Trypanosoma; MmpL3
Uncoupler of mitochondrial membrane potential (Δψm) and pH gradient (ΔpH) in Trypanosoma cruzi
Inhibitor of T. cruzi squalene synthase (TcSQS) (IC50 ~100 μM)
Binds to active site of T. cruzi and human squalene synthase (structural evidence) [1]

With a selectivity index of approximately 10 to 20, SQ109 also inhibits clinically relevant intracellular amastigotes (IC50, ~0.5 to 1 μM) and extracellular epimastigotes (IC50, 4.6±1 μM). SQ109 performs poorly in an assay measuring hemolysis of red blood cells (EC50, ~80 μM). Furthermore, as shown by transmission electron microscopy (TEM), scanning electron microscopy (SEM), and light microscopy (LMC), SQ109 significantly alters the ultrastructural characteristics of all three life cycle forms[1].

---

Researchers tested the antituberculosis drug SQ109, which is currently in advanced clinical trials for the treatment of drug-susceptible and drug-resistant tuberculosis, for its in vitro activity against the trypanosomatid parasite Trypanosoma cruzi, the causative agent of Chagas disease. SQ109 was found to be a potent inhibitor of the trypomastigote form of the parasite, with a 50% inhibitory concentration (IC50) for cell killing of 50 ± 8 nM, but it had little effect (50% effective concentration [EC50], ∼80 μM) in a red blood cell hemolysis assay. It also inhibited extracellular epimastigotes (IC50, 4.6 ± 1 μM) and the clinically relevant intracellular amastigotes (IC50, ∼0.5 to 1 μM), with a selectivity index of ∼10 to 20. SQ109 caused major ultrastructural changes in all three life cycle forms, as observed by light microscopy, scanning electron microscopy (SEM), and transmission electron microscopy (TEM). It rapidly collapsed the inner mitochondrial membrane potential (Δψm) in succinate-energized mitochondria, acting in the same manner as the uncoupler FCCP [carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone], and it caused the alkalinization of internal acidic compartments, effects that are likely to make major contributions to its mechanism of action. The compound also had activity against squalene synthase, binding to its active site; it inhibited sterol side-chain reduction and, in the amastigote assay, acted synergistically with the antifungal drug posaconazole, with a fractional inhibitory concentration index (FICI) of 0.48, but these effects are unlikely to account for the rapid effects seen on cell morphology and cell killing. SQ109 thus most likely acts, at least in part, by collapsing Δψ/ΔpH, one of the major mechanisms demonstrated previously for its action against Mycobacterium tuberculosis. Overall, the results suggest that SQ109, which is currently in advanced clinical trials for the treatment of drug-susceptible and drug-resistant tuberculosis, may also have potential as a drug lead against Chagas disease [1].

---

SQ109 exhibits potent activity against the trypomastigote form of T. cruzi with an IC50 of 50 ± 8 nM for cell killing, while showing little hemolytic effect on red blood cells (EC50 > 80 μM). It also inhibits extracellular epimastigotes (IC50 4.6 ± 1 μM) and intracellular amastigotes (IC50 ~0.5 to 1 μM), with a selectivity index of ~10 to 20. The compound causes major ultrastructural changes in all three life cycle forms, collapses the inner mitochondrial membrane potential in succinate-energized mitochondria, alkalinizes internal acidic compartments, and inhibits sterol side-chain reduction. It acts synergistically with posaconazole against intracellular amastigotes (FICI = 0.48). [1]

For 28 days, mice given SQ109 orally (0.1–25 mg/kg daily) showed dose-dependent reductions in lung and spleen mycobacterial loads, which were similar to those of EMB given daily at 100 mg/kg, though less effective than isoniazid (INH) given daily at 25 mg/kg. After a single administration, the pharmacokinetic profiles of SQ109 in mice revealed a Cmax of 1038 for intravenous (i.v.) and 135 ng/mL for oral administration, along with an oral Tmax of 0.31 hours.Regarding SQ109, the elimination t1/2 is 3.5 (i.v.) and 5.2 h (p.o.). Oral bioavailability amounts to 4% [2]. Dogs have a significantly larger volume of distribution for SQ109 than rats do (7-8 h, mean 7.4 h), indicating that dogs have a longer terminal half-life (t1/2) of SQ109. It has been found that SQ109 has an oral bioavailability of 12% in rats and 5% in dogs[3].\n

---

\nSQ109 is a novel [1,2]-diamine-based ethambutol (EMB) analog developed from high-throughput combinatorial screening. The present study aimed at characterizing its pharmacodynamics and pharmacokinetics. The antimicrobial activity of SQ109 was confirmed in vitro (Mycobacterium tuberculosis-infected murine macrophages) and in vivo (M. tuberculosis-infected C57BL/6 mice) and compared to isoniazid (INH) and EMB. SQ109 showed potency and efficacy in inhibiting intracellular M. tuberculosis that was similar to INH, but superior to EMB. In vivo oral administration of SQ109 (0.1-25 mg kg(-1) day(-1)) to the mice for 28 days resulted in dose-dependent reductions of mycobacterial load in both spleen and lung comparable to that of EMB administered at 100 mg kg(-1) day(-1), but was less potent than INH at 25 mg kg(-1) day(-1). Monitoring of SQ109 levels in mouse tissues on days 1, 14 and 28 following 28-day oral administration (10 mg kg(-1) day(-1)) revealed that lungs and spleen contained the highest concentration of SQ109, at least 10 times above its MIC. Pharmacokinetic profiles of SQ109 in mice following a single administration showed its C(max) as 1038 (intravenous (i.v.)) and 135 ng ml(-1) (p.o.), with an oral T(max) of 0.31 h. The elimination t(1/2) of SQ109 was 3.5 (i.v.) and 5.2 h (p.o.). The oral bioavailability was 4%. However, SQ109 displayed a large volume of distribution into various tissues. The highest concentration of SQ109 was present in lung (>MIC), which was at least 120-fold (p.o.) and 180-fold (i.v.) higher than that in plasma. The next ranked tissues were spleen and kidney. SQ109 levels in most tissues after a single administration were significantly higher than that in blood. High tissue concentrations of SQ109 persisted for the observation period (10 h). This study demonstrated that SQ109 displays promising in vitro and in vivo antitubercular activity with favorable targeted tissue distribution properties. [2]

