencies and, with the exception of MCP‐2, acted as full agonists in stimulating [35S]‐GTPγS exchange. We then assessed the utility of HL‐60 cells cultured with known differentiating agents (PMA, DMSO, dibutyryl‐cAMP or retinoic acid) for investigating CCR1 pharmacology. In [35S]‐GTPγS exchange assays, membranes from cells cultured with retinoic acid (4–6 days) were the most responsive to activation by MIP‐1α and MPIF‐1. FACS analysis and comparative pharmacology confirmed that these activities were mediated by CCR1. Using [35S]‐GTPγS exchange assays, intracellular calcium flux and/or whole cell chemotaxis assays in HL‐60(Rx) cells, we validated that MIP‐1α was the most potent CCR1 ligand (MIP‐1α>MPIF‐1>RANTESMIP‐1β) although the ligands differed in their efficacy as agonists. MPIF‐1 was the more efficacious (MPIF‐1>RANTES=MIP‐1α>>MIP‐1β). 125I‐MIP‐1β binding in Ba/F3‐hCCR1 and HL‐60(Rx) membranes was competitively displaced by MIP‐1α, MPIF‐1 and MIP‐1β. The binding Ki for these chemokines with 125I‐MIP‐1β were essentially identical in the two membrane systems. Lastly, MIP‐1β antagonized [35S]‐GTPγS exchange, Ca2+ flux and chemotaxis in HL‐60(Rx) cells in response to robust agonists such as MIP‐1α, RANTES and MPIF‐1. Based on our results, we propose that MIP‐1β could function as an endogenous inhibitor of CCR1 function.