Immunotherapy of breast cancer,malignant melanoma,and acute leukemia with BCG: Prolongation of disease free interval and survival

Summary Results of immunotherapy with BCG in patients with malignant melanoma, breast cancer, and acute leukemia are described. The first study demonstrated that high doses of living BCG organisms (6×10⁸ viable units) delivered by scarification in the upper arms and legs prolonged the disease-free interval and survival of 52 malignant melanoma patients with regional lymph node metastases compared to 218 comparable surgical control patients. Patients with trunk and extremity, but not head and neck melanoma, benefited from BCG, suggesting the importance of the delivery of BCG into the tumor-involved lymphatics.The second study evaluated the therapeutic efficacy of living BCG organisms by scarification in a group of adult acute leukemia patients after the cessation of chemotherapy. Thirty-seven patients had been in remission on intermittent chemotherapy for 12–24 months. Following late intensive consolidation chemotherapy, 7 consecutive patients received no further therapy and then 30 consecutive patients received BCG. Patients maintained on BCG have had a prolonged disease-free interval compared to those given on no further therapy (P=0.07) or compared to a group of similar patients maintained on chemotherapy alone (P=0.001). Similarly, the survival has been improved for patients maintained on BCG compared to those left unmaintained (P=0.009), or those maintained on chemotherapy (P=0.001).The principles of intermittent chemotherapy combined with BCG immunotherapy, first developed in patients with disseminated melanoma and acute myelogenous leukemia, have been confirmed in a series of patients with disseminated breast cancer. Forty-five patients treated with a combination of 5-FU, adriamycin, and cyclophosphamide (FAC) plus BCG by scarification showed prolongation of remission as well as survival compared to a comparable group of 44 patients treated with FAC chemotherapy without immunotherapy. Thus, 23/44 patients treated with FAC have died (median=14 months) compared to only 5/45 patients on FAC-BCG (median=12⁺ months), P=0.005. The limitations of BCG immunotherapy as well as speculations for future developments of immunotherapy are discussed.This work was supported by Contract No1-CB 33888 from the National Institutes of Health, Public Health Service, Bethesda, Maryland 20014. Drs. Gutterman and Mavligit are the recipients of Career Development Awards (Ca 71007-02 and CA 00130-01, respectively) from the National Institutes of Health, Education, and Welfare, Bethesda, Maryland 20014.     

A controlled trial of BCG immunotherapy in bronchogenic carcinoma treated by surgical resection

Summary Ninety-two patients with bronchogenic carcinoma who were treated by surgical resection of the tumour were subsequently given immunotherapy with BCG (Glaxo). The study was strictly randomised into three groups. Twenty-nine patients received multipuncture BCG (50–250×10⁶ viable units) and 26 patients intradermal BCG (0.4–0.9×10⁶ viable units) treatment being given at 1, 2, 5, 9, 13 and 26 weeks after operation and every 26 weeks thereafter. Thirty-seven control patients did not receive BCG. The patients have been observed for 15–33 months. There was no significant difference in survival between the control group and the two immunotherapy groups or between the two immunotherapy groups. The tumour cell type and presence of mediastinal nodes significantly influenced overall survival but not the response to BCG immunotherapy. The possible reasons for the failure of BCG to prolong survival in this study are discussed.     

Various uses of BCG and allogeneic acute leukemia cells to treat patients with acute myelogenous leukemia

