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PURPOSE Patients with colon or rectal cancer were entered onto a prospectively randomized, controlled clinical trial of active specific immunotherapy (ASI) with an autologous tumor cell-bacillus Calmette-Guérin (BCG) vaccine. We investigated whether ASI could improve disease-free status and survival. PATIENTS AND METHODS Ninety-eight patients with Dukes' stage B2-C3 colon or rectal cancer were randomized into groups treated by resection alone or resection plus ASI. Eighty patients met all eligibility criteria. All patients with rectal cancer were to receive 50 Gy of pelvic irradiation. Analysis of distribution of survival and disease-free survival was made on all eligible patients until December 31, 1990. RESULTS As a single study, no statistically significant differences were detected in survival or disease-free survival for all 80 eligible patients. However, since it was recognized at the outset that there were treatment differences, in that rectal cancer patients were to receive postimmunotherapy radiation, it was considered that a cohort analysis of the colon and rectal cancer patients might be informative. With a median follow-up of 93 months, there is a significant improvement in survival (two-sided P = .02; hazards ratio, 3.97) and disease-free survival (two-sided P = .039; hazards ratio, 2.67) in all eligible colon cancer patients who received ASI. With a median follow-up of 58 months, no benefits were seen in patients with rectal cancer who received ASI. CONCLUSION This study suggests that ASI may be beneficial to patients with colon cancer.

Conventional "staging " of traumatic brain injury (TBI) typically uses depth of coma assessed by the Glasgow Coma Scale (GCS) score near admission as a severity marker. Frequently, early GCS scores are contaminated by alcohol or drug ingestion and other, nonneurological organ system trauma. As well, this measure's usefulness is limited due to its restricted range in survivors. Here we explore the utility of using length of posttraumatic amnesia, coma duration, and age as indirect markers of severity. Cluster analytic techniques were used to examine the relationship of severity so defined to long-term outcome in 106 mild, moderate, and severe TBI patients. Results indicate complex relationships between cluster groups with the influence of age of patient being highlighted as an important moderator in determining severity of injury and later psychosocial outcome.

Research has begun to identify early markers that predict survival after traumatic brain injury. In this study, trauma and biochemical indicators of severity were used to predict quality of life in 61 adults with traumatic brain injury and no damage to other organ systems. Severity markers available within 24 hours of injury were predictive of later psychosocial, behavioural and social role functioning. Multiple regression analyses demonstrated that the Glasgow Coma Scale, plasma glucose levels, leukocyte cell count and serum potassium concentration accounted for 12% to 66% of variance in certain measures of later quality of life. The importance of health-care resource allocation and psychosocial and behavioural intervention to the outcome after moderate traumatic brain injury is discussed.

Intradermal vaccinations with Bacillus Calmette-Guerin (BCG) and viable but nontumorigenic syngeneic hepatocarcinoma cell vaccines were used successfully to treat micrometastatic disease experimentally induced in inbred guinea pigs at 100 times the minimal lethal dose. Several complications have been associated with the use of viable BCG organisms in the treatment of cancer patients and, in this animal model, intradermal administration of BCG uniformly results in severe ulceration and eschar formation at the injection sites leading to secondary microbial invasion and regional lymphadenopathy. We now report the use of isoniazid (Nydrazid) as part of an active specific immunization regimen. Incorporation of isoniazid into the immunization procedure for two weeks alleviates or reduces the side effects of BCG infection. Moreover, with proper consideration of BCG dosage, isoniazid does not impair the efficacy of the BCG plus tumor cell vaccines.

