Phase I-II study of the addition of alpha-2a interferon to 5-fluorouracil/leucovorin. Pharmacokinetic interaction of alpha-2a interferon and leucovorin.

5-Fluorouracil (5-FU) activity has been improved by the use of leucovorin (LV) or alpha-2a interferon (alpha-IF). We investigated the feasibility and activity of addition of alpha-IF to a 5-FU/LV regimen. A phase I study with 26 patients (14 previously untreated, 12 previously treated) with disseminated cancer was conducted. 15 patients were treated with 5-FU/LV and 11 with 5-FU/LV/alpha-IF. The 5-FU/LV regimen consisted of escalating doses of 5-FU bolus intravenously on days 2 and 3, combined with repeated oral LV on days 1, 2 and 3. Treatment was every 2 weeks. In the 5-FU/LV/alpha-IF schedule, 18 x 10(6) U alpha-IF subcutaneously daily was added on days 1, 2 and 3. The phase I study was followed by a phase II study of 5-FU/LV/alpha-IF at the established 5-FU dose in 29 previously untreated patients with disseminated colorectal cancer. The optimal 5-FU dose in both parts of the phase I study was 750 mg/m2/day. Mucositis, diarrhea and leucopenia were dose limiting. Although alpha-IF added its own toxicity (fever, flu-like symptoms, fatigue), it did not decrease the optimal dose of 5-FU. In the phase II study 28 patients were evaluable for response: three complete responses and 12 partial responses were observed (response rate 54%, 95% confidence interval, 34 to 72%). Pharmacokinetics of oral LV was performed in patients treated with and without alpha-IF: significantly higher serum levels of LV and 5-methyltetrahydrofolate were found after alpha-IF addition. Influence of alpha-IF on gastrointestinal absorption or renal clearance could be excluded. In conclusion, this 5-FU/LV/alpha-IF combination seems active in metastatic colorectal cancer. The pharmacokinetic interaction between alpha-IF and LV may play a role in the activity of this regimen. Controlled studies are necessary to establish the value of addition of alpha-IF to 5-FU/LV regimens.

The cellular immune response to cell-associated and cell-free cytomegalovirus (CMV) antigens after primary CMV-infection in non-immunocompromised hosts: development and maintenance of CMV-latency and its influence on immunocompetence.

Development and maintenance of cellular immunity to cell-associated CMV-antigens (CMVFF) was investigated in non-immunocompromised hosts during the first year (group I, n = 11) and 1 to 5 years (group II, n = 9) after a symptomatic primary CMV infection, as well as in healthy CMV-seropositive controls without a history of CMV disease (group III, n = 28). During the acute phase (0-2 months) of a primary CMV-infection CMVFF-induced lymphocyte proliferation was severely decreased compared to that in the post-illness phase (5-12 months), and to that in groups II and III. After the reconvalescent period (3-4 months) it gradually increased to levels seen in group III. In group II higher responses to CMVFF were found than in group III or during the post-illness phase of a primary infection. In general CMV-virion induced lymphocyte proliferation showed the same pattern of development as the CMVFF-induced lymphocyte response, but during the acute phase (0-2 months) a lack of correlation was observed between lymphocyte proliferation to CMVFF and CMV-virions. Lymphocyte proliferation to PHA, Con A, allogeneic lymphocytes and recall antigens were severely depressed in the acute phase of a primary CMV-infection and restored gradually to levels seen in groups II and III, with the exception of Con A-reactivity. The latter response remained depressed when compared to healthy seropositive controls, not only during the post-illness period, but also later on (group II).

HLA-DRw6 as a risk factor for active cytomegalovirus but not for herpes simplex virus infection after renal allograft transplantation.

To study genetically determined susceptibility to cytomegalovirus and herpes simplex virus infections in patients given renal transplants a prospective study was performed of 68 consecutive patients receiving their first cadaveric kidney allograft. The recipients positive for HLA-DRw6 showed a significantly increased incidence of active cytomegalovirus infection as early as the 10th week after transplantation (p less than 0.05). No relation with other human leucocyte antigens was found, nor did a correlation exist between HLA typing and the incidence of herpes simplex virus infections. Furthermore, recipients positive for HLA-DRw6 with secondary cytomegalovirus infections excreted infectious virus more often (p less than 0.01) and showed more clinical symptoms (p less than 0.01) than a comparable group of recipients negative for HLA-DRw6. These observations may have practical implications for the treatment of patients who have had renal transplant operations.

Maintenance of cytomegalovirus (CMV) latency and host immune responses of long term renal allograft survivors. II. Secondary CMV infections associated with impaired in vitro proliferative responses to mitogens, allogeneic lymphocytes and CMV infected cells.