Summary Ninety-six remission patients with acute myelogenous leukemia have been treated with various forms of immunotherapy and chemotherapy in three distinct studies and the clinical outcome of these patients has been reported. In the first study 22 patients were maintained on chemotherapy alone and 28 patients were given the same chemotherapy and additional immunotherapy consisting of BCG and irradiated allogeneic AML cells given at separate sites weekly. It was found that there was a significant increase in survival time of the patients who received immunotherapy (median 510 days) compared with the chemotherapy alone patients (270 days). The p value for this was 0.03. The reason for this prolongation of survival was mainly due to a markedly increased survival time of immunotherapy patients after they relapsed when compared with the chemotherapy patients (165 days compared with 75 days median, p equal to 0.0005). In the second sequential study 24 patients were given immunotherapy alone consisting of irradiated allogeneic AML cells and BCG given at separate sites, and this was compared with unirradiated allogeneic cells and BCG given to 22 patients. There was no difference in the remission length or survival between these two groups. In the third study 13 patients received irradiated cells and BCG as in Study 1 and a further 11 patients received the same immunotherapy but also received a mixture of cells and BCG given during the first three months. There was no difference in the remission and survival of these two groups. The significance of these results is discussed.     

Adjuvant BCG immunotherapy for stage I and II malignant melanoma.

Initial adjuvant immunotherapy trials have demonstrated a greater disease-free interval in patients treated with bacille Calmette-Guérin (BCG) compared with historical controls. In this study 149 patients at high risk of recurrence after surgical treatment of local or regional malignant melanoma were given BCG for 2 years and were followed up for a median of 28 months from the start of immunotherapy. The 36 patients in the comparison group had a higher rate of recurrence than the patients treated with BCG, and the rate in the treatment group was close to that reported from a similar study at the University of California at Los Angeles. The relatively long disease-free interval for the high-risk comparison patients in this study suggests that the control groups at other centres may have included patients with unrecognized additional risk. The rates of survival in the Canadian treatment group were also comparable to those reported by other centres. However, reports of a favourable BCG-mediated pattern of recurrence could not be confirmed. Therefore, the routine use of adjuvant BCG immunotherapy is not recommended.     

Improvement of monocyte chemotaxis with BCG immunotherapy in acute lymphoblastic leukemia: Comparison of chemotherapy versus chemotherapy plus immunotherapy as therapeutic regimens

Summary Thirty-six patients with acute lymphoblastic leukemia (ALL) were divided into a group receiving chemotherapy and a group receiving identical chemotherapy plus BCG immunotherapy. Monocyte cellular chemotaxis in both groups was initially significantly depressed compared with that in a group of age-matched controls (P<0.001). Chemotaxis in both patient groups improved with therapy. The mean improvement in chemotaxis per day of the group treated with chemotherapy plus BCG was over 3 times that of the group receiving chemotherapy alone (P<0.01). This change was not due solely to quantitative increases in monocyte numbers. The T-cell subset of ALL patients showed the same response in chemotaxis to chemotherapy plus immunotherapy as the null-cell ALL patients. In vitro monocyte chemotaxis in ALL patients was significantly increased by the addition of BCG immunotherapy to a standard chemotherapy regimen.     

Adjuvant BCG immunotherapy for malignant melanoma.

A total of 199 patients with stage I malignant melanoma at Clark''s level 3 to 5 of invasion were entered into a prospectively controlled randomized clinical trial that attempted to assess the value of local and systemic immunotherapy with BCG (bacille Calmette-Guérin) after surgery. The patients were randomly assigned, with stratification by Clark''s level, to receive either routine follow-up or immunotherapy with BCG, administered intradermally with a Heaf gun around the site of wide excision and then given orally for 2 years. Intradermal administration of BCG was repeated after 1 year''s oral therapy with BCG. Of the 99 patients in the treatment group 66 had Clark''s level 3, 28 had level 4, and 5 had level 5 invasion. Of the 100 patients in the control group, 61 had level 3, 36 had level 4, and 3 had level 5 invasion. Other prognostic factors, such as sex, depth of invasion, histologic features, site of disease and type of surgery, were evenly distributed. There were 57 recurrences of the melanoma, 24 in the treatment group and 33 in the control group. However, this trend was not statistically significant (p = 0.194). The suggestion that BCG may reduce the likelihood of local/regional recurrence has not been confirmed with longer follow-up. There were 13 such recurrences in the BCG group, compared with 21 in the control group; the proportions of patients in each group who had such a recurrence were not significantly different. Of the 199 patients 41 died, 24 in the control group and 17 in the treatment group; again, this difference was not significant. While there may be minor activity in selected patients, there appeared to be no benefit from this form of adjuvant BCG therapy in patients with malignant melanoma.     

