Sirolimus is effective for primary refractory/relapsed warm autoimmune haemolytic anaemia/Evans syndrome: a retrospective single-center study

Figures & data

Figure 1. Study profile.

A total of 68 patients were screened. Among those, 1 patient was excluded with an age of less than 18; 2 in combination with other drugs (except glucocorticoids); 3 with incomplete data; 4 with anaemia in chronic kidney disease or other types of anaemia; 3 discontinued sirolimus due to uncontrolled infection or tumour recurrence (treated with sirolimus for less than 6 months); 11 lost during the first 12 months of follow-up (7 lost with less than 3 months of follow-up, and 4 lost with 3–6 months of follow-up). The remaining 44 patients all have been treated with sirolimus for at least 6 months and followed up for at least 12 months.

Figure 2. Sirolimus plasma trough concentration in patients with response or no response to sirolimus. A line chart was conducted to assess intraindividual variability of sirolimus plasma trough concentration in each patient. One patient did not have a sirolimus plasma concentration test during the administration of sirolimus.

Table 1. Baseline characteristics.

Characteristics	Total (N = 44)	AIHA (N = 37)	ES (N = 7)	P value
Male sex, No. (%)	9 (20.5)	9 (24.3)	0 (0)	0.148
Age, y, median (range)	44 (18–86)	45 (18–86)	43 (30–72)	0.441
Time from diagnosis to sirolimus treatment, m, median (range)	15 (1–212)	15 (1–212)	58 (8–83)	0.150
Diagnosis, No. (%)
wAIHA	37 (84.1)	–	–	–
ES	7 (15.9)	–	–	–
Type of antibody, No. (%)				0.480
Warm (IgG)	33 (75.0)	27 (73.0)	6 (85.7)
Negative	11 (25.0)	10 (27.0)	1 (14.3)
Disease status, No. (%)				0.104
Refractory to glucocorticoid	25 (56.8)	23 (62.2)	2 (28.6)
Relapsed to glucocorticoid	19 (43.2)	14 (37.8)	5 (71.4)
Previous treatment, No. (%)
Glucocorticoid	44 (100)	37 (100)	7 (100)	1.000
Rituximab	6 (13.6)	5 (13.5)	1 (14.3)	0.975
CsA	13 (29.5)	10 (27.0)	3 (42.9)	0.405
CTX	10 (22.7)	7 (18.9)	3 (42.9)	0.171
IVIG	5 (11.4)	4 (10.8)	1 (14.3)	0.793
Azathioprine	5 (11.4)	4 (10.8)	1 (14.3)	0.793
Two or more lines of previous therapies	26 (59.1)	20 (54.1)	6 (85.7)	0.122
Haemoglobin, g/L, median (range)	85 (30–107)	85 (30–106)	83 (44–107)	0.759
Platelet, ×10^9/L, median (range)	189 (17–414)	211 (102–414)	57 (17–69)	3.55 × 10^-5
Absolute neutrophil count, ×10^9/L, median (range)	2.95 (0.90–12.61)	2.95 (0.90–8.73)	3.40 (1.21–12.61)	0.360
Absolute reticulocyte count, ×10^9/L, median (range)	195.2 (75.0–875.2)	202.0 (75.0–875.0)	188.4 (111.4–371.6)	0.656
Lactate dehydrogenase, U/L, median (range)	256 (114–491)	259 (138–405)	224 (114–491)	0.890
Serum total bilirubin, μmol/L, median (range)	30.1 (8.5–286.6)	34.5 (8.5–137.3)	18.7 (12.5–286.6)	0.285
Serum indirect bilirubin, μmol/L, median (range)	20.3 (5.7–251.3)	24.3 (5.7–128.1)	13.4 (9.0–251.3)	0.292
Serum ferritin, ng/mL, median (range)	364 (43–1661)	419 (43–1661)	171 (47–600)	0.188
With glucocorticoid tapering, No. (%)	13 (29.5)	13 (35.1)	0	0.065
Duration of glucocorticoid tapering, wks, median (range)	8 (2–19)	8 (2–19)	–	–
Sirolimus plasma trough concentration, ng/mL, median (range)*	4.5 (2.0–8.1)	4.5 (2.0–8.1)	4.3 (3.5–8.1)	0.742
Duration of sirolimus treatment, m, median (range)	23 (6–80)	23 (6–80)	23 (6–38)	0.712
Duration of follow-up, m, median (range)	25 (12–80)	26 (12–80)	23 (12–38)	0.688

