LETTER
Tecovirimat (also known as TPOXX or ST-246) is a drug available for the treatment of mpox through the Centers for Disease Control and Prevention’s Expanded Access Investigational New Drug “compassionate use” protocol (https://www.cdc.gov/poxvirus/monkeypox/clinicians/Tecovirimat.html). In Los Angeles County, a fatal case of mpox with tecovirimat resistance was previously reported (1). Epidemiologic surveillance in Los Angeles County has since identified additional cases of severe mpox that did not improve after multiple rounds of tecovirimat treatment, including one involving a person who succumbed to infection (Table 1). Consistent with reports describing severe manifestations of mpox within the current global outbreak (1, 2), the identified cases involved host immunodeficiency due to advanced HIV infection.
TABLE 1.
Clinical data for mpox cases
| Patient | CD4+ T-cell count (cells/mm3)a | HIV viral load (copies/mL)a | Mpox treatment and Specimen Collection History | Clinical outcome |
|---|---|---|---|---|
| A | <35 | 130,565 | Tecovirimat POc 600 mg: BIDd, 1 wk Tecovirimat PO 600 mg: BID, 1 wk Tecovirimat IVe 200 mg: 3 doses Tecovirimat PO 600 mg: BID, 2 wks Tecovirimat IV 200 mg: qf 12 hours, 2 days Specimen A.1 collected VIGIVg (unknown dose): single infusion Tecovirimat IV 200 mg: q 12 hours, 2 wks Specimens A.2-A.26 collected |
Deceased |
| B | 73 | <20 | Tecovirimat PO 600 mg: BID, 2 wks Tecovirimat PO 600 mg: BID, 2 wks Specimens B.1-B.2 collected Tecovirimat IV 200 mg: q 12 hours, 2 wks VIGIV (unknown dose): single infusion Brincidofovir PO (unknown dose): single dose |
Unknown |
| C | 136 | 178,982 | Tecovirimat PO 200-600 mg: QDh-BID, 2 wks Tecovirimat IV 200 mg: q 12 hours, 3 wks Specimens C.1-C.2 collected VIGIV (unknown dose): single infusion |
Ongoing lesions (after 6 mo) |
| D | 96 | 2,620 | Tecovirimat PO 600 mg: BID, 2 wks Tecovirimat PO 600 mg: BID, 11 days Specimen D.1 collectedb Cidofovir IV 5 mg/kg: single infusion Tecovirimat PO 600 mg: BID, 4 days VIGIV 9000 units/kg: single infusion Tecovirimat IV 200 mg: q 12 hours, 2 wks Cidofovir IV 5 mg/kg: single infusion |
Recovered |
| E | 191 | 606 | Tecovirimat PO 600 mg: BID, 2 wks Tecovirimat PO 600 mg: BID, 4 wks Specimens E.1-E.8 collected Tecovirimat PO 600 mg: BID, 6 wks Tecovirimat PO 600 mg: BID, 10 wks Specimens E.9-E.14 collected |
Ongoing lesions (after 6 mo) |
| F | 54 | 485,298 |
Specimen F.1 collected Tecovirimat PO 600 mg: BID, 1 wk Tecovirimat PO 600 mg: BID, 2 days Tecovirimat IV 200 mg: q 12 hours, 4 days Specimens F.2-F.5 collected |
Ongoing lesions (after 7 mo) |
Results are most proximal to case identification and management. Specimen collection history is in bold text.
Specimen D.1 was collected 5 days into preceding tecovirimat treatment course.
PO = Per Oral.
BID = Twice daily.
IV = Intravenous.
q = every.
VIGIV = Vaccinia Immune Globulin Intravenous.
QD = Once daily.
