Humoral anti-idiotypic and anti-anti-idiotypic immune response in cancer patients treated with monoclonal antibody 17-1A.

A group of 96 patients with advanced colorectal carcinoma were treated with the mouse (m) or chimeric (c) (mouse variable regions x human IgG1 constant regions) monoclonal antibody (mAb) 17-1A recognizing the tumour-associated antigen GA733-2. Eighty-two of the 83 patients treated with mmAb 17-1A and 69% of the patients given cmAb17-1A (n = 13) developed anti-idiotypic antibodies (ab2). Auto-antibodies binding to tumour cells expressing GA733-2 were found in 7% of the patients. In a further 38 patients (40%) antitumour-cell antibodies, i.e. anti-anti-idiotypic antibodies (ab3), were induced by the mAb17-1A therapy. Patients with detectable ab3 after treatment had significantly higher ab2 levels than those not developing ab3. Addition of granulocyte/macrophage-colony-stimulating factor (GM-CSF) to mmAb17-1A significantly enhanced the induction of ab2 as well as induction of anti-anti-idiotypic antibodies (ab3) compared to mmAb17-1A alone. Patients with a high increase in antitumour-cell antibodies (ab3) induced by the therapy lived significantly longer than patients with no or a low level of induction of ab3 (P = 0.016). The results indicate that induction of an idiotypic network response might be an important effector mechanism in mAb therapy.

Tumor regression in monoclonal antibody-treated patients correlates with the presence of anti-idiotype-reactive T lymphocytes.

Treatment of cancer patients with unconjugated mAbs directed against tumor-associated antigens is considered passive immunotherapy due to the main suggested effector mechanisms: antibody-dependent cellular cytotoxicity, complement-dependent cytolysis, and apoptosis. The therapeutic antibody (ab1) may, however, also give rise to an idiotypic network response, i.e., an immunizing effect. Induced anti-idiotypic antibodies (ab2) mimicking the epitope that ab1 recognizes might subsequently induce an anti-anti-idiotypic humoral (ab3) and T-cell (T3) response recognizing the nominal tumor-associated antigen. Twenty-four patients with metastatic colorectal carcinoma were treated with MAb17-1A against the tumor associated antigen GA733-2 and were analyzed for the induction of T3 cells. Five of the patients responded to mAb therapy with tumor regression. These five patients all had T cells specifically recognizing human ab2 (DNA synthesis) after treatment, while all nonresponding patients lacked such T cells. Four of the five patients with ab2-reactive T cells also showed induction of T cells recognizing GA733-2. The association between T3 cells and tumor regression was highly significant (P = 0.0005). Thus, induction of T3 cells might be an important secondary antitumor effector function of therapy with unconjugated mAbs. Antibody therapy may therefore also be considered active specific immunotherapy.

Subcutaneous interleukin-2 and alpha-interferon in advanced colorectal carcinoma. A phase II study.

Subcutaneous administration of low doses of recombinant interleukin-2 (IL-2) and interferon-alpha (IFN-alpha) on an out-patient basis has been reported not to significantly compromise the response frequency compared to intravenous IL-2 in patients with renal cell carcinoma and melanoma. As part of an ongoing program to develop a biotherapeutic concept in patients with colorectal carcinoma (CRC) we studied the clinical effects of such a regimen in 15 patients with metastatic CRC. The daily dose of IL-2 varied between 4.8-14.4 x 10(6) U/m2 and of IFN-alpha between 3-6 x 10(6) U/m2. The cycle length was 6 weeks. The course was repeated every 8 weeks until disease progression. Maximum 4 cycles were administered. Maintenance therapy was given to responding patients once a week every month. No patient showed a major response (CR or PR). Six patients had a stable disease ranging from 3 months to 18 months with a median duration time of 5 months. The median survival of all patients was 13 months. The main adverse reactions were fever, chills, anorexia and shortness of breath. No treatment related deaths occurred. 6/14 patients developed abnormal concentration of serum levels of thyroid hormones. It is concluded that the present treatment schedule using IL-2 and IFN-alpha in advanced CRC seemed not to be of clinical benefit.