A vaccine of Bacillus Calmette-Guerin (BCG) admixed with tumor cells induced systemic immunity and had a therapeutic effect on subclinical, disseminated micrometastasis. Inbred strain-2 guinea-pigs given IV injections of 5×103 to 106 syngeneic L10 hepatocarcinoma cells were vaccinated after metastatic foci were established in the lung parenchyma. The purpose of this study was to establish the variables that can be manipulated to assure optimal immunotherapy while minimizing deleterious side effects of the BCG. In the present study we examined the variables of source, dose, and ratio of BCG to tumor cells. Four BCG sources (lyophilized Tice and Connaught; fresh-frozen Phipps and Tice) were compared. No significant differences among these BCG preparations could be detected with respect to adjuvant potential when they were admixed with attenuated tumor cells in a vaccine. The dose study clearly demonstrated that a BCG dose dependency exists with relation to induction of effective cell-mediated immunity or survival from disseminated micrometastatic disease. Furthermore, evaluations of dose versus ratio of BCG to tumor cells also supported a BCG dose dependency, with the lowest effective BCG dose being directly influenced by tumor burden of the host. Cutaneous reactivity and hypersensitivity of the primary and secondary immunization sites of tumor-bearing animals treated with effective and ineffective vaccines supported the direct association of reaction to BCG and specific tumor immunity. However, when an in vitro leukocyte migration inhibition assay was used, the degree of reactivity to BCG could not be exploited as a quantitative, diagnostic monitor of effective systemic tumor immunity.

Active-specific immunotherapy, which involves activation of host defenses toward antigenic factors distinct for each tumor, is a potentially useful antitumor strategy. Experimental studies have demonstrated the efficacy of autologous tumor cell vaccines supplemented with an adjuvant (Bacillus Calmette-Guerin, BCG) against intravenously injected tumor cells. In this study, we evaluated the vaccine's efficacy against a more clinically relevant model, spontaneously occurring metastases. Viable syngeneic hepatocarcinoma cells were injected intradermally into the left shoulder area of inbred guinea pigs. On Day 25, the primary tumor (1.5 cm in diameter) and the first draining node were excised. Animals were randomized into groups that received no further therapy or two, three, or four vaccinations with an autologous preparation of cryopreserved, irradiated tumor cells mixed with live BCG. Vaccinations were started 4 days after surgery and were given weekly on alternate sides of the shoulder and flank regions. Of those animals that received surgery alone, 5% survived, whereas of those that received two, three, or four vaccines, 35% ( P = 0.03), 45% ( P = 0.004), and 55% ( P = 0.002), respectively, were disease free at 200 days. That the effect was systemic and not due to BCG draining into the regionally involved lymph nodes was demonstrated in a repeat experiment in which all vaccinations were given in distant (contralateral) sites with equally effective results. Tumor cells in the lymph nodes beyond those excised were found in animals necropsied on the first day of therapy. Therefore, established distant lymph node metastases were controlled. Human tumors with tumor-specific antigens may respond similarly and should be studied.

Over the last four years a guinea pig model of active-specific immunotherapy (ASI) with a syngeneic tumor celhbacillus calmette-Guerin (BCG) vaccine was translated into a prospectively randomized, controlled clinical trial in patients with colorectal cancer. Primary tumors from patients undergoing standard surgical resection were dissociated enzymatically and cryopreserved by techniques that maintain cell viability. Patients with transmural extension of tumor or nodal metastases were randomized into groups treated by resection alone (control) or resection plus ASI. With a mean follow-up of 28 months (range, 14–24), only 3 of 20 treatment patients had recurrences and none have died, whereas 9 of 20 control patients had recurrences and 4 died. These differences are statistically significant and are sufficiently encouraging to warrant expansion of these studies into other research centers. Cancer 55:1236-1243, 1985.

In inbred guinea pigs, administration of Mycobacterium bovis strain BCG by scarification at a site distant from an excised skin tumor, but in the regional lymph node drainage, was evaluated for its immunotherapeutic effect on the development of lymph node metastases. Scarification was performed after surgical excision of intradermally transplanted syngeneic (line-10) hepatocarcinoma at a time when microscopic foci of tumor cells were present in regional lymph nodes. Various strains of BCG were evaluated for their immunotherapeutic potential: fresh-frozen Phipps, Pasteur, and Tice; and lyophilized Pasteur, Tice, and Connaught. Scarification commenced 3 days after surgical removal of the tumor and continued once a week for 5 weeks. Only lymph nodes from fresh-frozen Phipps- and Pasteur-scarified animals were significantly smaller than those in the control groups. Differences in lymph node weight correlated histologically with less detectable metastases. This cytostatic effect was short lived; eventually, the metastatic tumor growth was not significantly different from that of control animals. No significant differences were observed in mean survival time: All animals died as a result of metastases 3 months after tumor inoculation. These results demonstrated that limited scarification with BCG of certain strains temporarily inhibits the growth and proliferation of metastases in regional lymph nodes after removal of the primary tumor.