---

\nThis study aimed at characterizing the interspecies absorption, distribution, metabolism and elimination (ADME) profile of N-geranyl-N'-(2-adamantyl)ethane-1,2-diamine (SQ109), a new diamine-based antitubercular drug. Single doses of SQ109 were administered (intravenously (i.v.) and per os (p.o.)) to rodents and dogs and blood samples were analyzed by liquid chromatography tandem mass spectrometry (LC/MS/MS). Based on i.v. equivalent body surface area dose, the terminal half-life (t1/2) of SQ109 in dogs was longer than that in rodents, reflected by a larger volume of distribution (Vss) and a higher clearance rate of SQ109 in dogs, compared to that in rodents. The oral bioavailability of SQ109 in dogs, rats and mice were 2.4-5, 12 and 3.8%, respectively. After oral administration of [14C]SQ109 to rats, the highest level of radioactivity was in the liver, followed by the lung, spleen and kidney. Tissue-to-blood ratios of [14C]SQ109 were greater than 1. Fecal elimination of [14C]SQ109 accounted for 22.2% of the total dose of [14C]SQ109, while urinary excretion accounted for only 5.6%. The binding of [14C]SQ109 (0.1-2.5 microg ml-1) to plasma proteins varied from 6 to 23% depending on the species (human, mouse, rat and dog). SQ109 was metabolized by rat, mouse, dog and human liver microsomes, resulting in 22.8, 48.4, 50.8 or 58.3%, respectively, of SQ109 remaining after a 10-min incubation at 37 degrees C. The predominant metabolites in the human liver microsomes gave intense ion signals at 195, 347 and 363m/z, suggesting the oxidation, epoxidation and N-dealkylation of SQ109. P450 reaction phenotyping using recombinant cDNA-expressed human CYPs in conjunction with specific CYP inhibitors indicated that CYP2D6 and CYP2C19 were the predominant CYPs involved in SQ109 metabolism[3].

---

In a Mycobacterium tuberculosis H37Rv-infected C57BL/6 mouse model, oral administration of SQ109 for 28 days resulted in dose-dependent reductions of bacterial load (colony-forming units, CFU) in the spleen and lungs. At doses of 10 and 25 mg kg⁻¹ day⁻¹, SQ109 reduced bacterial CFU in both organs to a level comparable to EMB administered at 100 mg kg⁻¹ day⁻¹. The reduction was less potent than INH at 25 mg kg⁻¹ day⁻¹. SQ109 at a low dose of 0.1 mg kg⁻¹ day⁻¹ still produced a significant decrease in bacterial load in the spleen. All drug-treated mice survived the experiment, whereas some untreated mice died. [2]

Microsomal metabolism of SQ109 [3]
\nThe microsomal assay was similar to that described previously (Jia et al., 2003). Briefly, SQ109 (10 μM) was incubated with mouse, rat, dog and human liver microsomes, respectively, in an NADPH-generating system containing 1.3 mM NADP, 3.3 mM glucose-6-phosphate, 0.4 U/ml glucose-6-phosphate dehydrogenase and 3.3 mM MgCl2 in 100 mM potassium phosphate buffer (pH 7.4). Reaction mixtures were prepared in duplicate and were preincubated for 5 min at 37°C. The reactions were then initiated by the addition of microsomes (30 μl of a 20 mg ml−1 solution in 250 mM sucrose, yielding a final protein concentration of 0.5 mg ml−1). The final volume of each reaction mixture was 1.2 ml. Negative control reactions were prepared by incubating mixtures that excluded either microsomes or SQ109 from the mixture. For negative control incubation where microsomes were excluded, they were added back to the reaction mixture after quenching with acetonitrile. Samples were removed at 0, 10, 20, 40 or 80 min and vortex-mixed with cold acetonitrile to stop the reaction. After centrifugation, a portion of each resulting supernatant was analyzed by mass spectrometry for unchanged SQ109.\n

---

\n\nMetabolism of SQ109 by cDNA-expressed recombinant human CYPs [3]
\nSQ109 metabolism was also evaluated in microsomes prepared from insect cells transfected with cDNAs encoding for human CYP1A2, CYP2A6, CYP3A4, CYP2B6, CYP2C8, CYP2C9, CYP2C19 or CYP2D6. The recombinant enzymes and microsomes from untransfected insect cells were used in parallel as a control. SQ109 (10 μM) was preincubated in duplicate with the above-mentioned NADPH-generating system for 5 min at 37°C. The reactions were then initiated by the addition of the individual CYPs (final 100 pmol CYP ml−1) or corresponding untransfected cells. Samples were mixed by inversion, removed at 0 and 30 min and mixed with ice-cold acetonitrile to stop the reactions. After centrifugation (14,000 × g for 20 min at 4°C), each extract was analyzed by using the mass spectrometry to monitor metabolite formation or SQ109 depletion. Electrospray ionization (ESI) full mass scans were performed to obtain the ion chromatograms of the expected metabolites according to predicted mass gains and losses as compared with the molecular mass of SQ109. The ESI as a gentle ionization technique is preferred in metabolite analysis, since ESI usually does not dissociate compounds extensively. The metabolite profiling was based on the detection of protonated, deprotonated or adduct ions, but not on the fragment ions (Kostiainen et al., 2003). Samples were assayed in both the negative and positive ion to ensure detection of all potential metabolite(s), and define the structure(s). Metabolite quantitation was based on percentages of peak areas of each metabolite as a function of incubation time compared to the total area of all chromatographic peaks.\n

---

\n\nRadioactivity determination [3]
\nThe radioactivity of all samples was measured in the Tri-Carb 2100TR liquid scintillation analyzer. All counts were converted to absolute radioactivity (d.p.m.) by automatic chemiluminescence and quench correction. Samples having radioactivity (d.p.m.) less than or equal to twice background d.p.m. were considered to be below the limit of quantitation, and therefore the reading was considered zero d.p.m. for calculation purposes. SQ109 equivalents in biological samples were determined by dividing the sample d.p.m. by the specific activity of [14C]SQ109 in d.p.m. per microgram, and expressed in microgram per gram of tissue. [14C]SQ109 equivalents were also expressed as a percentage of [14C]SQ109 amount in organs or tissues over the administered total [14C]SQ109 amount per animal. The radioactivity in rat urine and feces was expressed as a percentage of the administered dose for each time interval and as a cumulative percentage.\n