wAIHA: warm autoimmune haemolytic anaemia; ES: Evans syndrome; IgG: immunoglobulin G; CsA: cyclosporin A; CTX: cyclophosphamide; IVIG: intravenous immunoglobulin.

* Based on the average sirolimus plasma trough concentration of the first 6 months after sirolimus of each patient.

Figure 3. Cumulative response rate and response rate at 3, 6, 12 months, and at the end of follow-up. A. The cumulative or rate curves. Patients who did not respond to sirolimus were censored at the end of follow-up. B. The cumulative CR rate curves. Patients who did not respond to sirolimus were censored at the end of follow-up. C. The efficacy of sirolimus at 3, 6, 12 months of treatment and at the end of follow-up.

Table 2. Response to sirolimus.

Response		Total (N = 44)	AIHA (N = 37)	ES (N = 7)
3m	CR	38.6%	43.2%	14.3%
	PR	22.7%	13.5%	71.4%
	NR	38.6%	43.2%	14.3%
6m	CR	43.2%	48.6%	14.3%
	PR	29.5%	18.9%	85.7%
	NR	27.3%	32.4%	0
12m	CR	50.0%	56.8%	14.3%
	PR	29.5%	18.9%	85.7%
	NR	20.5%	24.3%	0
Follow-up	CR	50.0%	56.8%	14.3%
	PR	22.7%	13.5%	71.4%
	NR	27.3%	25.0%	14.3%

AIHA: autoimmune haemolytic anaemia; ES: Evans syndrome; N: number; CR: complete response; PR: partial response; NR: no response.

Figure 4. Key laboratory parameter changes at 3, 6, 12 months, and at the end of follow-up. A. The haemoglobin levels at baseline, 3, 6, 12 months of treatment, and at the end of follow-up. The haemoglobin levels at 3, 6, 12 months, and at the end of follow-up were significantly higher than the baseline level (all p <0.001). B. The absolute reticulocyte counts at baseline, 3, 6, 12 months of treatment, and at the end of follow-up. The absolute reticulocyte counts at 12 months and at the end of follow-up were significantly lower than the baseline level (p = 0.005 and p < 0.001, respectively). C. The lactate dehydrogenase levels at baseline, 3, 6, 12 months of treatment, and at the end of follow-up. The lactate dehydrogenase levels at 12 months and at the end of follow-up were significantly lower than the baseline level (p = 0.006 and 0.036, respectively). D. The serum total bilirubin levels at baseline, 3, 6, 12 months, and at the end of follow-up. The serum total bilirubin levels at 3, 6, 12 months, and at the end of follow-up were significantly lower than the baseline (all p <0.001). E. The serum indirect bilirubin levels at baseline, 3, 6, 12 months, and at the end of follow-up. The serum indirect bilirubin levels at 3, 6, 12 months, and at the end of follow-up were significantly lower than the baseline level (all p <0.001). F. The platelet levels at baseline, 3, 6, 12 months of treatment, and at the end of follow-up. The platelet levels at 3 and 12 months were significantly higher than the baseline level (p = 0.037, 0.029, respectively), and there was a trend that the platelet levels at 6 months and at the end of follow-up were higher than the baseline level (p = 0.058, 0.055, respectively). In panels A-E, the horizontal line within each box represented the median, the lower and upper borders of each box represented the 25^th and the 75^th percentiles, respectively, and the I bars represented the adjusted minimum and maximum range. Dots represented outlier values.

Table 3. Adverse events during the treatment period.

