Are Basic and Lipophilic Chain Groups Highly Required in Leishmanicidal Quinolines to Favor the Phagolysosome Accumulation?

Angel Romero^*

• Universidad de la República, Montevideo, Uruguay

Phagolysosome is defined as a cytoplasmic body formed through the fusion of a phagosome with a lysosome during the phagocytosis process (Alexander and Vickerman, 1975). The phagolysosome is characterized by having an acidic environment (pH 4.5-5.0) and internal temperature of 37°C. The acidic condition plays an important role for the intracellular destruction of pathogens via enzymatic hydrolytic degradation (Nguyen and Yates, 2021). That body is essential in the survival of Leishmania parasites within the host cell (Zilberstein, 2021).Leishmania is an intracellular parasite that cycles between the midgut of female sandfly vectors and phagolysosomes of mammalian hosts. The infection initiated with the parasite found in the midgut of sandfly under the flagellated promastigote form. Then, parasites are deposited into human skin during the sandfly bloodmeal and then, they are phagocytosed by macrophages until to form the phagolysosome (Zilberstein, 2021). It is documented that the presence of a chemical component like the lipophosphoglycan (LPG) could be essential in the recognition of promastigote parasite by macrophage cells (Desjardins and Descoteaux, 1997;Moradin and Descoteaux, 2012). Once within the phagolysosome, promastigote are differentiated to smaller aflagellated intracellular amastigote form, which is favored as consequence of the extremely harsh environment inside phagolysosome (Chang and Dwyer, 1976;Berman et al., 1979). That parasite stage survives and elude the host defense mechanism within phagolysosome (Chang and Dwyer, 1976;Moradin and Descoteaux, 2012) and then, proliferates by binary cell division and invade other macrophages or phagocytic (i.e. dendritic cells) or no-professional phagocytic (i.e. fibroblasts) cells. To elude the host immune defense, Leishmania parasite developed a mechanism directed to promote change in the macrophage polarization, from a defensive macrophage M1 to an attenuated macrophage M2 (Naderer et al., 2008;Carneiro et al. 2025;Tomiotto-Pellissier, et al., 2018), which allows the survival and proliferation inside phagolysosome. Thus, the phagolysosome emerges as an attractive target for the development of leishmanicidal agent, being essential to design chemical structures that will be able to accumulate into the phagolysosome taking advantage on their internal acidic characteristic and highly lipophilic membrane. With this prelude in hand, the present article seeks to show the role of some physicochemical properties [e.g. ionization constant (pKa) and lipophilicity (Log P)] to favor the accumulation of quinoline systems into lysosome and then, to correlate these parameters with the in vitro leishmanicidal response against intracellular amastigote. From a physicochemical point of view, it is possible to predict the ability of a quinoline or any type of compound to accumulate into phagolysosome and/or analogue organelle (e.g. lysosome). S. Trapp and co-workers demonstrated, in general terms, the importance of the basicity to accumulate into the lysosome (Trapp et al., 2008). They predicted the accumulation of organic compounds in the cell studying the diffusion from the external solution to the cell organelle (e.g. cytosol, lysosome or mitochondria) using the Fick-Nernst-Planck-equation. To the best of the present analysis, most of the studied compounds were based on quinolines including amodiaquine, chloroquine, quinine, mefloquine, primacrine, quinidine and quinacrine. The rest of the tested compounds included cycloguanil, artemesinine, halofantrine and pyrimethamine. From the study, a high and selective accumulation in lysosome was found for weak mono-and bivalent bases having from intermediate to high values of Log KOW. Authors proposed that the selective accumulation into lysosomes over other organelles (e.g. cytosol or mitochondria) can be mediated by an "ion trapping" mechanism, where the protonation of the basic moiety captures the compounds forming a more hydrophilic specie whose outer diffusion is minimized. Physico-chemical properties like ionization constant (pKa) and equilibrium constant (Log KOW) are keys to understand the accumulation via "ion trapping". For monovalent weak bases, the optimal parameter for good lysosome accumulation consists of pKa and Log KOW values between 6 and 10 and