A Swedish study of chemoradiation in squamous cell carcinoma of the esophagus.

This multicenter study describes the development of a chemoradiation protocol for the treatment of non-metastatic squamous cell carcinoma of the esophagus. Eighty patients were treated with three courses of chemotherapy (cisplatinum and 5-fluorouracil) with concomitant radiotherapy (40 Gy) during the last two courses of chemotherapy. Esophagectomy was performed, when feasible. If no operation was performed, patients were planned to receive a target dose of 64 Gy. Toxicity was mainly attributable to hematological impairment and led to two adjustments of the treatment protocol (addition of filgrastim and lowering of the 5-fluorouracil dose). These changes made it possible to administer the planned treatment in a gradually higher proportion of patients (13/23 [57%] before changes of treatment compared with 30/36 [83%] after changes). Treatment-related mortality was 3.75% (3 patients, associated with leucopenic septicemia after chemotherapy). Fifty-four patients were resected. No per- or postoperative mortality was encountered. The complete response (pathological CR) rate in operated patients was 46% (27/59 patients) after chemoradiation. In the whole series the CR rate (including clinical CR for non-resected patients) was 44%. With a minimum follow-up of 37 months, the 3-year survival for the whole group was 31% compared with 57% for the CR patients. Total 5-year survival thus far (July 1999) is 26%.

Percutaneous endoscopic gastrostomy for nutrition in patients with oesophageal cancer.

OBJECTIVE: To evaluate the technical aspects and risks of using percutaneous endoscopic gastrostomy (PEG) in the treatment of patients with oesophageal cancer. DESIGN: Retrospective study. SETTING: Teaching hospital, Sweden. SUBJECTS: 229 consecutive patients who presented with oesophageal cancer between January 1990 and the end of December 1999. INTERVENTION: Insertion of a PEG after diagnosis and before treatment. MAIN OUTCOME MEASURES: Morbidity and mortality. RESULTS: PEGs were successfully inserted in 222/229 (97%), and the tumour required dilatation in 103 (45%). There was 1 oesophageal perforation and 1 tear of the stomach wall, both of which resulted in death (mortality 0.9%). In 1 operated patient the right gastroepiploic artery was injured by the PEG, but this did not prevent the stomach being used successfully as the oesophageal substitute. PEGs were removed because of leaks in 2 patients. There was 1 possible implantation metastasis. CONCLUSION: PEG is a safe and a well tolerated way of ensuring enteral nutrition in patients with oesophageal cancer. The risk of the PEG complicating any later operation is minimal.

Ga733/EpCAM as a target for passive and active specific immunotherapy in patients with colorectal carcinoma.

GA733/EpCAM is an oncofetal antigen abundantly expressed in colorectal carcinoma. This antigen can spontaneously induce a humoral and cellular antitumor immunity and may therefore be a suitable target structure for immunotherapy. Patients with advanced colorectal carcinoma have been treated with monoclonal antibodies (MAb17-1A) against this structure. The data indicate that the chimeric variant was not superior to the original mouse MAb. Addition of cytokines and chemotherapeutics may improve the therapeutic effect of the MAb. A particularly interesting regimen is a combination of MAb17-1A/GM-CSF/alpha-IFN/5-Fu. The GA733 protein antigen can also be used as a vaccine. Patients with colorectal carcinoma stages B and C were vaccinated with this protein antigen in combination with GM-CSF as an adjuvant cytokine. A strong type I T cell response was induced that seemed to be MHC class I as well as class II restricted. No systemic side effects were noted.

Clinical effects of monoclonal antibody 17-1A combined with granulocyte/macrophage-colony-stimulating factor and interleukin-2 for treatment of patients with advanced colorectal carcinoma.

