Downregulation of NKG2DLs by TGF-ß in human lung cancer cells.

BACKGROUND: Transforming growth factor beta (TGF-ß) is a typical immuno-inhibitory cytokine and highly secreted by lung cancer cells. It was supposed that its immunosuppressive effects to NK cell might be related with the altered expression of activating and inhibitory molecules in lung cancer cells. In this study, we examined the expression of NKG2DLs, PD-L1 and PD-L2 in lung cancer cells after treatment of TGF-ß and a TGF-ß inhibitor, Galunisertib (LY2157299). RESULTS: TGF-ß reduced the level of surface proteins of five NKG2DLs without altered transcription levels in lung cancer cells. Galunisertib reversed the effect of TGF-ß on the expression of NKG2DLs. Since MMP inhibitors, MMPi III and MMP2 inhibitor I, restored the reduced expression of NKG2DLs after treatment of TGF-ß, it was thought that TGF-ß induced the expression of MMP2 which facilitated the shedding of the NKG2DLs in cancer cells. However, the expression of PD-L1, L2 were not changed by treatment with TGF-ß or Galunisertib. CONCLUSIONS: Therefore, inhibition of TGF-ß might reverse the immunosuppressive status on immune cells and restore NK cell mediated anticancer immune responses by upregulation of NKG2DLs in cancer cells.

Using macrophage activation to augment immunotherapy of established tumours.

BACKGROUND: Successful immunotherapy will require alteration of the tumour microenvironment and/or decreased immune suppression. Tumour-associated macrophages (TAMs) are one major factor affecting tumour microenvironment. We hypothesised that altering TAM phenotype would augment the efficacy of immunotherapy. METHODS: We and others have reported that 5,6-Dimethylxanthenone-4-acetic-acid (DMXAA, Vadimezan) has the ability to change TAM phenotypes, inducing a tumour microenvironment conducive to antitumour immune responses. We therefore combined DMXAA with active immunotherapies, and evaluated anti-tumour efficacy, immune cell phenotypes (flow cytometry), and tumour microenvironment (RT-PCR). RESULTS: In several different murine models of immunotherapy for lung cancer, DMXAA-induced macrophage activation significantly augmented the therapeutic effects of immunotherapy. By increasing influx of neutrophils and anti-tumour (M1) macrophages to the tumour, DMXAA altered myeloid cell phenotypes, thus changing the intratumoural M2/non-M2 TAM immunoinhibitory ratio. It also altered the tumour microenvironment to be more pro-inflammatory. Modulating macrophages during immunotherapy resulted in increased numbers, activity, and antigen-specificity of intratumoural CD8(+) T cells. Macrophage depletion reduced the effect of combining immunotherapy with macrophage activation, supporting the importance of TAMs in the combined effect. CONCLUSION: Modulating intratumoural macrophages dramatically augmented the effect of immunotherapy. Our observations suggest that addition of agents that activate TAMs to immunotherapy should be considered in future trials.

[Research on the relationship between chinese medical syndrome types and Th1/Th2 in bronchioloalveolar carcinoma by thoracoscopic technique].

OBJECTIVE: To study the relationship between Chinese medical syndrome types of bronchioloalveolar carcinoma (BAC) and Th1/Th2. METHODS: Totally 60 BAC patients were syndrome typed as qi and yin deficiency syndrome (QYDS) and qi stagnation and phlegm-blood stasis syndrome (QSPSS), 30 cases in each group. Meanwhile, 30 subjects with benign pulmonary nodules were recruited as the control group. The contents of interferon-gamma (INF-gamma), interleukin 4 (IL-4), IL-2, and IL-5 were detected using thoracoscopic technique. RESULTS: As for Th1 (INF-gamma and IL-2), it was ranked from high to low as the control group > the QSPSS group > the QYDS group (P < 0.05). As for Th2 (IL-4 and IL-5), it was ranked from high to low as the QYDS group > the QSPSS group >the control group (P < 0.05). As for Th1/Th2 (INF-gamma/lL-4, IL-2/IL-5), it was ranked from high to low as the control group > the QSPSS group >the QYDS group (P < 0.05). CONCLUSIONS: Compared with the tissue of benign nodules, Th1 function in tumor tissue of BAC patients was weaker and Th2 function stronger. Chinese medical syndrome types of BAC had correlation with Th1/Th2. Patients of excess syndrome had stronger immunity with Th1/Th2 shifting left,while those of deficiency syndrome were predispose to humoral immunity with Thl/Th2 shifting right.

