First-in-human high-cumulative-dose stem cell therapy (PDF)

Received: 3 February 2019 Accepted: 18 August 2019 DOI: 10.1002/sctm.19-0037 HUMAN CLINICAL ARTICLES First-in-human high-cumulative-dose stem cell therapy in idiopathic pulmonary fibrosis with rapid lung function decline Alexander Averyanov1,2 | Irina Koroleva1 | Mikhail Konoplyannikov1 |...

Received: 3 February 2019 Accepted: 18 August 2019 DOI: 10.1002/sctm.19-0037 HUMAN CLINICAL ARTICLES First-in-human high-cumulative-dose stem cell therapy in idiopathic pulmonary fibrosis with rapid lung function decline Alexander Averyanov1,2 | Irina Koroleva1 | Mikhail Konoplyannikov1 | Veronika Revkova1 | Victor Lesnyak1 | Vladimir Kalsin1 | Olesya Danilevskaya1,2 | Alexey Nikitin1,2 | Anna Sotnikova1,2 | Svetlana Kotova3,4 | Vladimir Baklaushev1,2 1 Federal Research and Clinical Center of Federal Medical-Biologic Agency, Moscow, Abstract Russia Previous phase I studies demonstrated safety and some beneficial effects of mesenchy- 2 Pulmonology Scientific Research Institute mal stem cells (MSCs) in patients with mild to moderate idiopathic pulmonary fibrosis under Federal Medical-Biologic Agency, Moscow, Russia (IPF). The aim of our study was to evaluate the safety, tolerability, and efficacy of high 3 Institute for Regenerative Medicine, I. M. cumulative dose of bone marrow MSCs in patients with rapid progressive course of Sechenov First Moscow State Medical University, Moscow, Russia severe to moderate IPF. Twenty patients with forced ventilation capacity (FVC) ≥40% 4 Semenov Institute of Chemical Physics, and diffusing capacity of the lung for carbon monoxide (DLCO) ≥20% with a decline of Moscow both >10% over the previous 12 months were randomized into two groups: one group Correspondence received two intravenous doses of allogeneic MSCs (2 × 108 cells) every 3 months, and Vladimir Baklaushev, M.D., Ph.D., Federal the second group received a placebo. A total amount of 1.6 × 109 MSCs had been Research and Clinical Center of Federal Medical-Biologic Agency, 28 Orekhovy Blvd., administered to each patient after the study completion. There were no significant 115682 Moscow, Russia. adverse effects after administration of MSCs in any patients. In the group of MSC ther- Email: [email protected] apy, we observed significantly better improvement for the 6-min walk distance in Funding information 13 weeks, for DLCO in 26 weeks, and for FVC in 39 weeks compared with placebo. FMBA of Russia Government Order, Grant/Award Number: 20.001.13.800 FVC for 12 months in the MSCs therapy group increased by 7.8% from baseline, whereas it declined by 5.9% in the placebo group. We did not find differences between the groups in mortality (two patients died in each group) or any changes in the high-resolution computed tomography fibrosis score. In patients with IPF and a rapid pulmonary function decline, therapy with high doses of allogeneic MSCs is a safe and promising method to reduce disease progression. KEYWORDS adult human bone marrow, adult stem cells, bone marrow stromal cells, cellular therapy, clinical trials, lung, stem cell transplantation, transplantation tolerance 1 | I N T RO D UC T I O N origin characterized by the development of fibrotic transformation of the lung parenchyma, predominantly in the older population.1 Idiopathic pulmonary fibrosis (IPF) is the most common of the inter- The prevalence of IPF appears to be increasing, potentially stitial lung diseases, a progressing chronic disease with unknown explained by population aging.2 The survival median of patients with This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. © 2019 The Authors. STEM CELLS TRANSLATIONAL MEDICINE published by Wiley Periodicals, Inc. on behalf of AlphaMed Press STEM CELLS Translational Medicine. 