---

\n\nChemical inhibition studies [3]
\nThe following inhibitors, at the concentrations shown, were incubated with the corresponding CYP isoforms. These concentrations were based on literature information (Parkinson, 1996; Tucker et al., 2001; Bjornsson et al., 2003): furafylline (CYP1A2; 0.1, 1 and 10 μM), quinidine (CYP2D6; 0.5, 1 and 10 μM), ticlopidine (CYP2C19; 5, 20 and 100 μM) (Donahue et al., 1997; Tateishi et al., 1999; Ha-Duong et al., 2001) and troleandomycin (CYP3A; 0.5, 1 and 10 μM). All inhibitors were dissolved in methanol prior to addition to the incubation mixtures. Reaction mixtures containing human cDNA expressed CYP2D6 or CYP2C19 (100 pmol P450 ml−1 reaction mixture), the NADPH-generating system, selective CYP inhibitors and 100 mM potassium phosphate buffer (pH 7.4) were preincubated for 15 min at 37°C. Each reaction was then started by the addition of SQ109 (10 μM) with subsequent mixing of each sample by inversion. The samples were immediately removed and mixed with cold acetonitrile to stop reaction at 0 and 30 min of incubation. SQ109 and its metabolites were identified by the LC/MS/MS method. Peak areas formed were used for quantitative analyses. Control incubation mixtures included mixtures without inhibitors, and mixtures with untransfected insect cell microsomes used as microsomal control, and mixtures that contained methanol instead of inhibitor (methanol control). Quantitative analyses were performed by comparing the peak areas of the inhibition reactions to their respective methanol controls. The total organic solvent content of the in vitro reaction mixtures was less than 2%.

---

T. cruzi squalene synthase (TcSQS) and human squalene synthase (HsSQS) were expressed and purified. Crystals of TcSQS were obtained in the presence of farnesyl-S-thiolodiphosphate (FSPP) and soaked with SQ109. HsSQS crystals complexed with SQ109 were obtained by co-crystallization. X-ray diffraction data were collected and structures were solved by molecular replacement, revealing SQ109 binding to the active sites of both enzymes. [1]

SQ109 (2.5–20 μM) is applied to the LLC-MK2 cells, and they are then incubated for 96 hours at 37°C. To the untreated samples, fresh RPMI 1640 medium containing 10% FBS is added as a control. The MTS/PMS test is used to assess toxicity. Based on its activities against the trypomastigote and intracellular amastigote forms of T.cruzi, SQ109's selectivity index is calculated as the ratio of the parasite's 50% lysing concentration (LC50) or IC50 to the 50% cytotoxic concentration (CC50) of mammalian cells. Every experiment is run in duplicate. Three or more experiments yield the means[1].\n

---

\nIn vitro infection model [2]
\nThe RAW 264.7 (ATCC TIB-71) murine macrophage cell line was seeded overnight at 5 × 105 cells per well in 24-well plates at 37°C and 5% CO2 in Dulbecco's modified Eagle's medium (DMEM) with essential amino acids and glutamine supplemented with 10% heat-inactivated fetal bovine serum (FBS). Log-phase cultures of recombinant M. tuberculosis H37Rv containing the luciferase reporter construct pSMT1 (hsp60 promoter-driven luciferase) (Snewin et al., 1999) were grown in Middlebrook 7H9 supplemented with bovine serum albumin (BSA), dextrose, catalase and 50 μg ml−1 of hygromycin B at 37°C with 5% CO2 from frozen stocks. Mycobacteria were harvested by centrifugation at 2500 × g for 10 min and washed twice with serum-free DMEM before re-suspension in DMEM supplemented with 5% heat-inactivated FBS at 5 × 106 cells ml−1. Macrophages were infected by incubation with the bacteria at a ratio of 10 : 1 (M. tuberculosis: cell) overnight at 37°C; they were then washed three times in Dulbecco's phosphate-buffered saline (PBS, pH 7.4). The infected cells were treated in triplicate with INH, EMB and SQ109 dissolved in DMEM containing 5% FBS at various MIC for 7 days. Macrophage cells were lysed by the addition of 1 ml per well sterile distilled H2O containing 0.1% Triton X-100 with stirring for 2 min. In all, 100 μl lysate was sampled from each well and an equal amount of 1% N-decyl aldehyde in ethanol was added. Luminescence was immediately quantified using a Luminiskan luminometer (Packard) with a dwell-time of 10 s per well to test the activities of each drug on the infected RAW 264.7 cells. The bioluminescence-based assay employs a reporter strain of M. tuberculosis, which endogenously expresses firefly luciferase that catalyzes the substrate to produce luminescence. Therefore, mycobacterial growth inside the infected cells can be estimated based on luminous intensity (Snewin et al., 1999).\n

---

\n\nPlasma protein binding [3]
\nThe percent binding of [14C]SQ109 to mouse, rat, dog and human plasma proteins was determined by using ultracentrifugation (Barre et al., 1985; Boulton et al., 1998). Briefly, spiking solutions of [14C]SQ109 were prepared by diluting the stock [14C]SQ109 (1 mCi of 5.9 mg−1 ml−1) with absolute ethanol to yield spiking solutions containing 5, 25 or 125 μg ml−1 of [14C]SQ109. For each species, a 10.8 ml aliquot of plasma was mixed with 0.22 ml of the appropriate spiking solution of [14C]SQ109 to yield final SQ109 concentrations of 0.1, 0.5 or 2.5 μg ml−1. The plasma mixtures were placed into individual polycarbonate ultracentrifuge tubes and centrifuged at 100,000 × g for 24 h at 4°C.\n [3]
\nAt the end of the centrifugation period, the upper chylomicron layer, middle aqueous layer and lower protein pellet were separated and the volume of each layer was determined. The protein pellets were dissolved in Soluene 350 tissue solubilizer. The radioactivity of duplicate portions of the chylomicron and aqueous layers as well as the solubilized protein layer was determined, and the total amount of radioactivity in each layer was calculated. The percentage of radioactivity in each layer was determined by comparing the amount of radioactivity in each layer with the sum of the total amount of radioactivity in all three postcentrifugation plasma samples. The percent of the total radioactivity in the aqueous layer was considered to represent the unbound fraction of SQ109, while the sum of the radioactivity in the chylomicron and lower (pellet) protein layers was considered to represent the bound fraction of the drug.