MHC II lung cancer vaccines prime and boost tumor-specific CD4+ T cells that cross-react with multiple histologic subtypes of nonsmall cell lung cancer cells.

Nonsmall cell lung cancer (NSCLC) is the major cause of lung cancer-related deaths in the United States. We are developing cell-based vaccines as a new approach for the treatment of NSCLC. NSCLC is broadly divided into 3 histologic subtypes: adenocarcinoma, squamous cell carcinoma and large cell carcinoma. Since these subtypes are derived from the same progenitor cells, we hypothesized that they share common tumor antigens, and vaccines that induce immune reactivity against 1 subtype may also induce immunity against other subtypes. Our vaccine strategy has focused on activating tumor-specific CD4(+) T cells, a population of lymphocytes that facilitates the optimal activation of effector and memory cytotoxic CD8(+) T cells. We now report that our NSCLC MHC II vaccines prepared from adeno, squamous or large cell carcinomas each activate CD4(+) T cells that cross-react with the other NSCLC subtypes and do not react with HLA-DR-matched normal lung fibroblasts or other HLA-DR-matched nonlung tumor cells. Using MHC II NSCLC vaccines expressing the DR1, DR4, DR7 or DR15 alleles, we also demonstrate that antigens shared among the different subtypes are presented by multiple HLA-DR alleles. Therefore, MHC II NSCLC vaccines expressing a single HLA-DR allele activate NSCLC-specific CD4(+) T cells that react with the 3 major classes of NSCLC, and the antigens recognized by the activated T cells are presented by several common HLA-DR alleles, suggesting that the MHC II NSCLC vaccines are potential immunotherapeutics for a range of NSCLC patients.

Intrapulmonary administration of CCL21 gene-modified dendritic cells reduces tumor burden in spontaneous murine bronchoalveolar cell carcinoma.

The antitumor efficiency of dendritic cells transduced with an adenovirus vector expressing secondary lymphoid chemokine (CCL21) was evaluated in a murine model of spontaneous bronchoalveolar cell carcinoma. The transgenic mice (CC-10 TAg) express the SV40 large T antigen (TAg) under the Clara cell promoter, develop bilateral, multifocal, and pulmonary adenocarcinomas, and die at 4 months as a result of progressive pulmonary tumor burden. A single intratracheal administration of CCL21 gene-modified dendritic cells (DC-AdCCL21) led to a marked reduction in tumor burden with extensive mononuclear cell infiltration of the tumors. The reduction in tumor burden was accompanied by the enhanced elaboration of type 1 cytokines [IFN-gamma, interleukin (IL)-12, and granulocyte macrophage colony-stimulating factor] and antiangiogenic chemokines (CXCL9 and CXCL10) but a concomitant decrease in the immunosuppressive molecules (IL-10, transforming growth factor-beta, prostaglandin E(2)) in the tumor microenvironment. The DC-AdCCL21 therapy group revealed a significantly greater frequency of tumor-specific T cells releasing IFN-gamma compared with the controls. Continuous therapy with weekly intranasal delivery of DC-AdCCL21 significantly prolonged median survival by >7 weeks in CC-10 TAg mice. Both innate natural killer and specific T-cell antitumor responses significantly increased following DC-AdCCL21 therapy. Significant reduction in tumor burden in a model in which tumors develop in an organ-specific manner provides a strong rationale for further evaluation of intrapulmonary-administered DC-AdCCL21 in regulation of tumor immunity and genetic immunotherapy for lung cancer.

Anti-Podocalyxin Monoclonal Antibody 47-mG2a Detects Lung Cancers by Immunohistochemistry.

