Medical genetics Expression analysis of activated protein kinase C gene (LACK1) in antimony sensitive and resistant Leishmania tropica clinical isolates using real-time RT-PCR Homa Hajjaran 1, *, PhD, Elham Kazemi-Rad 2, *, PhD, Mehdi Mohebali 1,3 , VMD, MPH, PhD, Mohammad A. Oshaghi 4 , PhD, Mohammad B. Khadem-Erfan 5 , PhD, Elham Hajaliloo 1 , PHD, Hossein Reisi Nafchi 1 , MSc, and Reza Raoofian 6 , MD, PhD 1 Department of Medical Parasitology and Mycology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran, 2 Department of Parasitology, Pasteur Institute of Iran, Tehran, Iran, 3 Research Center for Endemic Parasites of Iran, Tehran University of Medical Sciences, 4 Department of Medical Entomology and Vector Control, School of Public Health, Tehran University of Medical Sciences, Tehran, 5 Department of Medical Parasitology and Mycology, Kurdistan University of Medical Sciences, Sanandaj, and 6 Legal Medicine Research Center, Legal Medicine Organization, Tehran, Iran Correspondence Homa Hajjaran, PhD Department of Medical Parasitology and Mycology, School of Public Health Tehran University of Medical Sciences Tehran Iran E-mail: hajaranh@tums.ac.ir *These authors contributed equally. doi: 10.1111/ijd.13321 Abstract Background Resistance to pentavalent antimonial drugs has become a serious problem in the treatment of cutaneous leishmaniasis in some endemic areas. Investigations on molecular markers involved in drug resistance are essential for monitoring of the disease. Leishmania-activated C kinase gene (LACK1) is involved in multiple central processes such as signal transduction. According to the probable role of the LACK1 gene in antimony resistance, we used real-time reverse transcription–polymerase chain reaction (PCR) to investigate the expression of this gene in clinical L. tropica strains, which were resistant or sensitive to meglumine antimoniate. Methods We analyzed the expression level of LACK in 18 sensitive and 14 resistant L. tropica isolates collected from patients with anthroponotic cutaneous leishmaniasis. After cDNA synthesis, gene expression analysis was performed by quantitative real-time PCR using SYBR Green. In addition, the full length of the LACK gene from six reference strains was cloned and sequenced then deposited in the NCBI database to confirm our strains. Results Real-time reverse transcription-PCR revealed that the average RNA expression level of LACK in isolates from unresponsive and responsive patients were 0.479 and 4.583, respectively, and expression of LACK was significantly downregulated (9.56-fold) in resistant isolates compared to sensitive ones. Conclusion Results of the present study suggest the probable role of the LACK gene in antimony resistance. Moreover, it can be considered as a potential marker for monitoring antimony resistance in clinical isolates. However, further studies are required to exploit the biological functions of it in antimony resistance. Introduction Leishmaniasis is a protozoan infection with worldwide spread particularly in tropical and subtropical countries. Leishmania species as arthropod-transmitted parasites can infect and replicate in host macrophages. 1 Various species of Leishmania have a different clinical manifestation from self-curing cutaneous lesions to the potentially fatal vis- ceral disease. In addition, atypical infections have been reported. 2 In Iran, human leishmaniasis occurs from the northeast to the central, west, and covers the entire south- east and west region. Depending on the Leishmania spe- cies, several forms of disease including anthroponotic cuta- neous leishmaniasis (ACL; L. tropica), zoonotic CL (L. major), and visceral leishmaniasis (L. infantum) occur in Iran. 3 Cutaneous infection is limited to skin, but some studies show that L. tropica can infect visceral organs of humans and dogs. 4–6 The mainstay of therapy for leishma- niasis is meglumine antimoniate (Glucantime) in Iran. Unfortunately, during the recent decade, the increase in resistance of antileishmanial drugs has become a serious public health concern, particularly the ACL form induced by L. tropica in endemic areas of Iran. 7,8 Studies showed that the Leishmania parasites have been slowly acquiring mutations and/or modulating in gene expression, leading to decreased susceptibility to the drug. However, other fac- tors such as host immunity might be reasons for treatment failure. 9 In this context, determining some biomarkers for ª 2016 The International Society of Dermatology International Journal of Dermatology 2016 1