Patient Characteristics
A total of 56 patients with steroid-refractory GI-GvHD were enrolled. Clostridium difficile infection was not observed among all patients, and were not responders to methylprednisolone (mPSL) at≥2mg/kg per day. Immunosuppressant as the secondary-line therapy was given to all the patients. Eight patients were excluded, including four reluctant to participate in the study and four failing to meet inclusion criteria (one for primary disease recurrence, two for combined TMA, one for combined CMV before FMT). Of 27 patients in FMT group, the data of three patients with < grade IV GI-GVHD were not selected for statistical analysis. Of 21 patients in the control group, the data of three patients (one with missed follow-up and two with < grade IV GI-GVHD) were not used for statistical analysis (Fig.1).
The patients’ information were shown in Table 1 and supplement table. For 24 FMT patients, the median age was 29 years (range 13 to 55). The male/female ratio was 16/8. The median stool volume was 865 mL/day (range 360 to 2,100 mL/day). The median stool frequency was 6 times/day (range 3 to 21 times/day). The median abdominal pain score was 3 (range 1 to 4) (Table 1 and supplement table). For the 18 patients in the control group, the median age was 31.5 years old (range 13 to 59) (vs FMT group p>0.05). The male/female ratio was 7/11(vs FMT group p>0.05). The median stool volume was 510 mL/day (range 250 to 1,400 mL/day) (vs FMT group p<0.05). The median stool frequency was 5 times/day (range 3 to 20 times/day) (vs FMT group p>0.05). The median abdominal pain score was 2 (range 0 to 4) (vs FMT group p>0.05) (Table 1 and supplement table). No difference was observed in the occurrence of hematologic disease, stem cells donor gender match and stem cells donor relationship in two group. In FMT group, 11 patients received 2 times of FMT, 9 patients received 1 time of FMT, 3 patients received 3 times of FMT and 1 patients received 6 times of FMT (supplement table).
Table 1
Baseline characteristics of patients
|
FMT
|
control
|
p
|
median Age(min-max)
|
29(13-55)
|
31.5 (13-59)
|
>0.05
|
Gender
|
>0.05
|
male
|
16
|
7
|
|
female
|
8
|
11
|
|
Volume ml
median(min-max)
|
865(360-2100)
|
510(250-1400)
|
<0.05*
|
Frequencies
|
6(3-21)
|
5(3-20)
|
>0.05
|
Abdominal pain
|
3(1-4)
|
2(0-4)
|
>0.05
|
Hematologic Disease
|
AML
|
8
|
9
|
>0.05
|
ALL
|
4
|
2
|
>0.05
|
MDS
|
5
|
4
|
>0.05
|
AA
|
4
|
1
|
>0.05
|
CML
|
2
|
0
|
>0.05
|
Others
|
1
|
2
|
>0.05
|
Stem cells donor gender match
|
13
|
7
|
>0.05
|
Stem cells donor relationship
|
Haplo-HSCT
|
20
|
14
|
>0.05
|
SIB-HSCT
|
2
|
3
|
>0.05
|
URD-HSCT
|
2
|
1
|
>0.05
|
Clinical Outcomes
With Cox regression model, we first declared that immunosuppressant did not affect the outcomes of the two groups (supplement table).
At Day 14, 13 (27%) patients in FMT group and none in control group achieved clinical remission according to modified intention-to-treat analysis (p<0.05). Meanwhile, 20(83%) patients in FMT group and 7(39%) patients in control group showed effective response (clinical remission + partial remission) (RR 7.86, 95%CI 1.88–32.9; p=0.005). Three (12.5%) patients died in FMT group and one (5.5%) patient died in control group (RR 0.41, 95%CI 0.04–4.33; p=0.46)(Fig.2)(Table 2 and supplement table). No relapse of GI-GvHD was recorded in all patients at this time point.
