Presence of activated lymphocytes in the urine of patients with superficial bladder cancer after intravesical immunotherapy with bacillus Calmette-Guérin

Abstract

To study the mode of action of intravesical bacillus Calmette-Guérin (BCG) immunotherapy in the prevention and cure of superficial bladder cancer, flow-cytofluorometric analysis of the cellular immunological reaction in the urine of patients was performed. Fresh urinederived leucocytes were obtained from eight patients before (t ₀) and 24 h (t ₂₄) and 48 h (t ₄₈) after repeated intravesical BCG instillations (at least 5 instillations). For two patients urine-derived leucocytes were investigated at the first BCG instillation. The number of leucocytes in the urine was markedly increased 24 h after repeated BCG instillations, indicating a local cellular immunological reaction induced by BCG. The mean number of cells per milliliter of urine at that time was 2.9×10⁶±3.6×10⁶ (n = 8). These leucocytes consisted mainly of granulocytes (75±11%,n = 8). In addition monocytes/macrophages (4±2%,n = 8) and T lymphocytes were present (1±1%,n = 5). The relative increase of monocytes/macrophages in the urine after BCG application tended to be higher compared to the other leucocyte subtypes. As T lymphocytes may play an important role in the BCG-mediated antitumour activity, subsets of lymphocytes were further characterized att ₀,t ₂₄, andt ₄₈ after repeated BCG instillations. The lymphocyte population consisted mainly of T cells (86% CD3⁺,t ₀). Most of the T cells were CD4⁺ (helper/inducer) and were significantly decreased at 48 h (62±9% att ₀ vs 49±6% att ₄₈). Lymphocytes partly expressed HLA-DR antigens (44%,t ₀). The percentage of lymphocytes with interleukin-2 (IL-2) receptors (CD25⁺) was significantly increased at 24 h and 48 h, compared to pre-instillation values (19±11% and 10±4% vs 3±3% respectively). Natural killer cells (CD 16⁺ and/or CD56⁺) and B cells (CD 19⁺) were less numerous (10% and 19% att ₀ respectively). After the first BCG instillation the increase in the number of leucocytes in urine seemed to be less compared to the numbers after repeated BCG instillations. Lymphocytes could not be detected in the urine collected before or after the first BCG instillation. In conclusion, we demonstrated the presence of considerable numbers of leucocytes in the urine 24 h after repeated BCG instillations, i.e. shortly after immunological activation. The antigen expression of the lymphocytes suggested that they may represent the lymphocytes in the bladder wall. Expression of HLA-DR and IL-2 receptors on lymphocytes indicated activation of T cells by the intravesical BCG treatment. These leucocytes may be useful for functional studies, which are essential to elucidate the actual effector mechanism(s) in the mode of action of BCG against superficial bladder cancer in man.