2019;1–11. wileyonlinelibrary.com/journal/sct3 1 2 AVERYANOV ET AL. IPF is 3 to 5 years; at the same time, the individual survival of patients may be extremely variable.3–5 Conventionally, the IPF clini- cal course can be categorized into three types based on the rate of Significance statement The results of this first-in-human clinical trial revealed that the forced ventilation capacity (FVC) decline: slow deterioration (less high cumulative dose of mesenchymal stem cells (MSCs) is than 10% decline in FVC over 6 months or no changes in the dis- safe and well tolerated by patients' idiopathic pulmonary ease indicators for many months), intermittent deterioration with fibrosis with a rapid lung function decline. During the treat- episodes of exacerbations and rapid deterioration (more than 10% 3,6,7 ment period, the patients in the main group had their lung decline in FVC for 6 months). For the last type, before the era function increased, as compared to the placebo group, in of antifibrotic medications, the life expectancy of patients did not which the continued decline of the lung function was usually exceed 2 years from the time of diagnosis.6–8 Besides acute observed. Thus, this study states the safety, tolerability, and exacerbations, a poor prognosis might be suspected in patients with potential benefits of greater doses of MSCs than those used IPF with such predictors as low values of FVC and diffusing capacity earlier in patients with idiopathic pulmonary fibrosis, and of the lung for carbon monoxide (DLCO), old age, and a high rate of these findings might move future trials toward a new step in pulmonary function decline.3,9 The majority of clinical studies on the stem cells transplantation. drug therapy of IPF did not evaluate the pretreatment functional 10,11 reduction rate ; therefore, it is difficult to judge its efficiency in patients with a fast lung function decline. After a large number of successful preclinical studies, a treat- series included two intravenous infusions performed in a 7-day inter- ment with mesenchymal stem cells (MSCs) is considered a potential val. Each infusion contained a suspension of 200 million cells in new direction for lung fibrosis therapy.12,13 The potential effects of 400 ml of normal saline. The last MSCs administration was performed MSCs in pulmonary fibrosis are related to their ability to produce a at 39 weeks after the study beginning. The total number of MSCs large number of biologically active substances with anti-inflamma- administered in group 1 was 1.6 × 109 for the patients who com- tory, immunosuppressive, and angiogenic properties.12 The results pleted the protocol. Group 2 received a placebo (400 ml of normal of the published clinical trials on the cell therapy for IPF confirmed saline) with the same schedule. Sixteen patients completed the study the safety of MSCs; however, all of these studies included patients (Figure 2). with mild to moderate impairment of lung function, without taking The total characteristics of enrolled patients in each group are into account the rate of their preceding decline. Also, the total dose presented in Table 1. None of the patients took antifibrotic drugs of administered cells did not exceed 2 × 108.14–16 We performed because of their inaccessibility in the country during the study. Most the first clinical trial in this field in patients who had a fast lung of patients took prednisone as a single drug approved by the current function decline, using a significantly more intensive intervention national guidelines. The diagnosis of IPF was based on the results of a with respect to both the individual MSCs dose and the frequency of multidisciplinary discussion (MDD) according to American Thoracic dose administration. Society/European Respiratory Society criteria.1 The primary endpoint was the evaluation of treatment-related adverse events (AEs), related 2 | MATERIALS AND METHODS to timing of