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Abstract

Cynomolgus macaques (Macaca fascicularis) are commonly used in safety assessment and as translational models for drug development. Recent supply chain pressures, exportation bans, and increased demand for drug safety assessment studies exacerbated by the COVID-19 pandemic have prompted the investigation of utilizing macaques of different geographic origin in preclinical toxicity studies. This study compares routine hematology, coagulation, and clinical chemistry endpoints of 3 distinct subpopulations of mainland Asia origin (Cambodia, China, and Vietnam) with Mauritius origin macaques compiling results of 3,225 animals from 123 regulatory toxicology studies conducted at North American and European Union contract research organization facilities between 2016 and 2019. Results were generally similar amongst the subpopulations compared in this study. Few notable differences in hematology test results and several minor differences in serum biochemistry and coagulation test results were identified when 3 distinct subpopulations of mainland Asia origin macaques were compared with Mauritius origin macaques. Our findings support the use of different origin macaques in drug development programs; however, emphasizes the importance of maintaining consistency in geographic origin of animals within a study.

Keywords

clinical pathology hematology serum biochemistry Mauritius cynomolgus macaque safety assessment preclinical toxicology

Introduction

Refinement and innovation in preclinical toxicity assessment of candidate new drugs has greatly reduced the use of animals in pharmaceutical safety assessment; however, preclinical animal studies are still necessary and required by regulatory agencies in the development of safe and effective therapeutics for human medicine. The cynomolgus macaque (M. fascicularis), also known as the crab-eating or long-tailed macaque, is a cercopithecine primate and is the most frequently used nonhuman primate (NHP) species in biopharmaceutical research. Macaques originate from a wide range of geographic locations including India, China, Southeast Asia, and the island of Mauritius located in the Indian Ocean just east of the African continent.¹ Increased demand for cynomolgus macaques for use in biomedical research in North America and Europe has been exacerbated by the increased development of large molecule therapeutics (NHPs are often the only relevant species for assessing these therapeutics) and the most recent increasing need for resources in emerging disease research (eg, SARS-CoV-2 studies). Combined with NHP exportation bans, this increased demand has led to a global shortage of Chinese origin NHPs for preclinical toxicity studies and has impacted the breeding/sourcing practices for NHPs destined for biomedical research.² Given this recognized limited availability of mainland Asian origin NHPs and pressures on the supply chain, it is imperative to consider and investigate the utilization of NHPs from different geographical origins and their potential interchangeability, or lack thereof, in pharmaceutical safety assessment programs. Recognition and characterization of origin-related differences can lessen or even eliminate the possibility of erroneous interpretation of preclinical study data in pharmaceutical safety assessments.

Mainland Asian macaques including Cambodian, Chinese, and Vietnamese subpopulations originated in Southeast Asia from common ancestors and share a similar genetic background, which supports using these subpopulations interchangeably for biomedical research.³ In contrast to Asian macaques, Mauritian macaques are a population of NHP that have been geographically isolated for several centuries and based on mitochondrial DNA analysis, have expanded from a small number of founder animals originating from Indonesia.⁴ Genetic diversity between Asian and Mauritius macaques has been observed and described in the literature.^3,5,6 Mauritius macaques have a more homogenous genetic pool than mainland Asia origin macaques, are devoid of herpes B virus, respond differently to immune system challenges, and have lower incidences of background pathology based on previously published literature^1,3,5-8 although a recent study⁹ reports these lower incidences depend on the tissues and/or organs sampled with a higher incidence of inflammatory infiltrates identified in some tissues when Mauritius origin macaques were compared with other origin macaques.

Dedicated breeding colonies in China, the United States, and/or the European Union have led to predictable decreases in genetic diversity within each of the individual colonies, which mimics the decreases in genetic diversity observed in the naturally sequestered Mauritian cynomolgus macaques.^1,3,10 Cynomolgus macaques are now purpose-bred in a limited number of breeding centers in different geographical locations across the globe, but most European countries depend on NHP importation due to fewer breeding facilities in Europe compared with other countries.

Evaluation of clinical pathology data is an integral part of pharmaceutical candidate safety assessments and a robust database for standard clinical pathology endpoints is available for the cynomolgus macaque, including some subpopulations. Nevertheless, when interpreting clinical pathology data, it is imperative to consider the variability observed in cynomolgus macaques, not only as individuals but also mindful of the source and geographic background.^11-14 Limited information on the potential differences in routinely evaluated clinical pathology endpoints (ie, standard hematology, coagulation, and clinical chemistry parameters) of Mauritian macaques compared with unique Asian origin subpopulations (ie, Cambodia, China, and Vietnam) is available in the literature and many of the previous studies were comprised of data sets containing low animal numbers.^5,15,16 This study contributes to the literature with a large scale, well-powered, review of standard clinical pathology endpoints of cynomolgus macaques conducted at 3 contract research organization (CRO) facilities; 2 in North America and 1 in Europe. Comparisons were made to investigate and describe differences of toxicological relevance, if any, between Mauritius macaques and other Asian origin macaques, and to highlight the influence geographic isolation may have on the Mauritius population of macaques. A secondary objective was to determine if any differences would impact animal sourcing for a given study or drug development program.

Material and Methods

Animals

Cynomolgus macaques of mainland Asia (ie, Cambodia, China, and Vietnam), or Mauritius origin were procured from AAALAC-International accredited sources/suppliers and housed at one of 3 geographically distinct AAALAC-International accredited CRO facilities in North America (Madison, WI, USA [MSN], Greenfield, IN, USA [GRN]) or the European Union (Münster, DE [MUE]). Animals underwent required physical examinations, pathogen screenings, and at a minimum were serologically tested and confirmed negative for simian immunodeficiency virus (SIV), Cercopithecine herpesvirus 1 (B virus), simian retroviruses type D, rabies, simian T-cell leukemia virus (STLV), and filoviruses. Additional testing, including tuberculin and bacterial (eg, Shigella, Campylobacter, Yersinia, and Salmonella) and parasitological (eg, malaria smears) screening was performed. Upon receipt to the facility, animals underwent required quarantine and a minimum of 3 to 6 weeks acclimation per local facility standard operating procedures. Animals were pair or group-housed in climate-controlled rooms (a minimum of 8 air changes/hour, 19-26°C, 30%-70% relative humidity, 12-hour artificial light/dark cycles), received tap water ad libitum, and were fed a standardized diet across facilities once or twice daily (Certified lab diet for primates [LabDiet 5048, PMI Nutrition International, Inc.] supplemented by fresh fruit and vegetables). Animals were given a variety of room, cage, and dietary enrichment. Studies were performed in compliance with United States federal regulations and recommendations and institutional ethical committees or in accordance with the European Convention for the Protection of Vertebrate Animals Used for Experimental and Other Scientific Purposes (Strasbourg, Council of Europe); see Animal Use Statement for more detail.

Animals were fasted overnight for a minimum of 6 hours prior to nonsedated blood collection from the femoral (North America [NA] facilities) or cephalic or saphenous (European Union [EU] facility) vein. Hematology, coagulation, and serum biochemistry data used in this comparative assessment were obtained from scheduled intervals for blood collection prior to study initiation (naïve animals). Animal origin, study location, regulatory compliance, and animal numbers are summarized in Table 1.

