Non-clinical toxicology evaluation of BIA 10-2474

Figures & data

Table 1. Clinical findings observed in toxicology studies in laboratory species.

Species	Mouse	Mouse	Mouse	Rat	Rat	Rat	Dog	Dog	NHP	NHP
Type of study	14-day DRF	4-week	13-week	14-day DRF	4-week	13-/26-week	4-week	13-week	4-week	13-week
Dose (mg/kg/day)	100–300–600*	100–300–500	25–75–150	150–200–250^†	30–90–150	10–30–90	20–50–100	20–50–100	10–50–100	6.25–37.5–75
Death	600	500		200			–
Tremor							100		50	37.5
Weakness	600	500					100		50	37.5
Decreased activity	600	500		200(F); 250(M)			100
Unusual posture	600	500		150(F)			–		50
Ataxia/abnormal gait	600(F)	500		150	150		100	50	50
Ruffled fur	600	500		150(F)	150
Hair loss									10	6.25
Enlarged abdomen		500
Salivation							100	50	50	6.25
Vomiting							100	20(M); 50(M)	50	6,25(M); 37,5
Miosis							100		10(F)
Hypothermia		–							10	37.5
Hyperthermia							100
Abnormal breathing	600						100	20(F); 50(M)
Decreased food intake	600	–	25(M)		30(F)				50
Increased food intake		300(M)	25(F)		30(M)	30(M); 90(F)
Decreased BW or gain	600	300(M)		150(F)	30(F); 90(M)		50
Increased BW or gain		500(F)	25(M); 150(F)
Pasty/liquid feces									10	6,25(F); 37,5(M)

The dose indicated is the lowest dose at which those effects were seen.

*The starting high dose was 1000 mg/kg/day but following the death of 3 of the 6 animals within 4 days this was reduced to 600 mg/kg/day in the surviving 3 animals.

^†Only the 150 mg/kg/day dose was given for 14 days. Animals receiving 200 or 250 mg/kg/day were killed on day 4 or 5.

DRF: dose range finding; NHP: non-human primate.

Table 2. Hematology findings observed in toxicology studies in laboratory species.

Species	Mouse	Mouse	Rat	Rat	Rat	Dog	Dog	NHP	NHP
Type of study	4-week	13-week	4-week	13-week	26-week	4-week	13-week	4-week	13-week
Dose (mg/kg/day)	100–300–500	25–75–150	30–90–150	10–30–90	10–30–90	20–50–100	20–50–100	10–50–100	6.25–37.5–75
Decreased erythrocytes	500					100(F)
Decreased Hb	500	150(F)					50(M)
Decreased leukocytes	100(M)
Decreased platelets		75
Decreased leukocytes	100(M)								F
Increased leukocytes
Decreased lumphocytes	100(M)
Decreased monocytes	100(M)
Increased erythrocytes
Increased reticulocytes	500(M)		150(F)
Decreased basophils		25(F)
Decreased platelets		75
Increased protein	500(F)	75(M)			90
Urea decrease		150(M)
Urea increase	500(F)			90	10(F); 90(M)
Bilirubin increase				30(M); 90(F)	30(M); 90(F)				37.5
Cholesterol increase		75(M); 150(F)		10(M)	10(F); 30(M)
Triglycerides					90
Glucose increase					90		50(F)
ALAT increase	500(F)
ASAT decrease		150(F)				100(F)
Creatinine kinase decrease						100(F)
Globulin increase	500(F)	150(F)
Albumin increase					30	50(M)
Sodium increase	500(F)				10(F); 30(M)
Potassium increase	300(F)			30	10(F)
Potassium decrease						100(M)	20(M)
Chloride decrease				90(M)
Chloride increase	500(F)			90(M)	10(F)
Calcium increase				30(F); 90(M)	30
Phosphorus increase				30(F); 90(M)	30(F); 90(M)
Phospholipids increase				10(M)	10(F); 30(M)
Urine pH decrease			90(F); 150(M)

The dose indicated is the lowest dose at which those effects were seen. NHP: non-human primate.

Table 3. Necropsy findings observed in toxicology studies in laboratory species.