Tecovirimat targets the conserved orthopoxvirus VP37 envelope protein required for extracellular virus particle generation, and previous studies from monkeypox virus (MPXV) and other orthopoxviruses identified more than 20 mutations in VP37 associated with tecovirimat resistance (https://www.fda.gov/emergency-preparedness-and-response/mcm-issues/fda-mpox-response#therapeutics) (1, 3). To determine whether these identified cases involved tecovirimat resistance, MPXV specimens were subjected to whole-genome sequencing and examined for mutations in the homolog of the vaccinia virus Copenhagen F13L gene that encodes VP37 protein (1, 4). All specimens encoded lysine at amino acid position 353, which is prevalent within the current outbreak and does not affect tecovirimat sensitivity (5). Specimens from six cases harbored a cumulative total of eight VP37 mutations associated with resistance in prior studies: H238Q, P243S, N267D, A288P, A290V, D294V, A295E, and I372N (Table 2). Five previously undescribed mutations of unknown significance were also identified (T220A/I, T245I, A265D, and T289A), which may represent novel tecovirimat-interacting residues. Allele frequency analysis demonstrated resistance mutation heterogeneity within single lesion specimens, and different mutations were identified among distinct lesion specimens from the same person, suggesting mutations were acquired during treatment. Consistent with a de novo process, comparison of specimens collected before and after treatment in a single person (Patient F) showed resistance-associated mutations in VP37 only after treatment exposure (Table 2).
TABLE 2.
VP37 mutation and tecovirimat resistance data for human monkeypox virus specimensa
| Patient | Specimen (source) | VP37 mutations detected (allele frequencyb) | GISAIDc ID | GenBank ID | SRAd ID | ECe50 (μM) | Fold changef |
|---|---|---|---|---|---|---|---|
| A | A.1 (Unknown) | A290V (0.97) | EPI_ISL_15597062 | OP748968 | SRR22116985 | 0.2032 | 12 |
| A.2 (Unknown) | N267D (0.24), A288P (0.76) | EPI_ISL_15896341 | OP890565 | SRR22398844 | Pending | ||
| A.3 (Penis) | N267D (0.38), A288P (0.37), A295E (0.31), I372N (0.22) | EPI_ISL_15896342 | OP890566 | SRR22398843 | Culture contaminated | ||
| A.4 (Scrotum) | A288P (0.95) | EPI_ISL_15896343 | OP890567 | SRR22398842 | Culture contaminated | ||
| A.5 (Ltg Thigh) | A288P (1.0) | EPI_ISL_15896344 | OP890568 | SRR22398841 | Pending | ||
| A.6 (Rth Sole) | A295E (0.82) | EPI_ISL_15896345 | OP890569 | SRR22398840 | Culture contaminated | ||
| A.7 (Rt Heel) | I372N (0.84) | EPI_ISL_15896346 | OP890570 | SRR22398839 | Culture negative | ||
| A.8 (Lt Back Knee) | N267D (0.55), A288P (0.18), D294V (0.29) | EPI_ISL_15896347 | OP890571 | SRR22398838 | Pending | ||
| A.9 (Unknown) | N267D (0.91) | EPI_ISL_15896348 | OP890572 | SRR22398837 | Culture contaminated | ||
| A.10 (Unknown) | T220A (0.44)i, A265D (0.50)i, A295E (0.51) | EPI_ISL_15896349 | OP890573 | SRR22398836 | Culture contaminated | ||
| A.11 (Lt 2nd Finger) | A288P (0.62), I372N (0.10) | EPI_ISL_15896350 | OP890574 | SRR22398835 | Culture negative | ||
| A.12 (Lt Medial Thigh) | N267D (0.37), A288P (0.63) | EPI_ISL_15896351 | OP890575 | SRR22398833 | Culture contaminated | ||
| A.13 (Rt Eyelid) | A288P (1.0) | EPI_ISL_15896352 | OP890576 | SRR22398832 | Pending | ||
| A.14 (Tongue) | A288P (1.0) | EPI_ISL_15896353 | OP890577 | SRR22398831 | Pending | ||
| A.15 (Rt Forehead) | I372N (0.96) | EPI_ISL_15896329 | OP890553 | SRR22398858 | Pending | ||
| A.16 (Lt Forehead) | N267D (0.88), D294V (0.11) | EPI_ISL_15896330 | OP890554 | SRR22398857 | Culture contaminated | ||
| A.17 (Between Eyes) | A288P (0.88) | EPI_ISL_15896331 | OP890555 | SRR22398855 | 10.28 | 590 | |
