Figure 4.
FNA-Based Patient-Derived RCC Organoids Contain Immune Cells from the Tumor Microenvironment
(A) Immune cells are extracted and plated during FNA-PDO collection and generation. This representative RCC FNA-PDO demonstrates classic morphology by H&E with positive cytokeratin staining (AE1/AE3) and infiltration of B cells (CD20), T cells (CD3), and macrophages (CD163) by immunohistochemistry (IHC).
(B) Cell viability at collection was assessed using flow cytometry on fresh FNA material collected from RCCs versus fresh material following tumor digestion of RCCs, showing similar overall viability and CD45+ viability between FNA and digestion extraction methods (n = 4 tumors). Error bars represent standard error of the mean (SEM).
(C) Both FNA and digestion methods have similar lymphoid subset viability at collection (n = 4 tumors, error bars represent SEM).
(D) Both FNA and digestion methods have similar myeloid subset viability at collection (n = 4 tumors, error bars represent SEM).
(E) Absolute immune cell counts at collection for FNA (nine needle passes per tumor) and digestion (1 cm3 tissue per tumor completely digested, n = 4 tumors, error bars represent SEM).
(F) RCC FNA-PDO and digestion-PDO immune cell recovery counts at 2 weeks of disc culture demonstrate significantly improved survival of CD45+ cells from FNA-PDOs as compared with digestion-PDOs in both Wnt3A-enriched and non-Wnt-enriched complete media (n = 4 tumors, error bars represent SEM, ∗p < 0.05).
(G) FNA-PDO CD45+ cell recovery counts at 2 weeks in culture demonstrates improved CD45+ cell survival with both Wnt-enriched media and 100 IU/mL IL-2 supplementation (n = 2 tumors, error bars represent SEM, ∗p < 0.05).
(H) FNA-PDO lymphocyte recovery counts at 2 weeks in culture demonstrates CD3+ lymphocyte survival in all conditions and improved CD8+ lymphocyte survival with Wnt-enriched media and IL-2 supplementation (n = 2 tumors, error bars represent SEM, ∗p < 0.05).
(I) FNA-PDO myeloid recovery counts at 2 weeks in culture demonstrates improved survival of CD14+CD11b+ myeloid cells when cultured with Wnt-enriched media (n = 2 tumors, error bars represent SEM, ∗p=<0.05).