It has been previously demonstrated that transplanted syngeneic tumors established in the skin of inbred (strain-2) guinea pigs regressed and regional lymph node metastases were eliminated after intralesional injection of viable Mycobacterium bovis (BCG). During the course of this reaction there is the development of tumor-specific immunity. This experimental model was further manipulated in order that it would more closely approximate a clinical reality. In the present study an evaluation was made of the effectiveness of the developing tumor-specific immunity in this BCG therapy model, to abrogate artifically induced distant tumor deposits and to assess the requirement for tumor-specific immunity in the local BCG-mediated tumor regression. During BCG-mediated regression of established intradermal tumor, the developing tumor-specific immunity inhibited the growth of artificially induced vascular metastases in animals receiving a 10(4) or 10(5) tumor cell dose. However, there is a direct causal relationship between the distant tumor burden and the escape of skin tumor and regional lymph node metastases from BCG-mediated regression. Thus, multiple tumor deposits as low as 10(4) cells are capable of competing for or preempting tumor-specific immune reactivity, which must be a requirement during some phase of the intralesional BCG-mediated therapy of established tumor and regional lymph node metastases. Thus, a significant therapeutic effect could be achieved in guinea pigs with established skin tumors and limited vascular metastases when the modality of therapy included BCG intralesional injection, followed 6 weeks later by surgery of the treated skin tumor and regional lymph node.

A vaccine of Bacillus Calmette-Guérin (BCG) admixed with tumor cells induced systemic immunity and had a therapeutic effect on subclinical, disseminated micrometastases. Inbred strain 2 guinea pigs given intravenous injections of either 10(4), 10(5) or 10(6) syngeneic L10 hepatocarcinoma cells were vaccinated after metastatic foci were established in the lung parenchyma. The studies demonstrate that under defined conditions of vaccine preparation and regimen, nontumorigenic preparations of BCG and tumor cells can cure the majority of animals of otherwise lethal visceral metastases. Histopathologically it was determined that immunization with these vaccines prevented the progressive growth of pulmonary micrometastatic foci approximately 0.1 mm in diameter. However, in this micrometastasis therapy model, the number of metastatic tumor foci is a major limitation in the efficacy of vaccine therapy. No protection against L10 tumor was achieved when antigenically distinct but syngeneic L1 hepatocarcinoma was used in the vaccine, suggesting that this is a tumor-specific immunotherapeutic procedure. This BCG-L10 tumor vaccine was also effective in curing guinea pigs of minimal disseminated tumor burden when administered after surgery of an established skin tumor and draining lymph node.

The recognition that host immunocompetence can limit the development of malignant clones has arisen both from teliologic arguments and from data derived from investigations in experimental systems. Several familiar correlations have been cited as evidence for the role of immunosurveillance in cancer, such as the variability in tumor incidence that accompanies age-associated changes in immunocompetence, the increased tumor incidence associated with chronic or induced immune deficiencies, and the general suppression of immunity to alloantigens during treatment with certain chemical carcinogens.

In a transplantable hepatocarcinoma (L10) model of inbred Sewall Wright strain 2 guinea pigs, established micrometastatic tumor foci could be eliminated from the viscera by specific tumor immunity induced by the systemic effect of a BCG-L10 tumor cell vaccine. The conditions of vaccine preparation and regimen were rigid, and a dose dependency for both BCG and tumor cells existed. Comparison of the influence of cryobiologic preservation procedures on the tumor cells to be used in the vaccine showed that cells frozen by an optimized cryobiologic procedure, improved for maintenance of viable cells, were more effective than cells frozen by a conventional glycerol method. The glycerol method resulted in a lower percentage of viable cells, and the effectiveness of the vaccine was greatly diminished. Thus some past failures of active specific immunotherapy could be associated with acute antigenic exposures resulting from suboptimal cryobiologic preservation of tumor cells in the vaccine preparation instead of prolonged or chronic tumor cell antigenic exposure resulting from optimal cryobiologic procedures.

