key: cord-0767751-2xtrgkp8
authors: Shi, L.; Huang, H.; Lu, X.; Yan, X.; Jiang, X.; Xu, R.; Wang, S.; Zhang, C.; Yuan, X.; Xu, Z.; Huang, L.; Fu, J.; Li, Y.; Zhang, Y.; Liu, W.; Liu, T.; Song, J.-W.; Sun, L.; Yang, F.; Zhang, X.; Zhang, B.; Shi, M.; Meng, F.; Song, Y.; Yu, Y.; Wen, J.; Li, Q.; Mao, Q.; Maeurer, M.; Zumla, A.; Yao, C.; Xie, W.; Wang, F.-S.
title: Treatment with human umbilical cord-derived mesenchymal stem cells for COVID-19 patients with lung damage: a randomised, double-blind, placebo controlled phase 2 trial
date: 2020-10-20
journal: nan
DOI: 10.1101/2020.10.15.20213553
sha: a26dc4f4d6fa9ef1cf8113a899d038946055f356
doc_id: 767751
cord_uid: 2xtrgkp8

Objective To assess the safety and efficacy of human umbilical cord-derived MSCs (UC-MSCs) for severe COVID-19 patients with lung damage. Design, Multicentre , randomised, double-blind, placebo-controlled trial. Setting Two hospitals in Wuhan, China, 5 March 2020 to 28 March 2020. Participants 101 severe COVID-19 patients with lung damage aged between 18-74 years. Intervention Patients were randomly assigned at a 2:1 ratio to receive either UC-MSCs (40 million cells per infusion) or placebo on days 0, 3, and 6. Main outcome measures The primary endpoints were safety and an altered proportion of whole lung lesion size from baseline to day 28, measured by chest computed tomography. Secondary outcomes were reduction of consolidation lesion sizeand lung function improvement (6-minute walk test, maximum vital capacity, diffusing capacity). Primary analysis was done in the modified intention-to-treat (mITT) population and safety analysis was done in all patients who started their assigned treatment. Results 100 patients were finally recruited to receive either UC-MSCs (n = 65) or placebo (n = 35). The patients receiving UC-MSCs exhibited a trend of numerical improvement in whole lung lesion size from baseline to day 28 compared with the placebo cases (the median difference was -13.31%, 95%CI -29.14%, 2.13%, P=0.080). UC-MSCs administration significantly reduced the proportions of consolidation lesion size from baseline to day 28 compared with the placebo (median difference: -15.45%, 95% CI -30.82%, -0.39%, P=0.043). The 6-minute walk test showed an increased distance in patients treated with UC-MSCs (difference: 27.00 m, 95% CI 0.00, 57.00, P=0.057). The incidence of adverse events was similar, and no serious adverse events were observed in the two groups. Conclusions UC-MSCs treatment is a safe and potentially effective therapeutic approach for COVID-19 patients with lung damage. A phase 3 trial is required to evaluate effects on reducing mortality and preventing long-term pulmonary disability.

1. Department of Infectious Diseases, Fifth Medical Center of Chinese PLA General 1 1 . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020. product. Methods for data collection and in-study measurements are described in detail in 2 0 8 Supplement 1. 2 0 9

Imaging and clinical outcomes 2 1 0

All patients underwent high-resolution chest CT examination at baseline, day 10, and day 2 1 1 28. The primary outcome was gauged as a change in the total lesion proportion (%) of the 2 1 2 whole lung volume from baseline to day 28, as measured by chest CT. It was defined as 2 1 3 (total lesion proportion of the whole lung volume at day 28-total lesion proportion of the 2 1 4 whole lung volume at baseline) / total lesion proportion of the whole lung volume at 2 1 5 baseline. The secondary imaging outcomes were a change in the total lesion proportion (%) 2 1 6 of the whole lung volume from baseline to day 10, a change in consolidation and 2 1 7 ground-glass lesion proportion from baseline to day 10, 28 , and change in lung 2 1 8 densitometry at day 10, 28. Lung lesions were evaluated by using the changes in 2 1 9 high-resolution chest CT images and measured by centralised imaging interpretation based 2 2 0 on both lung radiologist analyses and imaging software. The imaging data were derived 2 2 1 from a software-assisted lung volumetry and densitometry procedure (Supplement 1).