Post-operative intrapleural BCG in lung cancer: A 5-year follow-up report

Summary Of a total study group of 99 patients 56 received intrapleural BCG in three different dosages (5 patients 16×10⁶, 25 patients 32×10⁶, and 26 patients 64×10⁶ culturable particles) following resection treatment for lung cancer. This study group was compared with an historical control population, very closely matched with regard to age, sex, stage of disease, histology and type of operation.Although the clinical condition of the patients selected for BCG treatment was above average, no survival benefit ensued from the intrapleural BCG. On the contrary, disease-free survival in BCG receivers with stage I and II squamous cell carcinoma was shortened significantly at 2 years follow-up due to the earlier appearance of local recurrences, with the same tendency still present after 5 years. This observation suggests an enhancement of tumour growth by the intrapleural BCG treatment. This alarming phenomenon could be a warning to clinicians when planning clinical immunotherapy trials to be aware of the potentially deleterious effects of such treatment.     

1975 current results of the first 100 cytologically typed acute lymphoid leukemia submitted to BCG active immunotherapy

Summary The first 100 acute lymphoid (and undifferentiated) leukemias, (of which the smears at the first presentation of the disease are still available for typing), treated successively with remission induction chemotherapy, complementary cell-reducing chemoradiotherapy and then active immunotherapy with irradiated pooled allogeneic leukemic cells and fresh Pasteur Institute BCG applied on scarifications, have been reviewed, especially in connection with BCG application.Tolerance of BCG has been good. Its application had to be stopped due to a side effect (choroiditis) in only one patient. This toxic cost is negligible compared to that of so-called maintenance chemotherapy.No subject of our first control trial started in 1963 has relapsed between 3 and 13 years.In the overall group of the 100 patients studied, no relapse has been observed after 48 months, which is quite different to the observations of frequent relapses after that time in patients submitted to maintenance chemotherapy.Moreover, second remissions are obtained in 94% of the patients who relapsed early under immunotherapy, and their life expectancy after a second remission is as high as it is after the first remission.The median of survival is longer than 5 years.The action of active immunotherapy on the immune machinery has been followed by several assays, of which the increase of null cells (which include K-cells) may be the most interesting.Several prognostic factors have been demonstrated among which are sex, the volume of the neoplasia, meningeal localizations, and the cytological types. Age has no prognostic value in immunotherapy patients, contrary to maintenance chemotherapy patients. Also the cytological types behave differently under immunotherapy and under maintenance chemotherapy. The disease-free survival of more than 85% of the microlymphoblastic patients submitted to immunotherapy is not observed in J. Bernard's patients submitted to maintenance chemotherapy, which suggests that this high cure rate is due to active immunotherapy. Hence, these prognostic factors are probably factors of sensitivity to active immunotherapy. A statistical computerized study has shown that there is a link between the cytological types and other prognostic factors and that they all depend on the cytological type.Hence, our present protocol is adapted to this immunotherapy sensitivity factor. It comprises a nonrisk preimmunotherapy chemotherapy for the microlymphoblastic type, and a longer and more intensive chemotherapy for less immunotherapy sensitive types.     

Adjuvant specific immunotherapy of resectable squamous cell lung carcinoma analysis at the eighth year

From June 1976 to June 1981, 86 patients with resectable (Stage I and II) squamous cell lung carcinoma were entered into a randomized controlled study with three arms: Control Group - no treatment postoperatively. Specific Immunotherapy Group - three monthly doses of 500 micrograms of tumor associated antigen (TAA) emulsified with complete Freund's adjuvant (CFA). Nonspecific Immunotherapy Group - three monthly doses of CFA emulsified in saline. All the patients in the study received skin tests with PPD (5TU) and 100 micrograms of the same TAA used for the immunotherapy at 1, 4, 6, 9, and 12 months postoperatively. Patients in both immunotherapy groups showed a tendency for a better disease-free interval and overall survival compared to those of the control, but these interval and beneficial therapeutic effects were statistically significant only in the Group III patients who had no hilar lymph node metastasis (T1N0 and T2N0). Although Group III was originally designated as a nonspecific immunotherapy group, retrospectively, it should be called a lowdose specific immunotherapy group because these patients actually received a total of 500 micrograms of TAA (as skin tests) and three doses of CFA at separate sites.     