AEs	Any grade	Grade 3*
	No. of patients with Aes (%)
Mucositis	5 (11.4)	0
Infection	4 (9.1)	1 (2.3)
Alanine aminotransferase elevation	4 (9.1)	0
Gastrointestinal disorder	2 (4.5)	0
Edema	2 (4.5)	0
Myalgia	2 (4.5)	0
Creatinine elevation	1 (2.3)	0
Dyspnea	1 (2.3)	0
Hyperuricemia	1 (2.3)	0
Acne	1 (2.3)	0

AEs: adverse events.

* Adjuvant therapies were used to alleviate adverse effects.

Figure 5. Relapse-free survival of sirolimus treatment. A. The relapse-free survival of 35 patients with response to sirolimus. The solid line represented relapse-free survival during treatment, and the dotted line represented the 95% CI range. B. Relapse-free survival in patients refractory or relapsed. The blue line represented relapse-free survival in refractory patients, and the red line represented relapse-free survival in relapsed patients. Patients who did not relapse were censored at the end of follow-up.

Table 4. Factors that may affect or/CR/relapse.

	Characteristics	P-value
		Univariate	Multivariate
OR vs. non-OR	Age, years	0.298	0.212
	Sex (male vs. female)	0.292	0.250
	Median time from diagnosis to sirolimus treatment, months	0.465	0.161
	Type of diseases (wAIHA vs. ES)	0.999	0.446
	Disease status (refractory vs. relapsed)	0.932	0.669
	Hemoglobin, g/L	0.980	0.686
	Absolute reticulocyte count, ×10^9/L	0.075	0.217
	Sirolimus plasma trough concentration, ng/mL*	0.009	0.028
	Previous response to CsA (refractory vs. relapsed)	0.186	–
CR vs. non-CR	Age, years	0.135	0.983
	Sex (male vs. female)	0.405	0.716
	Median time from diagnosis to sirolimus treatment, months	0.209	0.726
	Type of diseases (wAIHA vs. ES)	0.118	0.982
	Disease status (refractory vs. relapsed)	0.900	0.716
	Hemoglobin, g/L	0.065	0.994
	Absolute reticulocyte count, ×10^9/L	0.190	0.784
	Sirolimus plasma trough concentration, ng/mL^a	0.011	0.983
	Previous response to CsA (refractory vs. relapsed)	0.322	–
Relapse vs. non-relapse	Age, years	0.179	0.645
	Sex (male vs. female)	0.855	0.880
	Median time from diagnosis to sirolimus treatment, months	0.584	0.091
	Type of diseases (wAIHA vs. ES)	1.000	0.301
	Disease status (refractory vs. relapsed)	0.102	0.998
	Hemoglobin, g/L	0.344	0.763
	Absolute reticulocyte count, ×10^9/L	0.381	0.250
	Time to response	0.914	0.753

OR: overall response; CR: complete response; wAIHA: warm autoimmune haemolytic anaemia; ES: Evans syndrome; CsA: cyclosporine A.

^a Based on the average sirolimus plasma trough concentration of the first 6 months after sirolimus for each patient.

Table 5. Previous studies about sirolimus in patients with refractory primary AIHA/ES.

Reference	Disease	No. of patients	Period of follow-up	Efficacy
Jasinski et al. [22]	ES	5	2.25 (range: 1–6) years*	CR 2, PR 2
Bride et al. [35]	AIHA	2	1 and 4 years	CR 1, PR 1
	ES	8	15 weeks-2 years	CR 4, PR 2
Miano et al. [31]	AIHA	4	3.46 years*	CR
Miano et al. [20]	AIHA	3	9, 31, 42 months	CR
Li et al. [23]	AIHA	14	18(range: 10–40) months*	CR 57.1%, OR 85.7%
	ES	12		CR 25.0%, OR 33.3%
Williams et al. [21]	AIHA	2	14 and 16 months	CR

AIHA: autoimmune haemolytic anaemia; ES: Evans syndrome; CR: complete response; PR: partial response; OR: overall response.

* Follow-up time for all patients included in the study.

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Data availability statement

The datasets are not publicly available due to personal data protection reasons but are available from the corresponding author upon reasonable request.