The endolysosome system is a dynamic interconnected network with morphological and functional heterogeneity, and this system exhibits complex interactions with other organelles. Extracellular macromolecules and membrane components are up-taken into endosomes by a variety of endocytic pathways and can either be trafficked through early endosomes to recycling endosomes, which mediates receptor recycling to plasma membrane or Golgi apparatus, or can transition to late endosomes and fusion with lysosomes [26, [31] [32] [33] . As mentioned above, a hallmark feature of the endolysosome system is their acidic luminal pH [25] [26] [27] [28] that is critical for the activity of up to 60 different pH sensitive hydrolytic enzymes including proteases, lipases and nucleases [33] .
Early endosomes act as a major sorting station and their acidic nature enables internalized ligands to be dissociated from the endocytosed cell surface membrane receptors to which they were bound. The receptors unbound with ligand can then be recycled back to the cell surface or can traffic to Golgi apparatus via recycling endosomes; dissociated ligands are transported through late endosomes to lysosomes for degradation. The maturation of early endosomes to late endosomes is characterized by an increased number of intralumenal vesicles and the formation of multivesicular through the multidrug resistance protein p-glycoprotein [97] [98] [99] .Through their ability to deacidify acidic organelles along endocytic and biosynthetic secretory pathways, CQ and HCQ disturb many key aspects of cell biology including organellar biology and interorganellar signaling, many of which are linked to their antiviral and immunosuppressive effects. Decades of clinical usage have shown that CQ and HCQ are relatively safe drugs. They are even commonly used during pregnancy in patients with autoimmune disorders. Recent systematic reviews and meta-analyses suggest that maternal HCQ use during pregnancy does not increase risk of major congenital malformations [100, 101] . The reported tissue side effects of CQ and HCQ such as retinopathy [102, 103] , cardiomyopathy [104] , neuromyotoxicity [105] are likely due to abnormal accumulation of these drugs in tissues with chronic use [12, 106] . The unexpected high rate of side effects including cardiac arrhythmias during the COVID-19 pandemic may be related to higher doses of HCQ used, the older ages of the patients, and to drug-drug interactions 
Other enveloped viruses like influenza A and Ebola, also use the acidic environment of endosomes or endolysosome hydrolases to drive the fusion of viral membranes with endosome membranes and the release of viral genomic content into the cytoplasm [121] . As such, endolysosome de-acidification with a v-ATPase inhibitor [122] or CQ [123, 124] has been used frequently to inhibit cellular entry of enveloped virus in vitro. In the context of COVID-19, CQ and HCQ are often used in combination with antivirals and other drugs such as azithromycin and zinc [130, 131] . By de-acidifying acidic organelles, CQ and CQ could alter the cellular distribution of these drugs. Azithromycin, a macrolide antibiotic, has been used against Zika [132] and Ebola viruses [133] . In vitro evidence indicates that HCQ and azithromycin show synergistic effects on inhibiting SARS-CoV-2 [130] . As a weak base, azithromycin (pKa = 8.5) also tends to accumulate in acidic environments [134] . By de-acidifying acidic organelles, CQ and HCQ could prevent the accumulation of azithromycin in acidic organelles and increase its concentrations in cytosol where azithromycin exerts inhibitory effects on viral replication [130] . On the other hand, such CQ-and HCQ-induced cellular redistributions of azithromycin may exaggerate its QT prolongation effects [135] as observed in the context of COVID-19 [136] . In vitro evidence indicates that zinc inhibits SARS-CoV [137] .
In a preprint paper [131] , the authors reason that CQ, as a zinc ionophore [138] , could increase cytosolic concentrations of zinc to inhibit RNA-dependent RNA polymerase. This is contrary to the original observation that CQ is a zinc ionophore [138] , in which chloroquine was observed as a zinc ionophore that targets zinc to lysosomes. Nonetheless, zinc has been shown to inhibit the activity of furin [139] and cathepsins [140] . Interestingly, the activity of furin is also pH-dependent; furin's optimum pH is 6.0 [141] . Thus, in addition to its de-acidifying effects, CQ-induced accumulation of zinc in J o u r n a l P r e -p r o o f endolysosomes could further inhibit activities of furin and cathepsins that are responsible for cleavage of SARS-CoV spike proteins and viral entry.
De-acidifying acidic organelles may not be the only mechanism whereby CQ and HCQ interfere with the biology of SARS-CoV2. In human epithelial lung cells, CQ inhibits the phosphorylation of p38 mitogen-activated protein kinase (MAPK) and inhibits the release of human coronavirus from infected cells [142] . Thus, CQ and HCQ may exert its effects on SARS-CoV-2 via altering protein kinase activities.
Concerns about the possible antiviral effects of CQ and HCQ: Currently, there exist very limited evidence supporting the use of CQ and HCQ against SARS-CoV-2 and COVID- 19 . First, almost all in vitro studies reported to date were conducted in epithelioid cells derived from the kidney of African green monkeys (Vero E6 cells) or human hepatomas (Huh7 cells) [129, 143, 144] . Vero E6 cells are extremely permissiveness for viral replication including coronaviruses, which is due in part to genetic defects in interferon production [145, 146] and defects in innate antiviral responses [147] . Huh7 cells are also highly permissive for virus replication due to defective retinoic acid-inducible gene I signaling, impaired interferon signaling, and defects in innate antiviral responses [148, 149] . Thus, it is not clear how finding from these cells are translatable to human clinical trials. Second, CQ and HCQ change the pH of endolysosomes and Golgi and cause multiple morphological and functional changes to these organelles including swelling and altered exocytotic release of virus. These changes may result in the formation of intracellular SARS-CoV-2 reservoirs capable of being re-activated when pH normalizes.
Most recently (June 23, 2020), several clinical studies have reported on the favorable use of CQ and HCQ for the treatment of COVID-19 [150] [151] [152] [153] [154] [155] [156] [157] [158] [159] . However, other studies have reported no significant beneficial effects and some detrimental effects [160] [161] [162] .
Multiple clinical trials testing the possible effectiveness of CQ and HCQ against COVID-19 are ongoing [163] , the results of these studies will be important as more people require treatments for COVID-19.
The immunomodulatory properties of CQ and HCQ have long been recognized and these drugs continue to be used clinically for the treatment of rheumatoid arthritis, systemic lupus erythematosus and other inflammatory rheumatic diseases [164] .
Although their mechanisms of action remain under investigation, the immunomodulatory effects are likely due to their accumulation in and de-acidification of acidic compartments; endolysosomes and Golgi apparatus of immune cells. The importance of CQ-and HCQ-induced de-acidification as an important mediator in immune modulation may be due to impaired maturation of lysosomes and blockade of fusions between autophagosomes and lysosomes. Indeed, lysosomal degradation of endocytosed or autophagocytosed proteins affects antigen processing and MHC class II presentation [165, 166] . This is consistent with findings that CQ and HCQ inhibit MHC class II expression, antigen presentation and immune activation [167] . Further, RNA and DNA binding to toll-like receptor 7 (TLR7) and TLR9 in endosomes results in TLR signaling activation and production of pro-inflammatory cytokines [168] . Thus, CQ-and HCQ- Membrane bound vesicles in the endocytic pathway (early endosome, recycling endosome, late endosome, and lysosomes) and the biosynthetic secretory pathway (Golgi apparatus and secretory vesicles) all display varying degrees of acidity, and these vesicles rapidly acidify as they progress along the endocytic or secretory pathway.
As diprotic weak bases, CQ and HCQ are taken up by cells and trapped in these acidic organelles, where they neutralize pH and alter their structure, function, and trafficking. 