Clinical outcomes within 28 days included 6-minute walk test (6-MWT), status of 2 2 3 oxygen therapy maximum forced vital capacity (VC max ), diffusion lung capacity for carbon 2 2 4 monoxide (DL CO ), modified Medical Research Council Dyspnoea Scale (mMRC), changes 2 2 5 in absolute lymphocyte counts and subsets, as well as plasma cytokine and chemokine 2 2 6 levels. Safety evaluation included adverse events and all-cause mortality. Detailed 2 2 7 definitions and assessment procedures are described in Supplement 1. This study was designed as a phase 2 clinical trial. The limited efficacy information of the 2 3 0 medication in patients with COVID-19 and the exploratory nature of this study meant that 2 3 1 . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020. the original target sample size was not justified by statistical calculation and was set as 45 2 3 2 patients, with an allocation ratio of 2:1. Minimal serious adverse events were observed in 2 3 3 our phase 1 trial; therefore, the sample size was expanded to 90, and finalized at 101, to 2 3 4 obtain more data from this study. Sample size adjustments were made in a manner that 2 3 5 maintained the double-blind status of this study and were approved by the institutional 2 3 6 review boards of the two participating hospitals. There were no pre-defined hypotheses made in this study; therefore, we focused on 2 3 8 description instead of inference for statistical analyses: all statistical tests, confidence 2 3 9 intervals, and P-values were used for exploration, not for inference. For primary outcome 2 4 0 analysis -the change in the total lesion proportion (%) of the whole lung volume from 2 4 1 baseline to day 28 and the difference between the UC-MSC and placebo groups was tested 2 4 2 using wilcoxon rank sum test and the median differences were calculated using the 2 4 3

Hodges-Lehmann estimation (It was also applied to other secondary outcomes which were 2 4 4 not in accordance with normal distribution.). Six category scale and MMRC dyspnea score 2 4 5 were calculated by using ordinal logistic regression model. The modified intention-to-treat 2 4 6 (mITT) population was considered as the primary analysis population and safety analysis 2 4 7 was done in all patients who started their assigned treatment. If the patient missed a chest 2 4 8 CT scan, the last scan's results were carried forward to the missing visit for primary 2 4 9 endpoints in the mITT analysis. Other missing values of secondary endpoints and 2 5 0 per-protocol analyses were not imputed. Statistical analyses were performed using SAS 2 5 1 software, version 9.4 (Cary, NC, USA). The figures were generated using GraphPad Prism 2 5 2 7 software (GraphPad Inc., La Jolla, CA, USA). No patients were involved in setting the outcome measures, nor were they involved in 2 5 5 developing plans for recruitment, design, or implementation of the study. No patients were 2 5 6 . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)

The copyright holder for this preprint this version posted October 20, 2020. asked to advice on interpretation or writing up the results. All participants will be informed 2 5 7 of the trial results by telephone or email. The trial results will be disseminated to the public 2 5 8 through public media. 2 5 9 2 6 0 . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)