Granulocyte/macrophage-colony-stimulating factor (GM-CSF) has previously been indicated to enhance the therapeutic effect of the anti-colorectal carcinoma mAb17-1A as well as to augment in vivo immune effector functions. In vitro interleukin-2 (IL-2) augmented GM-CSF-induced antibody-dependent cellular cytotoxicity, a mechanism considered to be of significance for the therapeutic effect of mAb. A treatment regimen was elaborated that combined mAb17-1A (400 mg at day 3 of a 10-day treatment cycle) with the simultaneous administration of GM-CSF (250 microgram/m(2) once daily) and IL-2 (2.4 x 10(6) U/m(2) twice daily) for 10 days. The treatment cycle was repeated once a month. Twenty patients with advanced colorectal carcinoma were included in the study. One patient obtained a partial remission and 2 patients stable disease for 7 and 4 months respectively. The median survival time from the start of mAb therapy was 8 months. Owing to allergic reactions, the planned mAb17-1A dose had to be reduced by repeated infusions. At the fourth treatment cycle only 25% received the planned mAb dose. In 3 patients the GM-CSF and IL-2 dose was reduced because of side-effects. The subjective tolerability of the treatment was considered good or acceptable in more than 80% of the patients. The increment in white blood cell subsets induced by the cytokines decreased by increasing number of courses. This particular regimen did not augment the therapeutic effect of mAb17-1A anticipated from in vitro data but rather hampered the clinical effect of the antibody. The reason for this is not clear but a possibility might be the induction of immune suppression in vivo resulting from an impaired human anti-(mouse Ab) and anti-idiotypic antibody response as well as antibody-dependent cellular cytotoxicity, on the basis of a comparison of mAb17-1A/GM-CSF/IL-2- and mAb17-1A/GM-CSF-treated patients.

Induction of CD4(+) and CD8(+) Bordetella pertussis toxin subunit S1 specific T cells by immunization with synthetic peptides.

In this study two synthetic peptides from the Bordetella pertussis toxin subunit S1 were conjugated to human anti-idiotypic antibodies and used as an immunogen in cancer patients to induce immunity. The aims of the present report are to explain why no carrier or adjuvant effect of the conjugated pertussis peptides could be established regarding induction of responses against the anti-idiotype and to explore the type and quality of induced anti-pertussis immune responses. The lack of carrier and adjuvant effect of the peptides might be related to the fact that the anti-idiotypic antibodies by themselves include helper epitopes and that none of the patients had a detectable T cell response against any of the selected peptides before immunization, which might be a requirement for an adjuvant effect. However, three of four immunized patients mounted a humoral as well as cellular response against the pertussis peptides used. The induced T cell immunity was restricted to one of the two peptides in responding patients. Established T cell lines and MHC blocking studies indicated that the T cell epitopes of the two peptides had a different MHC restriction. The type of T cell response induced seemed to govern the humoral response. The only durable antibody response was accompanied by the presence of a CD4(+) T cell response against the same peptide. Immunization with an anti-idiotype conjugated to synthetic peptides might thus induce both a B and a T cell response against the peptides and the type of induced T cells (CD4 or CD8) governs the quality of the humoral response. Moreover, the possibility of boosting or inducing a response against the antigen from which the peptide sequences were deduced also seemed feasible.

Augmentation of the immune response with granulocyte-macrophage colony-stimulating factor and other hematopoietic growth factors.

Granulocyte-macrophage colony-stimulating factor is by far the most widely used hematopoietic growth factor to augment immune responses. At present, the best secured effect is as an adjuvant cytokine for vaccination. Granulocyte-macrophage colony-stimulating factor can be delivered as gene-transduced tumor cells, as plasmid DNA, or as the soluble free granulocyte-macrophage colony-stimulating factor protein. Granulocyte-macrophage colony-stimulating factor must be present at the same site as the vaccine component. Granulocyte-macrophage colony-stimulating factor may also augment the effect of therapeutic monoclonal antibodies by enhancing various effector functions such as antibody-dependent cellular cytotoxicity and amplifying an idiotypic network response (i.e., antitumor immunity). It may also be advantageous to combine granulocyte colony-stimulating factor with monoclonal antibodies (neutrophil and monocyte antibody-dependent cellular cytotoxicity) for tumor therapy. However, these growth factors might also induce immune suppression, which may hamper the contemplated effect of the growth factor. It is urgently warranted to better understand these dual effects on the immune system so that we can find optimal uses for the growth factors in various clinical settings.

Autoantibodies against the tumour-associated antigen GA733-2 in patients with colorectal carcinoma.