Lung cancer is one of the leading causes of cancer-related deaths in the world. Regardless of the advances in lung cancer treatments, the prognosis is still poor. Podocalyxin (PODXL) is a highly glycosylated type I transmembrane protein that is expressed in normal tissues, including the heart, pancreas, and breast. It is also found and used as a diagnostic marker in many cancers, such as renal, brain, breast, oral, and lung cancers. We previously developed specific and sensitive anti-PODXL monoclonal antibodies, PcMab-47 (mouse IgG1, kappa) and its mouse IgG2a-type (47-mG2a), both of which were suitable for immunohistochemical analyses of oral cancers. In this study, we investigated the utility of PcMab-47 and 47-mG2a for the immunohistochemical analyses of lung cancers. PcMab-47 stained 51/70 (72.9%) cases of lung cancer, whereas 47-mG2a stained 59/70 (84.3%) cases, indicating that the latter antibody is more sensitive and is useful for detecting PODXL in lung cancers.

Development of concomitant immunity in mice bearing the weakly immunogenic line 1 lung carcinoma.

The interaction between line 1 carcinomas growing s.c. and their spontaneous (or artificial) metastases has been studied in syngeneic BALB/c mice. In contrast to typical murine tumor systems, concomitant immunity, or the ability of primary tumors to suppress the growth of metastases, develops very slowly in this system, such that the metastases that are shed within the first week of tumor growth survive and ultimately prove lethal to the host. The rate of development of concomitant immunity can be accelerated by increasing the hosts' tumor burden or decelerated by exposure of the hosts to X-rays prior to tumor transplant. This cancer system offers, therefore, an excellent model for the therapy of metastases that cannot be obtained with typical murine tumors.

Antitumor immunity induced by hybrid tumor cells: comparison between hybrids and the parental tumor.

The ability of hybrid tumor cells to induce antitumor immunity has been evaluated in the line 1 alveolar cell carcinoma (L1) model of BALB/c mice. Hybrid tumor cells were produced by fusing freshly dissociated L1 cells isolated from in vivo tumors with the hypoxanthine:aminopterin:thymidine-sensitive cell line, GM 347, derived from C3H mice. Each hybrid was characterized by DNA content and expression of H-2 antigens using a fluorescence-activated cell sorter. Irradiated L1 cells in the presence or absence of Corynebacterium parvum were capable of immunizing BALB/c mice against a challenge of live L1 cells, provided the challenge dose was small (50% lethal dose was between 6 X 10(4) and 1.2 X 10(5) L1 cells). Testing of five hybrid clones and 1 uncloned hybrid line for their immunizing ability demonstrated a range in immunizing ability with none showing a statistically significant improvement in survival (p less than 0.0018) when compared to untreated controls. However, one hybrid clone, MoHb-L1-C2, was selected in which the survival of mice immunized with it compared to controls had a p value of 0.0255. A tumor (labeled L1/A) which grew in one of the mice immunized with this clone was removed and hybridized with GM 347 to yield a second set of hybrids. Both this variant of L1 cells and a hybrid clone made from it (MoHb-L1A-C18) were capable of immunizing mice against a challenge of live L1/A (p values of 0.0000 and 0.0028, respectively, when compared to controls). However, L1 cells were not able to immunize effectively against L1/A, and MoHb-L1A-C18 did not immunize against L1. This suggests that L1/A is a subpopulation of L1 cells with a different antigenic composition. The limited success of MoHb-L1A-C18 against L1/A is thought to be due to the narrower range of antigenic specificities in L1/A and the ability of MoHb-L1A-C18 to represent an important antigenic subpopulation of L1/A.

Characterization of a tumor cell surface protein with heterologous antisera to a spontaneous BALB/c lung carcinoma.

Line 1, a spontaneous alveolar carcinoma from a BALB/c mouse, is highly metastatic and weakly antigenic in the syngeneic host. Sera and enriched antibody preparations were made specific for line 1 cells by in vitro and in vivo absorptions. By lactoperoxidase-catalyzed radioiodination of cell surface protein followed by precipitation with specific antibodies, a protein with a molecular weight of about 180,000 (designated TSP-180) was identified that was present on line 1 cells but not on normal lung cells or Moloney sarcoma tumor cells. Studies of competition for immune precipitation of TSP-180 by unlabeled protein preparations from various sources indicate that (a) TSP-180 is present in tumor cells of various culture passage levels, (b) tumor cells grown i.m. also contain TSP-180, and (c) normal lung proteins compete weakly, and only at very high protein concentrations. A protein similar to TSP-180 was detected on Madison 109 carcinoma and on three other lung carcinomas adapted to culture. Experiments with specific antisera show that TSP-180 is not lactoperoxidase, fetal bovine serum protein, large external transformation-sensitive protein, or an antigen related to murine leukemia virus proteins.