Effect of monoclonal antibody 17-1A and GM-CSF in patients with advanced colorectal carcinoma--long-lasting, complete remissions can be induced.

Antibody-dependent cellular cytotoxicity (ADCC) is considered to be one of the effector functions of unconjugated monoclonal antibodies (MAbs) in tumor therapy. The antitumor activity of MAbs might therefore be augmented if the cytotoxic capability of the effector cells could be increased. In an in vitro system, the killing capacity of MAb was significantly enhanced by pre-treatment of the effector cells with granulocyte-macrophage colony-stimulating factor (GM-CSF). Based on these findings, the therapeutic effect of the combination of mouse MAb 17-1A (IgG2a) and GM-CSF was evaluated in 20 patients with metastatic colorectal carcinoma (CRC). The patients received GM-CSF for 10 days and a single i.v. infusion of MAb 17-1A on day 3 of the cycle. Four cycles were given at 1-monthly intervals. There was a continuous increase in blood monocytes and lymphocytes during all 4 GM-CSF cycles. Neutrophils and eosinophils were also significantly augmented but in a biphasic manner and the cell counts on day 10 of cycle IV were significantly lower than in cycles I and II. GM-CSF-related side-effects were of no major clinical importance. During the third cycle, an immediate-type allergic reaction (ITAR) against MAb 17-1A occurred in most patients, necessitating reduction of the MAb dose as well as of the infusion rate. Two patients achieved complete remission. One patient had a minor response, and 3 other patients were considered to have stable disease > 3 months.

Low dose cyclophosphamide, alpha-interferon and continuous infusions of interleukin-2 in advanced renal cell carcinoma.

Pretreatment with a low dose of cyclophosphamide (CY) has been claimed to inhibit suppressor functions and augment various immune functions. A combination of a low dose of CY, alpha-interferon (IFN-alpha) and continuous infusion of interleukin-2 (IL-2) was used to treat patients with advanced renal cell cancer (RCC) (stage IV). Sixteen patients received four cycles consisting of CY (500 mg m-2) three days prior to daily i.m. injections of alpha-IFN (3 x 10(6) U), and continuous infusion of 18 x 10(6) IU rIL-2 for five days. The cycle interval was three weeks. Two patients had partial response (13%) (26+ and 12+ months), two had a minor response (9+ and 4 months), and three patients achieved stable disease (19+, 14+ and 8+ months). No patients required intensive care. Side effects were mainly fever, malaise, capillary leak syndrome and diarrhoea. Non-responders showed significantly higher eosinophil and platelet counts compared to responders. Serum concentration of IL-2 was significantly higher in responders. 5/11 patients had abnormally low values of serum thyroxine after therapy. Two patients needed thyroid hormone substitution. The difference between the initial and the lowest thyroxine values correlated significantly to survival (p < 0.03). The addition of CY to rIL-2 and IFN-alpha in the present protocol did not contribute to an increased major response rate.

Cytotoxic functions of blood mononuclear cells in patients with colorectal carcinoma treated with mAb 17-1A and granulocyte/macrophage-colony-stimulating factor.

