2025.05.30
Peripheral blood mononuclear cells (PBMCs), as a core cell population in immunological research, have emerged as critical tools for preclinical in vitro experiments in new drug development due to their unique ability to efficiently mimic the in vivo immune microenvironment. This article focuses on animal-derived PBMC models and systematically reviews their application value in preclinical research, including drug screening, elucidation of immunomodulatory mechanisms, and toxicity evaluation.
Canine PBMCs: These are utilized in developing natural killer (NK) cell immunotherapy products. Co-culturing CD5-depleted cells with PBMCs from healthy beagle dogs in vitro generates highly active NK cell populations within 14 days. Their cytotoxicity, phenotypic characteristics, and cytokine secretion capacity can be evaluated via flow cytometry and RNA sequencing [1]. Comparable results observed in human PBMCs validate the translational potential of the canine model [1].
Transgenic mouse PBMCs: These are employed to assess the tumor-killing efficacy of bispecific antibodies (e.g., YG-003D3). In vitro experiments show that activated PBMCs achieve a tumor cell killing rate of up to 20%, significantly outperforming monotherapy groups [2].
Mouse PBMCs: These are used to evaluate the immunosuppressive effects of aluminum adjuvant (Al). In vitro data demonstrate that aluminum treatment significantly reduces activation levels of B cells and T cells (CD4+/CD8+) and suppresses IgM/IgG secretion, aligning with in vivo results and confirming the reliability of PBMCs as a model for immunosuppression research [3].
Mini-pig PBMCs: These facilitate cross-species comparisons of immune responses. Flow cytometry analysis of compound-induced PBMC proliferation reveals high similarity between mini-pig and human PBMCs in their responses to immunosuppressive drugs, supporting the applicability of mini-pigs as a translational medicine model [4].
[media-image-TbhQlY9SNck3JalXkbAp.png]
Supplementary evaluation of immune-related toxicity: Hepatic microsomes from dogs, mini-pigs, and monkeys (e.g., DLMs, PLMs, MyLMs) are commonly used in drug metabolism studies, while PBMCs serve as a supplementary model to assess immune-related toxicity. For example, PBMC experiments further reveal 2,4,6-tribromophenol (TBP)-induced interference with immune cell function, complementing findings from hepatic microsome studies on TBP’s effects on UGT metabolic enzymes [5][6].
Interspecies evaluation of metabolic stability: PBMCs are used to assess differences in drug metabolic stability. For instance, S-MRI-1867 exhibits low-to-moderate plasma clearance and high volume of distribution in monkeys, dogs, and rats. Its in vitro PBMC metabolic data complement hepatic microsome results, supporting its translational potential in developing neuroprotective agents [7].
Stimulating PBMCs in vitro (e.g., with LPS, PHA, or α-CD3/α-CD28) mimics drug-induced excessive immune responses. In psychiatric research, levels of IFN-γ, IL-6, and TNF-α secreted by stimulated PBMCs are used to evaluate drug-mediated immune regulation, providing a basis for CRS risk prediction [8].
[media-image-WMn2Kfdvcy0t8KpCVn6D.png]
Mini-pig PBMCs: These exhibit high consistency with human PBMCs in responses to immunosuppressive drugs, reinforcing their reliability as a translational model [4].
Non-human primate PBMCs: These show strong alignment with human pharmacokinetic data. For example, steady-state plasma concentrations of long-acting naloxone formulations in monkeys match human therapeutic levels [9].
Limitations of in vitro models: Traditional 2D-cultured PBMCs fail to fully replicate in vivo 3D tissue interactions and drug diffusion properties [10][11].
Correction of interspecies differences: Activity discrepancies in FMO enzymes (drug oxidases) between dogs and humans may affect the predictive value of PBMC metabolic experiments, necessitating integration of genomic data to optimize models [6].
Need for standardization: Current PBMC experiments lack uniform stimulation conditions and endpoint indicators, requiring standardized protocols to enhance data comparability [1][12].
