Discovery and Structure–Activity Relationship Studies of Thienopyrroles as Group 2 Influenza A Entry Inhibitors

Abstract

The influenza A virus has been detrimental to public health for centuries and continues to take the lives of tens of thousands of people annually. Although current therapeutics have been effective in the past, the emergence of drug-resistant strains underscores the need for agents with new mechanisms of action. Here, we report thieno­[3,2-b]­pyrroles as novel group 2 influenza A virus entry inhibitors. In this study, we used a HA-based surrogate entry assay to perform the structure–activity relationship (SAR) studies to optimize the lead compounds. Docking studies in the predicted hemagglutinin binding site provided insight into potential binding interactions. Mutagenesis studies supported the proposed binding site. We validated the lead compounds that were potent in pseudovirus against infectious H3N2 strains including an oseltamivir-resistant strain. Lead compounds were counter-screened against other pseudoviruses and found to be selective. Together, these findings establish thieno­[3,2-b]­pyrroles as a promising new scaffold for combating group 2 influenza A viruses.

Influenza A virus (IAV) remains a significant threat to public health, worldwide. With an estimated 290,000 to 650,000 respiratory deaths annually, influenza takes a significant toll on humans, despite the availability of seasonal vaccines.

IAV is an enveloped, negative-sense, single-stranded RNA virus belonging to the Orthomyxoviridae family. The influenza A virus genome consists of eight RNA segments that encode 10 essential proteins and several strain-dependent accessory proteins. These include the polymerase complex (PB1, PB2, and PA), nucleoprotein, matrix proteins (M1 and M2), and surface glycoproteins hemagglutinin (HA) and neuraminidase (NA). There are four general types of influenza: A, B, C, and D. Of these types, A, B, and C are known to infect humans, and A is the only type that has caused pandemics in the past. IAV is further divided into Groups 1 and 2 based on HA antigenic subtypes, described below. ,

Viral entry of IAV begins with HA-facilitated attachment to sialic acid residues on the surface of the host cell, followed by endocytosis and subsequent membrane fusion triggered by the acidification of the endosome. Viral ribonucleoproteins are released into the cytoplasm and imported into the nucleus, where RNA-dependent RNA polymerase-catalyzed transcription occurs. The newly synthesized viral RNAs and proteins are then assembled at the host cell plasma membrane, and progeny virions are released via neuraminidase-facilitated cleavage from the host cell sialic acid residues. ,

Although vaccines remain the primary strategy for preventing influenza, several factors limit their effectiveness. Antigenic drift and shift in the viral surface glycoproteins reduce the match between circulating strains and vaccine formulations. For instance, vaccine effectiveness estimates for the interim 2025–2026 season are only ∼20–30%. Vaccines also provide reduced protection in immunocompromised individuals, who mount weaker immune responses. Moreover, the long time period required to develop vaccines and the difficulty of predicting dominant strains diminish vaccine performance. Historically, there have been four major IAV pandemics (1918, 1957, 1968, and 2009) that have occurred, with the first three resulting in millions of deaths. The challenges associated with vaccines highlight the need for effective influenza antivirals.

Oseltamivir (Tamiflu), the primary influenza therapeutic for more than two decades, inhibits neuraminidase-mediated cleavage of sialic acid residues on the host cell surface, thereby preventing the release of newly formed virions; however, oseltamivir-resistant influenza strains are becoming increasingly prevalent, reducing the effectiveness of the therapeutic. Other approved compounds, such as amantadine and rimantadine, block the M2 ion channel, which normally acidifies the interior of the virion following endocytosis to trigger viral uncoating; however, current M2 ion channel inhibitors are no longer recommended for use due to widespread resistance. The recently approved class of influenza therapeutics comprises the polymerase acidic endonuclease inhibitors, such as baloxavir marboxil (Xofluza). Although baloxavir marboxil was approved in 2018, resistance has already been detected to be around 10% in adults and around 20% in children. − Due to these limitations and alarming rates of resistance of currently approved therapeutics, there is a great need for influenza therapeutics with new mechanisms of action.