The tumour-associated antigen (TAA) GA733-2 is expressed as a non-secreted surface molecule on the majority of human colorectal carcinoma cells. The antigen has been used as a target for passive and active immunotherapy during the last decade. To determine the incidence of autoantibodies against this antigen, sera from 1068 patients with colorectal carcinoma were analysed for naturally occurring IgG antibodies against the baculovirus-produced GA733-2E protein. A total of 14.5% of the patients had IgG antibodies against the antigen. In 519 patients, sera were collected at the time of diagnosis and 15% of those patients had anti-GA733-2E IgG antibodies. There was a tendency to a higher frequency of patients with antibodies among those in the advanced Dukes stages: 11% in stage A and 32% in stage D respectively (P = 0.06). Antibodies could be detected for up to 10 years after the diagnosis. Patients with Crohn's disease or colitis ulcerosa (n = 20) did not elicit anti-GA733-2E antibodies. No healthy control donor (n = 45) had detectable antibodies against the antigen. The specificity of GA733-2E-reactive serum IgG was indicated by significant inhibition of mAb17-1A (originally used to define GA733-2) binding to the GA733-2E antigen. Sera of positive patients bound to the GA733-2-expressing human colorectal carcinoma cell line, SW948. No significant correlation was found between the presence of antibodies and survival in the present patient population. However, the high incidence of autoantibodies against this tumour antigen in colorectal carcinoma patients confirms its antigenicity in humans and supports the use of the GA733-2 antigen as a target for immunotherapy.

T-cell-epitope mapping of the idiotypic monoclonal IgG heavy and light chains in multiple myeloma.

The idiotypic structures of the myeloma protein might be regarded as tumor-specific antigens. The present study was designed to map T-cell epitopes of the idiotypic myeloma protein to prove the existence of naturally occurring major-histocompatibility-complex-dependent idiotype (peptide)-specific T cells in multiple myeloma. The fine specificity of idiotype-reactive, interferon-gamma-producing blood T cells of a patient with multiple myeloma stage I was characterized by identification of idiotype (heavy and light chains)-derived MHC-restricted T-cell epitopes. T cells specifically reacting with peptides corresponding to each of the 3 complementarity-determining regions (CDRs) of the heavy-chain variable part (V(H)) of the autologous idiotype were found. In contrast, none of the peptides corresponding to the 3 CDRs of the light chain (V(L)) induced a specific T-cell response. The idiotype amino-acid sequence corresponding to the junction of the V(H), diversity (D), and joining (J) gene segments of the VH appeared to be an important target for T cells, since the sequence expressed MHC-class-I- as well as MHC-class-II-restricted epitopes. The study provides further support for the existence of MHC-restricted idiotype-specific T cells, which may target immunogenic CDR peptides in multiple myeloma. Such T cells could be an important part of the specific anti-tumor immune responses induced in idiotype vaccination protocols.

Pharmacological administration of granulocyte/macrophage-colony-stimulating factor is of significant importance for the induction of a strong humoral and cellular response in patients immunized with recombinant carcinoembryonic antigen.

Eighteen colorectal carcinoma patients without macroscopic disease after surgery were immunized using recombinant (r) human (h) carcinoembryonic antigen (CEA) with (n=9) or without (n=9) the addition of soluble granulocyte/macrophage-colony-stimulating factor (GM-CSF). The dose of rhCEA per immunization was 100 microg (n=6), 316 microg (n=6) or 1000 microg (n=6). rhCEA was given s.c. on day 1 and 80 microg/day of GM-CSF s.c. on days 1-4. The schedule was repeated six times during a period of 9 months. All patients in the GM-CSF group developed a strong rhCEA-dose-dependent IgG antibody response while only one-third of the non-GM-CSF patients mounted a weak antibody response. All patients (9/9) in the GM-CSF group developed a strong rhCEA-specific proliferative T cell response as well as type I T cells (interferon gamma secretion). In 45% of the patients also a weak type II T cell response (interleukin-4 secretion) was evoked. Both MHC-class-I- and -II restricted rhCEA-specific T cells were noted. A specific cellular response (proliferation and/or cytokine secretion) against native hCEA could be found in 8/9 patients in the GM-CSF group, although at a significantly lower level than against rhCEA. In the non-GM-CSF group a weak rhCEA-specific T cell response was induced. Three patients had a proliferative response, 4 patients type I T cells and 6 patients type II T cells. No signs of autoimmune reactions were noted. Local pharmacological administration of GM-CSF seemed to be a prerequisite for the induction of a strong immunity against baculovirus-produced hCEA protein. However, the cellular response against native CEA was of a significantly lower magnitude.

Idiotype immunization combined with granulocyte-macrophage colony-stimulating factor in myeloma patients induced type I, major histocompatibility complex-restricted, CD8- and CD4-specific T-cell responses.

