Pharmacological profile of a novel cyclic AMP-linked P2 receptor on undifferentiated HL-60 leukemia cells

Abstract

1. Extracellular ATP (EC₅₀=146±57 μM) and various ATP analogues activated cyclic AMP production in undifferentiated HL-60 cells.

2. The order of agonist potency was: ATPγS (adenosine 5′-O-[3-thiotriphosphate])⩾BzATP (2′&3′O-(4-benzoylbenzoyl)-adenosine-5′-triphosphate)⩾dATP>ATP. The following agonists (in order of effectiveness at 1 mM) were all less effective than ATP at concentrations up to 1 mM: β,γ methylene ATP⩾2-methylthioATP>ADP⩾Ap4A (P¹, P⁴-di(adenosine-5′) tetraphosphate)⩾Adenosine>UTP. The poor response to UTP indicates that P2Y₂ receptors are not responsible for ATP-dependent activation of adenylyl cyclase.

3. Several thiophosphorylated analogs of ATP were more potent activators of cyclic AMP production than ATP. Of these, ATPγS (EC₅₀=30.4±6.9 μM) was a full agonist. However, adenosine 5′-O-[1-thiotriphosphate] (ATPαS; EC₅₀=45±15 μM) and adenosine 5′-O-[2-thiodiphosphate] (ADPβS; EC₅₀=33.3±5.0 μM) were partial agonists.

4. ADPβS (IC₅₀=146±32 μM) and adenosine 5′-O-thiomonophosphate (AMPS; IC₅₀=343±142 μM) inhibited cyclic AMP production by a submaximal concentration of ATP (100 μM). Consistent with its partial agonist activity, ADPβS was estimated to maximally suppress ATP-induced cyclic AMP production by about 65%. AMPS has not been previously reported to inhibit P2 receptors.

5. The broad spectrum P2 receptor antagonist, suramin (500 μM), abolished ATP-stimulated cyclic AMP production by HL-60 cells but the adenosine receptor antagonists xanthine amine congener (XAC; 20 μM) and 8-sulpho-phenyltheophylline (8-SPT; 100 μM) were without effect.

6. Extracellular ATP also activated protein kinase A (PK-A) consistent with previous findings that PK-A activation is involved in ATP-induced differentiation of HL-60 cells (Jiang et al., 1997).

7. Taken together, the data indicate the presence of a novel cyclic AMP-linked P2 receptor on undifferentiated HL-60 cells.

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