each dosing regimen of the MSCs suspension. Vital signs (including body temperature, pulse rate, respiration rate, and blood 2.1 | Study design pressure), oxygen saturation, 12-lead electrocardiogram, and clinical The phase I/IIA study reported here was randomized, open-label, and laboratory tests (including hematology, chemistry, and urinalysis) were placebo-controlled in two groups and was conducted at the Federal assessed before and during 3 days after each infusion. The AE fre- Research Clinical Center of Federal Medical-Biologic Agency of quency and severity were coded according to the Medical Dictionary Russia. The study was approved by the local ethical committee for Regulatory Activities, version 16.1. All AEs were recorded from the (approval 5-11-03-2013 of March 03, 2013) and registered at time the subject signed the informed consent until 12 weeks after the ClinicalTrials.gov (identifier NCT02594839). last dose of MSCs or placebo. The secondary endpoints were as Twenty patients were included into the study (age 33-74 years, follows: (a) changes in the lung function (FVC, DLCO), (b) physical tol- 11 males, 9 females), all of whom had a histologically or radiologically erance (6-min walk test distance (6MWTD) at week 13, 26, 39, and confirmed pattern of usual interstitial pneumonia (UIP), with a history 52 of the therapy, and (c) a semiquantitative estimation of pulmonary of a lung function decline (FVC or DLCO) ≥10% over the last fibrosis by high-resolution computed tomography (HRCT) data at 12 months and a current FVC ≥40% of predicted and DLCO ≥20% of week 26 and 52. Spirometry and diffusion tests were conducted in predicted. The complete list of inclusion/exclusion criteria is shown in accordance with european respiratory society (ERS)/american thoracic Supporting Information Appendix S1. According to the study protocol, society (ATS) regulations.17,18 The 6MWTD was performed according those patients that met the eligibility criteria were randomized into to the АТS protocol.19 A semiquantitative assessment of the HRCT two groups, with 10 patients per group: group 1 received four series pulmonary fibrosis score was performed by two independent radiolo- of MSCs intravenous infusions, repeated after 12 weeks. Each of the gists using the method of Oda et al.20 STEM CELL THERAPY IN IDIOPATHIC PULMONARY FIBROSIS 3 TABLE 1 Descriptive statistics for the groups at the baseline point* Disease FVC DLCO Age duration BMI Prednisone (% of (% of UIP biopsy Smoking Group Sex (years) (months) (kg/m2) (dose mg/daily) predicted) predicted) proved history Group 1: MSC treatment 1 M 61 21 27 20 67 41 + + 2 F 60 30 23 20 75 22 + + 3 M 50 27 30 25 36 24 + − 4 F 74 51 19 0 79 28 − − 5 F 58 84 18 10 43 28 − + 6 M 33 16 27 15 52 31 + + 7 M 68 27 25 10 48 33 + − 8 M 61 45 23 20 55 37 − + 9 F 59 48 22 20 51 22 + − 10 F 70 66 19 25 40 20 + + Summary 5/5 59.4 ± 11.5* 41.5 ± 21.5* 23.3 ± 4.0* 16.5 ± 7.8* 54.6 ± 14.6 28.6 ± 6.9 7* 6* Group 2: Placebo 1 M 56 30 20 20 61 38 + + 2 F 59 18 26 25 53 29 − − 3 F 65 21 26 20 48 26 + − 4 M 69 51 24 15 61 35 + − 5 M 52 24 28 12.5 50 31 + − 6 M 71 45 20 15 45 24 − + 7 M 69 48 26 15 52 30 − + 8 F 60 54 29 15 64 39 + − 9 M 58 18 21 25 45 21 + + 10 F 66 12 17 25 49 21 − − Summary 6/4 62.5 ± 6.4* 32.1 ± 15.8* 23.7 ± 4.0* 18.8 ± 4.9* 52.8 ± 6.9* 29.4 ± 6.5* 6* 4* Note: Data are presented as mean ± SD. Abbreviations: BMI, body mass index; DLCO, diffusing capacity of the lung for carbon monoxide; F, female; FVC, forced ventilation capacity; M, male; UIP, usual interstitial pneumonia. *There were no significant differences between the study groups (p >.05, using general linear model). 