Reduction of neutrophilic lung inflammation by inhalation of the compatible solute ectoine: a randomized trial with elderly individuals

Figures & data

Figure 1 CONSORT flow scheme of the EFECT study.

Abbreviations: EFECT, An efficacy and feasibility study to investigate the effect of EIL in subjects with inflammation and airway obstruction; SAE, serious adverse events; EIL, ectoine inhalation solution.

Figure 2 Study design of the double-blinded placebo-controlled crossover study.

Abbreviations: V1–V4, visits at the study center; EIL, ectoine inhalation solution.

Table 1 Volunteer characteristics (PP population)a

	Placebo first	EIL first	Total
Subjects (n)	17	16	33
Age (years)	76.61 (73.83–80.88)	76.08 (71.53–80.26)	76.35 (71.53–80.88)
Females (%)	100	100	100
Height (cm)	158.24 (149.00–170.00)	157.69 (149.00–165.00)	157.97 (149.00–170.00)
Weight (kg)	72.80 (49.70–99.60)	70.33 (60.30–92.00)	71.60 (49.70–99.60)
Never smoker (n)	16	15	31
Exsmoker (n)	1	1	2
FEV1 (l)	1.97 (1.14–2.99)	2.03 (1.31–2.68)	2.00 (1.14–2.99)
FVC (l)	2.70 (1.93–3.86)	2.75 (1.81–3.54)	2.72 (1.81–3.86)
FEV1/FVC (Tiffeneau)	0.73 (0.59–0.81)	0.74 (0.68–0.82)	0.73 (0.59–0.82)

Note:

a Arithmetic mean (minimum – maximum).

Abbreviations: EIL, ectoine inhalation solution; FEV₁, forced expiratory volume in 1 second; FVC, forced vital capacity; PP, per-protocol.

Table 2 Descriptive statistics given as geometric means (SD) (PP population)

	EIL first				Placebo first
	EIL		Placebo		Placebo		EIL
	V1	V2	V3	V4	V1	V2	V3	V4
Subjects (n)	16	16	16	16	17	17	17	17
IL-8 (pg/mL) in sputum	711.1 (2.8)	452.1 (4.2)	382.4 (6.1)	491.2 (5.6)	903.3 (3.4)	819.6 (2.8)	840.6 (5.9)	1,206.6 (5.3)
Neutrophils (%)	17.6 (4.0)	8.9 (4.5)	5.2 (4.8)	6.6 (3.9)	21.5 (2.5)	19.4 (4.0)	18.7 (4.9)	21.6 (4.1)
Neutrophils/macrophagesc	3.0 (2.5)	2.1 (2.8)	3.7 (3.7)	4.6 (2.0)	2.1 (2.8)	2.8 (2.5)	4.7 (2.8)	5.5 (3.1)
TNF-α (pg/mL) in sputum	19.5 (1.9)	33.7 (2.3)	20.8 (1.5)	23.5 (2.3)	20.5 (2.7)	28.8 (2.4)	28.0 (2.5)	27.8 (3.2)
Nitrogen oxides (μM) in sputum (nitrate + nitrite)	45.3 (1.8)	32.4 (2.1)	42.8 (1.8)	43.6 (2.6)	34.0 (1.9)	36.8 (2.1)	54.7 (1.9)	35.3 (1.9)
CRP (μg/mL) in serum	1.0 (2.9)	1.1 (2.7)	1.1 (3.0)	1.1 (2.8)	0.9 (3.6)	0.7 (3.0)	0.9 (3.1)	0.9 (3.1)
FEV1 (l)a	2.0 (1.2)	2.0 (1.2)	2.0 (1.2)	1.9 (1.3)	1.9 (1.3)	1.9 (1.3)	1.9 (1.2)	1.8 (1.3)
FVC (l)a	2.7 (1.2)	2.7 (1.2)	2.6 (1.2)	2.6 (1.2)	2.7 (1.2)	2.7 (1.2)	2.6 (1.2)	2.6 (1.2)
Tiffeneaua,b	0.74 (0.04)	0.74 (0.05)	0.74 (0.05)	0.73 (0.04)	0.73 (0.05)	0.72 (0.06)	0.73 (0.06)	0.71 (0.06)
QoL coughb	1.4 (1.3)	1.6 (1.2)	1.8 (1.4)	1.2 (1.3)	2.7 (1.8)	1.9 (1.6)	2.1 (1.0)	1.8 (1.4)
QoL phlegmb	0.7 (1.1)	1.3 (1.1)	1.3 (1.6)	0.9 (1.0)	2.2 (1.7)	1.8 (1.3)	2.0 (1.1)	1.5 (1.3)
QoL tightness in chestb	0.6 (0.9)	0.2 (0.5)	0.5 (0.9)	0.3 (0.6)	1.3 (1.8)	0.7 (1.2)	0.9 (1.3)	1.1 (1.3)
QoL breathlessnessb	1.4 (1.6)	1.6 (1.5)	1.6 (1.5)	1.5 (1.5)	2.8 (1.6)	2.7 (1.5)	2.7 (1.5)	2.5 (1.3)