An underexplored therapeutic strategy for targeting influenza is blocking its entry into host cells. Attachment and endosomal membrane fusion of influenza virus are mediated by HA, a homotrimeric class 1 fusion glycoprotein. Each HA monomer is composed of two subunits: HA1 and HA2. HA1, also known as the head domain, is the receptor binding domain. HA2, also known as the stalk domain, contains a fusion peptide. After the virion enters the endosome, the subsequent reduction in pH causes a conformational change to occur, exposing the hydrophobic fusion peptide that attaches to the host membrane to facilitate fusion. Group 1 IAV is composed of 12 hemagglutinin subtypes (H1, H2, H5, H6, H8, H9, H11, H12, H13, H16, H17, and H18), and Group 2 is composed of six (H3, H4, H7, H10, H14, and H15). Of these subtypes, the stalk region is more highly conserved than the receptor-binding head region, making the stalk region an attractive target.

We and others have advanced several scaffolds targeting group 2 hemagglutinins in the past. − In this work, we report thieno­[3,2-b]­pyrroles as a new class of group 2 hemagglutinin inhibitors. These compounds are derived from a high-throughput screen of a 10,000-compound ChemDiv chemical library against pseudotyped H7N1. Through structure–activity relationships, we subsequently found the thieno­[3,2-b]­pyrrole scaffold to be a potent, group 2-selective hemagglutinin inhibitor with nanomolar activity. These studies guided further investigation into selectivity, binding, and antiviral activity of the top analogues. Overall, this work describes the discovery and structural optimization of the thienopyrrole scaffold as a targeted inhibitor of group 2 IAV hemagglutinin.

Docking studies were performed to gain insight into the potential binding mode of the original hit, C703-0617 (1), and to guide SAR studies. The cryoEM structure of a group 2 IAV hemagglutinin in complex with the imidazo­[2,1-a]­pyrimidine-based entry inhibitor SA-67 (PDB: 9ONZ), , previously developed by our group, was used as the structural template. Docking studies indicated that hit compound 1 (Figure ) occupies the same hemagglutinin pocket targeted by SA-67 and Arbidol. These studies further suggested that the N-substituted region 2 of hit compound 1 extends more deeply into a hydrophobic pocket of hemagglutinin near residues Leu19 and Ala101 (Figure ). This observation motivated us to incorporate substituents with greater hydrophobicity in region 2. In contrast, region 1 was closer to a solvent-exposed area and adjacent to polar/charged residues Gln302 and Glu98. We also observed a hydrogen bonding interaction between the methoxy group of compound 1 and Gln302. These observations suggested that incorporation of substituents with hydrogen-bond-accepting capabilities in region 1 could further enhance binding affinity. The thieno­[3,2-b]­pyrrole core appears to fit tightly within the pocket, suggesting limited space for expansion in this region of the molecule; however, modifications to the core heteroatoms may still be explored. Additionally, overlaying SA-67 with compound 1 revealed that both adopt similar poses in the binding site.

1.

Hit compound C703-0617 (1) with selected regions for modification, Arbidol, and SA-67.

2.

(A) Predicted binding of C703-0617 (1) to HA. The hit compound 1 (shown in green), docked into the cryoEM structure of group 2 influenza strain H7N9 hemagglutinin in complex with the imidazopyrimidine entry inhibitor SA-67 (PDB: 9ONZ). , Subunit HA1 is shown in blue and subunit HA2 is shown in orange. Enhanced images show (B) relevant residues and (C) a 3D surface to depict regions of 1 predicted to reside more deeply within the protein. Hit compound 1 is also overlaid with SA-67, the ligand bound in the cryoEM structure, to demonstrate similarities in binding poses.

Three regions of the thieno­[3,2-b]­pyrrole scaffold were selected for SAR modifications (Figure ). The new analogues 2–46 (Tables –) were synthesized as described below. Most derivatives were prepared using the synthetic route outlined in Scheme . The starting methyl 4H-thieno­[3,2-b]­pyrrole-5-carboxylate (47a) or 4H-furo­[3,2-b]­pyrrole-5-carboxylate (47b) was first alkylated on the nitrogen with various alkyl bromides 48a–j. After completion of the alkylation, the methyl ester was saponified under basic conditions to afford the corresponding carboxylic acids 49a–j. These intermediates were coupled with a series of amines 51 using N,N-diisopropylethylamine (DIPEA) and 2-(7-azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluronium hexafluorophosphate (HATU) to furnish the final compounds 1–44 in moderate to high yields (40–85%).