---

Trypomastigotes obtained from infected LLC-MK2 cell supernatants were incubated with SQ109 (0.05 to 5 μM) for 24 h at 37°C. Cell lysis was assessed by counting in a Neubauer chamber. Epimastigotes were cultivated in LIT medium with 10% FBS and treated with SQ109 (0.5 to 10 μM) for 120 h at 28°C; cells were counted every 24 h. For intracellular amastigotes, LLC-MK2 cells were infected with trypomastigotes, washed, and after 24 h treated with SQ109 (0.5 to 6 μM) for 96 h. Cells were fixed, stained with Giemsa, and parasites counted. Infection index was calculated as percentage of infected host cells multiplied by average number of amastigotes per infected cell. [1]
Mitochondrial membrane potential was measured using safranine as a probe. Epimastigotes were added to buffer containing succinate and safranine, with or without digitonin, and fluorescence changes were monitored after addition of SQ109 or FCCP. [1]
Hemolysis assay was performed using a 4% suspension of fresh defibrinated human blood treated with SQ109 (10 to 80 μM) for 24 h at 37°C. Absorbance at 540 nm was measured to determine percent hemolysis. [1]
Sterol biosynthesis inhibition was assessed by growing epimastigotes in the absence or presence of 6 μM SQ109 for 96 h. Lipids were extracted, saponified, and analyzed by gas chromatography-mass spectrometry (GC/MS) to identify sterol species. [1]

Mice [2]
\n\\nEight-week-old female C57BL/6 mice are utilized. Twenty days after inoculation, mice are given oral doses of INH (25 mg/kg), ethambutol (EMB) (100 mg/kg), and SQ109 (0.1, 10 and 25 mg/kg). Mice infected but untreated control groups are euthanized either at the start of therapy (early controls) or at the conclusion of the treatment period. Every group consists of six mice. Four weeks after the start of treatment, the mice receive chemotherapy five days a week until they die. Weighing and aseptic removal of the lungs and spleen are performed.Two milliliters of sterile PBS containing 0.05% Tween-80 are used to homogenize the organs. After being diluted ten times in PBS, homogenate samples from distinct tissues are plated on 7H10 agar plates. Before calculating CFU, inoculated dishes are incubated for three weeks at 37°C in room temperature. A logarithmic scale is used to convert viable counts, and readings are adjusted to reflect totals for all organs. The survival rate and the mean number of CFU in mouse organs are used to evaluate the degree of infection and the efficacy of the treatments.\\n

---

\n\\nPharmacokinetic studies [2]
\n\\nMale CD2F1 mice (23–27 g) were administered SQ109 at 3 mg kg−1 (intravenous (i.v.)), or 25 mg kg−1 (p.o.). Four or five mice were anesthetized with isoflurane at the following times after administration of SQ109 to collect blood from the brachial region of each animal: 2, 5, 10, 15 and 30 min and 1, 3, 6, 10 and 24 h after a single i.v. dosing; 5, 15 and 30 min and 1, 2, 4, 6, 10 and 24 h after a single oral dosing. Each blood sample was collected into a tube containing EDTA and centrifuged (2000 × g, 10 min) to separate plasma and red blood cells. To each 200 μl of plasma sample, 10 μl of internal standard solution (10 μg ml−1) was added. SQ109 was then separated and analyzed according to the previously described procedures. Peak area ratios of SQ109 to the internal standard were plotted against theoretical concentrations. Drug concentrations in samples were calculated from the standard calibration curves. Pharmacokinetic parameters were calculated using the computer program WinNonlin (Pharsight Co., Mountain View, CA, U.S.A.), and bioavailability was calculated as (AUCp.o. AUCi.v.−1) × (dosei.v. × dosep.o.−1) × 100%.\\n

---

\n\\nSQ109 levels in vital tissues following multiple dosing for 28 days [2]
\n\\nIn order to determine whether tissue levels of SQ109 correlate with its efficacy in the H37Rv-infected mouse model, and whether SQ109 accumulates in the targeted tissues over a long period of multiple dose administration, SQ109 levels in the lung, spleen, liver, kidney and heart were monitored during the period of multiple dose administration. Briefly, C57BL/6 mice were orally given SQ109 by gavage at 10 mg kg−1 day−1 for 28 days. Groups of five mice each were anesthetized at 1 h after oral administration on days 1, 14 and 28 with CO2/O2. Blood and the five vital tissues were collected. Plasma and tissue homogenates were prepared for analysis using the below-mentioned procedures. Standard curves for SQ109 in different tissue matrices were plotted for quantitative purposes.\\n

---

\n\\nTissue distribution and elimination of SQ109 after a single administration [2]
\n\\nMale CD2F1 mice (25–27 g) were housed in suspended wire metabolism cages in order to collect their urine and feces to determine SQ109 excretion. The mice were fasted overnight before dosing. Water was provided throughout the study. Mice were dosed with SQ109 at either 3 mg kg−1 (i.v.) or 25 mg kg−1 (p.o.). Groups of four mice each were anesthetized with isoflurane at 1, 4 and 10 h after dosing, and blood was collected from the brachial region of each mouse into a tube containing EDTA. Tissues and organs were immediately removed, individually weighed, washed with cold saline and stored at −20°C prior to analysis for levels of SQ109. On the day of analysis, tissues and organs were minced with scissors and homogenized in ice-cold 5 mM ammonium acetate buffer (1 : 5, w : w). Aliquots of the homogenate (200 μl) were extracted as described previously.\\n[2]
\n\\nFor elimination studies with SQ109, groups of four mice resided in metabolism cages, where urine and feces were separated by a cone-shaped device. Pooled urine and feces were cumulatively collected prior to drug administration, and at 4, 8, 24 and 32 h after a single dose (3 mg kg−1, i.v.; 25 mg kg−1, p.o.). Feces were homogenized in 12 volumes by fecal weight of 5 mM ammonium acetate buffer, and 200 μl aliquots were extracted after centrifugation as described previously. Urine samples were diluted 1 : 10 with 5 mM ammonium acetate without further preparation.\\n

---

\n\\nRat and dog pharmacokinetic studies with SQ109 [3]
\n\\nRats with an indwelling jugular vein catheter were used for the pharmacokinetic studies. Rats were given either a single intravenous (i.v.) bolus dose of 1.5 mg kg−1 (9 mg m−2) or an oral dose of 13 mg kg−1 (78 mg m−2) of SQ109 (n=8 per dose group); rats were divided into subgroups consisting of four rats per subgroup. Rat blood (0.7 ml) was withdrawn from the jugular vein catheter at alternating time points from individual rats in each subgroup. Blood samples were collected at 2, 5, 10, 15 and 30 min and 1, 3, 6, 10 and 24 h after a single i.v. administration, or 5, 15 and 30 min and 1, 2, 4, 6, 10 and 24 h after a single oral administration. Each blood sample was centrifuged to separate plasma, which was then stored at −70°C until analysis.\\n [3]
\n\\nBeagle dogs were dosed by gavage at either 3.75 or 15 mg kg−1 (75 or 300 mg m−2), or intravenously at either 0.45 or 4.5 mg kg−1 (9 and 90 mg m−2). Dog blood (0.7 ml) was withdrawn from the jugular vein at 2, 5, 10, 20 and 30 min and 1, 2, 4, 8, 12, 18 and 24 h after a single i.v. administration, or 10, 20 and 30 min and 1, 2, 4, 8, 12, 18 and 24 h after a single oral administration.\\n [3]
\n\\nEach blood sample was collected into a tube containing EDTA and centrifuged (2000 × g, 10 min) to separate plasma and red blood cells. To each 200 μl of plasma sample, 10 μl of internal standard solution (10 μg ml−1) was added. SQ109 was then separated and analyzed by the LC/MS/MS method according to the previously described procedures (Jia et al., 2005b). Peak area ratios of SQ109 to the internal standard were plotted against theoretical concentrations. Drug concentrations in the plasma samples were calculated from the standard calibration curves. Pharmacokinetic parameters were calculated using the computer program WinNonlin, and bioavailability was calculated as (AUCp.o. AUCi.v.−1) × (dosei.v. dosep.o.−1) × 100%.\\n