Table 2
Clinical results of 14th day and 21th day
|
|
FMT
(n=24)
|
Control
(n=18)
|
p
|
0 day
|
Stool volume ml
median(min-max)
|
865(360-2100)
|
510(250-1400)
|
<0.05*
|
Stool frequencies
|
6(3-21)
|
5(3-20)
|
>0.05
|
Abdominal pain score
|
3(1-4)
|
2(0-4)
|
>0.05
|
14th day
|
Stool volume ml
median(min-max)
|
200(0-1300)
|
500(0-1700)
|
<0.05*
|
Stool frequencies
|
2(0-9)
|
5(0-12)
|
<0.05*
|
Abdominal pain score
|
0(0-3)
|
2(0-4)
|
<0.05*
|
CR
|
13(54.2%)
|
0(0%)
|
<0.05*
|
Efficiency(CR+PR)
|
20(83.3%)
|
7(39%)
|
<0.05*
|
Die
|
3(12.5%)
|
1(5.5%)
|
>0.05
|
21th day
|
Stool volume ml
median(min-max)
|
180(0-2365)
|
450(0-1400)
|
>0.05
|
Stool frequencies
|
2(0-8)
|
4(0-15)
|
>0.05
|
Abdominal pain score
|
0(0-3)
|
2(0-4)
|
>0.05
|
CR
|
14(58.3%)
|
3(16%)
|
<0.05*
|
Efficiency(CR+PR)
|
16(66.7%)
|
9(50%)
|
>0.05
|
Die
|
5(20.8%)
|
2(11%)
|
>0.05
|
GI-GVHD Relapse
|
2(8.3%)
|
2(11%)
|
>0.05
|
CR(clinical remission); PR(partial remission) |
At Day 21, clinical assessments showed that clinical remission was significantly more obvious in FMT group than in control group (14[58.3%] of 24 vs 3[16%] of 18; RR 6.0, 95%CI 1.22–29.45; p=0.027), but clinical response did not difference (16 [66.7%] of 24 vs 9 [50%] of 18; RR 4.0, 95%CI 0.84–19.16; p=0.083). Five patients died in FMT group and two patients died in control group (5 [20.8%] of 24 vs 2 [11%] of 18; RR 0.48, 95%CI 0.08–2.79; p=0.41). Two patients in FMT group and two patients in control group showed disease relapse, but no significant difference (2 [8.3%] of 24 vs 2 [11%] of 18; RR 0.73, 95%CI 0.09–5.72; p=0.76). (Fig.2) (Table 2 and supplement table)
Within 90 days of follow-up, PFS showed no significant difference between two groups (HR 0.36, 95%CI, 0.1-1.28; p=0.11) (Fig.3A). The FMT group showed better OS (HR 7.0, 95%CI, 1.53-32.08; p=0.012) (Fig.3B). At the end of the research, the median survival time was >600 days in FMT group and 107 days in control group (HR 4.73, 95%CI, 1.58-14.14; p=0.005). Both the PFS (HR 0.24, 95%CI, 0.06-0.95; p=0.055) and OS (HR 5.97, 95%CI, 1.52-23.43; p=0.01) kept increasing during the follow-up time in FMT group(Fig.3CD).
Safety of FMT
Only one patient experienced thrombocytopenia after FMT and one patient developed cardiac event at day 3 after FMT. Although we were not sure whether there was a possible association between these events and FMT, it cannot be completely exclude. No other severe adverse events were observed in FMT group during seven days of follow-up after FMT. Other common adverse events include incomplete ileus in one patient,fever in one patient, vomiting and low fever in two patients, and grade-3 rash in two patients, no special treatment only symptomatic treatment need for these patients.
In overall results, the mortality rate was low in FMT group (HR 5.97, 95%CI, 1.52-23.43; p=0.01). No significant difference was observed in the occurrence of hemorrhagic cystitis(p>0.05), bacteria & fungi infection (p>0.05), CMV&EBV infection(p>0.05), septicemia (p>0.05), TMA (p>0.05), cardiac events (p>0.05), thrombocytopenia(p>0.05) and epilepsy (p>0.05). (Table 3 and supplement table)
Fecal Microbiota and Immunity functions Analysis
Available fecal samples at baseline and weeks 1 after FMT were used for microbiota analyses (N=10). Compared with that of the donors, the diversity of fecal microbiota in fecal samples of the patients was decreased (Fig.4A). Proteobacteria increased while firmicutes decreased at phylum level in the microbiota of the patients(Fig.4B). After Week 1, the microbiota composition was reconstructed in FMT patients, showing a trend back to normal(Fig.4CD). The bacterial diversity improved at Week 1 after FMT in half patients (5/10) (Figure S1A). Similar to the results of our prior study, the ratio of firmicutes to proteobacteria was restored (7/10), proteobacteria decreased(9/10), and firmicutes increased (6/10) after FMT(Figure S1B). Bacteroidetes increased (7/10) in the fecal microbiota of patients with steroid-refractory GI-GvHD (Figure S1C) after FMT.
We also recorded the change in the levels of peripheral immunity cells (CD4+ cells, CD8+ cells or CD16+CD56+ cells, CD19+ cells, Treg cells) of four patients during four weeks’ follow-up after FMT. Unfortunately, we did not find any obvious trends(Figure S2).