2.2 | MSCs preparation bone marrow-derived mononuclear cells according to the standard protocol.21 2.2.1 | Isolation of human MSCs For half of the patients (n = 5), the bone marrow for MSCs separa- 2.2.2 | Flow cytometry analysis tion was obtained from young (20-35 years of age), healthy blood relatives. In cases of the absence of young, healthy relatives or MSCs (passage 3-5) were washed with phosphate-buffered saline containing 1% fetal bovine serum (FBS). Fluorescein isothiocyanate- their refusal (n = 5), we used the bone marrow from healthy donors conjugated anti-human CD34, CD45, and CD105 antibodies and with the same sex as the patient. In these cases, donors for each phycoerythrin-conjugated antihuman CD29, CD 44, CD73, and CD90 MSC injection were randomized. Each donor signed the informed antibodies were used for staining the cells. All antibodies were pur- consent, and the protocol of bone marrow harvesting was approved chased from Miltenyi Biotec. The analysis was performed with a by the local ethics committee of the Federal Research Clinical Cen- CyFlow Space flow cytometer (Sysmex Partec) using the Partec ter of FMBA of Russia (4-11-03-2013 of March 03, 2013). FloMax flow cytometry Data Acquisition and Analysis Software. A total of 100 to 150 ml of bone marrow was collected from donors pretested for cytomegalovirus (CMV), human immunodeficiency virus 2.2.3 | Osteogenic, adipogenic, and chondrogenic 1 (HIV-1) and 2, human T-lymphotropic virus 1 (HTLV-1) and differentiation 2, Epstein–Barr virus (EBV), B19, hepatitis B virus, and hepatitis C virus into heparin-containing test tubes (100 U/ml of punctate; Sigma, The cultured MSCs (passage 5) were differentiated into the osteo- Sigma-Aldrich Corp.St. Louis, MO, USA). MSCs were obtained from genic, adipogenic, and chondrogenic lineage by culturing in the 4 AVERYANOV ET AL. osteogenic medium (Dulbecco's modified Eagle's medium [DMEM] was prepared from the patients' plasma by freezing–thawing of the supplemented with 10−8 M dexamethasone [Sigma, D4902], 10 mM platelet fraction. The cells were kept in the hypoxic (5% О2) condi- b-glycerophosphate [Sigma, G9422], and 50 μg/ml ascorbic acid), tions for the entire duration of culturing that, besides the activation of adipogenic medium (DMEM supplemented with 10 mM 3-isobutyl- glycolytic processes characteristic of multipotent stem cells, allowed 1-methylxanthine [Sigma, 17018], 0.1 mM indomethacin [Sigma, minimization of oxidative damage to the cell membrane systems. 17378], 10 μg/ml insulin [Sigma, I6634], 10−6 dexamethasone), and When making each preparation, the following was controlled: (a) cell chondrogenic medium (Stempro, Invitrogen, Life Technologies Ltd concentration (2 × 106 for each preparation); (b) survival—using a Luna Paisley PA4 9RF, UK), with the subsequent staining with alizarin red 2 cell counter (it was no less than 95% in each preparation directly before (Sigma), Oil Red O (Sigma), and Alcian blue (Sigma), respectively. the injection); and (c) the absence of mycoplasma contamination (using A Nikon Eclipse Ci microscope (Nikon Instruments Inc., NY, USA) was the real-time PCR analysis, Mycoplasma Detection Kit; ThermoFisher). used for the image capture. 2.3 | Statistical analysis 2.2.4 | Karyotyping A statistical power analysis showed 76% and 94% for FVC and DLCO, Karyotypes were analyzed in MSCs from the first and the last (4 or 5) respectively, based on a sample size of 20 patients in two comparable passages that were isolated and cultured in vitro using the chromosome groups. The primary efficacy endpoint of the study was the rate of multicolor fluorescence in situ hybridization (mFISH) technique.22 The decline in percent of predicted FVC value from baseline to week 52. A cells were incubated for 4 h with colchicine (40 μg/ml), and then the generalized linear repeated measures model was used as the primary cells were incubated in 0.075% KCl fixed with Carnoy's solution efficacy analysis. The secondary efficacy endpoints of the study were (3:1 vol/vol absolute ethanol: glacial acetic