In vivo enhancement of antitumor immunity by interleukin 2-rich lymphokines.

The ability of interleukin 2 (IL-2) to enhance in vivo antitumor immunity has been evaluated in the line 1 alveolar cell carcinoma (L1) model of BALB/c mice. A crude supernatant from phorbol myristate acetate exposed EL-4 cells rich in IL-2 plus other lymphokines (EL-4 IL-2), a concanavalin A-induced supernatant from murine splenocytes (Con A IL-2), and recombinant IL-2 (rIL-2) provided by Biogen were tested. Mice were immunized with a cloned population of L1 cells (10(6) irradiated L1 cells given s.c. in the left inguinal region) followed by s.c. injections of EL-4 IL-2, Con A IL-2, or rIL-2 given to the same site. Two immunizations of L1 cells each followed by IL-2 administration were given prior to challenge with live L1 cells s.c. on the right chest wall. Mice receiving EL-4 IL-2 survived significantly longer than those receiving L1 cells only. Daily administration of EL-4 IL-2 for 7 days after the last L1 immunization was significantly better than 3 days (P less than 0.01) which in turn was significantly better than 1 day (P less than 0.05). Among the doses tested (normalized in vitro to the Biologic Response Modifiers Program IL-2 standard) 404 units of IL-2/injection was optimal. The EL-4 IL-2 had to be injected adjacent to the site of L1 cells; s.c. injection at a distant site or i.p. was not effective. When rIL-2 or Con A IL-2 was substituted for EL-4 IL-2, survival was not prolonged; however, if Con A IL-2 (low IL-2 levels) was supplemented with rIL-2 to 404 units of IL-2, it augmented immunity as well as 404 units of EL-4 IL-2. The data suggest that IL-2 is not the only lymphokine active in augmenting antitumor immunity induced by L1 cells. Some preliminary experiments indicate that a multilymphokine approach may have potential clinical relevance.

Hepatocyte growth factor production by neutrophils infiltrating bronchioloalveolar subtype pulmonary adenocarcinoma: role in tumor progression and death.

Increased numbers of tumor-infiltrating neutrophils are linked to poorer outcome in patients with adenocarcinoma of the bronchioloalveolar carcinoma (BAC) subtype. Hepatocyte growth factor (HGF) is a pleiotropic cytokine operating through activation of the proto-oncogene c-met and is a factor of poor prognosis in various cancers. Reports that neutrophils produce HGF led us to investigate their participation in the aerogenous spread of tumor cells and the prognosis of BAC, through the effect of HGF on c-met-expressing tumor cells. Immunoreactive HGF was detected in bronchoalveolar lavage fluid (BALF) supernatants from 34 of 36 patients, whereas it was undetectable in BALF from healthy controls. The HGF thus detected was locally produced, because HGF mRNA was expressed by the patients' fresh alveolar cells, and HGF protein was detected in 24-h culture supernatants. In immunocytochemical studies of BALF cytospin preparations and tumor specimens from the patients, neutrophils were always HGF-positive, whereas alveolar macrophages and tumor cells gave inconsistent results. Alveolar neutrophil-derived HGFs induced significant, concentration-dependent migration of BAC-derived tumor cells in vitro, and this effect was inhibited by anti-HGF neutralizing antibodies. Granulocyte-macrophage colony-stimulating factor and tumor necrosis factor alpha (present in the lung tumor microenvironment) provoked HGF release from neutrophil intracellular stocks, and the capacity of blood neutrophils from BAC patients to produce HGF was unaltered. Immunochemical studies of c-met expression in BALF cytospin preparations and tumor sections showed that most HGF receptor-bearing cells were tumor cells. High HGF levels in BALF supernatants were significantly associated with poorer outcome in patients with BAC and were an independent predictor of clinical outcome in multivariate analysis. Altogether, our results support the notion that BAC generates a local environment that attracts functionally normal neutrophils from peripheral blood and leads to neutrophil release of biologically active HGF on contact with HGF receptor-expressing tumor cells, thereby contributing to poorer patient outcome.

Development of antibodies against p53 in lung cancer patients appears to be dependent on the type of p53 mutation.