The copyright holder for this preprint this version posted October 20, 2020. From March 5, 2020, to March 28, 2020, a total of 288 patients were screened at two 2 6 3 hospitals in Wuhan city. The majority of severe hospitalized COVID-19 patients were at 2 6 4 the convalescent stage and some of them were with progression stage. Among them, 101 2 6 5 eligible patients previously diagnosed as severe COVID-19 type, being referred as the ITT 2 6 6 placebo group. Only one case experienced a grade 3 adverse event (pneumothorax) in the 3 3 4 . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020. other adverse events at grade 1 or 2 in both groups. All adverse events during the 3 3 6 observation period were judged by the site investigators and found to be unrelated to 3 3 7 UC-MSC intervention. No deaths were observed in this trial. Whilst several trials of the therapeutic use of MSCs for patients with COVID-19 have been 3 4 0 registered at Clinicaltrial.gov, there are no data available to date from randomised 3 4 1 placebo-controlled clinical trials yet. This is the first report of a double-blind, randomised, 3 4 2 and controlled phase 2 trial, in COVID-19 patients with lung damage. Our data found that under what is known as 'expanded access compassionate use' for COVID-19 patients. A 3 5 1 phase 3 trial is now required to further evaluate effects on mortality and long term 3 5 2 pulmonary disability, and to determine the underlying mechanisms of UC-MSC treatment 3 5 3 for COVID-19 disease. 22 3 5 4 COVID-19 is characterized by pathological lung changes in both the parenchyma 3 5 5 and interstitium 2 , including ground glass opacity, consolidation, traction bronchiectasis, 3 5 6 reticulation, and thickening of the bronchovascular bundles, as imaged using chest CT 23 . 3 5 7

Notably, the improvement of pulmonary lesions, especially the improvement of pulmonary 3 5 8 . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020. interstitial lesions, directly affects the recovery of lung function and the remission of 3 5 9 clinical symptoms. 24 The outbreak of COVID-19 occurred in China in January and early 3 6 0 February, and was basically brought under control in March. As enrolment ensured, many 3 6 1 severe hospitalized COVID-19 patients were at the convalescent stage and some of these 3 6 2 patients in the progression stage, which presented a challenge to explore clinical 3 6 3 improvement. Considering lung damage was still a common characteristic in these patients 3 6 4 at convalescent phase which affected their recovery and life quality, we modified the 3 6 5 primary outcome to the change in the total lesion proportion (%) of the whole lung volume 3 6 6 as measured by CT from baseline to day 28. The patients enrolled in this study were all 3 6 7 previously diagnosed as severe types, with a longer disease course and older age, 3 6 8 compared with the recent studies. 25, 26 In particular, all the patients suffered from serious 3 6 9 pulmonary damage and needed oxygen inhalation support during the course of disease. Our 3 7 0 current trial showed that UC-MSCs therapy improved the resolution of the whole lung 3 7 1 damage size, as detected by CT scanning, particularly the consolidation lesions. This finding 3 7 2 indicated that UC-MSC administration has a therapeutic benefit for patients with 3 7 3 COVID-19, even in the convalescent stage. It is known that the consolidation lesions in the 3 7 4 lung include the interstitial fibrosis. Thus, the improvements of consolidation lesions might 3 7 5 also imply the alleviation of lung fibrosis. 3 7 6

The 6-MWT has been used to evaluate patients suffering from a variety of 3 7 7 cardiopulmonary diseases. The results reflect the integrated reserve capability of complex 3 7 8 physiology, involving the pulmonary and cardiovascular systems, and neuromuscular 3 7 9 circulation 27 . In this trial, the 6-MWT was numerically, but not statistically, improved in 3 8 0 the MSC group compared with that in the placebo group. Given that there was no 3 8 1 significant difference in cardiovascular diseases between the two groups and no 3 8 2 . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020. It is hypothesized that the beneficial effect of MSC treatment for patients with severe 3 8 5 COVID-19 is mediated via reduction of pro-inflammatory cytokines, that jointly mediate 3 8 6 immune pathology and worsen clinical COVID-19 outcomes. 7,28-30 Cytokines such as 3 8 7 serum IL-6 are considered as biologically relevant biomarkers associated with disease 3 8 8 progression of COVID-19. In this study, UC-MSC infusion did not result in a significant 3 8 9 reduction in the duration of oxygen therapy, mMRC, cytokine, or chemokine levels, which 3 9 0 might be in part attributed to the status of the enrolled population, since most of them were 3 9 1 not in the acute progressive stage. Other mechanisms of actions have to be explored for 3 9 2

MSCs that are measurable in the systemic circulation. We could not, however, measure the 3 9 3 local, i.e. intrapulmonary, effects of MSC delivery. It could very well be that the local 3 9 4 MSC-mediated effects were not measurable in the systemic circulation, a similar scenario 3 9 5 as in MSC treatment of patients with corticosteroid-resistant graft-versus-host-disease 3 9 6 (GVHD). 3 9 7

