Escalating Regulation of 5T4-Specific IFN-γ+ CD4+ T Cells Distinguishes Colorectal Cancer Patients from Healthy Controls and Provides a Target for In Vivo Therapy

Abstract

The relationship between the adaptive CD4 þ T-cell response and human cancer is unclear. The oncofetal antigen 5T4 is expressed in many human carcinomas, including colorectal cancer cells, but has limited expression on normal tissues. We previously identified anti-5T4 CD4 þ T cells in a proportion of patients with colorectal cancer, and we extended this study to examine whether the quality or quantity of the T-cell response reflects tumor stage. An overlapping peptide library spanning 5T4 was used as a target to enumerate cognate IFN-g cells [measured as spot-forming cells (SFC)/10 5 cultured T cells] in peripheral blood-derived lympho-cytes following a 14-day in vitro culture period comparing patients preoperatively (n ¼ 27) to healthy controls (n ¼ 17). Robust 5T4-specific T-cell responses were present in 100% of healthy donors. There was a steady loss of T-cell responses with advancing tumors with a significant negative correlation from stage I to III (P ¼ 0.008). The predictability of the decline meant <200 SFC/10 5 were only found in subjects with stage III colorectal cancer.

T‐cell modulation by cyclophosphamide for tumour therapy

“…Again, these results contrast with another study using 300 mg/m 2 CY in melanoma patients, which reported small, transient depletions of Treg cells . The differential effect could be due to the use of CY in patients with earlier stage cancers and hence lower tumour burdens; baseline frequencies of Treg cells in this patient group, although greater than healthy controls, was statistically lower than in patients with late‐stage melanoma, a finding also observed in colorectal cancer . A recent randomized controlled phase I/II study performed by our group administered low‐dose CY with or without a modified vaccinia Ankara‐based vaccine encoding the oncofetal antigen 5T4 (TroVax) to stage IV colorectal cancer patients, a notoriously poorly responsive tumour to immunotherapy .…”

“…determined that using CY in this manner actually augments anti‐tumour immunity in such patients through selective effects on CD4 + CD25 + Treg cells, allowing for greater tumour control . A number of studies have since used similar metronomic dosing regimens to limit Treg cell function to unmask tumour‐specific T‐cell responses . The dose of CY appears crucial to its immune‐potentiating effects as demonstrated in a clinical trial involving 28 patients with metastatic breast cancer: HER2‐specific antibody responses were enhanced following vaccination with a granulocyte–macrophage colony‐stimulating factor‐secreting breast tumour vaccine and CY administered at a dose < 200 mg/m 2 but higher dosages of CY suppressed these responses .…”

“…59 A number of studies have since used similar metronomic dosing regimens to limit Treg cell function to unmask tumour-specific T-cell responses. 60,61 The dose of CY appears crucial to its immune-potentiating effects as demonstrated in a clinical trial involving 28 patients with metastatic breast cancer: HER2-specific antibody responses were enhanced following vaccination with a granulocyte-macrophage colony-stimulating factor-secreting breast tumour vaccine and CY administered at a dose < 200 mg/m 2 but higher dosages of CY suppressed these responses. 62 With oral administration, 100 mg given twice-a-day depleted all lymphocyte sub-populations, whereas 50 mg twice-a-day was selective for Treg cells.…”

“…70 The differential effect could be due to the use of CY in patients with earlier stage cancers and hence lower tumour burdens; baseline frequencies of Treg cells in this patient group, although greater than healthy controls, was statistically lower than in patients with late-stage melanoma, 70 a finding also observed in colorectal cancer. 61 A recent randomized controlled phase I/II study performed by our group administered low-dose CY with or without a modified vaccinia Ankara-based vaccine encoding the oncofetal antigen 5T4 (TroVax) to stage IV colorectal cancer patients, 71,72 a notoriously poorly responsive tumour to immunotherapy. 73 Compared with no treatment control patients, Treg cell depletion was observed in 24/27 (89%) patients receiving 50 mg CY twice a day, although unlike previous reports, significant B-cell and natural killer cell depletion was also detected.…”

Immunotherapy of Metastatic Colorectal Cancer: Prevailing Challenges and New Perspectives

“…Lymphodepleting chemotherapy prior to immunotherapy reduces the number of circulating T regs , promotes successful engraftment of adoptively transferred T-cells, and improves the overall survival of melanoma patients [101]. Cyclophosphamide decreased peripheral FoxP3 + T regs in all six metastatic CRC patients and increased IFN-γ producing T-cells after 22 days [102]. Local ablation of immune cells using fludarabine, cyclophosphamide, and/or temozomide may dramatically increase the durability of ACT by reducing immunosuppressive pressures and widening a niche for tumor-specific T-cells.…”