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⁎ A.M. and RG. contributed equally to the manuscript.
Roberta Gagliardini
Correspondence
Corresponding author: Dr. Roberta Gagliardini, Clinical and Research Infectious Diseases Department, National Institute for Infectious Diseases Lazzaro Spallanzani IRCCS, Via Portuense 292, 00149 Rome, Italy, 0039 3331045103, 0039 0655170368
We read with interest the manuscript by Li D. and colleagues, recently published in this Journal, in which the authors revealed the potential binding mode for tecovirimat with a poxvirus phospholipase from monkeypox (MPX) virus [
Here we retrospectively describe clinical presentation, evolution, management and viral kinetics of the first 19 MPX cases treated with antivirals at the INMI Lazzaro Spallanzani IRCCS in Rome, Italy. The decision regarding treatments was based on international medical consensus and availability of drugs.
Viral DNA was extracted by the automatic extractor QIAsymphony (Qiagen, Hilden, Germany), and amplified using the real-time PCR method targeting the tumor necrosis factor receptor gene, G2R. Monkeypox virus (MPXV) DNA concentration was measured using threshold cycles (Ct) values of the MPXV-specific PCR. To obtain an absolute quantification of MPXV DNA in the clinical samples, the PCR assay was adapted to run in digital droplet PCR (ddPCR). The nucleic acid extracted from each sample was loaded into specific nanoplate and distributed, amplified and read in each one of the 26,000 partitions of each well, with a detection limit of the assay of 5 copies/µL.
The study was conducted as a part of biological studies on emerging infections approved by the Ethical Committee of the Lazzaro Spallanzani Institute (approval number 14/2015 and amendments). Patients provided written informed consent.
As of September 19, 2022, 19/128 (15%) diagnosed cases of MPXV infection at INMI L. Spallanzani received antiviral treatment. All patients were males aged between 27 and 50 years, all but one patients self-identified as men who have sex with men or bisexual and seven patients (37%) were HIV-positive. Systemic symptoms were reported in all but one patient. Muco-cutaneous lesions were observed in all patients (skin lesions in 89% and mucosal lesions in 95%) and in half of them preceded systemic symptoms.
The majority (79%) of patients complained of a painful lymphadenopathy. Patients were admitted to hospital within a median of 8 days (IQR 5-10) from date of symptoms onset (OD), mainly for mucosal inflammation caused by MPXV and/or superinfection of the lesions and/or management of severe pain due to the lesions. Specifically, proctitis was diagnosed in four patients (21%) and severe pharyngo-tonsillitis in six patients (32%). One patient presented ocular localization complicated by periorbital edema and conjunctival hyperemia. Nine patients (47%) presented with superinfection of the soft tissues, one of which was complicated by abscess of a finger. Finally, one patient was admitted and treated for worsening of genital lesions.
Antiviral treatment was started with a median time of 11 days (IQR 8-12) from OD with oral tecovirimat in 15 (79%) patients and intravenous (IV) cidofovir in 4 (21%) patients All patients treated with oral tecovirimat completed a 14-day course of therapy. Similarly, IV cidofovir was well tolerated. Symptoms improvement and no new lesion appearance were observed 72 hours after the start of treatment in all but one patient treated with cidofovir.
No significative alterations of blood tests were observed, apart from a transient increase of alanine aminotransferase after cidofovir. Complete recovery was observed in all patients with a median of 15 days (IQR 11-19) from treatment start. Three patients had still persistence of signs of MPX-mucosal involvement after the resolution of lesions (Table 1).
Table 1Patients’ characteristics and clinical course.