Notes:

a All values are given as measures before salbutamol, no significant changes were observed after salbutamol inhalation;

b arithmetic means (SD), as these values exhibited normal distributions;

c monocytes and macrophages.

Abbreviations: CRP, C-reactive protein; EIL, ectoine inhalation solution; PP, per-protocol; QoL, quality of life; SD, standard deviation; TNF-α, tumor necrosis factor-alpha; FEV₁, forced expiratory volume in 1 second; FVC, forced vital capacity; IL, interleukin; V1–V4, visits at the study center.

Table 3 Changes after application of EIL inhalation solution

End point	Mean quotienta/differencec	95% CI (LL)	95% CI (UL)	P-value
IL-8 (pg/mL) in sputuma	0.88	0.48	1.63	0.68
% neutrophilsa	0.71	0.37	1.39	0.31
Neutrophils/macrophagesa,d	0.69	0.41	1.16	0.15
TNF-α (pg/mL) in sputuma	1.04	0.58	1.84	0.9
Nitrogen oxides (μM) in sputum (nitrate + nitrite)a	0.65	0.45	0.93	0.02
CRP (μg/mL) in seruma	1.17	0.80	1.71	0.40
FEV1 (l)a,b	0.98	0.95	1.02	0.32
FVC (l)a,b	0.98	0.96	1.00	0.10
Tiffeneaua,b	0.003	−0.017	0.023	0.757
QoL coughc	0.69	−0.09	1.37	0.08
QoL phlegmc	0.44	−0.14	1.02	0.13
QoL tightness in chestc	0.35	−0.15	0.84	0.16
QoL breathlessnessc	0.16	−0.33	0.64	0.51

Notes:

a A quotient <1 indicates a reduction by EIL treatment compared to placebo;

b before salbutamol;

c mean differences <1 would indicate an improvement by EIL treatment compared to placebo;

d monocytes and macrophages.

Abbreviations: EIL, ectoine inhalation solution; FEV₁, forced expiratory volume in 1 second; FVC, forced vital capacity; TNF-α, tumor necrosis factor-alpha; CI, confidence interval; IL, interleukin; CRP, C-reactive protein; QoL, quality of life; UL, upper limit; LL, lower limit.

Figure 3 Effect of EIL in comparison with the effect of placebo inhalation on inflammatory markers. (A) Effects after the end of the first inhalation period (V1/V2). (B) Effects after the end of the washout phase before starting the second period (V1/V3). Dark gray bars, EIL; light gray bars, placebo. Geometric mean values of individual quotients with 95% confidence intervals are presented. P-values of Student’s t-tests for the EIL–placebo comparison are given if P<0.05.