Unconjugated monoclonal antibodies (mAb) may induce tumour regression in patients. The mechanisms of action are complex. Antibody-dependent cellular cytotoxicity (ADCC) is considered one of the effector functions. Augmentation of the killing capacity of cytotoxic cells may thus be a way to increase the therapeutic potential of mAb. Granulocyte/macrophage-colony-stimulating factor (GM-CSF) has been shown to enhance this function in vitro. Eighteen patients with metastatic colorectal carcinoma received GM-CSF (250 micrograms m-2 day-1 s.c.) for 10 days and a single infusion of the anti-(colon carcinoma) mAb 17-1A (mouse IgG2A) (400 mg) on day 3 of the cycle. The cycles were repeated once a month four times. Neutrophils, eosinophils, monocytes and lymphocytes increased significantly in a biphasic way. However, at the fourth cycle the rise in white blood cells was significantly lower compared to the preceding courses. ADCC (SW948, a human CRC cell line,+mAb 17-1A) or peripheral blood mononuclear cells (PBMC) was significantly (P less than 0.05) augmented by day 6 of a cycle and then declined gradually and, at the end of a cycle, the ADCC activity had returned to the pretreatment level. The spontaneous cytotoxicity of PBMC against the natural-killer-resistant cell line, SW948, varied in a similar way. During GM-CSF treatment there was also a significant increase in FcRI+ (CD64), FcRII+ (CD32), FcRIII+ (CD16) and CD14+ cells but not of CD56+ cells.

Therapy of colorectal carcinoma with monoclonal antibodies (MAb17-1A) alone and in combination with granulocyte monocyte-colony stimulating factor (GM-CSF).

A mouse monoclonal antibody (MAb17-1A) (IgG2A) against colorectal carcinoma cells was used to treat patients with metastatic disease. Major direct effector functions of MAb seem to be ADCC (antibody dependent cellular cytotoxicity), CDC (complement dependent cytolysis) and apoptosis ('programmed cell death'). Thus, a high tumor cell saturation of the MAb should be achieved. Increasing doses of MAb to the patients increased the total area under the concentration curve and thus the exposure of tumor cells to MAb. However, the response rate (with complete + partial + minor response + stable disease defined as response) was not augmented. In total, 10/52 (19%) patients responded and in fact lower doses (less than 2 g) might induce a higher response frequency (9/52) than higher doses (greater than 2 g) (1/52). During treatment, the numbers of cytotoxic cells (lymphocytes and monocytes) increases in the tumor lesion and complement components were deposited. As ADCC may be important, effector mechanism attempts were made to augment the cytolytic capability of the effector cells by simultaneously giving the patients GM-CSF. The combination of MAb17-1A + GM-CSF augmented the ADCC activity of blood mononuclear cells and a heavy infiltration of monocytes could be noted in the tumor. Out of 15 available patients 6 (40%) showed a response.

Chemotherapy and immunotherapy of colorectal cancer.

More than 50% of the patients with large bowel cancer develop disseminated disease and invariably succumb. Adjuvant chemotherapy with 5-FU and levamisole have been shown to be more efficient than 5-FU alone or in combination with cytostatics. The combination of 5-FU, leukovorin and methotrexate induces prolonged survival with a good quality of life in metastatic colorectal cancer (CRC). During the last decade tumor immunotherapy has been an alternative facilitated by isolation and large scale production of cytokines and monoclonal antibodies. The mouse monoclonal antibody (MAb) 17-1A recognizes a tumor-associated antigen (TAA), present in high concentrations on the surface of gastrointestinal tumor cells. Injections of MAb 17-1A in patients with metastatic CRC induced generation of anti-idiotypic (ab2) in 90% and anti-anti-idiotypic (ab3) antibodies in 47% of the treated patients. The development of ab3 correlated significantly with survival (mean 80 weeks) while ab3- patients survive only 38 weeks. One of 52 patients treated with MAb 17-1A is a complete remission after 66 months, 3 had minor regression and 6 had a stable disease (19% RR). Based on in vitro findings showing increased antibody-dependent cellular cytotoxicity (ADCC) by the combination of granulocyte-macrophage colony stimulating factor (GM-CSF) and MAb 17-1A, 16 CRC patients have been treated with subcutaneously injections of GM-CSF for 10 days and intravenous infusions of MAb 17-1A at day 3. Two of 16 are in CR, 1 in MR and 3 in SD (37.5% RR). Minor side-effects were registered. A further development of immunotherapy of CRC might imply vaccination by injection of specific human anti-idiotypic antibodies (ab2) which mimics the nominal antigen, in order to induce a specific immunity.