between 0 and 3, respectively. An optimal accumulation was corresponded for bases with a pKa of 8. For bivalent bases, the optimal pKa2 (aliphatic amine) value was between 8 and 10 and a pKa1 value between 4 and 8, whereas the optimal Log KOW value was found between 3 and 6. Neutral compounds (e.g. artemesinine) showed a negligible accumulation into lysosome. Regarding the lipophilicity parameter, Marceau identified the correlation between the Log P and the optimal accumulation into lysosome (Marceau et al., 2012). Based on a series of cationic triethylamine derivatives including triethanolamine, procainamide, triethylamine, lidocaine, imatinib, chloroquine, astenizole, quinacrine dronedarone and amiodarone, which displayed pKa values between 8 and 10, compounds having a Log P value between 1 and 4 showed the highest accumulation in lysosome, whereas a decrease in the lysosome accumulation was found with the increase of the Log P for values higher than 4. Thus, the combination and appropriate control of these two variables could be pivotal in the rational design of leishmanicidal agents, whose goal is to guarantee the quinoline accumulation within the phagolysosome. Then, the incorporation of lipophilic group, that provide to the molecule a general Log P ~ 1-4, seeks to facilitate the penetration of the quinoline drug through the lipophilic phagolysosome membrane, whereas the incorporation of basic moiety, that provide to the molecule pKa ~ 4-10 (pKa1 and pKa2 correspond to the quinolinic and alkyl-amine chain), seeks to guarantee the accumulation inside the lysosome through an "ion trapping" mechanism, which consisted of the capture of the molecule by the generation of a polar and cationic form that is derived from the protonation of the basic moieties under acidic environment as depicted in Scheme 1. Scheme 1. Tentative mechanism of accumulation of quinoline inside phagolysosome via lipophilic penetration and subsequent ion trapping mechanism through protonation of basic moieties under acidic environment. ), it should be noted that most of the potent and selective 4aminoquinolines (e.g. compounds 1-6) are characterized by incorporating in their structures either a basic group (e.g. tertiary dialkylamine, N-heteroarene) or a lipophilic group (e.g. aryl, alkyl chain) that disclose appropriate or acceptable Log P and pKa parameters. For example, the most selective compound (compound 1), that displayed an IC50 value of 0.023 µM against amastigote of L. donovani and a selectivity index (S.I.) of 1739, showed an optimal pKa2 value of 8.32 and a discretely high Log P value of 5.29. The compound 2, that displayed a high antiamastigote response against L. donovani (IC50 = 0.36 µM) and selectivity (S.I.˃ 1111), showed an optimal Log P of 4.10 and an appropriate pKa2 of 3.98. Meanwhile the compound 3, that presented appropriate values of Log P (4.15) and pKa2 (9.78), exhibited a high leishmanicidal response and selectivity against amastigote models of L. amazonensis (IC50 = 0.34 µM; S.I.=145) and L. infantum (IC50 = 0.18 µM; S.I.= 392). Other interesting example is the compound 4, which showed moderate potency (IC50 = 5.48 µM) and selectivity (S.I. =41) against amastigote of L. amazonensis, which could be more correlated with its lower lipophilicity (Log P= 3.31) than with the basicity because it presented an optimal pKa2 (6.61). Compound 5, that was identified as the most potent metallic complexes quinoline-based by their excellent in vitro (IC50 = 0.5 vs. L. donovani amastigote) and in vivo responses (Del Carpio et al. 2025), displayed appropriate Log P and pKa2 values of 4.10 and 3.68, respectively. Finally, within the analyzed 4-aminoquinolines (1-6), the compound 6 (Log P= 6.58; pKa2= 9.17) generated a highly potent but highly toxic, which could be associated with its extremely high lipophilicity. In general, based on examples of 4aminoquinolines, it is consistently documented that the inclusion either of several basic or extended lipophilic moieties into the quinoline structure compromises the selectivity and leishmanicidal potency of the 4-aminoquinolines (Romero and Delgado, 2025a). Then, it seems that the most active and selective leishmanicidal dibasic 4-aminoquinolines are characterized by having a Log P between 4 and 5.3 and a pKa2 between 4 and 9. Regarding the monobasic 4-aminoquinolines (having a pKa1 ~4 by quinolinic nitrogen), some compounds (e.g. 7, 8, 9, 10 and 11) exhibited good antiamastigote in similar range to the most active 4-aminoquinolines mentioned above, but less selective. Then, the selectivity of these compounds seems to have a correlation with its lipophilic characteristic. For example, the most active monobasic quinoline (compound 7), that showed an excellent potency against L. infantum (IC50 = 0.20 µM) and a good selectivity (S.I.