Table 1.

Distribution of 123 Studies (3,225 Animals) Surveyed to Compile Data Used in This Study.

	Number of studies per facility	Regulatory compliance	Age range of animals included	Number of studies by geographic origin	Final number of males/origin	Final number of females/origin
North America	55 MSN 1 GRN	55 GLP 1 non-GLP	2-5 years	10 Mauritius 39 China 7 Cambodia	110 Mauritius 662 China 111 Cambodia	105 Mauritius 660 China 113 Cambodia
European Union	67 MUE	66 GLP 1 non-GLP	2-5 years	19 Mauritius 48 Vietnam	251 Mauritius 541 Vietnam	236 Mauritius 442 Vietnam

Abbreviations: GLP, Good Laboratory Practice; Greenfield, IN, USA (GRN); Madison, WI, USA (MSN); Muenster, DE, EU (MUE).

Clinical Pathology

Clinical pathology data from 123 preclinical toxicology studies (assembling data from 3,225 animals) conducted predominantly under Good Laboratory Practice (GLP) regulations from the 3 previously described CRO facilities between 2016 and 2019 were tabulated. Clinical pathology parameters assessed for this comparison are listed in Table 2.

Table 2.

Clinical Pathology Parameters Assessed.

Parameter	Abbreviation		Unit
Parameter	NA	EU	NAUnited States Conventional Units	EUSystem International [SI] Units
Hematology
Red blood cell count	RBC	RBC	10⁶/µL	10¹²/L
Hemoglobin concentration	HGB	HGB	g/dL	mmol/L
Hematocrit	HCT	HCT	%	%
Mean cell volume	MCV	MCV	fL	fL
Mean corpuscular hemoglobin	MCH	MCH	Pg	fmol
Mean corpuscular hemoglobin concentration	MCHC	MCHC	g/dL	mmol/L
Red cell distribution width	RDW	—	%	—
Absolute reticulocyte count	RETIC	TRET	10³/µL	10¹²/L
Platelet count	PLT	PLT	10³/µL	10⁹/L
White blood cell count	WBC	WBC	10³/µL	10⁹/L
Absolute neutrophil count	NEUT	ANEU	10³/µL	10⁹/L
Absolute lymphocyte count	LYM	ALYM	10³/µL	10⁹/L
Absolute monocyte count	MONO	AMON	10³/µL	10⁹/L
Absolute eosinophil count	EOS	AEOS	10³/µL	10⁹/L
Absolute basophil count	BASO	ABAS	10³/µL	10⁹/L
Absolute large unstained cell count	LUC	—	10³/µL	—
Coagulation
Prothrombin time	PT	PT	sec	sec
Activated partial thromboplastin time	APTT	APTT	sec	sec
Fibrinogen concentration	FIB	FIB	mg/dL	g/L
Serum biochemistry
Glucose concentration	GLU	GLU	mg/dL	mmol/L
Urea nitrogen concentration	UN	BU	mg/dL	µmol/L
Creatinine concentration	CREAT	CREA	mg/dL	µmol/L
Total protein concentration	TP	TP	g/dL	g/L
Albumin concentration	ALB	ALB	g/dL	g/L
Globulin concentration	GLOB	GLOB	g/dL	g/L
Albumin: Globulin ratio	A: G	A: G	ratio	ratio
Total cholesterol concentration	CHOL	CHOL	mg/dL	mmol/L
Triglyceride concentration	TRIG	TRIG	mg/dL	mmol/L
Total bilirubin concentration	TBIL	TBIL	mg/dL	µmol/L
Aspartate aminotransferase activity	AST^†	AST*	U/L	U/L
Alanine aminotransferase activity	ALT^†	ALT*	U/L	U/L
Alkaline phosphatase activity	ALP	ALP	U/L	U/L
Gamma glutamyl transferase activity	GGT	GGT	U/L	U/L
Glutamate dehydrogenase activity	—	GLDH	—	U/L
Creatine kinase activity	CK	CK	U/L	U/L
Calcium concentration	Ca	CA	mg/dL	mmol/L
Inorganic phosphorus concentration	PHOS	PHOS	mg/dL	mmol/L
Sodium concentration	Na	NA	mmol/L	mmol/L
Potassium concentration	K	K	mmol/L	mmol/L
Chloride concentration	Cl	CL	mmol/L	mmol/L

Abbreviations: NA, North America; EU, European Union.

— Not tabulated.

†

analyzed with pyridoxal 5’-phosphate (P5P) cofactor (NA beginning 2018).

analyzed with pyridoxal 5’-phosphate (P5P) cofactor (EU).

Blood samples were collected into EDTA tubes for hematology tests and analyzed using an Advia 2120i analyzer (Siemens, USA)—(NA) or a Sysmex 2000 analyzer (Sysmex Europe, DE)—(EU). Samples for hematology tests were analyzed on the day of collection. Blood samples were collected into 3.2% sodium citrated tubes for coagulation tests and plasma was harvested following centrifugation (3000 RPM for 10 minutes at room temperature) within 75 minutes of collection. Plasma samples for coagulation tests were analyzed using a Destiny Amax analyzer (Tcoag, Wicklow, Ireland) or ACL TOP 500 analyzer (Instrumentation Laboratory, MA, USA)—(NA) or using a Destiny Max GUI analyzer (Tcoag, Wicklow, IR)—(EU). Plasma samples not analyzed on the day of collection were stored at −20°C for up to 13 days pending analysis. Blood samples were collected into tubes without anticoagulant for serum biochemistry tests. These samples were allowed to clot and serum was harvested following centrifugation (3000 RPM for 10 minutes at room temperature) within 4 hours of collection and analyzed using a Roche Modular Analytics P analyzer (Roche Diagnostics, IN, USA), Cobas 8000 analyzer (Roche Diagnostics, IN, USA)—(NA) or a Konelab Prime analyzer (Thermo Scientific, MA, USA)—(EU). Serum samples not analyzed on the day of collection were stored at −60 to −80°C for up to 7 days pending analysis. Quality Control (QC) was performed with appropriate levels of QC following specific manufacturer’s instructions at a minimum of once daily prior to the first analytical run of each day. Quality control was reviewed to ensure results were within established ranges and results were documented. In addition, prior to analysis, samples were assessed for visible clots, hemolysis, lipemia, and/or icterus.