Trial of BCG immunotherapy in the treatment of resectable squamous cell carcinoma of the bronchus (stages I and II)

Summary Thirty-nine patients with squamous cell carcinoma of the bronchus were randomized into two groups after radical surgery. The first group of 18 patients received 75 mg of living BCG on a scratched area once a week; the second group of 21 patients received no further treatment. After 2 years 3 patients of the group treated with BCG are dead and 1 is in relapse; 8 patients of the control group are dead and 2 are in relapse.In this study the hope of survival for more than 2 years for patients with squamous cell carcinoma of the bronchus is 66% in the group of patients treated with BCG, and 38% in the control group. From these results we can see the beneficial effect of BCG as a complementary treatment of surgery for patients with squamous cell carcinoma of the bronchus.     

Cancer immunotherapy.

Important contributions that stimulated studies in cancer immunotherapy included: (1) the discovery of tumour-associated antigens; (2) the observation that infection with bacille Calmette-Guérin (BCG) in animals was protective against tumour challenge; and (3) the observation that immunodepression due either to malignant disease or to treatment of the disease, was, in some instances, related to prognosis. Immunotherapy trials with microbial agents have involved attempts to obtain a local effect by injecting the agent into the tumour or into the region of the tumour and to obtain a "systemic" effect distant from the site of injection. Trials with active specific immunotherapy involving tumour cells or tumour cell extracts have frequently involved the combination of these specific agents with a nonspecific adjuvant such as BCG. Recent studies with thymosin and levamisole in patients with lung cancer and other types of malignant disease have shown prolonged survival in the groups receiving immunotherapy.     

BCG immunotherapy in stage I melanoma patients

Summary Previously, we have provided evidence for a positive correlation between HLA-DR expression in primary melanoma and early metastasis [3, 4]. In the present study we investigated whether this relationship was modified by adjuvant BCG immunotherapy. The study comprised 107 patients with a stage I high-risk melanoma; 44 patients had been treated with BCG, whereas the remaining patients had not received any adjuvant therapy. There was no difference in disease-free survival between BCG-treated and untreated patients. Disease-free survival was significantly shorter in patients with high expression of HLA-DR antigens in the primary tumor.Subgrouping BCG-treated and control patients according to HLA-DR phenotype of the melanoma revealed a prolongation of disease-free survival in the subgroup of BCG-treated patients with no or low expression of HLA-DR antigens in the primary melanoma. BCG therapy apparently did not influence prognosis of patients with high expression of HLA-DR antigens in the tumor.     

Intratumoral BCG and Corynebacterium parvum therapy of canine mammary tumours before radical mastectomy

Summary In two parallel studies, bitches with mammary tumour received single intralesional injections of BCG (1 mg: 10⁷ living bacteria) and Corybacterium parvum (10⁹ killed bacteria) (53 bitches) or C. parvum alone (129 bitches) at the same dosage. Control groups received injections, following the same protocol, of 1 ml BCG suspension medium diluted in saline in the first study (51 bitches) or no injections at all (120 bitches in the second study).A block dissection, including mammary tumours, adjacent mammary glands, and regional lymph nodes, was performed 2 weeks later in all animals. On the basis of histologically confirmed malignant tumours, 48 bitches (25 treated by immunotherapy and 23 controls) in the first study and 67 bitches (30 treated by immunotherapy and 37 controls) in the second study remained for postsurgical follow-up.The clinical tolerance of the treatment was generally good. No significant differences were found in cumulative survival rates between treated and control group in either studies.     

