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Corona Virus Disease 2019 (COVID-19) has spread all over the world and brings significantly negative effects on human health. To fight against COVID-19 in a more efficient way, drug-drug or drug-herb combinations are frequently used in clinical settings. The concomitant use of multiple medications may trigger clinically relevant drug/herb-drug interactions. This study aims to assay the inhibitory potentials of Qingfei Paidu decoction (QPD, a Chinese medicine compound formula recommended for combating COVID-19 in China) against human drug-metabolizing enzymes and to assess the pharmacokinetic interactions in vivo. The results demonstrated that QPD dose-dependently inhibited CYPs1A, 2A6, 2C8, 2C9, 2C19, 2D6 and 2E1 but inhibited CYP3A in a time- and NADPH-dependent manner. In vivo test showed that QPD prolonged the half-life of lopinavir (a CYP3A substrate-drug) by 1.40-fold and increased the AUC of lopinavir by 2.04-fold, when QPD (6 g/kg) was co-administrated with lopinavir (160 mg/kg) to rats. Further investigation revealed that Fructus Aurantii Immaturus (Zhishi) in QPD caused significant loss of CYP3A activity in NADPH-generating system. Collectively, our findings revealed that QPD potently inactivated CYP3A and significantly modulated the pharmacokinetics of CYP3A substrate-drugs, which would be very helpful for the patients and clinicians to avoid potential drug-interaction risks in COVID-19 treatment., Graphical abstract Image 1, Highlights 1. Qingfei Paidu decoction (QPD) dose-dependently inhibits CYPs1A, 2A6, 2C8, 2C9, 2C19, 2D6 and 2E1 in a reversible manner. 2. QPD inhibits CYP3A in a time- and NADPH-dependent manner in liver microsomes from both humans and rats. 3. QPD significantly increases the plasma exposure and half-life of lopinavir in rats. 4. Fructus Aurantii Immaturus (Zhishi) in QPD plays a crucial role in CYP3A inactivation.

Quebec platelet disorder (QPD) is a bleeding disorder characterized by a gain of function defect in fibrinolysis. The hallmark feature of QPD is the marked overexpression of urokinase plasminogen activator (uPA) in megakaryocytes (MK) and platelets. The genetic cause of QPD is a tandem duplication of a ~78 kb region that encompasses the uPA gene, PLAU. As the mechanism of PLAU overexpression is unknown, gene regulatory mechanisms specifically epigenetics were evaluated at the PLAU locus in QPD MK and granulocytes, a QPD unaffected lineage. The aims of the thesis were to assess if QPD is associated with 1) genome wide methylation changes of promoter CpG islands, particularly at PLAU and 2) genome wide changes of active histone modifications H3K27Ac, H3K36me3 and H3K4me2, particularly at the region of PLAU duplication. Methylation and active histone enrichment analysis revealed that in QPD and control subjects, PLAU promoter CpG island was characterized by unaltered hypo-methylation and changes in active histone peak enrichments that were within the realm of having one extra copy of PLAU in both MK and granulocytes. The findings imply that the PLAU CNV mutation does not induce altered promoter methylation status and/or significantly alter active histone markers as the reason for the marked PLAU overexpression in QPD MK. Instead, the rearrangement of an active enhancer element, particularly an H3K27Ac enhancer expressed in MK but not granulocytes, that is upstream of the second copy of PLAU might underlie the marked PLAU expression by differentiated QPD MK. The thesis provides novel insights into the epigenetic regulation of PLAU that will be crucial to identifying the mechanism underlying the aberrant PLAU expression in QPD. Thesis Master of Science (MSc)

Inherited bleeding disorders can be caused by mutations affecting platelet, coagulation, or fibrinolytic proteins. Quebec platelet disorder (QPD) is a rare, autosomal dominant disorder associated with increased expression of the fibrinolytic enzyme urokinase plasminogen activator (uPA) in platelets. Individuals with QPD experience delayed-onset bleeding after hemostatic challenges that is attenuated with fibrinolytic inhibitor therapy. The aims of this thesis were to: 1) determine if increased platelet uPA contributes to QPD clot lysis in vitro; 2) investigate whether QPD individuals have increased urinary uPA, as some individuals experience hematuria; and 3) map the genetic locus of QPD, and look for the putative mutation. Studies of clot lysis indicated that QPD platelets induce a gain-of-function defect in fibrinolysis when platelets are incorporated into clots. This suggests that accelerated fibrinolysis may contribute to QPD bleeding. Studies of urinary uPA in QPD showed that uPA is not increased, indicating that hematuria in QPD is likely a consequence of increased platelet uPA. This finding also suggests that uPA overexpression in QPD may be megakaryocyte-specific. Linkage studies showed that QPD is strongly linked to a 2 megabase region on chromosome 10 that harbors the uPA gene, PLA U. No mutations in PLA U or its regulatory regions were identified; however, a common haplotype for a 32.5 kilobase region around PLA U, including inheritance of a rare, linked polymorphism, suggests this is the most likely locus for QPD. mRNA studies in QPD platelets showed that QPD selectively increases expression of the linked PLAU allele, without similar increases in megakaryocyte progenitors or in saliva. These findings implicate a cis-mutation near PLA U as the cause of QPD. This thesis provides novel insights on the fibrinolytic abnormality in QPD blood, and on the QPD genetic locus. which will be important for identifying the precise mutation that converts normally prohemostatic platelets to profibrinolytic cells. Thesis Doctor of Philosophy (PhD)