The CD276 Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the near-haploid human HAP1 cell line, featuring targeted disruption of the CD276 gene. This heterogeneous pool of cells harbors a diverse array of indels at the CD276 locus, creating a loss-of-function model for robust immunological studies. The polyclonal format circumvents clonal bias, ensuring phenotypic assessments reflect global gene disruption effects. This ready-to-use product streamlines functional investigations into immune checkpoint mechanisms, providing a reliable platform for downstream assays.
The HAP1 cell line originates from the KBM-7 chronic myeloid leukemia line and maintains a near-haploid karyotype, with only chromosome 8 present in disomic form. This genetic simplicity eliminates allelic redundancy, allowing direct genotype-phenotype linkage following gene editing. HAP1 cells exhibit fibroblast-like morphology and retain crucial signaling pathways pertinent to oncology and immunology. Their haploid nature is particularly beneficial for CRISPR-based functional genomics, enabling unambiguous interpretation of gene disruption experiments without masking by a second allele.
CD276 (B7-H3) is an immune checkpoint protein that modulates T-cell responses, frequently overexpressed in tumors to facilitate immune evasion. Its expression is upregulated by IFN-??, LPS, CD40L, IL-4, and GM-CSF. Upon binding to an unidentified receptor, possibly via homodimerization, CD276 recruits SHP-2 to inhibit TCR signaling, thereby attenuating the PI3K/AKT/mTOR and MAPK/ERK cascades. This leads to reduced IL-2 production, impaired T-cell proliferation, and induction of T-cell anergy, while also promoting IL-10 secretion to foster an immunosuppressive milieu. Additionally, CD276 may interact with CD28/CTLA-4 family members, fine-tuning T-cell costimulation.
In the HAP1 context, CD276 knockout provides a genetically defined system for dissecting its role in immune checkpoint regulation. The near-haploid background ensures that CRISPR-mediated disruption directly ablates CD276 function, avoiding confounding effects from genetic redundancy. This model is instrumental for investigating CD276??s influence on PI3K/AKT and ERK signaling, and its impact on cytokine networks such as IL-2 and IL-10. The polyclonal population averages out off-target events, enhancing the reliability of target-specific conclusions and making it suitable for rigorous comparative studies.
Research applications encompass cancer immunotherapy, immune checkpoint blockade studies, and target validation for CD276-directed therapies. This product enables functional genomics screens and drug discovery, with compatibility in T-cell co-culture experiments for proliferation, cytokine release (ELISA), and cytotoxicity assays. Additional assays include flow cytometry for CD276 expression, western blotting, RT-qPCR, phospho-signaling analysis (e.g., phospho-AKT, phospho-ERK), RNA-seq, and in vivo tumor models. ADCC assays can assess CD276-targeted biologics. For inquiries, please contact Ascent Research.