The principles and procedures of active specific immunotherapy developed from studies with the inbred guinea pig hepatocarcinoma model were used as the basis of a randomized, controlled prospective trial of active specific immunotherapy of colorectal cancer patients. The goal was to determine whether colorectal cancer patients treated with vaccines made of autologous tumor cells plus Bacillus Calmette-Guérin as adjuvant would have an increased reaction to their autologous tumor cells as measured by delayed-type cutaneous hypersensitivity (DCH) responses. Our results demonstrate that the active specific immunotherapy significantly increased the DCH responses to autologous tumor cells in 16 of 24 patients (67%). The DCH response of immunized patients to autologous normal mucosa, used as a normal tissue control, did not increase significantly. Furthermore, no significant DCH responses against autologous tumor or mucosa cells were detected in a group of nonimmunized control patients. The induced DCH responses were not correlated with other factors, such as the presence of bacteria in the cell preparation or the protein concentration of the cell preparations. The qualitative and quantitative differences in DCH responses to tumor cells and to normal mucosa cells suggest that the immunizations are targeted mainly to tumor-associated antigens with tissue-associated antigens playing a secondary role.

We evaluated the use of Bacillus Calmette-Guérin admixed with tumor cells as a vaccine to induce systemic tumor immunity for therapy of subclinical (micrometastatic) disease. In several experiments inbred strain 2 guinea pigs were given i.v. injections of either 10(4), 10(5), or 10(6) syngeneic L10 hepatocarcinoma cells, and initial vaccinations were administered either 1 or 4 days after tumor inocluation. Variables in vaccine preparation, such as ratio of viable Bacillus Calmette-Guérin organisms to tumor cells, procedures for freezing the tumor cells, X-ray treatment of tumor cells, and vaccination regimen were evaluated. The studies demonstrated that under defined conditions nontumorigenic vaccines of Bacillus Calmette-Guérin and tumor cells can cure the majority of animals of otherwise lethal visceral micrometastases.

Intradermal vaccinations with Bacillus Calmette-Guérin (BCG) and viable but nontumorigenic syngeneic hepatocarcinoma cell vaccines were used successfully to treat micrometastatic disease experimentally induced in inbred guinea pigs at 100 times the minimal lethal dose. Several complications have been associated with the use of viable BCG organisms in the treatment of cancer patients and, in this animal model, intradermal administration of BCG uniformly results in severe ulceration and eschar formation at the injection sites leading to secondary microbial invasion and regional lymphadenopathy. We now report the use of isoniazid (Nydrazid) as part of an active specific immunization regimen. Incorporation of isoniazid into the immunization procedure for two weeks alleviates or reduces the side effects of BCG infection. Moreover, with proper consideration of BCG dosage, isoniazid does not impair the efficacy of the BCG plus tumor cell vaccines.

We studied the pharmacokinetic properties of two human monoclonal antibodies to colon carcinoma cells and their ability to detect tumors in nude mice bearing primary human colon carcinoma xenografts. The 16-88 and 28A32 monoclonal antibodies are immunoglobulin M class human antibodies produced by cell lines derived from peripheral blood lymphocytes from patients with colon carcinoma. The patients received an autologous tumor cell vaccine as part of an active specific immunotherapy protocol. The 125I-labeled antibodies were cleared from the circulation of non-tumor-bearing and tumor-bearing nude mice with a 6-8-h half-life. The half-life of the antibodies in tumor tissue was 48 to 72 h compared to 8 to 12 h for normal tissues. Tumor:normal tissue ratios were highest 4 to 7 days postinjection with tumor:blood ratios of 12:1 for 16-88 and 10:1 for 28A32 antibody. Experiments with a control human immunoglobulin M myeloma protein confirmed the specificity of the human monoclonal antibodies. Radioimmunoscintigraphic studies using nude mice bearing contralateral antibody-reactive and nonreactive colon tumor xenografts further confirmed that the antibodies specifically localized in tumor tissues. The antibody-reactive tumors were clearly visible by radioimmunoscintigraphy within 4 days of injection. These experiments, undertaken as a preliminary step to clinical trials, demonstrated for the first time that i.v. administered human immunoglobulin M monoclonal antibodies could be taken up by human colon tumor tissue and retained to a sufficient extent to easily permit tumor detection by external radioimmunoscintigraphy. These studies also demonstrated that the nude mouse human colon tumor xenograft model is a useful in vivo system for comparison studies of human monoclonal antibodies as part of a selection process for clinical trials and for evaluating immunoconjugates containing these antibodies for relative pharmacokinetic properties and potential diagnostic or therapeutic efficacy.