acid), dropped onto slides, changes in percent of predicted DLCO value, 6MWD test (meters), and dried. mFISH for chromosome karyotyping visualization was then and chest HRCT fibrosis score (points) from baseline to week 52. performed using the 24 XCyte color kit (Meta-Systems, Altlussheim, Another secondary efficacy endpoint included the change in a rate of Germany) according to the manufacturer's recommendations. FVC decline (percentage of the predicted value) from 12 months before the treatment to baseline and from baseline to week 52. To compare the study groups, we used multivariate tests and nonpara- 2.2.5 | MSC preparation for their injection to metric Mann–Whitney U test. The interobserver agreement for the patients: Management of cell therapy-related risks HRCT-based fibrosis score was calculated in accordance with the rec- At the first stage of the study, we performed an exam of each donor for ommendations by Kundel and Polansky, using Cohen's kappa coeffi- the absence of bloodborne diseases (HIV-1 and -2; hepatitis C virus; cient with 95% confidence limits.23 A safety analysis was performed hepatitis B, D, and E; syphilis; CMV, EBV, HTLV, acute myeloid leukemia, in the safety population. All AEs were compared in two groups by and chronic myeloid leukemia) to comply with the safety requirements. Fisher's exact criteria. A two-sided probability threshold of.05 was For injection to patients, MSCs of a third to fifth passage were used, considered statistically significant. The descriptive data are presented characterized by CD markers, karyotype, sterility tests, and apyrogenicity as mean ± SD and the other data as median and interquartile range. (for each MSC series). The preparations' sterility was tested via the seeding on culture media and using the matrix-assisted laser desorption/ 3 | RESULTS ionization (MALDI)-time-of-flight mass spectroscopy analysis (Microflex LRF, Bruker) in the federal research and clinical center (the main 3.1 | MSCs characterization affilation) bacteriological laboratory. For the mass spectroscopy, we used MALDI Sepsityper Kit, modified for the conditioned media (CM). Briefly, The results from the flow cytometric analysis revealed that the stan- all CM from MSCs was collected and incubated for 5 days in bacteriologi- dardized culture of human MSCs in vitro resulted in stable expression cal thermostat. At the same time, the samples of CM were seeded in the of the surface markers following a serial passage. The flow cytometry bacteriological media. Additionally, the cells from each batch were propa- analysis showed that the cultured MSCs had a CD29+ CD44+ CD73+ CD90+ CD105+ CD34− CD45− phenotype, the one characteristic of 9 gated up to 10 , centrifuged, and prepared for mass spectroscopy according to the manufacturer protocol. The taxonomic dataset consist of adult human MSCs (Figure 1A). The differentiation potential of MSCs 7854 spectra of prokaryotic and fungi organisms. No cases of microbial or was identified by their osteogenic, adipogenic, and chondrogenic dif- fungal contamination were detected. Although the contamination has not ferentiation and demonstrated by positive staining with alizarin red, been detected by mass spectroscopy, the CM samples from each cell Oil Red O, and Alcian blue, respectively (Figure 1B–D). A normal dip- batch were also tested using chromogenic limulus amebocyte lysate-test loid karyotype with 46 chromosomes and no abnormal changes in (Hycult biotech, HIT302), which is more sensitive for bacterial endotoxin the chromosome structure was observed by the analysis of 30 meta- detection. In each case, the level of endotoxin was less than 0.1 UE/ml. phase cells by mFISH visualization method (Figure 1E). In order to minimize the negative effect of the medium compo- Thus, the characterization of cells prior to administration revealed nents, MSCs were