Tumor-Infiltrating Lymphocyte Function Predicts Response to Neoadjuvant Chemoradiotherapy in Locally Advanced Rectal Cancer

Abstract

Purpose The presence of tumor-infiltrating lymphocytes (TILs) in tumors is superior to conventional pathologic staging in predicting patient outcome. However, their presence does not define TIL functionality. Here we developed an assay that tests TIL cytotoxicity in patients with locally advanced rectal cancer before definitive treatment, identifying those who will obtain a pathologic complete response (pCR). We also used the assay to demonstrate the rescue of TIL function after checkpoint inhibition blockade (CIB). Patients and Methods Thirty-four consecutive patients were identified initially, with successful completion of the assay before surgery in those 17 patients who underwent full treatment. An in vitro cytotoxic assay of rectal cancer tumoroids cocultured with patient-matched TILs was established and validated. Newly diagnosed patients were recruited with pretreatment biopsy specimens processed within 1 month. Evaluation of TIL-mediated tumoroid lysis was performed by measuring the mean fluorescence intensity of cell death marker, propidium iodide. CIB (anti–programmed cell death protein 1 [anti–PD-1] antibody) response was also assessed in a subset of patient specimens. Results Six of the 17 patients achieved an objective pCR on final evaluation of the resected specimen after neoadjuvant chemoradiotherapy. Cytotoxic killing identified the pCR group with a higher mean fluorescence intensity (27,982 [95% CI, 25,340 to 30,625]) compared with the non-pCR cohort (12,428 [95% CI, 9,434 to 15,423]; p < .001). Assessment of the effectiveness of CIB revealed partial restoration of cytotoxicity in TILs with increased PD-1 expression with anti–PD-1 antibody exposure. Conclusion Evaluating TIL function can be undertaken within weeks of the diagnostic biopsy, affording the potential to alter patient management decisions and refine selection for a watch-and-wait protocol. This cytotoxic assay also has the potential to serve as a platform to assist in the additional development of CIB.

Organoid Models of Tumor Immunology

“…As one solution, immune checkpoint treatment has been performed on epithelial-only submerged Matrigel organoids reconstituted with exogenous immune components [ 72 , 73 ]. Notably, co-culture of patient derived-tumor-only organoids with autologous TILs after expansion of organoids and TILs separately, enabled TIL migration toward PDOs through Matrigel, and tumor cell cytotoxicity, suggesting the use of this co-culture system to assess cytotoxic TIL function [ 72 ].…”

“…As one solution, immune checkpoint treatment has been performed on epithelial-only submerged Matrigel organoids reconstituted with exogenous immune components [ 72 , 73 ]. Notably, co-culture of patient derived-tumor-only organoids with autologous TILs after expansion of organoids and TILs separately, enabled TIL migration toward PDOs through Matrigel, and tumor cell cytotoxicity, suggesting the use of this co-culture system to assess cytotoxic TIL function [ 72 ]. The potential application for immunotherapy screening was suggested by co-culture of patient-derived tumor spheroids with autologous TILs treated with immunomodulatory antibodies targeting MICA/B and NKG2A antigens in colorectal cancer (CRC) [ 73 ].…”

Cancer research using organoid technology

“…Importantly, co-cultures of expanded T cells with organoid generated from adjacent healthy epithelial tissue resulted in undisturbed organoid expansion without significant levels of organoid cytotoxicity [107]. In another approach, tumour-infiltrating T cells and CRC organoids were separately expanded using their respective gold standard methods and then combined to assess T cell-mediated organoid killing [111]. Interestingly, the extent of cell death in the in vitro co-culture assay correlated well with the patient's response to chemotherapy and immune checkpoint blockade [111].…”

“…In another approach, tumour-infiltrating T cells and CRC organoids were separately expanded using their respective gold standard methods and then combined to assess T cell-mediated organoid killing [111]. Interestingly, the extent of cell death in the in vitro co-culture assay correlated well with the patient's response to chemotherapy and immune checkpoint blockade [111]. Furthermore, the authors tested immune checkpoint blockade using PD1 antibodies in their coculture model system and demonstrated that organoid killing by PD1 high T cells was improved upon antibody treatment.…”

Organoids: the new kid in cancer research

“…Organoids can be used to help predict and prognosticate patient responses to therapy. Early studies with rectal cancer have shown that organoids can help predict rectal cancer responses to chemoradiotherapy . This could potentially change how we treat rectal cancer in the future.…”