Abbreviations: EIL, ectoine inhalation solution; IL, interleukin; V1–V3, visits at the study center.

Table S1 List of adverse events reported during V1–V4 periods of the EFECT study

Treatment	Reported adverse event; ICD-10 code; verbatim term	Severitya	Relation to treatmentb
EIL	Diseases of the eye and adnexa; H57.9; inflammation of the eye	1	4
EIL	Diseases of the genitourinary system; N30.9; cystitis	1	4
EIL	Diseases of the genitourinary system; N30.9; cystitis	2	4
EIL	Diseases of the respiratory system; J06.9; cold, cough	1	4
EIL	Symptoms, signs, and abnormal clinical and laboratory findings, not elsewhere classified; R05; cough and irritation	1	3
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the circulatory system; I10; arterial hypertension (>200) and cardiac arrhythmia	2	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Pressure and pain in chest area. According to the subject known for a long time and no therapy possible	3	4
EIL	Symptoms, signs, and abnormal clinical and laboratory findings, not elsewhere classified; R05; cough and irritation during inhalation of EIL or placebo, whichever applies	1	2
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J30.1; hay fever	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
EIL	Diseases of the respiratory system; J06.9; cold	1	4
PLAC	Symptoms, signs, and abnormal clinical and laboratory findings, not elsewhere classified; R05; cough	1	6
PLAC	Diseases of the musculoskeletal system and connective tissue; M19.94; arthrosis in right hand	2	4
PLAC	Diseases of the digestive system; K52.9; diarrhea	1	4
PLAC	Diseases of the respiratory system; J06.9; cold	1	4
PLAC	Unclear. Volunteer reports on mild irritation. A fast recovery was achieved by sage tea	1	6
PLAC	Diseases of the respiratory system; J06.9; cold	1	4
PLAC	Diseases of the respiratory system; J06.9; cold and cough	1	6
PLAC	Diseases of the respiratory system; J06.9; cough and cold	1	4
PLAC	Diseases of the respiratory system; J06.9; cold	1	4
PLAC	Diseases of the respiratory system; J06.9; cold	1	4
PLAC	Diseases of the respiratory system; J06.9; cold	1	4
PLAC	Diseases of the respiratory system; J06.9; cold	1	4
PLAC	Diseases of the respiratory system; J06.9; cold	1	4
PLAC	Diseases of the respiratory system; J06.9; cold	1	4
PLAC	Diseases of the respiratory system; J06.9; cold	1	4
PLAC	Diseases of the musculoskeletal system and connective tissue; M54.99; back pain	1	4
PLAC	Diseases of the musculoskeletal system and connective tissue; M25.59; pain in joint	1	4
PLAC	Diseases of the respiratory system; J06.9; cold	1	4
PLAC	Diseases of the respiratory system; J06.9; cold	1	4
PLAC	Diseases of the respiratory system; J06.9; cold	1	4
PLAC	Diseases of the respiratory system; J06.9; cold, sniffles, flu	2	4
EIL	Diseases of the digestive system; K31.9; gastric disorders Neoplasm; C16.0	4	4
PLAC	Diseases of the circulatory system; I48; cardiac arrhythmia and atrial fibrillation	4	4

Notes: AEs in the washout phase are assigned to period 1 and AEs on the last study day (V4) are assigned to period 2.

a Severity: 1= mild, 2= moderate, 3= severe, 4= life-threatening, 5= lethal;

b Relation to study treatment: 1= definitely, 2= probably, 3= possibly, 4= not related, 5= not assessed, 6= not assessable;

c Outcome: 4= in-patient hospitalization or prolongation of existing hospitalization.

Abbreviations: AE, adverse event; EIL, ectoin inhalation solution; EFECT, an efficacy and feasibility study to investigate the effect of EIL in subjects with inflammation and airway obstruction; PLAC, placebo inhalation solution; V1–V4, visits at the study center; ICD-10, International Classification of Diseases-10th Revision.