---

\n\\nTissue distribution and elimination of [14C]SQ109 in rats [3]
\n\\n[14C]SQ109 (5.8 mg ml−1) was diluted 4.4-fold with 0.9% sterile saline to yield a formulation containing 1.3 mg ml−1 SQ109 (225 μCi ml−1). Male Fischer rats (271–289 g) were individually housed in metabolism cages from which urine and feces were cumulatively collected to determine [14C]SQ109 excretion rate. The rats were orally dosed by gavage with 13 mg kg−1 of [14C]SQ109. Rats were killed at 0.5, 5, 10 and 24 h after dosing (n=3 per time point) in order to collect blood, tissues and organs for quantitative analysis. Tissues, intestinal tract contents and feces were homogenized in 10 volumes of water. Duplicate aliquots of whole blood, homogenates of tissues, intestinal contents and feces were digested with tissue solubilizer Soluene 350, and decolorized with 30% hydrogen peroxide to eliminate chemiluminescence. The samples were radioassayed after mixing with glacial acetic acid and the scintillation cocktail. Duplicate aliquots of urine and cage rinses were radioassayed after mixing with scintillation cocktail.\\n [3]
\n\\nPieces of carcasses were digested with 650 ml of 10 N sodium hydroxide, maintained at 37°C for 3 days, and then at room temperature until complete dissolution of the carcasses (∼2 weeks). Quadruplicate aliquots of each dissolved carcass were diluted with water (1 : 20, v v−1); portions of each diluted sample were radioassayed after the addition of an appropriate scintillator. After correction for volume by dilution and volume assayed, the radioactivity expressed as disintegrations per min (d.p.m.) of [14C]SQ109 in each sample was determined.\\n\\n

---

\nIn vivo efficacy study: Female C57BL/6 mice were infected via lateral tail vein injection with Mycobacterium tuberculosis H37Rv (approximately 10⁵ CFU per mouse). Oral treatment with SQ109 (0.1, 10, and 25 mg kg⁻¹), INH (25 mg kg⁻¹), or EMB (100 mg kg⁻¹) was initiated 20 days post-infection. Treatment was administered daily, 5 days per week, for 4 weeks. Control groups of infected but untreated mice were included. At the end of treatment, mice were euthanized, and spleen and lungs were aseptically removed, homogenized, and plated on agar to determine bacterial CFU counts. [2]
\nPharmacokinetic/Tissue distribution study: Male CD2F1 or C57BL/6 mice were administered SQ109 via single intravenous (3 mg kg⁻¹) or oral (25 mg kg⁻¹) dose, or via repeated oral dosing (10 mg kg⁻¹ day⁻¹ for 28 days). Blood and tissues (lung, spleen, kidney, liver, heart) were collected at specified time points post-dose. Tissues were homogenized, and drug concentrations in plasma and tissue homogenates were determined using a validated LC/MS/MS method. [2]