In our trial, a total of three doses of 4 × 10 7 UC-MSCs were transfused for each 3 9 8 patient. No MSC-related predefined haemodynamic or respiratory adverse events were 3 9 9

observed. The incidences of adverse events were similar between the MSC group and the 4 0 0 placebo group. Only one patient in the MSC group suffered a pneumothorax that was 4 0 1 judged to be unrelated to UC-MSC medication. No patient died during the follow-up period. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020. There were several operational limitations to our study. A larger sample size could 4 0 8 have improved efficacy analyses. According to management guidelines issued by the 4 0 9

Chinese National Health Commission (7 th edition), 21 patients with COVID-19 require 4 1 0 further centralized isolation for 14 days after discharge. In this setting, some of the patients 4 1 1 missed the follow-up data at day 28, but they did receive a follow-up check around 7-10 4 1 2 days after the 28-day follow-up window. Importantly, our PP population analysis also 4 1 3 revealed similar results compared with mITT population analysis. Whether the cell dosage, 4 1 4 interval duration, and cycles of UC-MSC medication were the best regimen for patients 4 1 5 with severe COVID-19 were not fully investigated in this study. To the best of our knowledge, this is the first randomised, double-blind, 4 1 8 placebo-controlled trial evaluating the safety and preliminary efficacy of UC-MSCs as a 4 1 9 potential treatment for patients with COVID-19 with lung damage, even at the 4 2 0 convalescent stage. UC-MSC administration was safe and accelerated resolution of lung 4 2 1 consolidation lesions and improvement in the integrated reserve capability after UC-MSC 4 2 2 administration. UC-MSCs treatment offers a safe and potentially effective therapeutic 4 2 3 approach for COVID-19 patients with lung damage. A phase 3 trial is required to further 4 2 4 evaluate effects on preventing long-term pulmonary disability, reducing mortality and 4 2 5 determining the underlying mechanisms of UC-MSC treatment for COVID-19 disease. 4 2 6 . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020. 

All authors declare no competing interests. 4 4 7 . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020. Hospital of Hubei Province(2020IEC001).Written informed consent was obtained from all 4 5 2 the enrolled patients or their legal representatives if they were unable to provide consent. The lead author* affirms that this manuscript is an honest, accurate, and transparent 4 5 9 account of the study being reported; that no important aspects of the study have been 4 6 0 omitted; and that any discrepancies from the study as planned (and, if relevant, registered) 4 6 1 have been explained. 4 6 2 Dissemination to participants and related patient and public communities 4 6 3

Participants will be informed of the results of the study by telephone or mail. Dissemination to the public will be achieved through media outreach. Diseases consortium for Host-Directed therapies (https://www.fchampalimaud.org/covid19/aci/) 4 6 8

We thank Drs. Sibing Zhang, Dixiong Xu, Fangguo Dai and Bin Liu for logistic support 4 7 0 . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020. We thank randomization system provider (Chengdu Cims-medtech), PIATS provider 4 7 3 (Alibaba Health Technology) and academic secretaries (Yufeng Guo, Hairui Si, Lvshuai 4 7 4

Huang, Junqing Luan) for their services. We also thank the People's Government of 4 7 5

Wuhan Municipality for the help in organizing the follow-up work. 4 7 6 4 7 7

. CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020. A. Panel A shows the between-group median difference in the change in total lesion 5 9 1 proportion (%) and consolidation lesion proportion (%) of the whole lung volume from 5 9 2 baseline to day 28. I bars indicate the 95%CI described by Hahn and Meeker (1991) . consolidation lesion proportion (%) of the whole lung volume from baseline to day 28. 5 9 5 Q1 denotes the first quartile, and Q3 the third quartile. I bars indicate the minimum and 5 9 6 maximum. 5 9 7

