Tehran University of Medical Sciences Iranian Journal of Parasitology 1735-7020 17 4 2022 12 04 525 534 Sepand Razavi Vakhshourpour Department of Medical Parasitology & Mycology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran Mehdi Nateghpour 1. Department of Medical Parasitology & Mycology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran 2. Research Center of Quran, Hadith, and Medicine, Tehran University of Medical Sciences, Tehran, Nader Shahrokhi Department of Molecular Biology, Pasteur Institute of Iran, Tehran, Iran Afsaneh Motevalli Haghi Department of Medical Parasitology & Mycology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran Mehdi Mohebali Department of Medical Parasitology & Mycology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran Haleh Hanifian Department of Medical Parasitology & Mycology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran 2021 08 30 2022 04 10 Background: Infections by Plasmodium falciparum, are becoming increasingly difficult to treat. Therefore, there is an urgent need for novel antimalarial agents’ discovery against infection. In present study, we described a 2’-O-Methyl gapmer phosphorothioate oligonucleotide antisense targeting translation initiation region of 3D7 strain RH5 gene. Methods: The study was conducted in Pasteur Institute of Iran in 2020. ODNs effects were measured by microscopic examination and real time RT-PCR. For microscopy, microplates were charged with 2’-OMe ODNs at different dilutions. Unsynchronized parasites were added to a total of 0.4 ml (0.4% parasitemia, 5% red blood cells), and slides were prepared. Proportion of infected cells was measured by counting at least 500 red blood cells. Results: RH5 genes start codon regions selected as conserved region besed on alignment results. Gap-RH5-As which was complementary to sequence surrounding AUG RH5 start codon significantly reduced parasite growth (>90% at 50 nM) compared to sense sequence control (Gap-RH5-Se) (17%), (P<0.001). RH5 transcripts were dramatically reduced after exposed to ODNs at a concentration of 5-500 nM for 48 h. Conclusion: Gemnosis delivery of a chimeric gapmer PS-ODN with 2’-OMe modifications at both sides had high antisense activity at low concentrations (10-100 nM) and shown a good efficiency to reach to target mRNA in human RBCs. Anti-parasite effect was correlated to reduction of target gene mRNA level. In addition, 2’-OMe ODNs free delivery is an effective way and does not need any carrier molecules or particles. https://ijpa.tums.ac.ir/index.php/ijpa/article/view/3439 https://ijpa.tums.ac.ir/index.php/ijpa/article/download/3439/1252