1. Pseudovirus Anti-influenza Activity of Thienopyrroles 1–9 .

#	R	pH7N1 EC50 (μM)	A549 CC50 (μM)	SI
1	4-OMe	2.1 ± 0.2	>100	>48
2	3-OMe	2.0 ± 0.4	>100	>50
3	2-OMe	5.4 ± 3	>100	>18
4	3,4-OMe	1.1 ± 0.1	>100	>91
5	1,3-dioxole	1.2 ± 0.1	49 ± 4	40
6	4-OEt	7.7 ± 5	>100	>13
7	4-CO2Et	6.2 ± 1	>100	>16
8	3-CO2Et	29 ± 4	>100	>3
9	4-Cl,3-OMe	1.7 ± 0.1	>100	>59

^aEC₅₀ values were calculated by four parameter nonlinear regression analysis. Results are from three replicates. EC₅₀ data are presented as mean ± SD.

4. Chemical Structures and Pseudovirus Anti-influenza Activity of Thienopyrroles 30–46 .

^aEC₅₀ values were calculated by four-parameter nonlinear regression analysis. Results are from three replicates. EC₅₀ data are presented as mean ± SD.

1. General Procedure for the Synthesis of Thieno­[3,2-b]­pyrroles 1–43 and Furo­[3,2-b]­pyrrole 44 .

^a Reagents and conditions: (a) NaH, DMF, 4 °C to RT, 16–24 h; (b) LiOH, NaOH, H₂O/DMF, RT, 16–24 h; (c) HATU, DIPEA, DMF, 4 °C to RT, 10–12 h.

A Group 2 H7N1 pseudovirus (pH7N1) with an HIV-luciferase core was used to evaluate derivatives. Modifications to Region 1 were first carried out with attempts to improve interactions with polar and hydrogen-bonding residues (Table ), inspired by docking studies (see above). Changing the position of the methoxy group to the 3-position to confirm optimal position of the group (2) resulted in retention of activity, while switching to the 2-position (3) resulted in a 2.6-fold decrease in activity; however, when a 3,4-dimethoxy (4) or benzodioxole (5) was incorporated, there was a ∼2-fold increase in activity, suggesting that interactions with polar and charged residues may be better achieved with substituents in the 3- and 4-positions of the phenyl ring. Although speculative, the higher toxicity of compound 5 may be due to the formation of a catechol group by oxidative metabolism. When the 4-methoxy group was replaced with a 4-ethoxy group (6) to probe the impact of longer carbon chains, a 3.7-fold decrease in activity was observed. When an ethyl ester was placed in the 4-position (7) and the 3-position (8), 3-fold and 14-fold decreases in activity were observed, respectively. These results suggest that longer carbon chains are not tolerated close to the solvent exposed region of the binding site. A 4-chloro-3-methoxy substitution (9) did not result in a significant change in activity, suggesting hydrogen bond acceptors like the methoxy group provide more favorable interactions than halogens. Next, we determined the effect of replacing the phenyl ring with nitrogen heterocycles (Table ). A pyridin-4-yl (10) substitution resulted in a 2-fold increase in activity. Although a pryidin-3-yl (11) substituent maintained the same activity, a pyridin-2-yl (12) substituent resulted in a 2-fold decrease in activity. Compounds 11 and 12 also showed increased toxicity. Because substituted pyridines generally are not thought of as structural alerts, we speculate that off-target receptor inhibition may be responsible for this increase in toxicity. These results further support the hypothesis that interactions with polar or charged residues are best achieved in the 3- or 4-position of the substituent. A pyrazine substituent (13) was also tested but resulted in a 2.3-fold decrease in activity. Attempts to combine favored interactions by incorporating a 2-methoxy pyridine (14) resulted in a 1.5-fold increase in activity, which unfortunately is not as potent as the 4-pyridyl-substituted 10 or 3,4,-dimethoxy-substituted 4. A 2-methylpyridine substitution (15) decreased activity by 1.7-fold, suggesting hydrophobic interactions are not beneficial in Region 1. The increase in toxicity of compound 15 may be due to off-target effects, similar to compounds 11 and 12; however, it is also possible the 3-methyl group undergoes CYP oxidation. 2-Chloropyridinyl (16) and 2-bromopyridinyl (17) substitutions were also tested and resulted in 3- and 3.5-fold increases in activity, respectively; however, the electrophilic nature of these substituents may imply nonspecific interactions. A benzyl substitution (18) resulted in a 1.5-fold decrease in activity, while a cyclohexyl group (19) resulted in a complete loss of activity. This finding suggests that the planarity and aromaticity of the substituent is essential, and that Region 1 is not tolerant of cyclic aliphatic groups. Increasing the linker length (20) and decreasing it (21) resulted in a complete loss of activity, suggesting a methylene unit is the optimal linker length for Region 1. In conclusion, benzyl substituents with small, polar, H-bond acceptors in the 4-position result in the strongest potency.