PT1
PT2
PT3
PT4
PT5
PT6
PT7
PT8
PT9
PT10
PT 11
PT 12
PT 13
PT 14
PT 15
PT 16
PT 17
PT 18
PT 19
Gender/Age/ Ethnicity
M/28 y/ Caucasian
M/33 y/Caucasian
M/35 y/Caucasian
M/46 y/ Caucasian
M/33 y/ Caucasian
M/33 y/Hispanic
M/42y/Asian
M/40y/ Caucasian
M/47y/ Caucasian
M/27y/Caucasian
M/36 y/ Caucasian
M/38y/Hispanic
M/48y/ Caucasian
M/45y/Caucasian
M/35y/ Caucasian
M/50y/African
M/36y/Hispanic
M/38y/ Caucasian
M/47y/ Caucasian
Sexual orientation
Bisexual
MSM
MSM
MSM
MSM
MSM
MSM
MSM
MSM
MSM
MSM
MSM
MSM
MSM
MSM
Hetero-sexual
MSM
MSM
MSM
HIV status (ART; last CD4 (cell/mm3)/VL)
Neg
Neg
Neg
Pos(BIC/ TAF/FTC; 1622/ND)
Pos (3TC/DT; 872 /ND)
Pos* (TDF/FTC +DTG;526/26 cp/mL)
Neg
Neg
Pos (3TC/DTG; 828 /ND)
Pos**(BIC/ TAF/FTC;140/ <30cp/mL)
Neg
Neg
Neg
Neg
Neg
Neg
Pos(TAF/ FTC/DRV/ c+DTG; 253/22 cp/mL)***
Pos(TDF/ FTC/EFV; 1323; ND)
Neg
HbsAg/HCVAb
Neg/Neg
Neg/Neg
Neg/Neg
Neg/Neg
Neg/Neg
NA/Neg
Neg/Neg
Neg/Neg
Neg/Neg
Neg/Neg
Neg/Neg
Neg/Neg
Neg/NA
Pos/Neg
NA/NA
Neg/Neg
Neg/Neg
NA/Neg
Neg/Neg
PREP
No
Yes
Yes
No
No
No
No
Yes
No
No
No
Yes
No
No
No
No
No
No
No
Smallpox vaccination
No
No
No
No
No
No
No
No
No
Yes
No
No
Yes
No
No
No
No
No
No
Systemic symptoms
Fever, headache
Fever, sore throat, myalgias, diarrhoea
Fever
Fever, myalgias, rectal pain with discharg, bleeding
Super-infection of cutaneous lesions, pain management
Pain management
Compli- cated proctitis
Phary- ngoton- sillitis
Super- infection of cut- aneous lesions
Soft-tissue superinfection (upper lip)
Super-infection of cut-aneous lesions
Super-infection of cut-aneous lesions/Proctitis
Proctitis
Soft-tissue superinfection (upper lip)
Days from OD to admission/treatment
5/12
10/18
7/10
9/11
11/13
6/7
3/6
9/12
4/11
8/11
11/12
10/11
9/10
7/9
5/6
9/9
10/12
7/8
5/7
Days from treatment to recovery
13
10
21
9
12
11
14
4
7
18
6
21
14
20
7
18
17
27
15
* HIV diagnosis 2 months before MPX; ** AIDS presenters with HIV/AIDS diagnosis six months before MPX (multidrug resistant disseminated tuberculosis on treatment); *** recent virological failure. Abbreviations: M, male; y, years, MSM men who have sex with men; Neg, negative; Pos, positive; Unk, unknown; ART, antiretroviral therapy; VL, viral load; ND, not detectable, PREP, pre-exposure prophylaxis; BIC, bictegravir; TAF, tenofovir alafenamide fumarate; FTC, emtricitabine; 3TC, lamivudine; DTF dolutegravir, TDF, tenofovir disoproxil fumarate; DRV/c, darunavir/cobicistat; EFV, efavirenz; NA, not available; STD, sexual transmitted disease; CT computed tomography; OD, onset date.
Finally, viral kinetics have been evaluated in 12 patients (Fig. 1). In all of them, MPXV-DNA was detected in at least one sample from at least one compartment. Particularly, during the follow-up, MPXV-DNA was detected by real-time PCR in: 10/12 patients on oropharyngeal swab (OPS), including 9 at the start of antiviral treatment, with a median Ct of 36 (IQR 33-41); 8/9 patients on blood samples with a median Ct of 41 (IQR 37-41); 6/6 patients on feces with a median Ct of 41 (IQR 36-41); 3/3 patients on saliva with a median Ct of 38 (IQR 32-40); 3/3 patients on seminal fluids with a median Ct of 39 (IQR 37-41). In almost all patients, a progressive decline in viral load was observed over the course of treatment. Most biological samples were negative at the last available observation. DdPCR results approximately mirrored the viral shedding expressed with real-time PCR. It is worth noting that, given the low threshold used for the ddPCR, several samples with high Ct values in real-timePCR, resulted negative in ddPCR.
Fig. 1Kinetics of MPXV DNA shedding in different biological samples from starting of the antiviral therapy. A) MPXV DNA levels detected in different longitudinal samples are shown for the 12 MPX patients followed up during infection. B) MPXV DNA levels detected in the different type of samples: blood, oropharyngeal swabs (OPS), stool, sperm and saliva. MPXV DNA levels are expressed as cycle threshold (Ct) values. In grey are shown digital droplet PCR (ddPCR) results express as Log10 copies/µL with a detection limit of the assay of 5 copies/µL.