| A.18 (Lt Ear) | I372N (0.94) | EPI_ISL_15896332 | OP890556 | SRR22398854 | Pending | ||
| A.19 (Lt Nose) | A288P (0.19), A290V (0.25), I372N (0.35) | EPI_ISL_15896333 | OP890557 | SRR22398853 | Pending | ||
| A.20 (Rt Cheek) | A288P (0.12), I372N (0.87) | EPI_ISL_15896334 | OP890558 | SRR22398852 | Culture negative | ||
| A.21 (Lower Lip) | N267D (0.70), A288P (0.22) | EPI_ISL_15896335 | OP890559 | SRR22398851 | Pending | ||
| A.22 (Lt Neck) | A288P (0.85) | EPI_ISL_15896336 | OP890560 | SRR22398850 | Culture negative | ||
| A.23 (Lt Chest) | T220I (0.49)i, P243S (0.50), A295E (0.51) | EPI_ISL_15896337 | OP890561 | SRR22398849 | Culture negative | ||
| A.24 (Rt Hand) | A290V (0.88) | EPI_ISL_15896338 | OP890562 | SRR22398848 | Culture negative | ||
| A.25 (Lt Elbow) | D294V (1.0) | EPI_ISL_15896339 | OP890563 | SRR22398847 | Pending | ||
| A.26 (Lt Shoulder) | H238Q (0.99) | EPI_ISL_15896340 | OP890564 | SRR22398846 | 0.5803 | 33 | |
| B | B.1 (Unknown) | T245I (0.12)i, D294V (0.91) | EPI_ISL_15325434 | OP615280 | SRR21864633 | 0.3820 | 22 |
| B.2 (Rt Ear) | H238Q (0.27), A288P (0.21), D294V (0.15), I372N (0.33) | EPI_ISL_15417996 | OP680496 | SRR21966131 | ~5.150 | ~290 | |
| C | C.1 (Unknown) | A290V (1.0) | EPI_ISL_15896305 | OP890529 | SRR22398845 | Culture negative | |
| C.2 (Unknown) | A290V (1.0) | EPI_ISL_15955339 | OP920682 | SRR22459664 | Culture negative | ||
| D | D.1 (Penis) | N267D (0.68), A288P (0.29) | EPI_ISL_17582852 | OQ888843 | SRR24326324 | 15.89 | 910 |
| E | E.1 (Lt Foot Between Toes) | I372N (1.0) | EPI_ISL_15418000 | OP680500 | SRR21966127 | 0.07286 | 3.5 |
| E.2 (Rt Cheek) | D294V (1.0) | EPI_ISL_15418001 | OP680501 | SRR21966126 | 0.3364 | 19 | |
| E.3 (Lt Hand by Thumb) | A290V (1.0) | EPI_ISL_15418002 | OP680502 | SRR21966125 | 0.1670 | 7.9 | |
| E.4 (Lt Foot) | A290V (0.30), I372N (0.86) | EPI_ISL_15418003 | OP680503 | SRR21966124 | 0.05551 | 2.6 | |
| E.5 (Lt Ring Finger) | H238Q (1.0) | EPI_ISL_15418004 | OP680504 | SRR21966123 | 0.5390 | 28 | |
| E.6 (Rt Foot) | N267D (0.97) | EPI_ISL_15418005 | OP680505 | SRR21966122 | 11.05 | 480 | |
| E.7 (Lt Foot, Scab) | A290V (0.56), I372N (0.77) | EPI_ISL_15418007 | OP680507 | SRR21966119 | 77.37 | 4400 | |
| E.8 (Lt Chin, Scab) | D294V (1.0) | EPI_ISL_15418008 | OP680508 | SRR21966118 | 0.2344 | 13 | |
| E.9 (Lt Great Toe) | I372N (1.0) | EPI_ISL_16997407 | OQ503839 | SRR23580660 | Pending | ||
| E.10 (Lt 4th Finger) | H238Q (1.0) | EPI_ISL_16997408 | OQ503840 | SRR23580659 | Pending | ||
| E.11 (Rt Medial Foot) | N267D (1.0) | EPI_ISL_16997409 | OQ503841 | SRR23580658 | Pending | ||
| E.12 (Lt Great Toe, Tissue) | I372N (1.0) | EPI_ISL_16997410 | OQ503842 | SRR23580657 | Pending | ||
| E.13 (Rt Medial Foot, Tissue) | N267D (0.45), I372N (0.56) | EPI_ISL_16997411 | OQ503843 | SRR23580656 | Pending | ||
| E.14 (Lt 4th Finger, Tissue) | H238Q (1.0) | EPI_ISL_16997412 | OQ503844 | SRR23580655 | Pending | ||
| F | F.1 (Unknown)# | none | EPI_ISL_16679205 | OQ330972 | SRR23238068 | 0.007144 | 0.41 |
| F.2 (Face) | A290V (0.68) | EPI_ISL_16997464 | OQ503820 | SRR23580586 | Culture contaminated | ||
| F.3 (Lt Ear) | T289A (0.41)i, I372N (0.81) | EPI_ISL_16679238 | OQ331005 | SRR23238051 | Pending | ||
| F.4 (Lt Buttock) | A290V (1.0) | EPI_ISL_16679239 | OQ331006 | SRR23238050 | 0.2257 | 13 | |
| F.5 (Rt Foot) | A290V (0.52) | EPI_ISL_16679240 | OQ331007 | SRR23238049 | Culture negative |
All specimens are from lesion swabs unless noted otherwise. Consensus mutations with an allele frequency of 0.6 or greater are in bold text. Specimens with a 2- to 9-fold increase in resistance are considered partially resistant, while a 10-fold or greater increase is considered resistant.