2. Pseudovirus Anti-influenza Activity of Thienopyrroles 10–21 .

^aEC₅₀ values were calculated by four-parameter nonlinear regression analysis. Results are from three replicates. EC₅₀ data are presented as mean ± SD.

^bEC₅₀ could not be determined because the compound showed ∼50% inhibition from 3.7 μM to 100 μM.

Next, modifications to region 2 were explored (Table ). First, further increasing the polar character of the 4-fluoro substitution was attempted. A 3,4-difluoro substitution (22) resulted in a 1.8-fold decrease in the activity. Replacing the 4-fluoro with a 4-chloro group (23) resulted in a 3.4-fold decrease in activity. Incorporating a 4-trifluoromethyl substitution (24) resulted in a complete loss of activity. This finding suggests that larger groups are not well tolerated in the 4-position of region 2. When the trifluoromethyl group was moved to the 3-position (25), some activity was regained, but there was still a 3.1-fold decrease in activity. This result suggests that there is more free space in the binding site surrounding the 3-position. Replacing the 4-fluorobenzyl group with a benzyl substituent (26) resulted in a 3.6-fold decrease in activity, suggesting that the 4-fluoro has a beneficial interaction; however, when a cyclohexyl group was introduced (27), a slight increase in activity was observed. In an attempt to gain hydrophobic interactions deeper in the pocket around Region 2, an additional methylene was added to make the cyclohexylethyl substituent (28), which resulted in a slight increase in activity. To confirm Region 1 and 2 substituents were selective toward their respective binding pocket, Region 1 and 2 substituents were flipped (29), resulting in a complete loss of activity. In conclusion, less rigid, aliphatic, hydrophobic groups are preferred over aromatic polar substituents for region 2.

3. Chemical Structures and Pseudovirus Anti-influenza Activity of Thienopyrroles 22–29 .

^aEC₅₀ values were calculated by four-parameter nonlinear regression analysis. Results are from three replicates. EC₅₀ data are presented as mean ± SD.

Having found the cyclohexylethyl substituent to be beneficial in Region 2, we made additional modifications to Region 1 to further improve activity (Table ). Compared to the cyclohexylethyl-substituted 28, shortening the linker with a directly bound dimethylaminopiperidine (30) resulted in a 3.6-fold decrease in activity. This change was made to further confirm shortening the linker of Region 1 damages activity. An imidazole substitution with a propyl linker (31) resulted in a 3.9-fold decrease in activity, suggesting that some substitutions do not completely lose activity with longer linkers. Additionally, substituting in an even larger 4-hydroxyphenylpiperazine (32) resulted in a 4.1-fold decrease in activity. Substituting in a 2-furylmethyl (33) resulted in a 1.5-fold decrease in activity. Methylating the amine linker (35) resulted in a 4.3-fold decrease in activity, compared to its unmethylated 4-fluoro counterpart (34). This result suggests the amide has a favorable H-bonding interaction or there is not suitable space in the binding site to expand in that region. Compounds with substitutions previously tested with a 4-fluorobenzyl in Region 2 resulted in comparable activity (36, 37). A 3-bromopyridine substitution (38) resulted in a 2.2-fold decrease in activity compared to a 2-bromopyridine (39), a result that could be due to the electrophilicity of the latter. It was not until a 4-dimethylaminobenzyl substitution (40), which was synthesized based on previous success of substituents with H-bond acceptors in the 4-position, was incorporated that the highest potency was achieved (0.71 μM). This result is in good agreement with the beneficial activity seen with the 4-methoxy substitution. To confirm the optimal position of the dimethylamino group, the position was switched to the meta position (41), which resulted in a slight decrease in potency. This effect is similar to what was observed previously by changing the position of other substituents. Derivatives found to be potent with cyclohexylethyl were also made with a cyclohexylmethyl in Region 2 to confirm the longer linker also resulted in higher potency. The dimethylamino derivative (42) maintained similar activity to the dimethylamino-substituted 40; however, the pyridine substitution (43) saw a 1.8-fold increase in potency compared to its cyclohexylethyl counterpart (37). Overall, a 4-dimethylaminobenzyl, 3,4-dimethoxybenzyl, and 4-pyridiylmethyl substitution in Region 1 paired with a cyclohexylmethyl or cyclohexylethyl group in Region 2 produced the greatest potency (36, 40, 43).

