We conducted a phase II clinical trial of anti-CTLA-4 antibody (ipilimumab) and granulocyte-macrophage colony-stimulating factor (GM-CSF) in 22 patients with metastatic melanoma and determined clinical outcomes and immunologic responses. The treatment consisted of a 3-mo induction with ipilimumab at 10 mg/kg administered every 3 weeks for four doses in combination with GM-CSF at 125 µg/m² for 14 d beginning on the day of the ipilimumab infusion and then GM-CSF for 3 mo on the same schedule without ipilimumab. This was followed by maintenance therapy with the combination every 3 mo for up to 2 y or until disease progression or unacceptable toxicity. Blood samples for determination of immune subsets were obtained before treatment, at week 3 (end of cycle 1) and at week 6 (end of cycle 2). Blood samples were also obtained from seven subjects who were cancer-free. The immune response disease control (irDC) rate at 24 weeks was 41% and the overall response rate (ORR) was 32%. The median progression free-survival (PFS) was 3.5 mo and the median overall survival (OS) was 21.1 mo. 41% of the patients experienced Grade 3 to 4 adverse events. We conclude that this combination is safe and the results suggest the combination may be more effective than ipilimumab monotherapy. Further, the results suggest that lower levels of CD4⁺ effector T cells but higher levels of CD8⁺ T cells expressing PD-1 at pre-treatment could be a potential biomarker for disease control in patients who receive immunotherapy with ipilimumab and GM-CSF. Further trials of this combination are warranted.

Cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) blockade can induce tumor regression and improved survival in cancer patients. This treatment can enhance adaptive immune responses without an exogenous vaccine, but the immunologic biomarkers associated with improved clinical outcome in cancer patients are not fully established. A phase Ib trial in patients with metastatic, castration-resistant prostate cancer was performed combining ipilimumab with sargramostim (GM-CSF). In addition to evaluating ipilimumab dose, patients were followed clinically for response and overall survival, and for immunomodulation of circulating T cells. PSA declines of ≥50% and radiographic responses were observed at doses of ≥3 mg/kg/dose. Timing of clinical responses could be either immediate or delayed. Durable responses were also observed off treatment. A subset of patients experienced long-term survival with or without objective clinical responses. The relationship between T-cell phenotype in peripheral blood and overall survival was examined retrospectively. We found that the treatment induced an increase in the levels of CD4(+) effector T (Teff) cells, regulatory T cells, PD-1(+) CD4 Teff cells, and PD-1(+) CD8 T cells. However, these increased levels were not associated with overall survival. Instead, low pretreatment baseline levels of PD-1(+) CD4 Teff cells were found to correlate with longer overall survival. Furthermore, baseline levels of PD-1(+) CD4 Teff cells from patients with shorter overall survival were higher than from cancer-free male control subjects. These results suggest that preexisting expression of immunologic checkpoint marker PD-1 on CD4 Teff cells may help identify patients that may benefit from ipilimumab treatment.

Sipuleucel-T is a US Food and Drug Administration-approved immunotherapy for asymptomatic or minimally symptomatic metastatic castration-resistant prostate cancer (mCRPC). Its mechanism of action is not fully understood. This prospective trial evaluated the direct immune effects of systemically administered sipuleucel-T on prostatic cancer tissue in the preoperative setting. Patients with untreated localized prostate cancer were treated on an open-label Phase II study of sipuleucel-T prior to planned radical prostatectomy (RP). Immune infiltrates in RP specimens (posttreatment) and in paired pretreatment biopsies were evaluated by immunohistochemistry (IHC). Correlations between circulating immune response and IHC were assessed using Spearman rank order. Of the 42 enrolled patients, 37 were evaluable. Adverse events were primarily transient, mild-to-moderate and infusion related. Patients developed T cell proliferation and interferon-γ responses detectable in the blood following treatment. Furthermore, a greater-than-three-fold increase in infiltrating CD3(+), CD4(+) FOXP3(-), and CD8(+) T cells was observed in the RP tissues compared with the pretreatment biopsy (binomial proportions: all P < .001). This level of T cell infiltration was observed at the tumor interface, and was not seen in a control group consisting of 12 concurrent patients who did not receive any neoadjuvant treatment prior to RP. The majority of infiltrating T cells were PD-1(+) and Ki-67(+), consistent with activated T cells. Importantly, the magnitude of the circulating immune response did not directly correlate with T cell infiltration within the prostate based upon Spearman's rank order correlation. This study is the first to demonstrate a local immune effect from the administration of sipuleucel-T. Neoadjuvant sipuleucel-T elicits both a systemic antigen-specific T cell response and the recruitment of activated effector T cells into the prostate tumor microenvironment.

Granulocytic-macrophage colony-stimulating factor (GM-CSF) is used as an adjuvant in cancer vaccine trials and has the potential to enhance antitumor efficacy with immunotherapy; however, its immunologic effects are not fully understood. Here, we report results from a phase I study of neoadjuvant GM-CSF in patients with localized prostate cancer undergoing radical prostatectomy. Patients received subcutaneous injections of GM-CSF (250 μg/m²/day) daily for 2 weeks (cohort 1; n = 6), 3 weeks (cohort 2; n = 6), or 4 weeks (cohort 3; n = 6). Treatment was well tolerated with all grade 1 or 2 adverse events. Two patients had a decline in prostate-specific antigen (PSA) of more than 50%. GM-CSF treatment increased the numbers of circulating mature myeloid dendritic cells, proliferating conventional CD4 T cells, proliferating CD8 T cells, and to a lesser magnitude FoxP3⁺ regulatory CD4 T cells. Although GM-CSF treatment did not augment antigen-presenting cell localization to the prostate, treatment was associated with recruitment of CD8⁺ T cells to the tumor. These results suggest that systemic GM-CSF can modulate T-cell infiltration in the tumor microenvironment. Cancer Immunol Res; 4(11); 948-58. ©2016 AACR.