PcP is an infectious disease with significant morbidity and mortality attributable to the host inflammatory response as demonstrated in both patients (11
) and animal models (4
). T cells have been shown to be necessary to initiate the inflammatory immune response to Pc which in turn produces so called bystander injury to the lung. In most clinical scenarios patients lack CD4+
T cells, thereby leaving a predominantly CD8+
T cell-driven response which is ineffective in killing P. carinii
but which produces critical lung injury (4
). In other clinical situations, such as IRIS, CD4+
T cells also participate in the inflammatory response. In contrast to CD8+
T cells, the CD4+
T cell-driven immune response contributes to the eradication of Pc, although these responses may result in even greater lung damage (4
We have shown, previously, that the inflammatory injury associated with PcP-related IRIS can be significantly prevented by depleting CD4+
T cells prior to immune reconstitution (4
). While this is easily accomplished in mouse models, therapeutic agents to accomplish this in humans are not presently available. A pan T-cell antibody, OKT3 (Muromonab), is available for clinical use as an immunosuppressive agent especially in the control of transplant rejection. OKT3 and similar antibodies work by causing partial depletion of T cells and more importantly by disrupting the interaction of CD3 with the T cell receptor necessary for T cell antigen recognition and activation (25
). Because both CD4+
T cells express CD3 we reasoned that using an antibody directed against CD3 would result in T cell inactivation similar to that observed with specific monoclonal antibodies in mice.
SCID mice with PcP that were immune reconstituted and treated with anti-CD3 F(ab′)2
fragments showed a marked physiological benefit. Not only were these improvements statistically significant, they were also clinically relevant. Treated mice demonstrated reduced pulmonary inflammation, improved dynamic compliance, improved lung resistance and improved survival. Increased neutrophil numbers in pulmonary lavage fluid have been correlated with increased severity of PCP both in patients as well as in mouse models (12
). Using anti-CD3 F(ab′)2
in our model resulted in a significant reduction in neutrophil numbers in lavage fluid that was associated with less severe disease. Interestingly, macrophage numbers were also decreased although the significance of this observation with regards to the injury resulting from the Pc-driven inflammatory response is unknown.
The experimental approach described in this report differs from that described in our earlier publications in one important respect. In those prior experiments CD4+
T cell depletion was started before any signs of inflammation or pulmonary compromise were noted. That is, mice were treated at a point in time when they had PcP but were asymptomatic. While this approach has proved useful to identify mechanisms of immunopathogenesis in PcP, it may not be an ideal approach for modeling clinically-relevant therapeutic interventions. In the present study the disease process was allowed to progress to the point where the mice experienced a 10% acute weight loss and were tachypnic. That the mice had advanced PcP was evidenced by the fact that TMP-SMX did not consistently improve survival. We did not measure arterial oxygen tension in the current study but we have previously shown that mice in this condition would be hypoxic as well (8
). Thus, we feel the experimental approach used for these studies is a reasonable approximation of a patient presenting for treatment with moderate PcP. The fact that anti-CD3 F(ab′)2
administration could be delayed until after symptoms of PcP were obvious and still improve the course of disease, makes these studies highly clinically relevant.
The approach outlined in these experiments of further immunosupressing an immunosuppressed patient could be construed as counterproductive. However, it is important to keep in mind that suppressing the pulmonary inflammatory response would be done in conjunction with the administration of antibiotics. Importantly, TMP-SMX reduced the Pc burden when given with anti-CD3 F(ab′)2
fragments. For these experiments, anti-CD3 F(ab′)2
was given every other day until the termination of the experiments. This schedule was chosen because OKT3 is typically administered daily for 5–10 days when used to treat transplant rejection. However, the biologic effects of OKT3 are demonstrable within minutes of infusion (25
). This rapid onset of action may be why anti-CD3 treatment was effective in already symptomatic mice. Furthermore, as the experiment described in demonstrated, it may be possible to use fewer doses of anti-CD3 to suppress inflammation while TMP-SMX takes effect. Short term use of OKT3 would reduce its side effects and shorten the period of drug induced immunosuppression.
Although we chose to use an OKT3-like antibody our results suggest that other therapies which disrupt T cell function could be effective adjunctive therapy for PcP. For example, monoclonal antibody to the IL-2α receptor (CD25; basiliximab, Novartis Pharmaceuticals, Basel, Switzerland) may also be effective in this setting. In addition to global suppression of T cell function, it may be possible to identify and inhibit specific molecular pathways that lead to lung injury during PcP.
In summary, the pathogenesis of PcP includes an immune mediated inflammatory response which causes compromised lung function. T cells, especially CD8+ T cells, are a critical component of this inflammatory response. We have shown that inactivation of T cells with antibody directed against the CD3 molecule results in a marked physiologic benefit to mice with symptomatic PcP. Over the short term, this improvement is seen even in mice that are not treated with antibiotics to kill P. carinii, supporting the conclusion that lung injury during PcP is an immune mediated phenomenon. The clinical utility of using anti-CD3 as adjunctive therapy in patients with PcP will require clinical trials. Our results lend support to such an approach.