Statistical Analysis

This dataset consisting of data from Mauritian, Cambodian, Chinese, and Vietnamese origin macaques aged between 2 to 5 years was subjected to statistical evaluation. Due to differences in the clinical pathology instrumentation and methodology between the EU and NA facilities, data availability, and differences in reporting units (NA = United States Conventional Units and EU = System International [SI] Units), comparisons between Mauritian and Vietnamese macaques were carried out using data obtained from the EU facility and comparisons between Mauritian and Cambodian or Chinese macaques were carried out using data obtained from the NA facilities. Separate statistical evaluations were performed for each parameter and sex. To avoid any potentially erroneous or extreme data points from skewing statistical conclusions, outliers were removed using the Tukey box plot method.¹⁷ Only parameters with at least 40 individual values were subjected to statistical evaluation. Differences between origins were evaluated using Analysis of Variance and 2-sample t-tests.^18,19 For the purpose of this current investigation, data from the 2 North American laboratories (MSN and GRN) were combined (including coagulation data for the Destiny Amax and ACL TOP 500 analyzers) although direct correlations of the instruments were not performed between the 2 laboratories. The combined data was reviewed and the results were considered similar for the purpose of this investigation based on notable overlap of individual animal results. Differences were felt to have minimal impact on the comparisons and statistical analysis of the overall clinical pathology data. The use of data from a single study analyzed at the GRN location combined with the bulk of the results obtained from the MSN laboratory was felt to have minimal impact on the overall data. Statistical results are reported in Tables 3 –6 and describe sample size (“N”), arithmetic group means, observed range of minimum and maximum values, and 95% reference intervals (group mean +/- 1.96 standard deviations).

Table 3.

North America Hematology and Coagulation Results.

North AmericaHematology and Coagulation	Males			Females
Parameter	Mauritius	China	Cambodia	Mauritius	China	Cambodia
Red blood cell count RBC E6/µL	6.9^a (5.9-8.0)^b [6.0-7.8]^c N = 107	5.6* (4.6-6.7) [4.9-6.4] N = 637	5.8* (4.9-6.6) [5.1-6.4] N = 111	6.6 (5.5-7.9) [5.5-7.7] N = 105	5.5 (4.5-6.5) [4.7-6.2] N = 658	5.6 (4.7-6.3) [4.9-6.2] N = 113
Hemoglobin HGB g/dL	13.7 (12.4 –15.0) [12.6-14.8] N = 107	13.2* (11.2-15.2) [11.7-14.7] N = 635	13.5 (11.8-15.1) [11.9-15.0] N = 111	13.0 (11.2-15.0) [11.5-14.6] N = 104	12.9 (10.7-14.9) [11.4-14.4] N = 656	13.1 (11.5-14.9) [11.7-14.5] N = 113
Hematocrit HCT %	47.2 (41.5-53.9) [41.9-52.4) N = 106	43.6* (36.8-50.8) [38.5-48.8] N = 632	44.4* (38.9-51.6) [39.2-49.7] N = 111	45.5 (38.4-54.0) [39.0-52.0] N = 103	42.8 (35.4-50.0) [37.5-48.0] N = 654	43.7 (37.8-49.3) [38.7-48.6] N = 112
Mean corpuscular volume MCV fL	69.2 (62.9-75.9) [63.4-74.9] N = 107	77.2* (67.6-86.5) [70.4-84.0] N = 638	77.1* (69.2-84.1) [70.9-83.2] N = 110	69.4 (62.9-76.6) [64.2-74.6] N = 99	77.9 (67.9-88.2) [70.6-85.3] N = 659	78.5 (70.1-87.4) [71.3-85.6] N = 111
Mean corpuscular hemoglobin MCH pg	20.1 (17.4-22.5) [17.9-22.2] N = 108	23.4* (20.5-26.3) [21.3-25.6] N = 626	23.4* (20.7– 25.8) [21.4-25.5] N = 111	19.9 (17.1-21.9) [17.7-22.0] N = 104	23.5 (20.3-26.8) [21.1-25.8] N = 657	23.5 (21.0-25.8) [21.4-25.6] N = 111
Mean corpuscular hemoglobin concentration MCHC g/dL	29.0 (26.7-31.1) [27.1 -31.0] N = 106	30.3* (27.2-33.5) [28.1-32.5] N = 632	30.4* (27.9-33.1) [28.1-32.6] N = 111	28.7 (26.3-31.8) [26.5-30.9] N = 105	30.1 (27.3-33.0) [28.0-32.2] N = 653	29.9 (27.9-31.9) [28.4-31.5] N = 110
Reticulocyte count RETIC E3/µL	35.0 (10.5-74.3) [3.7-66.4] N = 107	75.4 (16.5-154.4) [20.7-130.2] N = 631	76.8 (14.4-152.7) [24.0-129.6] N = 111	55.2 (10.4-134.6) [0.0-110.6] N = 103	78.4 (15.2-158.8) [22.0-134.8] N = 650	79.1 (20.8-152.6) [21.8-136.5] N = 111
Red cell distribution width RDW %	14.2 (12.4-16.5) [12.4-15.9] N = 108	12.7* (10.8-14.7) [11.2-14.1] N = 569	12.6* (11.3-14.6) [11.2-14.1] N = 111	14.5 (12.2-17.4) [12.3-16.7] N = 103	12.5 (10.8-14.5) [11.2-13.9] N = 593	12.4 (11.1-13.8) [11.2-13.7] N = 110
Platelet count PLT E3/ µl	378.5 (206.0-561.0) [222.6-534.3] N = 108	450.3* (230.0-693.0) [267.8-632.8] N = 634	422.5 (253.0-620.0) [263.4-581.6] N = 110	395.6 (217.0-594.0) [246.3-544.9] N = 103	452.6 (225.0-684.0) [281.1-624.1] N = 650	444.4 (235.0 -704.0) [277.6-611.1] N = 110
White blood cell count WBC E3/µL	9.3 (4.82-16.49) [4.47-14.13] N = 107	11.2* (4.04-19.93) [5.04-17.33] N = 620	10.9 (5.24-19.21) [5.01-16.74] N = 110	9.8 (4.40-15.97) [5.01-14.50] N = 105	11.5 (4.23-20.61) [5.18-17.79] N = 645	10.8 (6.08-18.13) [5.43-16.24] N = 110
Neutrophil count NEUT E3/µL	3.9 (0.56-9.34) [0.00-7.93] N = 106	5.2* (0.95-11.35) [0.75-9.59] N = 619	4.0 (0.88-9.29) [0.55-7.36] N = 109	5.1 (1.19-10.87) [1.03-9.14] N = 105	6.3 (1.60-13.29) [1.59-10.97] N = 641	5.4 (2.10-10.13) [1.60-9.11] N = 108
Lymphocyte count LYM E3/µL	4.6 (2.36-7.73) [2.17-7.10] N = 108	5.3 (1.21-10.72) [1.60-9.06] N = 627	6.0* (2.20-10.75) [2.03-10.0] N = 109	4.1 (1.50-6.95) [1.80-6.33] N = 105	4.4 (1.38-8.50) [1.53-7.26] N = 635	4.6 (2.02-8.68) [1.40-7.87] N = 109
Fibrinogen FIB mg/dL	193.4 (123.0-319.0) [99.1-287.6] N = 108	219.6* (127.0-318.0) [150.5-288.6] N = 606	213.6* (148.0-282.0) [154.7-272.6] N = 109	177.6 (136.0-267.0) [125.6-229.7] N = 95	213.2 (131.0-298.0) [152.5-273.8] N = 622	199.3 (141.0-276.0) [140.8-257.8] N = 111
Prothrombin PT seconds	9.5 (8.0-10.9) [8.4-10.7] N = 106	9.4 (7.6-11.1) [8.0-10.7] N = 632	9.4 (8.2-10.7) [8.4-10.3] N = 110	9.3 (7.9-10.8) [8.1-10.6] N = 101	9.3 (7.8-10.8) [8.1-10.5] N = 648	9.3 (8.4-10.5) [8.4-10.2] N = 110
Activated partial thromboplastin time APTT seconds	19.9 (17.5-22.3) [17.9-22.0] N = 105	19.9 (15.9-24.1) [16.9-22.9] N = 625	19.8 (16.8-23.0) [17.4-22.3] N = 110	19.7 (18.1-21.9) [18.1-21.4] N = 102	20.0 (16.1-24.0) [17.1-22.9] N = 651	20.1 (17.0-24.4) [17.0-23.1] N = 110

For these data, mean values are rounded to one significant digit.

mean.