cultured without the FBS addition; instead, the that we obtained three potent MSCs with normal diploid karyotype autologous serum and platelet lysate were used. The platelet lysate and a high level of expression of MSC-specific surface markers. STEM CELL THERAPY IN IDIOPATHIC PULMONARY FIBROSIS 5 F I G U R E 1 Characterization of cultured human bone marrow-derived mesenchymal stem cell (MSCs; passage 4) before the transplantation. A, CD-immunophenotyping of MSCs using flow cytometry. Red histograms represent isotype specific Ig control; blue histograms: fluorescein isothiocyanate/PE-conjugated antihuman CD antibodies. B–D, Differentiation analysis. MSCs were characterized by their differentiation potential by staining with Oil Red O—adipogenic lineage, B, Alcian blue—specific to sulfated GAGs, C, and alizarin red—osteogenic lineage, D. Magnification ×200, B–D. E, Karyotype analysis of human MSCs, mFISH visualization method TABLE 2 Adverse effects Subjects, n (%)/events, n 1 2 Total Events (n = 10) (n = 10) (n = 20) p value All AEs 8 (80.0) // 28 7 (70.0) // 16 15 (75.0) // 44 >.999^ Death 2 (20.0) // 2 2 (20.0) // 2 4 (20.0) // 4 >.999^ Transient fever 4 (40.0) // 6 1 (10.0) // 1 5 (25.0) // 7.303^ Weakness 4 (40.0) // 4 1 (10.0) // 1 5 (25.0) // 5.303^ Cough 2 (20.0) // 2 2 (20.0) // 2 4 (20.0) // 4 >.999^ Headache 2 (20.0) // 4 2 (20.0) // 2 4 (20.0) // 6 >.999^ LRTI 2 (20.0) // 2 2 (20.0) // 2 4 (20.0) // 4 >.999^ Nausea 2 (20.0) // 2 2 (20.0) // 2 4 (20.0) // 4 >.999^ URTI 2 (20.0) // 3 2 (20.0) // 2 4 (20.0) // 5 >.999^ Chill 2 (20.0) // 2 0 (0.0) // 0 2 (10.0) // 2.474^ IPF exacerbation 0 (0.0) // 0 1 (10.0) // 1 1 (5.0) // 1 >.999^ Ischemic stroke 1 (10.0) // 1 0 (0.0) // 0 1 (5.0) // 1 >.999^ Skin rash 0 (0.0) // 0 1 (10.0) // 1 1 (5.0) // 1 >.999^ Note: There were no significant differences between the study groups compared using the general linear model. Abbreviations: AEs, adverse events; IPF, idiopathic pulmonary fibrosis; LRTI, lower respiratory tract infection; URTI, upper respiratory tract infection. 3.2 | Safety died from respiratory failure progression due to IPF (one was Of the 20 initially recruited patients, 16 completed the study. Four autopsied) between weeks 14 and 34. All of these four patients had patients (patients 9 and 10 from each group, 3 females and 1 male) initially low indices of DLCO (20%–22% of predicted), FVC (40%– 6 AVERYANOV ET AL. TABLE 3 Dynamics of basic study points MSCs, Placebo 12 months before Baseline 13th week 26th week 39th week 52nd week FVC (% of predicted) MSCs 60.0 (52-72) 51.5 (43-67) 46.5 (41-62) 51.5 (44-67) 56* (43-69) 55.5* (42-68) Placebo 58.0 (55-68) 51.0 (48-61) 49.0 (45-56) 49.5 (45-57) 49.0 (44-53) 48.0 (43-52) DLCO (% of predicted) MSCs 29.0 (26-42) 28.0 (22-33) 28.5 (24-34) 29.0* (25-36) 28.0 (22-34) 28.0* (24-34) Placebo 30.0 (27-40) 29.5 (24-35) 27.5 (22-34) 28.5 (24-34) 27.5 (23-31) 26.5 (22-29) 6MWDT distance (m) MSCs ND 271.0 (150-381) 340* (240-390) 360** (261-423) 366.5** (270-404) 373.5** (273-429) Placebo ND 247.5 (197-346) 289 (229-377) 287 (223-370) 277 (220-362) 267.5 (212-359) HRCT score (points) MSCs ND 150.5 (135-172) ND 143.5 (134-157) ND 145.0 (135-165) Placebo ND 138.5 (130-149) ND 136.5 (131-146) ND 138.0 (133-143) Note: Data are presented as median (interquartile range). Abbreviations: 6MWTD, 6-min walk test difference; DLCO, diffusing capacity of the lung for carbon monoxide; FVC, forced ventilation capacity; HRCT, high-resolution computed tomography; ND, no data. *Statistically significant differences (p <.05) between the study groups. **Statistically significant differences (p <.01) between the study groups. FIGURE 2 CONSORT flow diagram 51%), and 6MWTD (89-150 m) and high scores of lung fibrosis mild and did not require study discontinuation. Light fever after an according to the HRCT data (167-190 points), and they took