Destruction of tumor cells by BCG-activated alqolar macrophages.

Alveolar macrophages obtained from Syrian golden hamsters were tested for their ability to destroy tumor cells. Only macrophages obtained from BCG immune animals rechallenged intratracheally with BCG five days before assay exhibited cytotoxic activity. Maximum destruction of tumor cells occurred after 5 days of incubation. Immunologic activation of macrophages was required to attain cytotoxic alveolar macrophages. Induction of inflammatory lung exudates by a variety of nonspecific irritants did not result in tumor cell destruction by macrophages. These observations may prove useful in designing an approach for immunotherapy of lung cancer.     

Immunotherapy for remission maintenance in acute myeloblastic leukemia

Summary Forty-eight patients with acute myeloblastic leukemia in remission were treated with immunotherapy in addition to remission-maintenance chemotherapy. The first 16 patients were treated with weekly BCG and a leukemia cell vaccine (group 1). The next 32 patients were randomly allocated to receive BCG and a leukemia cell vaccine given once monthly (group 2) or BCG given monthly with no leukemia cell vaccine (group 3). There was no significant difference in remission duration or survival between the randomly allocated groups (2 and 3).Comparisons with group 1 are limited by the non-random allocation to this group, but selection bias was unlikely and clinical features were similar in the three patient groups. No significant difference in remission duration or survival was seen amongst the three groups studied. There was no advantage in the addition of leukemia cell vaccine (groups 1 and 2) to BCG alone (group 3) and no advantage to weekly (group 1) versus monthly immunotherapy (groups 2 and 3). Only 7 of the 48 patients achieved a second remission, and 4 of these were short-term partial remissions.The following are contributing members of the Toronto Leukemia Study Group: Doctor's Hospital, Harvey Silver MD; Humber Memorial Hospital, Alan Seidenfeld MD; Mississauga Hospital, Michael King MD; Mount Sinai Hospital, Dominic Amato MD; Northwestern Hospital, Wilhelm Kwant MD; Oshawa General Hospital, Hak Chiu MD; St Michael's Hospital, Bernadette Garvey MD, Kenneth Butler MD; St Joseph's Hospital, H. James Watt MD, Murray Davidson MD; Toronto General Hospital, Gerald Scott MD, William Francombe MD, Kenneth Shumak MD; John Crookston MD, PhD; Toronto Western Hospital, James G. Watt MD, David Sutton MD; Michael Baker MD; Domenic Pantalony MD; Wellesley Hospital, Dale Dotten MD; Women's College Hospital, George Kutas MD; York Finch Hospital, Sam Berger MD     

Presence of interleukin-2 in urine of superficial bladder cancer patients after intravesical treatment with bacillus Calmette-Guérin

Summary Urine samples were obtained from patients with superficial bladder cancer after immunotherapy with bacillus Calmette-Guérin (BCG). The patients were repeatedly (once a week for 6 consecutive weeks) treated with intravesical administration of approximately 5 × 10⁸ culturable particles of BCG. Some patients received more than six BCG instillations. The urine samples were investigated for the presence of interleukin-2 (IL-2) in an in vitro bioassay using a murine cytotoxic T cell line (CTTL-16) that shows IL-2-dependent growth. Preliminary experiments indicated the presence of inhibitory factors in the urine. This inhibitory activity was abolished after 24 h dialysis. In a neutralization assay with both polyvalent and monoclonal anti-(human IL-2) antibody it was demonstrated that there was indeed IL-2 in the urine samples. In 8 of 11 patients the presence of IL-2 in the urine was demonstrated. The IL-2 production was directly related to the BCG administration as samples obtained just before the BCG instillation were always negative. In IL-2-positive samples a maximum level of IL-2 was observed between 2 h and 6 h after the BCG instillation. In urine samples obtained 24 h after the BCG IL-2 was not detected. In most patients the urine became positive after the third or fourth BCG instillation     