Limited data on clinical effectiveness of tecovirimat are available, however, several recent reports on its use has shown good tolerability and no evolution versus severe disease in treated subjects [
]. Additionally, preliminary results from the first 549 MPX-positive patients treated with tecovirimat in United States (US), showed median time to subjective improvement of 3 days [
]. To the best of our knowledge, this is the first report of the use of antivirals for MPX with both clinical and virological results in this current outbreak. One case series of patients treated in 2018-2021 reported viral decay in one patient during tecovirimat treatment showing a shorter duration of viral shedding compared to the other patients [
]. Additionally, in a pre-print publication, outcomes, including viral kinetics, of 14 patients treated before February 2022 with tecovirimat were reported [
Festus Mbrenga, Emmanuel Nakouné, Christian Malaka, Josephine Bourner, J ake Dunning, Guy Vernet, Peter Horby PO. Monkeypox treatment with tecovirimat in the Central African Republic under an Expanded Access Programme Author. medRxiv Prepr doi:10.1101/2022.08.24.22279177.
Festus Mbrenga, Emmanuel Nakouné, Christian Malaka, Josephine Bourner, J ake Dunning, Guy Vernet, Peter Horby PO. Monkeypox treatment with tecovirimat in the Central African Republic under an Expanded Access Programme Author. medRxiv Prepr doi:10.1101/2022.08.24.22279177.
], in our patients clinical improvement and no new lesions were reported in almost all patients 72 hours after tecovirimat initiation. The longer time elapsed from symptoms onset to treatment start (21 days) compared to our study (12 days) might partially explain this different result. Of note, in our case series, 15% of MPX cases diagnosed received antiviral treatment, consistently with US data [
Concerning viral kinetics, it should be noted that low viral loads were observed. Additionally, some patients had all available samples negative in ddPCR since antiviral starting, in line with previous evidence showing that viral shedding occurs mainly during the first two weeks of the disease [
]. Due to the median time of 12 days from symptoms onset to starting treatment in this series, we cannot exclude a reduced impact of antiviral therapy on viral shedding or clinical resolution.
The main limitations of this study was the lack of control group, so that any conclusions on the effectiveness of antiviral therapy cannot be drawn, the small number of patients included, the heterogeneity of samples and the impossibility to collect samples for all the patients at each timepoint.
Data collected on observational studies such as this can help improve our knowledge of the use of antivirals for MPXV, waiting more robust results from the placebo-controlled randomized trial of tecovirimat for MPX.
Funding
This study was supported by Ricerca Corrente Linea 1 and 2, funded by the Italian Ministry of Health.
Conflicts of interests
The authors declare no conflict of interest for the present study.
Acknowledgments
INMI Monkeypox Study Group: Isabella Abbate, Alessandro Agresta, Camilla Aguglia, Alessandra Amendola, Andrea Antinori, Francesco Baldini, Tommaso Ascoli Bartoli, Alessia Beccacece, Rita Bellagamba, Giulia Berno, Aurora Bettini, Nazario Bevilacqua, Licia Bordi, Marta Camici, Priscilla Caputi, Fabrizio Carletti, Angela Corpolongo, Stefania Cicalini, Francesca Colavita, Alessandra D'Abramo, Angela D'Urso, Gabriella De Carli, Patrizia De Marco, Federico De Zottis, Silvia Di Bari, Lavinia Fabeni, Francesca Faraglia, Valeria Ferraioli, Carla Fontana, Federica Forbici, Concetta Maria Fusco, Marisa Fusto, Roberta Gagliardini, Anna Rosa Garbuglia, Saba Gebremeskel Teklé, Maria Letizia Giancola, Giuseppina Giannico, Emanuela Giombini, Enrico Girardi, Giulia Gramigna, Elisabetta Grilli, Susanna Grisetti, Cesare Ernesto Maria Gruber, Eleonora Lalle, Simone Lanini, Daniele Lapa, Gaetano Maffongelli, Fabrizio Maggi, Alessandra Marani, Andrea Mariano, Ilaria Mastrorosa, Giulia Matusali, Silvia Meschi, Valentina Mazzotta, Sabrina Minicucci, Claudia Minosse, Klizia Mizzoni, Martina Moccione, Annalisa Mondi, Vanessa Mondillo, Giorgia Natalini, Nicoletta Orchi, Sandrine Ottou, Jessica Paulicelli, Elisabetta Petrivelli, Maria Maddalena Plazzi; Carmela Pinnetti, Silvia Pittalis, Gianluca Prota, Vincenzo Puro, Silvia Rosati, Alberto Rossi, Gabriella Rozera, Martina Rueca, Laura Scorzolini, Eliana Specchiarello, Maria Virginia Tomassi, Massimo Tempestilli, Francesco Vaia, Francesco Vairo, Beatrice Valli, Alessandra Vergori, Serena Vita.
Festus Mbrenga, Emmanuel Nakouné, Christian Malaka, Josephine Bourner, J ake Dunning, Guy Vernet, Peter Horby PO. Monkeypox treatment with tecovirimat in the Central African Republic under an Expanded Access Programme Author. medRxiv Prepr doi:10.1101/2022.08.24.22279177.
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