Following a single intravenous or oral administration, SQ109 rapidly reached peak concentrations in the lungs and spleen of mice after entering the systemic circulation, far exceeding its minimum inhibitory concentration (MIC) against Mycobacterium tuberculosis, without significant side effects such as injection site irritation, limited mobility, ruffled fur, ataxia, tremor, convulsions, vomiting, diarrhea, dyspnea, or acute death. After oral administration, the concentration of SQ109 in the respiratory tract remained above the MIC for more than 10 hours (Figure 5). This compound's rapid translocation from the bloodstream to vascularized pulmonary tissue may be related to the presence of an adamantane tricyclic fragment in its chemical structure (Figure 1). Commercially available adamantane-containing antiviral drugs (such as rimantadine and amantadine) combat viral pulmonary pathogens such as influenza A by inhibiting infection initiation and viral assembly (Hay et al., 1985) and blocking ion channels on viral lipid membranes (Griffin et al., 2003). These drugs are specifically distributed in the lungs and have a large steady-state volume of distribution (Vss) (Hoffman et al., 1988). The adamantane structure is likely also a key component of the in vivo and in vitro anti-tuberculosis activity: four of the seven most active compounds in vitro and in vivo contain the adamantane moiety (unpublished observations) [2]. SQ109, like other ethambutol analogs, has a large steady-state volume of distribution (Vss). Its large steady-state volume of distribution (Vss) may also be attributed to the hydrophobic moiety and diamine structure of the compound, which allows the drug to penetrate rapidly into the extravascular space and has good tissue kinetics, especially in the lungs and spleen (Figures 3 and 5). During daily administration, the concentration of SQ109 in the lungs and spleen was consistently higher than the minimum inhibitory concentration (MIC). This finding suggests that oral administration of SQ109 may be beneficial for drug accumulation in these organs in animal models and patients, where mycobacteria replicate during disease. The drug was readily detectable in mouse organs after oral administration of a 25 mg kg⁻¹ dose of SQ109, which clearly indicates that most of the dose was absorbed by tissues. [2] However, pharmacokinetic studies have shown that the oral bioavailability of SQ109 is lower than that shown by its in vivo efficacy and tissue distribution in mycobacterial diseases (Table 2). Reasons for low oral bioavailability of any compound include: [2] Poor oral absorption: At the same dose, the Cmax value of oral SQ109 is about 80 times lower than that of intravenous SQ109, indicating problems with intestinal absorption. Many candidate drugs fail due to intestinal absorption problems because molecules must be able to penetrate cell membranes composed of phospholipid bilayers to cross the intestine and enter the bloodstream. In addition, two findings in this study suggest that the intestinal permeability problem of SQ109 is not serious: (i) the drug is able to cross multiple cell membranes and reach effective concentrations in phagosomes of infected macrophages in vitro (Figure 2); (ii) even at the lowest dose tested in vivo, 0.1 mg kg−1, sufficient drug crosses the intestine and distributes to the lungs and spleen, resulting in concentrations far above the in vitro MIC, thus producing effective antibacterial effects (Table 1). [2]
First-pass effect: After intravenous injection, the concentration of SQ109 in the liver (the main organ of drug metabolism) was lower than that in most tissues (including the brain) (Fig. 5). However, the concentration of SQ109 in the liver of mice after oral administration was higher than that after intravenous administration relative to the concentration of SQ109 in other tissues (Figs. 3 and 5). These data suggest that SQ109 may undergo first-pass metabolism in the liver after oral administration. We have now identified four metabolites after incubating SQ109 with mouse and human liver microsomes and recombinant CYP450 isoenzymes. [2]
SQ109 has a high dissociation constant with blood proteins, which leads to its rapid disappearance from systemic circulation through tissue redistribution. The tissue concentration change of SQ109 after oral administration is consistent with the metabolic process in which the substance first remains in the liver and then redistributes to the lungs, spleen and kidneys (Figs. 3 and 5). [2]
Many drugs, despite significant bioavailability issues, exhibit good or even excellent efficacy against their target pathogens. For example, halofiberone is used to prevent coccidiosis in poultry, treat cryptosporidiasis and Theileria in bovine animals, and treat various human diseases. Its oral bioavailability is extremely low (Stecklair et al., 2001). Similar to SQ109, after oral administration, halofiberone concentrations in target tissues are 50–2000 times higher than in plasma. Most antibiotics do not exert their effects in plasma but rather on specific target tissues, requiring distribution from the central compartment to these tissues (Muller et al., 2004). Recent studies have further demonstrated that target site antibiotic concentrations can differ significantly from corresponding plasma concentrations, and that target site concentrations are more helpful in clinical trial design and are a crucial determinant of clinical efficacy than plasma concentrations (Ryan, 1993; Presant et al., 1994; Joukhadar et al., 2001). In this respect, SQ109 exhibits good tissue distribution characteristics compared to its anti-tuberculosis activity. [2] Interestingly, after 28 days of continuous administration, SQ109 appears to accumulate preferentially in the lungs and spleen. When we compared the steady-state drug concentrations on days 14 and 28 after a single dose with the concentration on day 1, we found that the accumulation of SQ109 in these two tissues was statistically significant, while the concentration of SQ109 in other tissues was slightly increased (Figure 3). When the drug disposition kinetics are first-order, the steady-state tissue concentration should be higher after multiple doses. This suggests that the upregulation or downregulation of SQ109 in tissues may not be related to enzymatic reactions. These results also explain the complex relationship between the SQ109 concentration-time curve and its observed anti-tuberculosis effect (Figures 4 and 5). [2] In summary, SQ109 exhibits antibacterial activity against Mycobacterium tuberculosis H37Rv strains grown in host mouse macrophages in vitro and in vivo against mouse models inoculated with H37Rv. Oral administration of SQ109 for 28 consecutive days significantly reduced the Mycobacterium tuberculosis load in the spleen and lungs of mice. Monitoring of SQ109 levels in key tissues of mice during the 28-day oral administration period showed that the concentration of SQ109 at potential sites of action (e.g., lungs and spleen) was at least 10 times its minimum inhibitory concentration (MIC). SQ109 exhibited a large volume of distribution in various tissues. Despite the low oral bioavailability of SQ109, its target tissue concentration was at least 120 times higher than that in plasma. This study provides important insights into integrating in vitro efficacy parameters into the in vivo pharmacokinetics and pharmacodynamics evaluation of SQ109 and should contribute to future clinical trials of this compound. [2]