A new side effect of BCG immunotherapy — BCG-induced arthritis in man

Summary Ten of 159 patients showed arthritic symptoms during the course of BCG immunotherapy for advanced cancer. The arthritic symptoms occurring after BCG injections had the following characteristics: (1) The incidence of arthritis was closely correlated with the host immunologic responsiveness to BCG; (2) These symptoms usually occurred 1–5 months after the first BCG injection (7/10); (3) The arthritic symptoms usually started with morning stiffness (9/10), which was followed by acute inflammatory signs in the affected joints. They gradually subsided in response to treatment with nonsteroid antiinflammatory drugs, but were not completely cured while the effectiveness of BCG continued; (4) The symptoms were aggravated by additional BCG injections (8/10). (5) This form of arthritis could be differentiated from rheumatoid arthritis, tuberculous, or purulent arthritis by its clinical course and by roentgenograms of the affected joints. It is thought to be induced by the adjuvant effect of BCG, and is a new side effect of BCG immunotherapy.     

Present status of BCG immunotherapy of malignant melanoma

Summary BCG systemic adjuvant immunotherapy may be effective for improving both the recurrence and survival rates in patients with regional metastases from malignant melanoma. Clinical trials show that many of the principles derived from the study of animal tumor systems are applicable to human cancer in that immunotherapy is most effective for a small residual number of tumor cells. BCG treatment fulfills many of the ideal criteria for adjuvant treatment following surgery when disease burden is lowest. It is relatively nontoxic; it is effective for disseminated melanoma; it has systemic activity in the adjuvant treatment of subclinical metastases. However, until clinical trials are complete, BCG adjuvant therapy must be considered investigational.Supported by USPHS grants CA05252, CA12582, and NIH 0732001 CB43852.     

Nonspecific and selective stimulation of the immune system in the treatment of carcinoma in humans.

The experience of the Toronto General Hospital in the use of nonspecific stimulation of the immune system with bacille Calmette-Guérin (BCG) for the treatment of cancer of the gastrointestinal tract, malignant melanoma and breast cancer is described. The results are presented in terms of survival curves. The use of BCG administered intraperitoneally in a randomized study of patients with gastric, pancreatic and colorectal cancer proved of no benefit. On the other hand, when BCG was given orally in a randomized study of patients with resectable cancer of the colon and in nonrandomized consecutive studies of patients with malignant melanoma and stage IV carcinoma of the breast survival was increased. In a group of patients with advanced gastrointestinal cancer selective stimulation of the immune system with NED 137 produced a significant increase in survival when compared with the survival of historical controls (the patients given BCG intraperitoneally along with 5-fluorouracil for gastrointestinal cancer). The results of these studies suggest the need for a more rational approach in manipulating the immune response that would combine chemotherapy with selective stimulation of the immune system.     

Efficacy of oral and parenteral routes of Mycobacterium bovis bacille Calmette-Guerin vaccination against experimental bovine tuberculosis in white-tailed deer (Odocoileus virginianus): a feasibility study

We investigated the efficacy of oral and parenteral Mycobacterium bovis bacille Calmette-Guerin Danish strain 1331 (BCG) in its ability to protect white-tailed deer (Odocoileus virginianus) against disease caused by M. bovis infection. Twenty-two white-tailed deer were divided into four groups. One group (n=5) received 10(9) colony-forming units (cfu) BCG via a lipid-formulated oral bait; one group (n=5) received 10(9) cfu BCG in culture directly to the oropharynx, one group (n=6) was vaccinated with 10(6) cfu BCG subcutaneously, and one group served as a control and received culture media directly to the oropharynx (n=6). All animals were challenged 3 mo after vaccination. Five months postchallenge the animals were examined for lesions. Results indicate that both oral forms of BCG and parenterally administered BCG offered significant protection against M. bovis challenge as compared to controls. This study suggests that oral BCG vaccination may be a feasible means of controlling bovine tuberculosis in wild white-tailed deer populations.