Species	Mouse	Mouse	Rat	Rat	Rat	Dog	Dog	NHP	NHP
Type of study	4-week	13-week	4-week	13-week	26-week	4-week	13-week	4-week	13-week
Dose (mg/kg/day)	100–300–500	25–75–150	30–90–150	10–30–90	10–30–90	20–50–100	20–50–100	10–50–100	6.25–37.5–75
Liver hypertrophy			30
Increased liver weight	100(M); 300(F)	75(M); 150(F)	90	30(M); 90(F)	90	100(F)
Increased kidney weight			90	90		50(F); 100(M)	100(F)	100(M)
GI tract changes	500(F)
Lung discoloration							100
Increased lung weights	500(M)	150(M)				100(M)
Decreased spleen weight					30(M)		20(M); 50(F)
Increased spleen weight		75(F); 150(M)							75(M)
Increased pancreas weight				90(F)	90(F)
Decreased thymus size		150(F)				100	20	50(M)
Decreased thyroid weight
Increased adrenal weight						100(M)	100(F)		37.5(M); 75(F)
Increased pituitary weight							100(F)
Reduced testes weight	500
Decreased epididymus weight			90(M)
Decreased prostate weight	500(M)
Decreased uterus weight	500(F)
Increased uterus weight							100(F)

All significant changes are shown and could indicate a change in absolute weight or relative to body weight (see Hayes, Hardisty, Harris, Okazaki, et al. 2020; Hayes, Hardisty, Harris, Weber 2020, Hardisty et al. 2020, Weber et al. 2020 for details). The dose indicated is the lowest dose at which those effects were seen. NHP: non-human primate.

Table 4. Histopathology findings observed in toxicology studies in laboratory species.

Species	Mouse	Mouse	Rat	Rat	Rat	Dog	Dog	NHP	NHP
Type of study	4-week	13-week	4-week	13-week	26-week	4-week	13-week	4-week	13-week
Dose (mg/kg/day)	100–300–500	25–75–150	30–90–150	10–30–90	10–30–90	20–50–100	20–50–100	10–50–100	6.25–37.5–75
Sciatic nerve degeneration	500	150
Hippocampus degeneration	500
Medulla oblongata, axonal swelling/dystrophy				30	90			50	75
Ventral commissure 3rd vent. vacuolation					90
Spinal cord axonal swelling					10(F); 30(M)
Myofibre degeneration		150
Hepatocellular hypertrophy	100	75		90	30(F); 90(M)
GI villus hypertrophy	500
GI enterocyte vacuolation/degeneration	100		30	30	30
Myelin edema enteric plexus			90
Vacuolar degeneration of ganglia			90	90	90			50	75
Pituitary gland edema			90	30	90			50	75
Testicular degeneration/atrophy	500		150	30	90	100
Ovary: maturation arrest	500
Bronchopneumonia						100	100
Granulopoiesis	500(F)
Thymus atrophy	500					100	100
Spleen extramedullary hempoiesis	500	150(M)	150(M)
Bladder: submucosal inflammatory foci		25
Nephropathy	300	150	90	30(M); 90(F)	90

The dose indicated is the lowest dose at which those effects were seen. NHP: non-human primate.

Figure 1. Dose levels and pharmacokinetic parameters (C_max and AUC_0–t) observed in toxicology studies in laboratory species and the clinical trial BIA-102474-101 (single ascending dose [SAD] part; multiple ascending dose [MAD] part).

Table 5. Dose levels (mg/kg/day, p.o.), dose at no-observed-adverse-effect level (NOAEL), corresponding human equivalent dose (HED) and pharmacokinetic parameters (C_max and AUC_0–t) observed in toxicology studies in laboratory species.

Dose	NOAEL	HED	Cmax (ng/mL)	Cmax (ng/mL)	AUC0–t (ng.h/mL)	AUC0–t (ng.h/mL)
(mg/kg)			Males	Females	Males	Females
Mouse 4-week
100	X	486	46,200	53,200	160,000	149,000
300			92,100	98,500	420,000	475,000
500			117,000	93,400	805,000	960,000
Mouse 13-week
25			11,500	12,100	30,700	24,900
75	X	365	36,400	34,800	90,000	83,800
150			72,300	178,000	202,000	178,000
Rat 4-week
30	X	288	10,300	17,400	41,300	64,900
90			29,200	38,900	13,900	276,000
150			38,600	52,900	309,000	424,000
Rat 13-week
10	X	96	7400	7920	18,900	23,100
30			18,300	19,400	60,900	74,100
90			31,000	40,300	134,000	238,000
Rat 26-week
10	X	96	4880	5603	16,751	20,109
30			10,377	8463	40,396	38,610
90			31,500	25,367	186,842	111,997
Dog 4-week
20			4890	5490	24,300	24,300
50	X	1623	11,500	8560	65,000	38,600
100			17,400	21,600	102,000	137,000
Dog 13-/4-week
20	X	649	6200	7260	24,200	31,600
50			16,200	17,600	97,600	96,300
100			26,200	15,600	153,000	90,700
NHP 4-week
10			5947	6390	36,034	30,152
50			23,133	20,600	143,513	125,097
100	X	1944	26,900	28,350	179,416	168,722
NHP 13-week
6.25			3743	4770	19,632	24,245
37.5			15,025	13,375	77,207	78,917
75	X	1458	19,950	21,250	137,999	130,702

HED = animal dose × conversion factor × 60.

