Pyrene-armed calix[4]arene conjugates non-covalently bind to DNA/RNA, as well as to mononucleotides, with an amazing affinity.

As the resistance of Plasmodium to the existing antimalarials increases, there is a crucial need to expand the antimalarial drug pipeline. We recently identified potent antimalarial compounds, namely harmiquins, hybrids derived from the β-carboline alkaloid harmine and 4-amino-7-chloroquinoline, a key structural motif of chloroquine (CQ). To further explore the structure−activity relationship, we synthesised 13 novel hybrid compounds at the position N-9 of the β-carboline ring and evaluated their efficacy in vitro against Plasmodium falciparum 3D7 and Dd2 strains (CQ sensitive and multi-drug resistant, respectively). All compounds exhibit persistent antimalarial activity against both strains of P. falciparum. The most interesting derivatives had low nanomolar activity against both strains (IC₅₀ (33) = 4.7 ± 1.3 nM against Pf3D7 and 6.5 ± 2.5 nM against PfDd2; IC₅₀ (37) = 4.6 ± 0.6 nM against 3D7 and 10.5 ± 0.4 nM against Dd2). Resistance indices (RIs) ranged from 0.9 to 5.3 compared to CQ (RI = 14.4), highlighting their superior consistency in activity against both strains. The cytotoxicity screening performed on HepG2 revealed over 3 orders of magnitude higher IC₅₀ for most of the compounds, with SIs from 711.0 to 8081.8. Spectroscopic studies explored the affinities of newly synthesised compounds for DNA, RNA, and HSA. Both tested hybrids, 34 and 39, were intrinsically fluorescent in an aqueous medium, characterised by remarkable Stokes shifts of emission maxima (Δλ = +103 and +93 nm for 34 and 39, respectively). Fluorimetric experiments revealed that compound 34, with its shorter and more flexible linker, exhibited at least an order of magnitude higher affinity toward ds-DNAs versus ds-RNA and two orders of magnitude higher affinity toward GC-DNAs compared to 39. The behaviour of the investigated compounds upon binding to HSA is very similar, showing a strong hypsochromic shift of the emission maximum (almost Δλ = −70 nm) and demonstrating their effectiveness as fluorimetric probes for distinguishing between DNA/RNA and proteins.

The synthesis of novel 6-chloro/morpholino/amino/-9-sulfonylpurine derivatives was accomplished in two ways, either (i) involving the condensation reaction of 6-chloropurine with commercially available arylsulfonyl chlorides in acetone and the presence of aqueous KOH at 0 °C, followed by the substitution of C6-chlorine with morpholine, or (ii) employing a reversed synthetic approach where 6-morpholinopurine and commercially available adenine bases were reacted with the corresponding alkyl, 2-arylethene and arylsulfonyl chlorides giving the N9 sulfonylated products, the latter particularly used where prior nonselective sulfonylation was observed. In both approaches, the sulfonylation reaction occurred regioselectively at the purine N9 position lacking any concurrent N7 derivatives, except in the case of a smaller methyl substituent on SO₂ and the free amino group at C6 of the purine ring. The tautomeric features of initial N9 unsubstituted purines, as well as stability trends among the prepared N-9-sulfonylpurine derivates, were investigated using DFT calculations with an important conclusion that electron-donating C6 substituents are beneficial for the synthesis as they both promote the predominance of the desired N9 tautomers and help to assure the stability of the final products. The newly synthesized 6-morpholino and 6-amino-9-sulfonylpurine derivatives showed antiproliferative activity on human carcinoma, lymphoma, and leukemia cells. Among the tested compounds, 6-morpholino 17 and 6-amino 22 derivatives, with trans-β-styrenesulfonyl group attached at the N9 position of purine, proved to be the most effective antiproliferative agents, causing accumulation of leukemia cells in subG0 cell cycle phase. Graphical Abstract