≥ 250), presented a high Log P of 6.75. The second more selective compound of this group (compound 8) (S.I.= 188; IC50 = 0.22 µM vs. L. donovani) (Loiseau, et al., 2022) presented a Log P value of 2.98. For this case, the inclusion of the halogens (F and Cl) as well as the morpholine moiety in the quinoline core could be essential to modulate conveniently the Log P and pKa magnitudes for favoring the accumulation inside macrophage phagolysosome in the infected model. Other potent (IC50 = 1.20 µM vs. L. pifanoi) and selective (S.I. ˃ 42) compound (compound 9) exhibited a high Log P of 5.89. Meanwhile the compounds 10 (Log P= 3.46) and 11 (Log P= 3.65), that showed a Log P lower 4, exhibited more limited leishmanicidal activities and selectivities.Finally, analyzing the leishmanicidal response of antimalarial quinolines (Avanzo et al., 2025), lipophilic amodiaquine (AQ), sitamaquine (SQ), mefloquine and tafenoquine that bears a second basic group displayed the highest antiamastigote response against Leishmania spp. Although it is important to mention that the mefloquine and tafenoquine displayed a high cytotoxicity, which reflects that the selection of the type of lipophilic chain and probably its location in the quinoline ring is not trivial. Then, SQ (Log P =4.30; pKa2 = 10.22) and AQ (Log P =5.29; pKa2 = 7.39) showed to be the most selective and potent antimalarial quinolines against intracellular amastigote, which could be associated with their appropriate pKa2 and Log P values. Further reports have demonstrated that the SQ is able to accumulate in membranous organelles such as acidocalcisomes (López-Martín et al., 2008) and parasite mitochondria (Vercesi and Docampo, 1992;Vercesi et al., 2000). This last supports that the internalization of the SQ into these organelles could be favored by their appropriate pKa2 and Log P parameters. In summary, the present opinion article pretends to introduce the importance of the phagolysosome and the mechanism to favor its drug accumulation as pivotal concepts in the design of potent and selective leishmanicidal agents, applicable not only for the design of quinolines but also for other type of leishmanicidal compound. Among the reported cases, it is possible to infer that the high antiamastigote response and selectivity of the quinoline compounds could be favored by their appropriate Log P and pKa parameters, which seeks facilitate the transmembrane penetration and lysosome accumulation via "ion trapping". It should be noted that the dibasic quinolines tend to generate more active and selective leishmanicidal compound than monobasic and, even more than tri-or polybasic quinolines. Within the dibasic quinolines, mainly based on 4-aminoquinolines, the most potent and selective compounds are characterized by having pKa2 between 6 and 9 and Log P values from 4 to 6. Meanwhile the most promising monobasic quinolines presented, in general, Log P values higher than 3 and lower than 6.5, with some exception such as the compound 8 whose incorporation of halogen and morpholine moieties could be key to modulate conveniently the lipophilicity and basicity into the quinoline for a good penetration and accumulation inside phagolysosome.In general, the ionization and llipophilic parameter requirements to reach a good leishmanicidal response in the quinoline compounds are in good concordance with the Trapp's and Marceau's findings, being only essential a higher lipophilicity (Log P from 4 to 6) than expected for typical lysosome accumulation (Log P between 1 and 4). Probably, the combination of both factors under appropriate magnitudes could be highly required for the design of potent and selective leishmanicidal agents targeted to the macrophage phagolysosome. Although further studies are highly needed either to demonstrate its effective accumulation into the macrophage phagolysosome or to elucidate from a screening study what are the most appropriate parameter magnitudes to achieve a good phagolysosome accumulation and control of the selectivity in the designed quinoline and other type of leishmanicidal agents.- Carneiro, M.B., Vaz, L.G., Afonso, L.C.C., Horta, M.F. and Vieira, L.Q. (2021)