Table 6. Summary of male vs female and acute vs chronic differences and dose proportionality for AUC values in BIA 10-2474 toxicology studies.

Study	Sex differences	Acute vs chronic	Dose-proportionality
Mouse 4-week	No (0.91–1.3)	Increased exposure after repeat dosing (6.7–8.8 at highest dose)	Acute: less than proportional Chronic: proportional
Mouse 13-week	No (0.73–1.5)	No differences (1.1–1.3)	Dose-proportional
Rat 4-week	F > M (1.1–2.0)	Decrease in M (0.78–0.91), stable in F (0.84–1.0)	Slightly greater than dose-proportional
Rat 13-week	F > M (1.1–1.8) but low dose F≫M (4.5)	Slight increase (1.1–1.7) except low-dose F (0.46)	Dose-proportional except F on day 1 (less than proportional)
Rat 26-week	Variable: F tended to be more exposed on day 1 and 97.	Variable: No difference at low dose, decrease at mid-dose (0.5–0.8), increase in F at high dose (3.2)	Dose-proportional in M, less than dose-proportional in F
Dog 4-week	No (0.59–1.5)	Slight increase (1.3–2.2)	Greater on day 1 but dose-proportional on day 28
Dog 13-week	No (0.44–1.44)	Slight increase (0.97–2.1)	Yes in M, less than dose-proportional in F
NHP 4-week	No (0.8–1.3)	No difference (0.6–2.1)	Less than dose-proportional, particularly between top two doses
NHP 13-week	No (0.8–1.2)	No difference (0.7–1.3)	Less than dose-proportional

Table 7. Dose levels, no-observed-adverse-effect level (NOAEL) and pharmacokinetic parameters (C_max and AUC_0–t) observed in toxicology studies in laboratory species and the clinical trial BIA-102474-101 (single ascending dose [SAD] part; multiple ascending dose [MAD] part).

	Laboratory species				Human healthy subjects^Table Footnote†
Species	Mouse	Rat	Dog	NHP
Duration of treatment	13-week	26-week	13-week	13-week	SAD – Day 1	MAD – Day 10	MAD – Day 1
NOAEL	75	10	20	75	(100 mg/day)	(20 mg/day)	(50 mg/day)
Corresponding Cmax (ng/mL)*	35,600	5242	9375	20,600	1772	396	667
Corresponding AUC0–t (ng.h/mL)*	86,900	18,430	45,150	134,351	22,991	4651	7768
NOAEL safety margin (Cmax)	18.7	4.0	9.7	28.9	–	–	–
NOAEL safety margin (AUC0–t)	89.9	13.2	23.7	52.0	–	–	–

*Values shown are the mean values obtained from males and females.

^†According to Gama et al. (2016).

Safety margins based on systemic exposure (AUC_0–t = 4,651 h.ng/mL; Cmax = 396 ng/mL) on healthy adult subjects after repeated daily administrations (day 10) of highest MAD completed dose (20 mg).

Table 8. Dose levels, abnormal findings and pharmacokinetic parameters (C_max and AUC_0–t) observed in toxicology studies in laboratory species and the clinical trial (single ascending dose [SAD] part; multiple ascending dose [MAD] part).

Species	Mouse	Mouse	Rat	Rat	Rat	Dog	Dog	NHP	NHP	Human (100 mg/day)	Human (20 mg/day)	Human (50 mg/day)
Duration of treatment	4-week	13-week	4-week	13-week	26-week	4-week	13-week	4-week	13-week	SAD – Day 1	MAD – Day 10	MAD – Day 1
Unusual posture	500	–	–	–	125	–	–	50	–	No*	Yes (1/6)^†	Yes^‡
Ataxia	500	–	150	–	–	100	50	50	–	No*	–	Yes^‡
Hippocampus degeneration	500	–	–	–	–	–	–	–	–	–	–	Yes^‡
Brainstem degenerative changes	–	–	–	30	90	–	–	50	75	–	–	Yes^‡
Corresponding Cmax (ng/mL)^¶	105,200	–	45,750	18,850	5242	19,500	12,450	21,867	20,600	1772	396	667
Corresponding AUC0–t (ng.h/mL)^¶	882,500	–	366,500	67,500	18,430	119,500	63,900	134,305	134,350	22,991	4651	7768

^* According to Gama et al. (2016).

^† 1 out 6 subjects reported mild “dizzness postural” according to Gama et al. (2016).

^‡ According to Kerbrat et al. (2016).

^¶ Values shown are the mean values obtained from males and females.

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