Idiotypic structures expressed on the myeloma Ig protein might be regarded as a tumor-specific antigen. Five patients with IgG myeloma were immunized with the purified serum M-component by repeated intradermal injections together with soluble granulocyte-macrophage colony-stimulating factor (GM-CSF). All patients developed an idiotype (Id)-specific T-cell immunity, defined as blood T cells predominantly secreting interferon-gamma (IFN-gamma) and interleukin-2 (IL-2) (type I cells). Id-specific DNA synthesis was induced in one patient. Delayed-type hypersensitivity against the Id was not evoked. The specific IFN-gamma/IL-2 T-cell response was inhibited (46% to 100%) by a major histocompatibility complex (MHC) class I monoclonal antibody (MoAb) in all five patients. A 5% to 37% inhibition by an MHC class II MoAb was seen in four patients. CD4+ as well as CD8+ T cells enriched by magnetic microbeads contained Id-specific cells. The T cells recognized peptides corresponding to the complementarity-determining regions 1, 2, and 3 of the heavy chain of the Id. There was a transient rise of B cells producing IgM anti-idiotypic antibodies in all patients. The results indicate that immunization of myeloma patients using the autologous M-component and soluble GM-CSF may evoke an Id-specific predominantly MHC class I-restricted type I T-cell response.

[Vaccination against cancer soon a therapeutic possibility. B-cell tumors, colonic cancer and melanoma may be suitable for this treatment]. / Vaccination mot cancer snart en behandlingsmöjlighet. B-cellstumörer, koloncancer och melanom kan lämpa sig för denna terapi.

The theoretical basis of cancer vaccination having been well established during the past two decades, the translation of this knowledge into clinically applicable immunisation procedures is now an urgent need. Numerous antigenic preparations are available that are capable of inducing specific anti-tumour immunity which can be augmented by appropriate cytokines. Promising tumour vaccination results have been obtained in B-cell malignancies, colorectal carcinoma, and melanoma; tumour regression has been noted in myeloma, non-Hodgkin lymphoma, colorectal carcinoma, and melanoma patients, and significantly prolonged disease-freed survival in non-Hodgkin lymphoma and colorectal carcinoma patients. The presence of only minimal residual disease would seem to be a clinical prerequisite for tumour vaccination.

Induction of a T- and B-cell response against a unique amino acid sequence of the mouse IgG2A hinge region in a MAb-treated patient.

A patient with malignant glioblastoma was treated with intratumoral infusions of the murine MAb425 (IgG2A) directed against the epidermal growth factor receptor. At the 10th infusion, the patient developed somnolence, fever and headache. The symptoms increased during the subsequent 48 hr but then gradually disappeared within a week. The cerebrospinal fluid (CSF) contained increased concentrations of interleukin-2. The main CSF cell subset was CD4 T-cells. A marked blood lymphocyte proliferative response against mouse IgG2A was noted. The reactive T-cell epitope(s) could be localized to a 14 amino acid (RGPTIKPCPPCKCP) long peptide of the hinge region. A B-cell response (IgG antibodies) against this peptide was also induced.

Granulocyte-macrophage colony-stimulating factor as an adjuvant in tumor immunotherapy.

Induction of specific anti-tumor immunity by active immunization has been the aim of researchers for decades. However, a generally applicable successful immunization strategy that could be used in the clinic has not yet been devised. Recent research has been directed at identifying and defining tumor-specific and tumor-associated antigens. Several good candidates are now at hand. If these antigens are to perform optimally at immunization, there is a need for proper adjuvants. This article focuses on one adjuvant, the cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF), and the possible application of this molecule to active specific immunotherapy.

Pharmacokinetics of cisplatin and its monohydrated complex in humans.

The pharmacokinetics of cisplatin and its cytotoxic hydrolysis product cis-diammineaquachloroplatinum(II) ion (monohydrated complex) were investigated in seven patients after they received a 1-h infusion of cisplatin in normal saline at 100 mg/m2. The concentrations of intact cisplatin and the monohydrated complex were determined in blood by liquid chromatography with post-column derivatization, using diethyldithiocarbamate as the reagent. A pharmacokinetic model was developed assuming that a fraction of the dose (2.3%) is present as the monohydrated complex in the infusion solution and that reversible reactions between cisplatin and its monohydrated complex prevail. The clearances of cisplatin and the monohydrated complex were 0.32 +/- 0.05 and 0.27 +/- 0.11 L/min/m2, respectively. The apparent volume of distribution was considerably smaller for the monohydrated complex (4 +/- 2 L/m2) than for cisplatin (11 +/- 2 L/m2). The elimination rate constants were 0.030 +/- 0.002 and 0.07 +/- 0.02 min-1 for cisplatin and the monohydrated complex, respectively. The area under the time-concentration curve for the monohydrated complex was approximately 15% of that for cisplatin. It is concluded that the significant amounts of the monohydrated complex present in blood are due to the fraction already present in the administered dose and to the fraction formed in blood.