In order to test the hypothesis that infanticidal tendencies are suppressed when rats become mothers, very young newborn pups, either naturally born or cesarean-delivered, were presented to virgin females and to newly delivered mothers. Provided that the pups were lively, uncleaned of fetal fluids or membranes, and presented without placentas, nearly all virgins killed and nearly all mothers did not. Newborns were also presented to Day 22 pregnant rats and to rats whose pregnancies had recently been surgically terminated. Large proportions of both groups either were nonkillers or were actively maternally responsive (and a smaller proportion were both) despite the fact that none of these rats had undergone parturition or cared for pups. These results indicate that, independent of its effect on maternal caretaking, pregnancy suppresses infanticide in previously infanticidal nulliparae even before they become mothers. Hence, infants are protected from their own parents. In addition, evidence was obtained in support of the idea that freshly delivered pups have stimulus properties that make them specially suited for promoting the onset of maternal caretaking.

Eighteen postoperative patients with non-small cell lung cancer were actively immunized with a vaccine that included autologous cryopreserved irradiated tumor cells admixed with bacillus Calmette-Guerin. Patients received three weekly intradermal immunizations beginning 1-3 months after surgery (15 patients) or after completion of postoperative radiotherapy (3 patients). There was marked heterogeneity in the relative proportion of tumor cells versus host infiltrating cells within individual vaccines (range of percent tumor cells 7-75%). Five patients exhibited positive delayed cutaneous skin test reactivity (DCR) to autologous irradiated tumor cells prior to immunization, whereas 8 of 13 converted from skin test negative to positive. There were no correlations between DCR reactivity and in vitro lymphoproliferative responses to autologous tumor cells or to clinical outcomes, i.e., freedom from relapse. Possible explanations for the heterogeneity of the lung cancer vaccine and approaches for improving its immunogenicity are discussed.

In inbred guinea pigs, administration of Mycobacterium bovis strain BCG by scarification at a site distant from an excised skin tumor, but in the regional lymph node drainage, was evaluated for its immunotherapeutic effect on the development of lymph node metastases. Scarification was performed after surgical excision of intradermally transplanted syngeneic (line-10) hepatocarcinoma at a time when microscopic foci of tumor cells were present in regional lymph nodes. Various strains of BCG were evaluated for their immunotherapeutic potential: fresh-frozen Phipps, Pasteur, and Tice; and lyophilized Pasteur, Tice, and Connaught. Scarification commenced 3 days after surgical removal of the tumor and continued once a week for 5 weeks. Only lymph nodes from fresh-frozen Phipps- and Pasteur-scarified animals were significantly smaller than those in the control groups. Differences in lymph node weight correlated histologically with less detectable metastases. This cytostatic effect was short lived; eventually, the metastatic tumor growth was not significantly different from that of control animals. No significant differences were observed in mean survival time: All animals died as a result of metastases 3 months after tumor inoculation. These results demonstrated that limited scarification with BCG of certain strains temporarily inhibits the growth and proliferation of metastases in regional lymph nodes after removal of the primary tumor.

Over the last four years a guinea pig model of active-specific immunotherapy (ASI) with a syngeneic tumor cell:bacillus calmette-Guerin (BCG) vaccine was translated into a prospectively randomized, controlled clinical trial in patients with colorectal cancer. Primary tumors from patients undergoing standard surgical resection were dissociated enzymatically and cryopreserved by techniques that maintain cell viability. Patients with transmural extension of tumor or nodal metastases were randomized into groups treated by resection alone (control) or resection plus ASI. With a mean follow-up of 28 months (range, 14-24), only 3 of 20 treatment patients had recurrences and none have died, whereas 9 of 20 control patients had recurrences and 4 died. These differences are statistically significant and are sufficiently encouraging to warrant expansion of these studies into other research centers.