Alelle frequency = proportion of mapped reads containing the corresponding mutation at each site.
GISAID = Global Initiative on Sharing Avian Influenza Data.
SRA = Sequence Read Archive.
EC50 = Half-maximal effective concentration.
Fold change = observed EC50 compared to EC50 for MPXV Clade IIa (US, 2003 strain).
Lt = Left.
Rt = Right.
Previously unassociated with tecovirimat resistance; #Tecovirimat-naïve specimen.
Phenotypic testing of MPXV cultured from a subset of specimens (1, 5) displayed a wide range of tecovirimat resistance levels in vitro compared to wild-type isolates (Table 2), arguing these mutations confer resistance within currently circulating strains. For heterogenous cultures with multiple VP37 mutations, it is unclear how much individual mutations contribute to resistance and whether resistance is due to single or multiple mutations, especially in cases with low allele frequencies. It is also possible that uncharacterized mutations outside of VP37 enhance or diminish tecovirimat resistance.
These findings demonstrate that tecovirimat resistance can arise in the setting of prolonged treatment exposure with host immunodeficiency and underscore the need for combined therapeutics with different viral targets (including VIGIV and cidofovir/brincidofovir) (6, 7) when treating mpox in certain people (8), as is the case with progressive vaccinia (9). Clinical management should ensure effective drug delivery/dosing (e.g., making sure oral tecovirimat is taken with a fatty meal) and optimization of immune function when treating with tecovirimat to prevent resistance from developing. Additionally, this work suggests that sequencing of MPXV to identify tecovirimat resistance could serve as a valuable clinical tool.
Use of trade names and commercial sources are for identification only and do not imply endorsement by the U.S. Department of Health and Human Services or the County of Los Angeles Department of Public Health. The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention or their institutions, or the County of Los Angeles Department of Public Health.
This work was supported by the Epidemiology and Laboratory Capacity for Prevention and Control of Emerging Infectious Diseases cooperative agreement of the Centers for Disease Control and Prevention (grant 6 NU50CK000498). A.K. was supported by the 2T32AI7502-26 Applied Genomics in Infectious Diseases training grant from Stanford University.
The Los Angeles County Department of Public Health Institutional Review Board (IRB) determined that this study met the criteria for not research per 45 CFR 46.102(e). Therefore, IRB review was not needed.