Unlike modifications to the substituents, modifications to the central core resulted in a loss of activity. Substituting the thienopyrrole with a furanopyrrole (44) resulted in a 5.7-fold decrease in activity. Methylating the 2-position of the core (45) was an attractive option to possibly prevent oxidation, but doing so resulted in a 9-fold decrease in activity. An indole derivative (46) was also tested but was significantly less active compared with its similar thienopyrrole counterpart (30). Overall, results from modifications to the core could suggest that the core occupies a tight binding pocket that does not allow for expansion or modification of the heteroatom.

Four compounds were identified as the most promising from pseudovirus assays (1, 36, 40, and 43). Authentic group 2 H3N2 infectious viruses were then used to validate the activity of lead derivatives (Table ). Specifically, compounds were tested against human H3N2 A/Hong Kong-11/1968 and human H3N2 A/Victoria/361/2011 strains. Oseltamivir carboxylate, the active metabolite of oseltamivir phosphate, was used as a positive control. The 2011 H3N2 strain is known to be oseltamivir-resistant, caused by its E119V mutation in neuraminidase. Because the compounds are predicted to bind to hemagglutinin, activity should be similar in both strains. Strong potency was observed in both strains, with activity ranging from 50 nM to 1.9 μM. The hit compound 1 showed the weakest potency, with optimized thienopyrroles generally being more potent. Oseltamivir carboxylate had a greater than 55-fold decrease in potency in the resistant 2011 strain, while thienopyrroles showed a slight increase in activity. Compound 36 showed the highest activity of the thienopyrrole derivatives, with an EC₅₀ of 50 nM in the 2011 strain and 0.16 μM in the 1968 strain. This is an approximate 10-fold increase in potency in both strains compared to hit compound 1. In docking studies, compound 36 showed similar binding interactions to those observed with compound 1; however, the flexibility and increased length of the ethyl cyclohexyl group allowed for stronger interactions with the hydrophobic residues around Region 2 of the scaffold. The increase in activity compared to pseudovirus results could be attributed to the difference in morphology of IAV and HIV viruses. HIV viruses are known to have a small, spherical shape; while IAV viruses have a variable pleomorphic structure. The pleomorphism of IAV is thought to have a significant impact on its pathogenesis. These data suggest that thienopyrroles have potent inhibitory activity against Group 2 IAV strains.

5. Anti-influenza Activity of Top Compounds against Infectious IAV.

		H3N2 1968		H3N2 2011
Compound	A549 CC50 (μM)	EC50 (μM)	SI (CC50/EC50)	EC50 (μM)	SI (CC50/EC50)
Oseltamivir carboxylate	>100	0.00018 ± 0.00014	>5.6 × 105	>0.01	NA
1	>100	1.9 ± 0.1	>53	0.53 ± 0.07	>190
36	>100	0.16 ± 0.02	>620	0.05 ± 0.01	>2,000
40	>100	0.24 ± 0.02	>420	0.23 ± 0.01	>420
43	54 ± 1.4	1.2 ± 0.3	45	0.07 ± 0.01	>770

^aEC₅₀ values were calculated by four-parameter nonlinear regression analysis. Results are from three replicates. EC₅₀ data are presented as mean ± SD.

Lead compounds were also counter-screened against other pseudoviruses containing an HIV core to confirm selectivity toward group 2 IAV (Table ). Vesicular stomatitis virus (VSV) is an enveloped RNA virus from the Rhabdoviridae family. Ebola (EBOV) and Marburg (MARV) are both enveloped RNA viruses of the Filoviridae family. A Group 1 influenza strain containing the same neuraminidase but different hemagglutinin (H5N1) was also included to confirm compounds were not active against both phylogenetic groups. Hit compound 1 showed weak activity of around 30 μM toward pEBOV and pMARV. All lead thienopyrroles showed no observed activity in H5N1, VSV, EBOV, and MARV pseudovirus assays. These findings indicate that, as structural modifications were made and potency increased, specificity toward Group 2 IAV increased as well.

6. Pseudovirus Selectivity Analysis of Top Compounds .

#	A549 CC50 (μM)	pH7N1 EC50 (μM)	pH5N1 EC50 (μM)	pVSV EC50 (μM)	pEBOV EC50 (μM)	pMARV EC50 (μM)
1	>100	2.1 ± 0.2	>100	>100	30 ± 1	30 ± 6
36	>100	1.4 ± 0.2	>100	>100
40	>100	0.71 ± 0.06	>100	>100

^aEC50 values were calculated by nonlinear regression analysis. Results are from three replicates. EC₅₀ data are presented as mean ± SD.