(observed interval).

[95% reference interval].

P ≤ .0001 compared with Mauritius origin N = sample size

Table 4.

European Union Hematology and Coagulation Results.

European UnionHematology and Coagulation	Males		Females
Parameter	Mauritius	Vietnam	Mauritius	Vietnam
Red blood cell count RBC E12/L	7.1^a (5.9-8.4)^b [6.2-8.1]^c N = 247	6.0* (4.9-7.1) [5.2-6.8] N = 541	7.0 (5.6-8.3) [5.9-8.1] N = 234	5.7* (4.7-6.6) [5.0-6.4] N = 437
Hemoglobin HGB mmol/L	8.7 (7.3-9.9) [7.7-9.6] N = 250	8.7 (7.6-9.8) 7.8-9.6] N = 525	8.4 (7.0-9.7) [7.3-9.4] N = 232	8.2 (6.9-9.6) [7.3-9.2] N = 440
Hematocrit HCT %	45.9 (40.2-51.4) [41.6-50.2] N = 244	45.0* (38.6-51.4) 40.4-49.6] N = 530	44.3 (38.2-50.5) [39.4-49.2] N = 232	43.0* (37.0-49.3) [38.6-47.4] N = 440
Mean corpuscular volume MCV fL	64.2 (54.5-73.8) [57.0-71.4] N = 250	74.7* (64.9-84.5) [67.7-81.7] N = 538	63.5 (53.7-71.9) [56.3-70.7] N = 234	75.7* (66.4-85.1) [68.9-82.5] N = 441
Mean corpuscular hemoglobin MCH fmol	1.2 (1.1-1.4) [1.1-1.3] N = 247	1.4* (1.3-1.6) [1.3-1.6] N = 535	1.2 (1.0-1.3) [1.1-1.3] N = 234	1.5* (1.3-1.6) [1.3-1.6] N = 442
Mean corpuscular hemoglobin concentration MCHC mmol/L	18.9 (17.6-20.3) [17.9-20.0] N = 249	19.3* (18.1-20.5) [18.4-20.2] N = 540	18.9 (17.5-20.2) [17.8-20.0] N = 228	19.2* (17.9-20.3) [18.3-20.1] N = 441
Reticulocyte count (absolute) TRET E12/L	0.033 (0.006-0.069) [0.007-0.059] N = 234	0.045* (0.014-0.091) [0.013-0.077] N = 534	0.047 (0.010-0.111) [0.000-0.093] N = 229	0.052 (0.014- 0.108) [0.012-0.091] N = 431
Platelet count PLT E6/L	325.3 (138.0-506.0) [179.6-471.1] N = 249	378.1* (179.0-590.0) [223.6-532.6] N = 534	335.7 (111.0-579.0) [171.1-500.4] N = 236	398.0* (187.0-611.0) [240.2-555.7] N = 437
White blood cell count WBC E9/L	10.5 (3.76-18.36) [4.69-16.29] N = 240	11.1 (4.41-19.44) [4.94-17.20] N = 536	10.3 (4.35-19.14) [4.50-16.15] N = 227	10.6 (4.29-18.91) [4.74-16.53] N = 435
Neutrophil count ANEU E9/L	4.9 (1.02-12.26) [0.39-9.46] N = 234	5.6 (0.84-12.15) [0.98-10.2] N = 529	5.3 (0.70-13.32) [0-10.80] N = 226	5.7 (1.29-11.65) [1.34-10.05] N = 416
Lymphocyte count ALYM E9/L	4.7 (1.88-9.81) [1.04-8.36] N = 246	4.7 (1.53-9.04) [1.53-7.82] N = 517	4.1 (1.27-7.81) [1.58-6.72] N = 227	4.1 (1.35-8.47) [1.15-7.12] N = 433
Fibrinogen FIB g/L	2.08 (1.60-2.60) [1.6 2– 2.53] N = 123	2.36* (1.40-3.30) [1.70-3.03] N = 375	1.95 (1.40-2.50) [1.47– 2.42] N = 137	2.16* (1.40-3.10) [1.51-2.82] N = 334
Prothrombin PT seconds	16.1 (13.4-18.7) [14.0-18.3] N = 249	14.1* (11.2-17.1) [11.9-16.3] N = 522	15.5 (12.5-18.7) [13.3-17.8] N = 233	13.7* (11.0-16.5) [11.7-15.7] N = 433
Activated partial thromboplastin time APTT seconds	20.6 (16.8-24.6) [18.0-23.4] N = 248	19.4* (14.8-23.8) [16.2-22.5] N = 517	20.2 (17.1-23.4) [17.7-22.7] N = 228	19.2* (14.8-23.6) [15.9-22.5] N = 432

For these data, mean values (with the exception of reticulocyte count and fibrinogen concentration) are rounded to one significant digit.

mean.

(observed interval).

[95% reference interval].

P ≤ .0001 compared with Mauritius origin N = sample size.

Table 5.

North America Serum Biochemistry Results.