C. Panel C shows the mean absolute change from baseline in the total lesion proportion (%) 5 9 8 and consolidation lesion proportion (%) of the whole lung volume. I bars indicate the 5 9 9 standard error. 6 0 0 . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020. ITT= intention-to treat population. mITT= modified intention-to treat population . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity (which was not certified by peer review) . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity (which was not certified by peer review) P a g e 1 o f 5 . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020 . . https://doi.org/10.1101 /2020 . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted October 20, 2020. Increased lactic acid dehydrogenase 9(13.85%) 0(0.00%) 7(20.00%) 0(0.00%) Increased alanine aminotransferase 7(10.77%) 0(0.00%) 4(11.43%) 0(0.00%) Hypokalaemia 6(9.23%) 0(0.00%) 1(2.86%) 0(0.00%)

Increased aspartate aminotransferase 5(7.69%) 0(0.00%) 4(11.43%) 0(0.00%) Increased serum uric acid 5(7.69%) 0(0.00%) 3(8.57%) 0(0.00%) Diarrhoea 4(6.15%) 0(0.00%) 0(0.00%) 0(0.00%) Palpitations 3(4.62%) 0(0.00%) 0(0.00%) 0(0.00%) Increased γ -glutamyl transferase 2(3.08%) 0(0.00%) 1(2.86%) 0(0.00%) Dizziness 2(3.08%) 0(0.00%) 0(0.00%) 0(0.00%) Cough 2(3.08%) 0(0.00%) 1(2.86%) 0(0.00%) abdominal distention 2(3.08%) 0(0.00%) 0(0.00%) 0(0.00%) Anemia 2(3.08%) 0(0.00%) 0(0.00%) 0(0.00%) Pneumothorax 0(0.00%) 1(1.54%) 0(0.00%) 0(0.00%)

Metabolic alkalosis 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Urinary tract infection 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Bacterial infection 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Pharyngitis 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Increased heart rate 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Creatine phosphate stimulation 1(1.54%) 0(0.00%) 1(2.86%) 0(0.00%) Elevated blood urea 1(1.54%) 0(0.00%) 1(2.86%) 0(0.00%) Poor sleep quality 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Taste reversal 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%)

Chest musculoskeletal 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%)

Pulmonary edema 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Pharyngeal diseases 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%)

. CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)

The copyright holder for this preprint this version posted October 20, 2020. . https://doi.org/10.1101/2020.10.15.20213553 doi: medRxiv preprint P a g e 5 o f 5 Anxious 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Nervous 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Rash 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Thirsty 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Nausea 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Countercurrent 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Non infectious gingivitis 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%)

Abdominal pain 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Functional gastrointestinal turbulence 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Vomit 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Gastroesophageal reflux disease 1(1.54%) 0(0.00%) 1(2.86%) 0(0.00%) Toothache 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) heart failure 1(1.54%) 0(0.00%) 0(0.00%) 0(0.00%) Hypocalcemia 0(0.00%) 0(0.00%) 2(5.71%) 0(0.00%)

Hepatic cyst 0(0.00%) 0(0.00%) 2(5.71%) 0(0.00%) Creatine phosphate stimulation 0(0.00%) 0(0.00%) 1(2.86%) 0(0.00%) Elevated serum creatinine 0(0.00%) 0(0.00%) 1(2.86%) 0(0.00%) Respiratory alkalosis 0(0.00%) 0(0.00%) 1(2.86%) 0(0.00%) Pleural effusion 0(0.00%) 0(0.00%) 1(2.86%) 0(0.00%) Difficulty in falling asleep 0(0.00%) 0(0.00%) 1(2.86%) 0(0.00%) Pruritus 0(0.00%) 0(0.00%) 3(8.57%) 0(0.00%)

. CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)

The copyright holder for this preprint this version posted October 20, 2020. . https://doi.org/10. 1101 /2020 

Data are median (interquartile range (IQR)), n (%),or mean (SD)

* BMI values were available for 59 patients in the UC-MSC group and 33 patients in the placebo group

10.1038/s41467-020-17240-2 [published Online First: 2020/07/10] 4 9 8