PBMC in vitro models are increasingly integrated with advanced technologies such as organoids and microfluidic chips. For example, combining mini-pig PBMCs with 3D hepatic microtissues enables the construction of more complex in vitro toxicity evaluation platforms [10][13]. Additionally, single-cell sequencing technologies (e.g., cross-species lung tissue transcriptome analysis) provide novel tools to dissect PBMC heterogeneity [14]. These advancements will further enhance the predictive value of PBMCs in new drug development and promote implementation of the 3R principles (replacement, reduction, refinement) in animal experiments [15][16].
References
[1] Preclinical evaluation and first-in-dog clinical trials of PBMC-expanded natural killer cells for adoptive immunotherapy in dogs with cancer. J Immunother Cancer. 2024 Apr 16; DOI: 10.1136/jitc-2023-007963.
[2] PD-1/LAG-3 bispecific antibody potentiates T cell activation and increases antitumor efficacy. Front Immunol. 2022 Nov 28; DOI: 10.3389/fimmu.2022.1047610.
[3] Aluminum promotes B1 cells to produce IL-10 and impairs adaptive immune system. Environ Pollut. 2025 Feb 4; DOI: 10.1016/j.envpol.2025.125791.
[4] Species comparison: human and minipig PBMC reactivity under the influence of immunomodulating compounds. Front Immunol. 2024 Jan 9; DOI: 10.3389/fimmu.2023.1327776.
[5] Identification of Bromophenols’ glucuronidation and its induction on UDP-glucuronosyltransferases isoforms. Ecotoxicol Environ Saf. 2024 Apr 5; DOI: 10.1016/j.ecoenv.2024.116281.
[6] Molecular and functional characterization of flavin-containing monooxygenases in pigs, dogs, and cats. Biochem Pharmacol. 2022 Jun 8; DOI: 10.1016/j.bcp.2022.115125.
[7] In vitro and in vivo pharmacokinetic characterization, chiral conversion and PBPK scaling towards human PK simulation of S-MRI-1867, a drug candidate for Hermansky-Pudlak syndrome pulmonary fibrosis. Biomed Pharmacother. 2023 Oct 25; DOI: 10.1016/j.biopha.2023.115178.
[8] Evidence of innate immune dysfunction in first-episode psychosis patients with accompanying mood disorder. J Neuroinflammation. 2022 Dec 3; DOI: 10.1186/s12974-022-02648-y.
[9] Development and preclinical testing of a naloxone prodrug depot for extended protection against opioid overdose. Nat Commun. 2025 Jan 23; DOI: 10.1038/s41467-025-55945-4.
[10] Pre-clinical 2D and 3D toxicity response to a panel of nanomaterials; comparative assessment of NBM-induced liver toxicity. Drug Deliv Transl Res. 2022 Jun 28; DOI: 10.1007/s13346-022-01170-1.
[11] Sensor-integrated brain-on-a-chip platforms: Improving the predictive validity in neurodegenerative research. Bioeng Transl Med. 2023 Oct 18; DOI: 10.1002/btm2.10604.
[12] Complementary strategies to be used in conjunction with animal models for multiple sclerosis drug discovery: adapting preclinical validation of drug candidates to the need of remyelinating strategies. Expert Opin Drug Discov. 2024 Jul 22; DOI: 10.1080/17460441.2024.2382180.
[13] A deep learning-based system for automatic detection of emesis with high accuracy in Suncus murinus. Commun Biol. 2025 Feb 10; DOI: 10.1038/s42003-025-07479-0.
[14] A pulmonologist’s guide to perform and analyse cross-species single lung cell transcriptomics. Eur Respir Rev. 2022 Jul 27; DOI: 10.1183/16000617.0056-2022.
[15] A preclinical platform for assessing long-term drug efficacy exploiting mechanically tunable scaffolds colonized by a three-dimensional tumor microenvironment. Biomater Res. 2023 Oct 18; DOI: 10.1186/s40824-023-00441-3.
[16] Animal Alternatives OK’d by New Law. Cancer Discov. 2023 Apr 3; DOI: 10.1158/2159-8290.CD-ND2023-0001.