North AmericaSerum Biochemistry	Males			Females
Parameter	Mauritius	China	Cambodia	Mauritius	China	Cambodia
Alanine aminotransferase ALT U/L	48.5^a (20.0-96.0)^b [19.7-77.3]^c N = 101	48.2 (16.0-86.0) [21.8-74.5] N = 618	44.8 (23.0-77.0) [19.3-70.4] N = 109	49.6 (18.0-104.0) [10.5-88.6] N = 99	45.0 (12.0-85.0) [17.5-72.4] N = 636	41.9 (7.0-81.0) [12.4-71.3] N = 108
Aspartate aminotransferase AST U/L	36.9 (21.0-53.0) [23.8-50.0] N = 97	42.9* (18.0-71.0) [22.6-63.2] N = 615	39.0 (21.0-60.0) [23.4-54.5] N = 107	36.2 (19.0-59.0) [18.7-53.7] N = 100	38.2 (18.0-70.0) [17.0-59.5] N = 636	33.5 (18.0-50.0) [19.8-47.2] N = 106
Alkaline phosphatase ALP U/L	691.1 (327.0-1096.0) [380.6-1001.6] N = 108	496.0* (206.0-903.0) [216.9-775.1] N = 629	465.0* (216.0-768.0) [225.3-704.7] N = 109	395.1 (187.0-697.0) [158.0-632.2] N = 102	390.1 (118.0-743.0) [140.5-639.7] N = 652	337.0 (150.0-564.0) [165.1-508.9] N = 109
Gamma glutamyltransferase GGT U/L	129.8 (85.0-192.0) [81.1-178.6] N = 106	67.3* (22.6-117.8) [31.7-102.9] N = 617	63.1* (25.0-109.9) [25.4-100.8] N = 108	83.5 (43.0-122.69) [52.6-114.4] N = 105	56.3* (20.7-99.6) [26.7-85.9] N = 636	50.2* (22.3-86.9) [22.7-77.7] N = 111
Creatine kinase CK U/L	218.9 (86.0-501.0) [15.3-422.5] N = 83	207.9 (74.0-434.0) [76.5-339.3] N = 521	187.2 (86.0-332.0) [88.2-286.2] N = 101	200.2 (90.0-507.0) [32.5-367.9] N = 71	202.6 (63.0-449.0) [58.6-346.7] N = 537	199.0 (106.0-399.0) [71.0-327.0] N = 104
Total bilirubin TBIL mg/dL	0.2 (0.1-0.4) [0.1-.04] N = 90	0.2 (0.1-0.5) [0.1-0.4] N = 575	0.2 (0.10-0.46) [0.1-0.4] N = 108	0.2 (0.1-0.4) [0.1-0.4] N = 98	0.2 (0.1-0.5) [0.1-0.4] N = 607	0.2 (0.1-0.5) [0.1 -0.4] N = 104
Total cholesterol CHOL mg/dL	115.6 (70.0-168.0) [74.8-156.3] N = 109	136.1* (68.0-207.0) [84.3-187.9] N = 638	139.9* (86.0-198.0) [91.6-188.3] N = 109	105.9 (62.0-153.0) [71.4-140.4] N = 105	132.4* (66.0-198.0) [84.1-180.7] N = 654	135.5* (70.0-202.0) [83.1-187.8] N = 113
Triglyceride TRIG mg/dL	46.4 (23.0-74.0) [23.9-68.9] N = 105	43.6 (16.0-89.0) [13.5-73.7] N = 635	53.1 (20.0-87.0) [25.4-80.9] N = 108	55.0 (25.0-85.0) [31.2-78.7] N = 103	46.1* (12.0-85.0) [17.5-74.8] N = 657	52.1 (26.0-78.0) [29.5-74.6] N = 106
Glucose GLU mg/dL	64.6 (42.0-104.0) [37.5-91.6] N = 107	66.1 (35.0-109.0) [37.1-95.2] N = 634	66.8 (39.0-96.0) [45.1-88.6] N = 107	69.0 (44.0-100.0) [45.9-92.0] N = 103	66.1 (33.0-130.0) [39.2-93.0] N = 648	67.8 (40.0-97.0) [45.1-90.4] N = 113
Urea nitrogen UN mg/dL	20.0 (12.0-28.0) [13.3-26.7] N = 108	20.4 (9.0-32.0) [12.4-28.4] N = 635	21.9 (13.0-31.0) [14.4-29.5] N = 109	19.0 (12.0-31.0) [11.2-26.8] N = 104	19.4 (10.0-29.0) [11.7-27.1] N = 648	21.0 (13.0-30.0) [13.8-28.3] N = 111
Creatinine CREAT mg/dL	0.8 (0.6-1.1) [0.6-1.0] N = 46	0.6* (0.5-0.8) [0.5-0.8] N = 389	0.7* (0.5-1.0) [0.5-0.9] N = 71	0.8 (0.6-1.0) [0.6-1.0] N = 48	0.6* (0.3-0.9) [0.4-0.8] N = 451	0.7* (0.5-1.0) [0.5-0.9] N = 73
Total protein TP g/dL	8.2 (7.1-9.3) [7.2-9.1] N = 108	7.6* (6.4-8.8) [6.7-8.4] N = 642	7.7* (7.1-8.5) [7.1-8.4] N = 106	8.2 (7.3-9.1) [7.5-8.9] N = 103	7.6* (6.4- 8.7) [6.7-8.4] N = 660	7.7* (6.9-8.5) [7.0-8.4] N = 108
Albumin ALB g/dL	4.8 (3.9-5.6) [4.0-5.5] N = 108	4.7 (3.8-5.7) [4.1-5.4] N = 638	4.8 (4.2-5.2) [4.3-5.2] N = 108	4.6 (4.1-5.3) [4.0-5.2] N = 99	4.6 (3.7-5.6) [3.9-5.4] N = 655	4.7 (3.9-5.3) [4.1-5.2] N = 112
Globulin GLOB g/dL	3.4 (2.7-4.3) [2.6-4.1] N = 107	2.8* (1.9-3.8) [2.1-3.5] N = 633	3.0* (2.2-3.8) [2.3-3.6] N = 109	3.6 (2.6-4.5) [2.8-4.3] N = 105	2.9* (1.8-4.1) [2.1-3.7] N = 653	3.0* (2.3-3.9) [2.6-3.7] N = 111
Albumin: Globulin A: G ratio	1.4 (1.0-2.0) [1.0-1.9] N = 109	1.7* (1.0-2.5) [1.2-2.2] N = 630	1.6* (1.2-2.1) [1.2-2.0] N = 107	1.3 (1.0-1.6) [1.0-1.6] N = 93	1.6* (0.9-2.4) [1.0-2.2] N = 648	1.6* (1.0-2.2) [1.0-2.1] N = 113
Calcium Ca mg/dL	10.6 (9.6-12.0) [9.6-11.6] N = 108	10.4* (9.0-11.7) [9.3-11.4] N = 634	10.5 (8.9-12.2) [9.3-11.8] N = 110	10.5 (9.7-11.5) [9.7-11.3] N = 100	10.4 (9.0-11.9) [9.4-11.4] N = 660	10.6 (9.6-12.2) [9.5-11.7] N = 112
Inorganic phosphorus PHOS mg/dL	6.2 (4.5-7.8) [4.9-7.5] N = 103	6.7* (4.3-9.2) [4.9-8.5] N = 635	7.1* (5.2-9.6) [5.4-8.8] N = 110	5.3 (3.3-7.4) [3.6-7.0] N = 105	6.2* (3.4-8.7) [4.3-8.0] N = 662	6.3* (4.4-8.6) [4.6-8.0] N = 112
Sodium Na mmol/L	150.3 (143.0-160.0) [143.9-156.8] N = 109	149.4 (142.0-157.0) [143.8-155.0] N = 634	150.1 (144.0-158.0) [144.3-155.8] N = 110	149.6 (143.0-158.0) [143.6-155.7] N = 102	149.2 (142.0-157.0) [143.5-154.9] N = 655	149.3 (141.0-157.0) [143.4-155.2] N = 112
Chloride Cl mmol/L	104.2 (99.0-109.0) [99.9-108.5] N = 106	104.5 (99.0-110.0) [100.1-108.9] N = 630	103.8 (99.0-109.0) [99.6-108.1] N = 109	105.7 (100.0-111.0) [101.5-109.9] N = 100	105.0 (98.0-113.0) [100.1-109.9] N = 664	104.8 (99.0-112.0) [99.9-109.6] N = 112
Potassium K mmol/L	5.1 (4.1-6.4) [4.1-6.1] N = 108	4.9 (3.6-6.3) [3.9-5.9] N = 635	4.8* (3.9-5.8) [3.9-5.7] N = 109	5.0 (4.1-6.4) [4.0-5.9] N = 104	4.9 (3.8-6.3) [4.0-5.8] N = 655	4.9 (3.7-6.1) [3.9-5.8] N = 113

For these data, mean values are rounded to one significant digit.

mean.