REFERENCES
- 1.Alarcón J, Kim M, Terashita D, Davar K, Garrigues JM, Guccione JP, Evans MG, Hemarajata P, Wald-Dickler N, Holtom P, Garcia Tome R, Anyanwu L, Shah NK, Miller M, Smith T, Matheny A, Davidson W, Hutson CL, Lucas J, Ukpo OC, Green NM, Balter SE. 2023. An mpox-related death in the United States. N Engl J Med 388:1246–1247. doi: 10.1056/NEJMc2214921. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 2.Mitjà O, Alemany A, Marks M, Lezama Mora JI, Rodríguez-Aldama JC, Torres Silva MS, Corral Herrera EA, Crabtree-Ramirez B, Blanco JL, Girometti N, Mazzotta V, Hazra A, Silva M, Montenegro-Idrogo JJ, Gebo K, Ghosn J, Peña Vázquez MF, Matos Prado E, Unigwe U, Villar-García J, Wald-Dickler N, Zucker J, Paredes R, Calmy A, Waters L, Galvan-Casas C, Walmsley S, Orkin CM, SHARE-NET writing group . 2023. Mpox in people with advanced HIV infection: a global case series. Lancet 401:939–949. doi: 10.1016/S0140-6736(23)00273-8. [DOI] [PubMed] [Google Scholar]
- 3.Duraffour S, Lorenzo MM, Zöller G, Topalis D, Grosenbach D, Hruby DE, Andrei G, Blasco R, Meyer H, Snoeck R. 2015. ST-246 is a key antiviral to inhibit the viral F13L phospholipase, one of the essential proteins for orthopoxvirus wrapping. J Antimicrob Chemother 70:1367–1380. doi: 10.1093/jac/dku545. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 4.Chen NFG, Chaguza C, Gagne L, Doucette M, Smole S, Buzby E, Hall J, Ash S, Harrington R, Cofsky S, Clancy S, Kapsak CJ, Sevinsky J, Libuit K, Park DJ, Hemarajata P, Garrigues JM, Green NM, Sierra-Patev S, Carpenter-Azevedo K, Huard RC, Pearson C, Incekara K, Nishimura C, Huang JP, Gagnon E, Reever E, Razeq J, Muyombwe A, Borges V, Ferreira R, Sobral D, Duarte S, Santos D, Vieira L, Gomes JP, Aquino C, Savino IM, Felton K, Bajwa M, Hayward N, Miller H, Naumann A, Allman R, Greer N, Fall A, Mostafa HH, McHugh MP, Maloney DM, Dewar R, et al. 2023. Development of an amplicon-based sequencing approach in response to the global emergence of mpox. PLoS Biol. 21:e3002151. doi: 10.1126/science.add4153 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 5.Gigante CM, Korber B, Seabolt MH, Wilkins K, Davidson W, Rao AK, Zhao H, Smith TG, Hughes CM, Minhaj F, Waltenburg MA, Theiler J, Smole S, Gallagher GR, Blythe D, Myers R, Schulte J, Stringer J, Lee P, Mendoza RM, Griffin-Thomas LA, Crain J, Murray J, Atkinson A, Gonzalez AH, Nash J, Batra D, Damon I, McQuiston J, Hutson CL, McCollum AM, Li Y. 2022. Multiple lineages of monkeypox virus detected in the United States, 2021–2022. Science 378:560–565. doi: 10.1126/science.add4153. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6.Siegrist EA, Sassine J. 2023. Antivirals with activity against mpox: a clinically oriented review. Clin Infect Dis 76:155–164. doi: 10.1093/cid/ciac622. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 7.Wang J, Shahed-Ai-Mahmud M, Chen A, Li K, Tan H, Joyce R. 2023. An overview of antivirals against monkeypox virus and other orthopoxviruses. J Med Chem 66:4468–4490. doi: 10.1021/acs.jmedchem.3c00069. [DOI] [PubMed] [Google Scholar]
- 8.Rao AK, Schrodt CA, Minhaj FS, Waltenburg MA, Cash-Goldwasser S, Yu Y, Petersen BW, Hutson C, Damon IK. 2023. Interim clinical treatment considerations for severe manifestations of mpox - United States, February 2023. MMWR Morb Mortal Wkly Rep 72:232–243. doi: 10.15585/mmwr.mm7209a4. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 9.Lederman ER, Davidson W, Groff HL, Smith SK, Warkentien T, Li Y, Wilkins KA, Karem KL, Akondy RS, Ahmed R, Frace M, Shieh WJ, Zaki S, Hruby DE, Painter WP, Bergman KL, Cohen JI, Damon IK. 2012. Progressive vaccinia: case description and laboratory-guided therapy with vaccinia immune globulin, ST-246, and CMX001. J Infect Dis 206:1372–1385. doi: 10.1093/infdis/jis510. [DOI] [PMC free article] [PubMed] [Google Scholar]