---

SQ109 is lipophilic and has low water solubility. Since the properties of intestinal epithelial cell biomembranes are similar across different species, and the absorption process (simple diffusion) is essentially an interaction between a specific drug and the biomembrane (Lin, 1995; Martinez et al., 2002), the absorption time in the test animal species should be comparable. The results of this study support this view, showing that the Tmax (approximately 0.5 hours) of SQ109 is similar in mice, rats, and dogs (Tables 1 and 2). [3] Like other ethambutol analogs, SQ109 has a large steady-state volume of distribution (Vss) in mice (Jia et al., 2005b, 2005c). The same phenomenon was observed in rats and dogs in this study. Although methods that predict Vss based on experimentally determined physicochemical parameters can predict Vss close to the actual value, it must be remembered that the process of drug permeation into a hypothetical homogeneous compartment based on drug or plasma models is called "tissue permeation". This concept is misleading because it does not take into account the uniqueness of different heterogeneous organ systems (Muller et al., 2004). Therefore, it rarely corresponds to actual volumes such as plasma volume, extracellular fluid, or total body fluid volume. Drug distribution can extend to any one or more tissues and body fluids in the body. Furthermore, the binding of drugs to tissue components can be so strong that the steady-state volume of distribution (Vss) can be several times the total body volume (Rowland & Tozer, 1995). Most importantly, the actual target space for anti-infective drugs must be taken into account as the interstitial fluid (Ryan, 1993) [3]. Many drugs are subject to the "first-pass effect" when they are absorbed from the gastrointestinal tract into the systemic circulation. The organs responsible for producing this effect (e.g., intestine, liver, and lung) are arranged in order and may reduce the bioavailability of the drug. The contribution of each organ can be indirectly assessed by comparing the AUC values obtained from different routes of administration (Pang & Gillette, 1978). Based on the tissue concentration-time data of SQ109 (Figure 3), it can be concluded that SQ109 can easily penetrate the biomembrane of intestinal epithelial cells due to its relatively rapid Tmax (approximately 0.5 hours). The tissue concentration-time data also indicate that the liver plays an important role in the clearance and metabolism of SQ109. This finding is based on the following data: (1) the highest concentration of [14C]SQ109 was found in the liver (excluding the [14C]SQ109 concentration in the gastrointestinal tract); (2) the concentration of [14C]SQ109 in the liver was at least 200 times higher than that in whole blood; and (3) direct determination of SQ109 itself using LC/MS/MS showed that the concentration of SQ109 in the liver was significantly lower than that in the lungs, spleen, kidneys, and heart (Jia et al., 2005b). However, radiometric assays showed that the liver had the highest content of radioactive material. This difference can be explained by the fact that the LC/MS/MS method was developed to detect the parent molecule of the drug with high specificity, but could not detect any metabolites. Radiometric assays can detect all components containing ¹⁴C, including the parent compound and any corresponding metabolites of SQ109. These data suggest that SQ109 undergoes first-pass metabolism in the liver before entering systemic circulation. [3] Several factors may affect the clearance (CL) of a particular drug, including plasma protein binding, metabolism, and hepatic blood flow. For drugs with high clearance, hepatic blood flow limits their clearance (Boxenbaum, 1980). This pattern may explain the species-specific differences in SQ109 clearance. Although there are slight differences in the metabolism of SQ109 by hepatic microsomes among different species (Figure 4), the clearance (CL) of SQ109 in dogs is four times that in mice and rats, which may be due to the higher hepatic blood flow in dogs (Davies & Morris, 1993). From a kinetic perspective, drug distribution is generally defined as the ratio of the total amount of drug in the body to the plasma or blood concentration. SQ109 has relatively low binding to plasma proteins (Table 5), but high binding to tissue proteins (Jia et al., 2005b) and high biomembrane permeability. Therefore, its volume of distribution is very large across different species (Table 3). The half-life of a drug is directly proportional to its volume of distribution, but inversely proportional to its clearance rate. The results of this study support this theory, showing that the half-life (t1/2) of SQ109 in dogs is longer than that in mice and rats (Table 3) [3]. In this study, we used ultracentrifugation to determine the binding rate of plasma proteins, which avoids nonspecific membrane binding compared to other plasma protein binding techniques (Barre et al., 1985). To determine the binding capacity and affinity of plasma proteins for drugs, it is recommended to test at least three drug concentrations (e.g., 0.1, 0.5, and 2.5 μg ml−1) (Kariv et al., 2001; Jia et al., 2003) to elucidate the degree of binding as a function of concentration. As shown in Table 5, a 5-fold and 25-fold increase in total [14C]SQ109 concentration resulted only in a slight increase in the [14C]SQ109 binding fraction. This result suggests that SQ109 has a low binding affinity but a high binding capacity, as the proportion of bound SQ109 remained relatively constant over the 25-fold concentration range and was independent of drug concentration. [3] Due to the first-pass effect, the oral bioavailability of SQ109 in the tested animal species was relatively low (4%–12%; Table 3), which may be related to the high hepatic clearance of SQ109. Using hepatic microsome and CYP response phenotype analysis from different species, we found that SQ109 was rapidly metabolized after incubation with hepatic microsomes (Figure 4), although the metabolic rate varied from species to species. Hepatic microsome-induced SQ109 metabolism was again observed when SQ109 was incubated with a single CYP2D6 or CYP2C19 isoenzyme. In addition, selective inhibitors of CYP2D6 and CYP2C19 inhibited drug metabolism in a dose-dependent manner (Figures 6 and 7). Therefore, both CYP2D6 and CYP2C19 can be identified as major CYP enzymes catalyzing the metabolism of SQ109. CYP2D6 and CYP2C are involved in approximately 25% and 15% of drug biotransformation, respectively (Parkinson, 1996; Schlichting et al., 2000; Tucker et al., 2001). Although the low oral bioavailability of many drugs can be attributed to the extensive metabolism of CYP3A4, this study shows that CYP3A4 has little effect on the metabolism of SQ109. [3] Based on the P450 response phenotype study, we proposed an in vitro metabolic pathway for SQ109, as shown in Figure 5. N-nitroso-SQ109 (M1) can be generated by metabolism catalyzed by CYP2D6 and CYP2C19. The SQ109 molecule contains two amino groups, and the site most susceptible to electrophilic attack by -NO is likely the second amino group, namely the gerany-substituted amino group. The bioreactivity and function of bionitrosation remain controversial (Williams, 1988; Jia et al., 1996). Possible metabolic pathways for SQ109 include the addition of an oxygen atom to the molecule, resulting in a metabolite M2 with m/z 347. One such metabolite (M2-1, m/z 347) may be produced via the oxidation of the adamantane ring, similar to the metabolic pathway of the commercially available adamantane-containing antiviral drug rimantadine. Another monooxygen SQ109 metabolite (M2-2, m/z 347) may be formed via the epoxidation of the N-allyl double bond in the gerany moiety, generating a corresponding epoxide, which may further undergo thermal rearrangement to ultimately produce ketodiamine SQ109 (M2-2). N-(2-adamantyl)ethylenediamine (M3, m/z 195) may be formed by the cleavage of the parent compound, first by hydroxylation of the active carbon atom in the geranyl fragment, followed by N-dealkylation of the unstable intermediate. The formation of M3 was confirmed by a single mass spectrometry peak (not shown). Due to the different positions of the two oxygen atoms added to SQ109, there are multiple pathways for the formation of M4, but their structures may be the same, both with m/z 363. Subsequent oxidation reactions, such as epoxidation and N-hydroxylation (M4-1, m/z 363) and/or epoxidation and epoxidation (M4-2, m/z 363), are likely to occur, which ultimately produce the corresponding hydroxy metabolites. [3] In metabolic events occurring in SQ109, such as C-oxidation and N-hydroxylation (Fig. 5), oxygen from the iron (FeO)3+ of CYP450 may be incorporated into SQ109, while SQ109 itself remains intact. In the case of N-dealkylation, SQ109 may undergo thermal rearrangement after oxidation, resulting in the cleavage of the N-adamantane (M3) structure in the remaining aliphatic structure, while oxygen in the (FeO)3+ complex is incorporated into the aliphatic structure. However, the actual metabolic pathway and the role of the final metabolites in vivo still need to be further studied and verified using real metabolites. [3] In summary, this study systematically demonstrated the differences and similarities of the ADME characteristics of SQ109 among different species, as well as its large steady-state volume of distribution (Vss) and relatively low oral bioavailability. The highest level of radioactivity was observed in the liver after oral administration of [14C]SQ109. This may indicate that SQ109 has a significant first-pass effect, as evidenced by the rapid metabolism of SQ109 in liver microsomes, which is mainly catalyzed by CYP2D6 and CYP2C19. Fecal and urinary excretion accounted for 22.2% and 5.6% of the total dose of [14C]SQ109, respectively. These results, along with our previous findings, do not guarantee that SQ109 will be a successful antibiotic. Like any drug, long-term use of SQ109 may lead to toxicity. Nevertheless, it is clearly a promising new anti-tuberculosis drug, having passed several preclinical trials (Lee et al., 2003; Jia et al., 2005a, 2005b, 2005c). Following a single intravenous injection (3 mg kg⁻¹) in mice, the peak plasma concentration (Cmax) of SQ109 was 1038 ng mL⁻¹, the elimination half-life (t₁/₂) was 3.5 h, and the volume of distribution (Vd) was 11,826 mL kg⁻¹. Following a single oral administration (25 mg kg⁻¹), the Cmax was 135 ng mL⁻¹, the time to peak concentration (Tmax) was 0.31 h, and the elimination half-life was 5.2 h. Oral bioavailability is low, approximately 4%. [2]
SQ109 exhibits extensive tissue distribution. Following a single intravenous or oral administration, the highest drug concentrations were observed in the lungs, followed by the spleen and kidneys. Lung drug concentrations were at least 25 times (intravenous) and 120 times (oral) higher than the corresponding plasma concentrations. [2]
During repeated oral administration over 28 days (10 mg kg⁻¹ day⁻¹),SQ109 accumulated in target tissues, particularly in the lungs and spleen. On day 28, drug concentrations in the lungs and spleen were approximately 6108 ng g⁻¹ and 7596 ng g⁻¹, respectively, at least 10 times higher than their in vitro MIC and at least 45 times higher than plasma concentrations. [2]
Excretion of unmetabolizedSQ109 was minimal. Following intravenous administration, less than 0.04% of the dose was excreted unchanged in the urine within 32 hours, and up to 1% was excreted in the feces. Following oral administration, less than 0.01% of the dose was excreted unchanged in the urine, and approximately 2% was excreted in the feces. [2]