^bNo activity.

Residue-specific mutagenesis studies were also performed to examine whether our compounds were binding to the same binding site as Arbidol (Table ). We examined the following mutations in the H7 pseudovirus plasmid: F285L, L19V, Y94H, and L98V. Docking studies displaying key mutated residues in blue with compound 1 can be seen in Figure . Y94H resulted in the greatest decrease in activity (13-fold with 1 and 27-fold with 40). This decrease is likely due to a poorer π-stacking interaction between the residue and the thienopyrrole core of the ligands. The F258L mutation also resulted in a decrease in activity (12-fold with 1 and 3.2-fold with 40). Conversion of phenylalanine to leucine would result in a complete loss of π-stacking capability with the thienopyrrole core. L19V resulted in a 12-fold decrease with compound 1 and 6.1-fold decrease with compound 40. Hydrophobic substitutions were found to be favored in Region 2 of the thienopyrrole scaffold, indicating hydrophobic interactions are beneficial to ligand affinity. The reduction in size of the lipophilic residue near Region 2 of the ligand is likely responsible for the decrease in activity. Leu98 is located at the top of the binding site, close to Tyr94, so the smaller fold-change of compound 40 relative to hit compound 1 could indicate that Region 2 is positioned more deeply within the pocket closer to Leu19. Overall, changes in ligand binding activity support that ligands are binding to the predicted binding site.

7. Effect of H7 Residue Mutations on Compound Potency.

Mutant (pH7N1)	Compound 1 EC50 (μM)	Compound 1 Fold Change	Compound 40 EC50 (μM)	Compound 40 Fold Change
WT	3.2 ± 0.2		0.94 ± 0.1
F285L	37 ± 10	12	2.9 ± 0.6	3.2
L19V	40 ± 6	12	5.7 ± 0.8	6.1
L98V	5.2 ± 3	1.6	0.91 ± 0.3	0.97
Y94H	42 ± 1	13	25 ± 1	27

^aEC₅₀ values were calculated by four-parameter nonlinear regression analysis. Results are from three replicates. EC₅₀ data are presented as mean ± SD.

^bFold change represents the ratio of EC₅₀ relative to that of the wild-type EC₅₀ (WT).

^cReference.

In summary, we have presented the synthesis and biological characterization of thieno­[3,2-b]­pyrroles as Group 2 influenza A entry inhibitors targeting hemagglutinin. Docking studies in the predicted binding site initially identified potential interactions to exploit. SAR studies in pseudovirus assays resulted in an increase in potency from 2.1 μM to 0.71 μM, which identified compounds for further analysis in infectious assays. Selected compounds displayed even higher potencies in infectious assays, with the most potent compound (36) displaying an EC₅₀ of 50 nM in the 2011 H3N2 strain and 0.16 μM in the 1968 strain. All compounds also retained potent activity in the oseltamivir-resistant 2011 strain. Testing against other pseudoviruses showed increased selectivity of optimized compounds against Group 2 IAV. Mutagenesis studies supported that compounds were binding to the predicted hydrophobic pocket in the stem region of HA, indicating compounds act as interprotomer stabilizers. Overall, optimized thienopyrroles display selective and remarkably strong potency, with some of the strongest potency reported against the 2011 H3N2 strain. With the H3N2 subtype dominating influenza infections in recent years, the success of the thienopyrrole scaffold warrants further development for therapeutic use.

Glossary

Abbreviations Used

HA

Hemagglutinin

IAV

Influenza A Virus

NA

Neuraminidase

pEBOV

Pseudoviral Ebola virus

pH5N1

Pseudoviral H5N1 Influenza Virus

pH7N1

Pseudoviral H7N1 Influenza Virus

pMARV

Pseudoviral Marburg virus

PVSV

Pseudoviral Vesicular stomatitis virus

SI

Selectivity Index

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acsmedchemlett.6c00239.

• Detailed synthetic procedures for all reported compounds, plasma stability assay results for select compounds, and NMR spectra and HPLC chromatograms for key compounds to confirm structural integrity and purity (PDF)

Safety Statement: No unexpected or unusually high safety hazards were encountered.

The authors declare the following competing financial interest(s): L.R. is the owner of Chicago BioSolutions, Inc., and thus declares potential financial interests, as do I.N.G. and N.P.P., who are employed by Chicago BioSolutions, Inc.