higher intravenous cell infusion was noted at least once over the whole study doses of prednisone (20-25 mg daily) than other patients did. duration in 4 out of 10 patients. One female patient in the MSCs Adverse effects were more frequently observed in the group therapy group developed an ischemic stroke, with almost all sensory receiving MSCs; among them were fever and chills, predominantly in and motor functions restored after anticoagulant and vascular the first day after infusion (Table 2). However, these reactions were therapy. In the placebo group 1, one male patient suffered an STEM CELL THERAPY IN IDIOPATHIC PULMONARY FIBROSIS 7 F I G U R E 3 Secondary endpoints dynamics. A–C, Differences (delta) of real values (y-axis) of forced ventilation capacity (FVC; A), diffusing capacity of the lung for carbon monoxide, B, and 6MWD, C, from the baseline level, during the treatment period (median, minimum and maximum, 25% and 75% quartiles are shown). D, Change from the baseline in FVC median (% of predicted) during 12 months before the treatment and during the treatment (by week 52) in the main group and the control. In the legend, 3, 6, 9, and 12 mean the values after 3, 6, 9, and 12 months, respectively exacerbation of IPF; no exacerbations were detected in the MSCs completion and reaching the total dose of 1.6 billion MSCs after group. The statistical analysis did not reveal any therapy-associated 12 months (by week 52), statistically significant differences were significant AEs (Table 2). observed in the dynamics of FVC (+3.7% vs. −9.5%), DLCO (−5.1% vs. −12.9%), and 6MWDT (+26.6% vs. −9.8%) from the baseline (Figure 3A–C). 3.3 | Overall effects of MSCs versus placebo The interobserver agreement (κ value) in the HRCT signs estimation After the first 3 months of treatment (administration of 400 million was in general high and determined as 0.69 for the reticular changes, MSCs), we did not observe a statistically significant difference between 0.75 for the traction bronchiectasis, 0.85 for the normal attenuation, the groups with regard to any of the studied indices, except for the and 0.88 for the honeycombing. The HRCT fibrosis score did not differ 6MWTD, which increased by 15.3% in the MSCs group and decreased significantly from the baseline by week 26 (0.7% for MSCs vs. 0% for by 2.7% in the placebo group (Table 3). By the sixth month of the ther- placebo) and by week 52 (1.7% for MSCs vs. 0.7% for placebo). apy, the MSCs group showed a significantly better DLCO change When evaluating the FVC decline for the 12 months preceding ran- (−1.2% in the MSCs group vs. −6.5% in the placebo group) and domization and during the 52 weeks of the therapy, we found a signifi- 6MWTD (+22.0% vs. −3.4% in the placebo group) compared with base- cant improvement in the group receiving MSCs (from −13.8% to +3.7%) line. After 9 months, and a total cell dose of 1.2 × 10 , the difference in 9 as compared with the placebo group (from −11.7% to −9.5%; the dynamics from baseline points was observed in FVC (+4.7% Figure 3D). We analyzed individual patients in the MSCs therapy group, vs. −7.6%) and 6MWDT (+24.2% vs. −6.7%). After the treatment who had a continuing decline of FVC and DLCO >10% over the 8 AVERYANOV ET AL. observation period (patients 4 and 5), as well as deceased patients four died after four cell (8 × 108) or placebo infusions. We believe that (patients 9 and 10). All patients with no response to the therapy were cell therapy might be ineffective at this advanced stage of the disease. females, had the longest disease history (48-84 months), and had low Our data differ from the results of Chambers et al.,14 who observed 2 body mass index (BMI; three out of four had a BMI of 18 to 19 kg/m , a significant increase in 6MWD and a marginal improvement in DLCO 2 one a BMI of 22 kg/m ) and a low initial level of DLCO (