SQ109 exhibits limited toxicity to LLC-MK2 mammalian cells, with a CC50 of approximately 9 ± 2.5 μM. It also shows low hemolytic activity against human erythrocytes, with an EC50 > 80 μM, resulting in a selectivity index > 1600 relative to trypanosome activity. [1]

[1]. SQ109, a new drug lead for Chagas disease. Antimicrob Agents Chemother. 2015 Apr;59(4):1950-61.

[2]. Pharmacodynamics and pharmacokinetics of SQ109, a new diamine-based antitubercular drug. Br J Pharmacol. 2005 Jan;144(1):80-7

[3]. Interspecies pharmacokinetics and in vitro metabolism of SQ109. Br J Pharmacol. 2006 Mar;147(5):476-85.

SQ-109 is an orally effective small-molecule antibiotic used to treat pulmonary tuberculosis. Currently in Phase I clinical trials, SQ-109 is expected to replace one or more drugs in current first-line anti-tuberculosis regimens, simplifying treatment and shortening the course of tuberculosis treatment. The antibiotic SQ-109 is an orally effective acid-resistant diamine antibiotic with potential antibacterial activity against a variety of bacteria, including Helicobacter pylori and Mycobacterium tuberculosis. As an ethambutol analog with an asymmetric structure, SQ-109 targets a different site than ethambutol. However, this drug interferes with cell wall synthesis, leading to weakened cell walls and ultimately cell lysis. Drug Indications It has been studied for the treatment of bacterial infections, infectious and parasitic diseases (not specified), and tuberculosis.

---

Mechanism of Action
The mechanism of action of SQ109 differs from other antibiotics used to treat tuberculosis. It inhibits cell wall synthesis and acts on multiple cellular pathways of a specific microbiome.
The pharmacological potency of antimicrobial drugs depends on the concentration of the drug in vivo, and more specifically, on the concentration achieved at the site of bacterial infection. In this study, we assessed the changes in SQ109 concentration over time using pharmacokinetic methods and compared these results with the static relationship between infection site concentration, activity intensity quantified by pharmacodynamic analysis, and efficacy against proliferating Mycobacterium tuberculosis.
We first validated the efficacy of SQ109 against Mycobacterium tuberculosis in an in vitro macrophage model, followed by further testing in an in vivo animal model. Both models were infected with the same Mycobacterium tuberculosis H37Rv strain. SQ109 was able to penetrate macrophage phagosomes (the site of Mycobacterium tuberculosis replication), thereby inhibiting bacteria that typically reside within mammalian host macrophages.
In this respect, the activity of SQ109 was comparable to that of isoniazid (INH) in the macrophage assay system, but superior to ethambutol (EMB) (Figure 2). SQ109 has been demonstrated to have in vitro activity and also exhibits dose-dependent anti-tuberculosis activity in vivo, reducing bacterial load in liver and spleen homogenates by 1–1.9 log units over 28 days as monitored by routine colony-forming unit (CFU) levels. In reducing mycobacterial load in the spleen and lungs, isoniazid (INH, 25 mg kg⁻¹ day⁻¹) was more potent than SQ109 (0.1, 10, and 25 mg kg⁻¹ day⁻¹) and ethambutol (EMB, 100 mg kg⁻¹ day⁻¹) (Table 1). The differences in the in vitro and in vivo activities of SQ109, INH, and EMB are currently unclear, but likely reflect differences in their molecular targets and pharmacokinetic properties. [2] SQ109 is an ethylenediamine derivative containing N-geranyl and N-adamantyl. It is currently in Phase II clinical trials for drug-sensitive and drug-resistant tuberculosis. Its main mechanism of action against Trypanosoma cruzi is believed to be uncoupling of mitochondrial membrane potential and disruption of pH gradient, similar to its mechanism of action in bacteria. The drug can also bind to squalene synthase and inhibit sterol side-chain reduction, but these effects are unlikely to explain its rapid antiparasitic activity. The drug has a synergistic effect with posaconazole, which may be achieved through combined effects on sterol biosynthesis and cation homeostasis. [1]

C22H38N2

330.55

330.303

C, 79.94; H, 11.59; N, 8.47

502487-67-4

5274428

Light yellow to yellow oily liquid

0.97±0.1 g/cm3

5.464

2

2

9

24

431

0

C/C(C)=C/CC/C(C)=C/CNCCNC1[C@H]2C[C@H]3C[C@@H]1C[C@H](C3)C2

JFIBVDBTCDTBRH-WUROFCERSA-N

InChI=1S/C22H38N2/c1-16(2)5-4-6-17(3)7-8-23-9-10-24-22-20-12-18-11-19(14-20)15-21(22)13-18/h5,7,18-24H,4,6,8-15H2,1-3H3/b17-7+/t18-,19+,20-,21?,22?

N1-((1r,5R,7S)-adamantan-2-yl)-N2-((E)-3,7-dimethylocta-2,6-dien-1-yl)ethane-1,2-diamine

SQ109; SQ-109; SQ 109; NSC 722041; 502487-67-4; N-Geranyl-N'-(2-adamantyl)ethane-1,2-diamine; N'-(2-adamantyl)-N-[(2E)-3,7-dimethylocta-2,6-dienyl]ethane-1,2-diamine; 9AU7XUV31A; CHEMBL561057; NSC-722041; NSC722041.

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ≥ ~25 mg/mL (~75.63 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (7.56 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

---

Solubility in Formulation 2: ≥ 2.5 mg/mL (7.56 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

---

View More

Solubility in Formulation 3: ≥ 2.5 mg/mL (7.56 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

---

 (Please use freshly prepared in vivo formulations for optimal results.)