(observed interval).

[95% reference interval].

P ≤ .0001 compared with Mauritius origin N = sample size.

Table 6.

European Union Serum Biochemistry Results.

European UnionSerum Biochemistry	Males		Females
Parameter	Mauritius	Vietnam	Mauritius	Vietnam
Alanine aminotransferase ALT U/L	56.3^a (24.0-105.6)^b [21.0-91.6]^c N = 242	47.5* (11.4-86.6) [21.4-73.7] N = 514	51.6 (18.9-104.4) [18.3-84.9] N = 220	43.2* (12.1-83.5) [16.5-70.0] N = 418
Aspartate aminotransferase AST U/L	45.5 (24.5-78.7) [22.8-68.2] N = 239	43.7 (22.5-75.0) [22.9-64.4] N = 518	42.3 (19.8-72.2) [21.6-63.0] N = 218	38.5* (17.9-69.2) [18.7-58.3] N = 421
Alkaline phosphatase ALP U/L	825.7 (252.5-1419.0) [318.7-1332.7] N = 240	546.0* (189.1-997.5) [236.1-856.0] N = 526	623.9 (162.3-1244.4) [205.3-1042.4] N = 231	324.5* (105.1-672.5) [83.5-565.5] N = 433
Gamma glutamyltransferase GGT U/L	164.2 (79.2-260.1) [90.3-238.1] N = 243	79.8* (22.9-140.4) [36.2-123.4] N = 534	119.4 (67.4-204.6) [62.3-176.5] N = 228	57.2* (21.3-95.7) [28.5-86.0] N = 421
Glutamate dehydrogenase GLDH U/L	20.0 (7.6-36.5) [8.0-32.1] N = 236	15.1* (6.7-26.9) [6.7-23.5] N = 489	17.7 (7.6 -34.1) [6.8-28.6] N = 220	14.4 (6.4-28.4) [5.2-23.7] N = 385
Creatine kinase CK U/L	212.2 (79.7-438.8) [56.6-367.7] N = 49	206.7 (69.3-419.7) [65.9-347.5] N = 336	250.9 (68.5-588.0) [9.4-492.4] N = 76	182.8* (56.9-389.4) [45.1-320.5] N = 256
Total bilirubin TBIL µmol/L	6.1 (3.1-10.6) [3.0-9.3] N = 244	6.1 (2.8-10.0) [3.4-8.7] N = 521	6.9 (3.0-12.6) [3.1-10.8] N = 226	5.9* (3.2-9.8) [3.3-8.6] N = 426
Total cholesterol CHOL mmol/L	3.0 (1.6 -4.2) [2.0-4.0] N = 249	3.5* (1.8-5.3) [2.2-4.7] N = 540	3.1 (1.9-4.3) [2.1-4.0] N = 234	3.5* (1.8-5.4) [2.2-4.9] N = 440
Triglyceride TRIG mmol/L	0.5 (0.2-0.8) [0.2-0.7] N = 243	0.4* (0.1-0.7) [0.2-0.6] N = 521	0.5 (0.2-0.8) [0.3-0.7] N = 227	0.4* (0.2-0.8) [0.2-0.7] N = 431
Glucose GLU mg/dL	3.4 (2.1-5.1) [2.2-4.6] N = 243	3.8* (2.1-5.8) [2.3-5.2] N = 533	3.0 (1.9-4.5) [2.0-4.0] N = 230	3.6* (2.1-5.8) [2.1-5.1] N = 435
Blood urea BU mmol/L	6.8 (3.6-10.0) [4.3-9.3] N = 248	8.0* (4.5-11.7) [5.3-10.6] N = 533	6.4 (3.0-9.8) [3.9-8.8] N = 234	7.9* (4.2-11.7) [5.0-10.7] N = 440
Creatinine CREA µmol/L	71.4 (51.8-96.4) [54.5-88.3] N = 227	75.5* (56.0-99.0) [59.1-91.9] N = 540	68.6 (54.9-85.2) [57.0-80.2] N = 229	70.9* (53.4-89.4) [57.5-84.3] N = 439
Total Protein TP g/L	79.7 (67.7-91.3) [70.8-88.5] N = 248	75.0* (64.0-87.7) [65.8-84.3] N = 538	79.6 (67.4-92.6) [70.2-89.0] N = 235	73.3* (62.0-85.3) [64.1-82.4] N = 438
Albumin ALB g/L	46.6 (37.1-59.2) [36.6-56.5] N = 251	42.6* (37.5-48.1) [39.4-45.9] N = 432	42.7 (38.7-45.0) [39.7-45.7] N = 191	41.4* (35.5-45.0) [37.2-45.6] N = 397
Globulin GLOB g/L	33.3 (22.4-44.6) [24.8-41.7] N = 250	30.5* (20.1-41.4) [22.4-38.6] N = 539	34.9 (23.0-45.3) [26.5-43.4] N = 232	30.8* (20.6-41.2) [23.3-38.2] N = 437
Albumin: Globulin A: G ratio	1.4 (0.9-2.3) [0.8-2.0] N = 248	1.4 (1.0-2.1) [1.0-1.9] N = 502	1.2 (0.8-1.7) [0.9-1.6] N = 211	1.4* (0.8-1.9) [1.0-1.7] N = 421
Calcium CA mmol/L	2.7 (2.4-3.0) [2.5-2.9] N = 246	2.7 (2.4-3.0) [2.4-2.9] N = 529	2.6 (2.4-2.9) [2.4-2.8] N = 231	2.6 (2.3-3.0) [2.4-2.9] N = 437
Inorganic Phosphorus PHOS mmol/L	1.9 (0.9-2.7) [1.2-2.6] N = 251	2.0 (1.1-2.8) [1.4-2.6] N = 540	1.8 (1.1-2.6) [1.3-2.4] N = 234	1.8 (1.0-2.5) [1.2-2.4] N = 440
Sodium NA mmol/L	148.2 (140.9-156.2) [142.3-154.1] N = 242	147.6 (139.2-156.5) [141.4-153.9] N = 526	146.9 (140.7-154.3) [141.3-152.4] N = 234	147.5 (140.3-155.3) [141.8-153.2] N = 440
Chloride CL mmol/L	110.0 (103.8-116.3) [105.0-115.1] N = 248	109.7 (103.5-116.4) [104.6-114.8] N = 529	110.0 (104.9-114.5) [106.3-113.6] N = 234	110.2 (103.7-116.9) [105.4-114.9] N = 437
Potassium K mmol/L	5.1 (4.0-6.3) [4.1-6.0] N = 245	5.0 (3.6-6.5) [4.0-6.1] N = 535	4.9 (3.7-6.2) [4.0-5.8] N = 233	4.9 (3.6-6.3) [3.9-6.0] N = 441

For these data, mean values are rounded to one significant digit.

mean.