Production of neutralizing granulocyte-macrophage colony-stimulating factor (GM-CSF) antibodies in carcinoma patients following GM-CSF combination therapy.

In this study, the development of neutralizing and non-neutralizing GM-CSF antibodies and the clinical consequences related to the induction of these antibodies were analysed in 20 patients with metastatic colorectal carcinoma receiving a combination therapy of Escherichia coli-derived GM-CSF and a colon carcinoma-reactive MoAb in the absence of any concomitant chemotherapy. The recombinant human GM-CSF was administered subcutaneously for 10 days every month for 4 months. Following the first cycle of treatment, no GM-CSF antibodies were detected, but during subsequent therapy, 19 of the 20 patients studied developed GM-CSF binding antibodies. However, only a proportion (40%) of the 19 antibody-positive patients developed antibodies that neutralized the biological activity of GM-CSF in an in vitro bioassay. The presence of GM-CSF neutralizing antibodies was associated with a significant reduction in GM-CSF-induced expansion of leucocytes, neutrophils and eosinophils. Such clinical effects were not apparent in patients with non-neutralizing antibodies. Further characterization of sera from patients with neutralizing antibodies showed that, in most cases, the antibodies neutralized the biological activity of GM-CSF preparations derived using different expression systems (Chinese hamster ovary cells and yeast), suggesting that these antibodies may have the potential to cross-react with endogenously produced GM-CSF. These effects should be considered before therapeutic use of cytokines, particularly in patients who are not immunosuppressed, and therefore capable of mounting an effective immune response. Our results indicate that assessment of production of neutralizing antibodies induced during cytokine therapy can be used to predict diminished clinical response to further therapy.

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.

Interferon-alpha/beta can impede development of carcinogen-induced squamous-cell tumors in the esophagus of C57B1 mice.

The effect of treatment with murine interferon-alpha/beta preparations on diethylnitrosamine-induced squamous-cell tumors in the esophagus of C57B1 mice was investigated. Diethylnitrosamine was administered in the drinking water for 18 weeks. Interferon was given intraperitoneally during the same 18 weeks or from the end of the period of carcinogen exposure until termination of the experiment. In mice given interferon and diethylnitrosamine synchronously, a significantly lower tumor index (number of tumors/cm of esophageal mucosa) was observed as compared to all control groups. Treatment with interferon after the administration of the carcinogen was terminated had no effect on the appearance of tumors. These data suggest that interferon therapy can exert certain effects on carcinogenesis.

Granulocyte/macrophage-colony-stimulating factor augments the induction of antibodies, especially anti-idiotypic antibodies, to therapeutic monoclonal antibodies.

A group of 86 patients with advanced colorectal carcinoma were treated with the mouse (m) (IgG2A) or chimeric (c) monoclonal antibody (mAb) 17-1A. Prior to therapy, no patient had detectable levels of antibodies to mAb17-1A. All mmAb17-1A-treated patients (n = 76) developed antibodies against both idiotypic and isotypic determinants. Addition of granulocyte/macrophage-colony-stimulating factor (GM-CSF) to mmAb17-1A significantly enhanced the induction of anti-idiotypic (ab2) as well as anti-isotypic antibodies. Of the mmAb17-1A-treated patients, 16 developed type I allergic reactions. These patients had significantly higher concentrations of anti-(mouse Ig) antibodies than patients without type I reactions. Of these 16 patients, 5 had received mmAb17-1A alone; they constituted 9% of this group (5/56). The remaining 11 patients had been given mmAb17-1A together with GM-CSF, and represented 55% of this treatment group (11/20). The difference was statistically significant (P < 0.001). Of 10 patients, 9 (90%) treated with cmAb17-1A and GM-CSF developed ab2. The ab2 concentration in this patient group was significantly lower compared to those treated with mmAb-17A. Anti-(mouse Ig) antibodies caused clinical symptoms requiring therapeutic intervention in fewer than 10% of the patients treated with mmAb17-1A alone. With the addition of GM-CSF, the antibody concentration as well as the frequency of allergic side-effects calling for medical action increased significantly. Significantly more patients with a high ab2 concentration (at least 15 micrograms/ml) 1 month after completion of mAb therapy responded to mAb treatment as compared to those with a low ab2 concentration (P < 0.05). Moreover, patients with a high ab2 concentration (at least 15 micrograms/ml) had a median survival time of 15 months while those with a lower concentration survived for a median time of 9 months (P = 0.01).

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.