Tumor-associated macrophages (TAMs) accumulate in the solid tumor microenvironment (TME) and have been shown to promote tumor growth and dampen antitumor immune responses. TAM-mediated suppression of T-cell antitumor reactivity is considered to be a major obstacle for many immunotherapies, including immune checkpoint blockade and adoptive T/CAR-T-cell therapies. An ex vivo culture system closely mimicking the TME can greatly facilitate the study of cancer immunotherapies. Here, we report the development of a 3D TME-mimicry culture that is comprised of the three major components of a human TME, including human tumor cells, TAMs, and tumor antigen-specific T cells. This TME-mimicry culture can readout the TAM-mediated suppression of T-cell antitumor reactivity, and therefore can be used to study TAM modulation of T-cell-based cancer immunotherapy. As a proof-of-principle, the studies of a PD-1/PD-L1 blockade therapy and a MAO-A blockade therapy were performed and validated.

Expressed by cancer stem cells of various epithelial cell origins, CD133 is an attractive therapeutic target for cancers. Autologous chimeric antigen receptor-modified T-cell directed CD133 (CART-133) was first tested in this trial. The anti-tumor specificity and the postulated toxicities of CART-133 were first assessed. Then, we conducted a phase I clinical study in which patients with advanced and CD133-positive tumors received CART-133 cell-infusion. We enrolled 23 patients (14 with hepatocellular carcinoma [HCC], 7 with pancreatic carcinomas, and 2 with colorectal carcinomas). The 8 initially enrolled patients with HCC were treated by a CART-133 cell dose escalation scheme (0.05-2 × 106/kg). The higher CAR-copy numbers and its reverse relationship with the count of CD133+ cells in peripheral blood led to the determination of an acceptable cell dose is 0.5-2 × 106/kg and reinfusion cycle in 23 patients. The primary toxicity is a decrease in hemoglobin/platelet (≤ grade 3) that is self-recovered within 1 week. Of 23 patients, three achieved partial remission, and 14 achieved stable disease. The 3-month disease control rate was 65.2%, and the median progression-free survival was 5 months. Repeated cell infusions seemed to provide a longer period of disease stability, especially in patients who achieved tumor reduction after the first cell-infusion. 21 out of 23 patients had not developed detectable de novo lesions during this term. Analysis of biopsied tissues by immunohistochemistry showed CD133+ cells were eliminated after CART-133 infusions. This trial showed the feasibility, controllable toxicities, and effective activity of CART-133 transfer for treating patients with CD133-postive and late-stage metastasis malignancies.