(observed interval).

[95% reference interval].

P ≤ .0001 compared with Mauritius originN = sample size

Qualitative (eg, minimal, mild, moderate, marked) and quantitative (% difference) descriptors are used in the Results section to give context to the magnitude of change between group means of Cambodian, Chinese, and Vietnamese origin macaques compared with Mauritius origin macaques. Percent difference was calculated by dividing the difference between Cambodian, Chinese, or Vietnamese origin group mean value and Mauritius origin group mean value by the Mauritius origin group mean value and then multiplying that by 100. Data are presented in box plots for select endpoints (Figures 1 -4). Box plots partition the data distribution into quartiles. The box indicates the positions of the upper and lower quartiles; the interior of the box indicates the inner quartile range, which is the area between the upper and lower quartiles and consists of 50% of the distribution. Vertical lines extending on either end of the box (referred to as “whiskers”) indicate the distribution, either minimum or maximum values in the dataset. Outliers are represented individually by symbols above or below the whiskers. The crossbar within the box indicates the median of the dataset. The mean value is denoted with a symbol within the box.

Figure 1.

Box plot representations of select endpoints (North America; males).

Figure 2.

Box plot representations of select endpoints (North America; females).

Figure 3.

Box plot representations of select endpoints (European Union [Münster/Muenster]; males).

Figure 4.

Box plot representations of select endpoints (European Union [Münster/Muenster]; females).

Results

Hematology and Coagulation

Differences in hematology values were identified between Mauritius origin macaques and those from Cambodian, Chinese, and Vietnamese origin (Tables 3 and 4, Figures 1 –4). One of the most notable and well documented differences in hematology parameters is that Mauritius origin macaques have smaller red blood cell size (lower mean corpuscular volume [MCV]) and higher red blood cell (RBC) count compared with mainland Asian macaques.^5,20 Similar differences in the RBC population were demonstrated in our data showing minimally to mildly lower RBC count (up to −19%) and red cell distribution width (RDW)—(up to −14%) and minimally to mildly higher absolute reticulocyte count (up to +119%), MCV (up to +19%), mean corpuscular hemoglobin (MCH)—(up to +25%), and mean corpuscular hemoglobin concentration (MCHC)—(up to +5%), when macaques of Cambodian, Chinese, and Vietnamese origin were compared with Mauritius origin macaques. These differences in red blood cell indices were consistent and similar in NA and EU data, but as RDW was not reported in the EU data, an RDW comparison of Vietnamese with Mauritian origin was not made. Inter-animal variability was noted in total and differential white blood cell (leukocyte) counts in animals of the origins compared, but no discernible/remarkable group differences were identified due to overlapping individual results amongst the groups. The only notable difference in standard coagulation test results was minimally to mildly shorter prothrombin time (PT)—(up to 2 seconds) in Vietnam origin macaques compared with Mauritius origin macaques.

Serum Biochemistry

Serum biochemistry differences were noted between macaques of Mauritius origin compared with the other origins (Tables 5 and 6, Figures 1 –4). Small differences in hepatocellular/hepatobiliary enzyme activities were noted. Vietnam origin macaques were noted with minimally to mildly lower glutamate dehydrogenase (GLDH)—(up to −25%) activity compared with Mauritius origin macaques; minimally to mildly lower alkaline phosphatase (ALP)—(up to −48%) activity was noted in Cambodian (males only), Chinese (males only), and Vietnamese (both sexes) origin macaques compared with Mauritius origin macaques; and minimally to mildly lower gamma glutamyl transferase (GGT)—(up to −51%) activity was noted in Cambodian, Chinese, and Vietnamese origin macaques compared with Mauritius origin macaques. Serum total protein (up to −8%) and globulin (up to −19%) concentrations were minimally to mildly lower in Cambodian, Chinese, and Vietnamese origin macaques compared with Mauritius origin macaques. Minimally to mildly lower serum albumin (up to -9%) concentration was noted in Vietnamese origin macaques compared with Mauritius origin macaques. Serum urea nitrogen (up to +23%) and creatinine (up to +6%) concentrations were minimally to mildly higher in Vietnam origin macaques compared with Mauritius origin macaques. Minimally to mildly lower creatinine (up to −25%) concentration was also noted in macaques of Cambodian or Chinese origin when compared with Mauritian macaques. Total cholesterol (up to +28%) concentration was mildly higher in Cambodian, Chinese, and Vietnamese origin macaques compared with Mauritian macaques, whereas triglyceride (up to −20%) concentration was minimally to mildly lower in Vietnamese origin macaques when compared with Mauritian macaques.

Discussion

Cynomolgus macaques are the most widely used nonhuman primate species in preclinical pharmaceutical safety assessment studies and are well suited and established as translational models for drug development.²¹ Given the increased demand for these animals in the development of safe and effective therapeutics for human medicine coupled with a global shortage of Chinese origin NHPs exacerbated by exportation bans, the supply chain has been impacted and macaques of non-mainland Asia origin have been considered for use, and macaques of Mauritius, Vietnamese, and Cambodian origin are being used in increasing numbers in preclinical drug development programs. The high level of genetic diversity between subpopulations of different geographic origin, such as between cynomolgus macaques from Mauritius and those from mainland Asia, is an important consideration for any scientific study with cynomolgus macaques.^21-23 Genetic divergence also affects physiological traits and knowing the geographic origin and genetic background of NHPs before beginning any study is a well-recognized advantage.²¹ This current investigation of clinical pathology data supports the published literature and in-press work and helps to better characterize differences, or lack thereof, between subpopulations of cynomolgus macaques.

Differences between cynomolgus macaque origins were identified and described in our data comparison of clinical pathology test results. These differences were of small magnitude and considered minor which supports the inclusion of different origin of macaques for use in preclinical toxicity studies. Distinctions in hematology parameters were most notable, whereas smaller differences were identified in coagulation and serum biochemistry parameters. Differences in red blood cell parameters in Cambodian, Chinese, and/or Vietnamese origin macaques compared with Mauritius origin macaques consisted of minimally to mildly lower RBC counts and RDW (NA only), and minimally to mildly higher absolute reticulocyte counts, MCV, MCH, and MCHC which were consistent with the literature.^5,16 These origin-specific differences reflect the genetic diversity of different origin subpopulations of cynomolgus macaques. Despite lower numbers of red blood cells of larger size in Cambodian, Chinese, and Vietnamese origin macaques compared with Mauritius origin macaques (indicated by lower RBC count and higher MCV noted in Cambodian, Chinese, and Vietnamese origin), red cell mass was considered similar amongst the 4 origins assessed because hemoglobin concentration and hematocrit remained generally comparable amongst the subpopulations which is also consistent with the literature.¹⁶

Inter-animal variability, which occasionally resulted in statistically significant differences between groups, was noted in total and differential white blood cell counts in animals of all origins; however, no biologically important group differences were identified in macaques of Cambodia, China, or Vietnam origin when compared with those originating from Mauritius, which is consistent with findings in the published literature.⁵ The shorter PT in macaques of Vietnam origin reached statistical significance when compared with those of Mauritius origin, although a similar difference was not identified when Chinese or Cambodian origin were compared with Mauritian origin animals. Minimally to mildly shorter PT has been previously described in Mauritius origin compared with Asian origin macaques but that study only compared mixed Asian (ie, animals of Cambodian, Vietnamese, and Indonesian origin), Cambodian, and Mauritian whereas our current study clearly defined animal origin.⁵ In our data, no impactful difference in activated partial thromboplastin time (APTT) or fibrinogen concentration was identified in the 3 Asian origin subpopulation origins when compared with Mauritius origin macaques.