The relationship between secondary cytomegalovirus (CMV) infections and host general cellular immunocompetence was investigated in 16 renal allograft recipients with minimal immunosuppressive treatment and excellent renal function. Results were compared with 19 CMV seropositive healthy controls. Significantly impaired immune responses were detected in the subgroup of nine recipients who experience at least 2 years before a secondary CMV infection. Their in vitro lymphocyte reactivity (LR) tests to phytohaemagglutinin (PHA, P = 0.01), pokeweed mitogen (PWM, P less than 0.05), microbial antigens (P less than 0.001) and to pooled allogeneic stimulator lymphocytes in the MLC test (P = 0.02) were lower than the controls. The MLC responses, however, increased with graft survival time (r = 0.8810, P = 0.01). This was positively correlated with the virus specific cellular immunity measurable by the LR responses to CMV infected target cells (r = 0.8333, P = 0.02). In contrast, long term renal allograft survivors who maintained their CMV infection in latency after transplantation (n = 7) showed normal responses to PWM, pooled lymphocytes and CMV infected target cells, whereas the responses to PHA and to bacterial antigens were less severely impaired (P less than 0.05 and P less than 0.001, respectively). This study of long term renal allograft survivors shows that a secondary CMV infection has a long lasting negative effect on immunity especially against alloantigens and CMV infected targets. However, in the data presented here it would be as acceptable to suggest that the patients are consistently relapsing with CMV because they initially had poor immune response and not vice versa.

Maintenance of cytomegalovirus (CMV) latency and host immune responses of long term renal allograft survivors. I. Prolonged suppression of in vitro lymphocyte responses against CMV infected fibroblasts related to previous secondary CMV infection.

Cytomegalovirus (CMV) specific humoral and cellular immunity was investigated in 16 renal allograft recipients with long term graft survival (26-122 months) who were shown to be CMV seropositive before transplantation. Results were compared with healthy individuals with latent CMV infections. Recipients (n = 9) who experienced a symptomatic secondary CMV infection shortly after transplantation (less than 6 months), showed a prolonged but finally temporary suppression of their in vitro lymphocyte memory responses against CMV infected fibroblasts (CMV-FF; median SI: 1.9), a persistence of high antibody titres against intracellular CMV antigens and most of them also had antibodies against CMV membrane antigens (CMV MA). In contrast the recipients (n = 7) who could maintain their CMV in latency after transplantation, had lower antibody titres and their in vitro memory lymphocyte responses against CMV-FF (median SI: 9.3) were comparable to those of the healthy controls (median SI: 11.6). The memory lymphocyte responses against purified CMV virions were depressed in both recipient groups. These results suggest that cellular immunity against CMV infected target cells constitute an important mechanism in maintaining CMV in latency after allografting.

A stimulatory role for cytomegalovirus (CMV) antibodies in CMV-specific lymphocyte proliferation in vitro.

Cytomegalovirus(CMV)-specific lymphocyte proliferation in vitro was tested in the presence or absence of CMV-specific antibodies. CMV antibodies clearly enhanced CMV-specific lymphocyte proliferation but not lymphocyte proliferation induced by other antigens. Enhancement was most consistently found with cell-bound CMV antigens and often with cell-free CMV as stimulating antigen. CMV antibodies only stimulated when they were added at an early stage of the lymphocyte culture, and the stimulating effect was most pronounced at optimal and suboptimal antigen concentrations. Our data suggest that CMV antibodies stimulate CMV-specific lymphocyte proliferation at the level of antigen presentation.

Comparison of in vitro lymphocyte proliferations induced by cytomegalovirus-infected human fibroblasts and cell-free cytomegalovirus.

In vitro lymphocyte reactivity (LR) to cytomegalovirus (CMV)-infected human fetal fibroblasts (CMVFF) and cell-free CMV were measured by using lymphocytes from healthy donors. Lymphocytes from all seropositive donors were stimulated by CMVFF, whereas lymphocytes from negative donors were not. The optimal stimulator cell-to-lymphocyte ratio was in the range of 1:5 to 1:50, dependent on the virus dose used. LR to cell-free CMV was positive for 15 out of 18 seropositive donors and negative for 14 out of 16 seronegative donors. In most cases LR to CMVFF was considerably higher than LR to cell-free CMV. Within the CMV seropositive group there was no significant correlation between the LR to either CMVFF or cell-free CMV and the levels of antibodies to CMV early antigens or CMV late antigens. There was no strict correlation between LR to CMVFF and to cell-free CMV, especially not in tests with lymphocytes from two patients with CMV mononucleosis. Our data suggest that CMVFF and cell-free CMV are recognized (partly) by different subpopulations of CMV-specific memory lymphocytes. We conclude that the use of CMV-infected cells, in addition to cell-free CMV, in LR tests gives more reproducible and possibly also additional information about CMV-specific cellular immunity.