Minor differences were identified in serum chemistry results from macaques of Cambodia, China, or Vietnam origin when compared with Mauritius origin macaques. When compared with Mauritius origin macaques, small differences (minimal to mild in magnitude) were identified in hepatocellular/hepatobiliary enzyme activities which consisted of lower GLDH activity in Vietnam origin macaques; lower ALP activity in Cambodia and China (males only) and Vietnam (both sexes) origin macaques; and lower GGT activity in Cambodian, Chinese, and Vietnamese origin animals. Additional clinical chemistry differences were minimal to mild in magnitude and somewhat inconsistent between Cambodian, Chinese, and Vietnamese origin macaques compared with Mauritius origin macaques. Serum total protein and globulin concentrations were lower in Cambodian, Chinese, and Vietnamese origin macaques whereas serum albumin concentration was lower only in Vietnamese origin macaques. Serum urea nitrogen and creatinine concentrations were higher in Vietnam origin macaques whereas lower creatinine concentration was noted in macaques of Cambodian or Chinese origin when compared with Mauritian macaques. Total cholesterol concentration was higher in Cambodian, Chinese, and Vietnamese origin macaques compared with Mauritian macaques while triglyceride concentration was lower in Vietnamese origin macaques when compared with Mauritian macaques. Although not an objective of this current study, appreciable differences were not noted between macaques of different mainland Asia origin in the NA dataset (China compared with Cambodia), which correlated with the literature.^11,24,25

Some of the differences between macaques of different origins previously reported in the literature were attributed to age, animal condition, and/or background findings.^5,16,24,25 In our current study, many variables were standardized (eg, age, sex, husbandry, and health status) to prevent such biases in the identification of origin-related differences. For example, higher ALP activity has been reported associated with younger aged animals, and therefore animals under 2 years old were excluded from the current study to prevent the influence of young age on clinical pathology results.⁵ A potential source of bias or variability in our current investigations was recognized becasue the data from the MSN and GRN lab were combined to make up the NA data set despite the lack of correlation of instruments between the 2 sites. Any impact was considered negligible to minimal given data from a single study analyzed at the GRN location was combined with data analyzed at the MSN laboratory (MSN data made up most of the NA data). In addition, the effect of animal source/supplier has been identified in other research species and should also be considered as a variable in assessing clinical pathology data of NHPs in biomedical research.²⁰ Future work should be considered to identify any effect of source/supplier of NHPs of different origins and investigate the impact of genetics and in-breeding within source/supplier colonies on clinical pathology parameters assessed for NHP used in preclinical studies.

Numerical differences were present and recognized in our data. These numerical differences occasionally reached statistical significance, but were considered not toxicologically relevant because the differences were small in magnitude, overlap was appreciated in the individual results and observed ranges, and such differences are not expected to impact the interpretation of preclinical toxicology data in the context of a well-designed study (ie, appropriate predose/baseline data and control group comparators).^26,27 Statistical significance can also be influenced by seemingly small differences in the assessment of clinical pathology data given the often tightly regulated parameters (eg, electrolytes and minerals) and/or parameters with narrow “physiologic ranges” (eg, total bilirubin, creatinine), large sample size “n”, and results easily impacted by preanalytical/analytical variability (eg, coagulation times or minor leukocyte populations impacted by sample quality or anticoagulant ratio). Statistical analysis of toxicological clinical pathology data should not be solely relied upon, or used in isolation, to interpret differences in data sets. Interpretations should be based on an understanding of each analyte/parameter employing a weight of evidence approach rather than to substitute statistical analysis for the scientific process of appropriate data interpretation.^26,27

Based on this study and published literature characterizing differences in clinical pathology data, macaques of different origin (eg, Cambodian, Chinese, Vietnamese, and Mauritian) can be used to support drug development programs.^5,24,25 While Mauritius origin macaques are genetically and physiologically different from mainland Asia origin macaques, the herein described differences in clinical pathology endpoints should not preclude their use in preclinical pharmaceutical safety programs. Given this, differences in clinical pathology data based on macaque origin should be considered when interpreting preclinical safety assessment toxicology data to avoid erroneous clinical pathology data interpretation. Same origin macaques should be used for individual studies, and ideally (but not always possible) across a drug development program, to minimize impacts on clinical pathology data. Furthermore, CRO facilities should maintain origin specific (separate macaque subpopulation) clinical pathology reference intervals, even though controls and predose/baseline data are best used, and preferred, for the interpretation of preclincial study data.^1,26,27 Finally, the importance of well-designed preclinical toxicity studies with predose/baseline clinical pathology assessments and concurrent controls for proper interpretation of study data cannot be overemphasized.^16,26,27

Animal Use Statement

Studies performed in US-based facilities were conducted at AAALAC International-accredited contract research organization (CRO) facilities, and in compliance with United States federal regulations and recommendations detailed in The Guide for the Care and Use of Laboratory Animals (National Research Council 1996), and in accordance with Labcorp (formerly Covance) policies on the care, welfare, and treatment of laboratory animals. In addition, all procedures involving the animals were reviewed and approved by the respective facilities Institutional Animal Care and Use Committees (IACUC) prior to dose administration. Studies performed in the EU were conducted in accordance with the European Convention for the Protection of Vertebrate Animals Used for Experimental and Other Scientific Purposes (Strasbourg, Council of Europe), in addition to institutional ethical committees.

Footnotes

Acknowledgements

The authors would like to recognize and thank the following for their contribution to this manuscript. Laura Boone and Niraj Tripathi for their critical scientific review; Scott Templeton, Andrew Newell, and Kelly Ashcroft-Hawley for data management; and Gerhard Weinbauer for his passionate approach to NHP data projects.

Declaration of Conflicting Interests

The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Funding

The author(s) received no financial support for the research, authorship, and/or publication of this article.

ORCID iD

Tara Arndt

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Comparison of Routine Hematology,Coagulation,and Clinical Chemistry Parameters of Cynomolgus Macaques of Mauritius Origin With Cynomolgus Macaques of Cambodia,China,and Vietnam Origin

Abstract

Keywords

Introduction

Material and Methods

Animals

Clinical Pathology

Statistical Analysis

Results

Hematology and Coagulation

Serum Biochemistry

Discussion

Animal Use Statement

Footnotes

Acknowledgements

Declaration of Conflicting Interests

Funding

ORCID iD

References