Using immunoblotting techniques we studied the sera from small cell lung cancer and non-small cell lung cancer patients for antibodies directed against p53. We have also characterized the majority of these patients' tumors for p53 mutations. In the sera of 13% of the patients (4 of 40 small cell lung cancer and 2 of 6 non-small cell lung cancer) we found antibodies specific for the p53 tumor suppressor gene product. All of the antibody-positive patients tested had p53 missense mutations and expressed detectable p53 antigen in their tumor cell lines. No anti-p53 antibodies were detected in sera from patients whose tumor had p53 stop, splice/stop, splice, or frameshift mutations (n = 10). Thus, while we find that the ability of lung cancer patients to develop anti-p53 antibodies is correlated with the type of p53 mutation, many patients have tumors with missense p53 mutations and did not develop anti-p53 antibodies. The presence of p53 antibodies was not correlated to stage, prior treatment, sex, or survival. None of these lung cancer patient sera had measurable amounts of p53 antigen. By immunoblotting all six anti-p53 antisera we were able to detect a variety of mutant p53 proteins (including those from antibody-negative patients) and detected wild-type p53 protein. The development of anti-p53 antibodies represents an interesting model system for studying immune responses in cancer patients against mutant oncogene products.

Secondary lymphoid organ chemokine reduces pulmonary tumor burden in spontaneous murine bronchoalveolar cell carcinoma.

The antitumor efficiency of secondary lymphoid organ chemokine (SLC), a CC chemokine that chemoattracts both dendritic cells (DCs) and T lymphocytes,was evaluated in SV40 large T-antigen transgenic mice that develop bilateral multifocal pulmonary adenocarcinomas. Injection of recombinant SLC in the axillary lymph node region led to a marked reduction in tumor burden with extensive lymphocytic and DC infiltration of the tumors and enhanced survival. SLC injection led to significant increases in CD4 and CD8 lymphocytes as well as DC at the tumor sites, lymph nodes, and spleen. The cellular infiltrates were accompanied by the enhanced elaboration of Type 1 cytokines and the antiangiogenic chemokines IFN-gamma inducible protein 10, and monokine induced by IFN-gamma (MIG). In contrast, lymph node and tumor site production of the immunosuppressive cytokine transforming growth factor beta was decreased in response to SLC treatment. In vitro, after stimulation with irradiated autologous tumor, splenocytes from SLC-treated mice secreted significantly more IFN-gamma and granulocyte macrophage colony-stimulating factor, but reduced levels of interleukin 10. Significant reduction in tumor burden in a model in which tumors develop in an organ-specific manner provides a strong rationale for additional evaluation of SLC in regulation of tumor immunity and its use in lung cancer immunotherapy.

Interleukin-7 gene-modified dendritic cells reduce pulmonary tumor burden in spontaneous murine bronchoalveolar cell carcinoma.

The antitumor efficiency of dendritic cells transduced with an adenovirus vector expressing interleukin (IL)-7 (DC-AdIL-7) was evaluated in a murine model of spontaneous bronchoalveolar cell carcinoma. These transgenic mice (CC-10 TAg), expressing the SV40 large T antigen under the Clara cell promoter, develop bilateral multifocal pulmonary adenocarcinomas and die at 4 months as a result of progressive pulmonary tumor burden. Injection of DC-AdIL-7 in the axillary lymph node region (ALNR) weekly for 3 weeks led to a marked reduction in tumor burden with extensive lymphocytic infiltration of the tumors and enhanced survival. The antitumor responses were accompanied by the enhanced elaboration of interferon (IFN)-gamma and IL-12 as well as an increase in the antiangiogenic chemokines, IFN-gamma-inducible protein 10 (IP-10/CXCL10) and monokine induced by IFN-gamma (MIG/CXCL9). In contrast, production of the immunosuppressive mediators IL-10, transforming growth factor (TGF)-beta, prostaglandin E(2) (PGE(2)), and the proangiogenic modulator vascular endothelial growth factor (VEGF) decreased in response to DC-AdIL-7 treatment. Significant reduction in tumor burden in a model in which tumors develop in an organ-specific manner provides a strong rationale for further evaluation of DC-AdIL-7 in regulation of tumor immunity and its use in lung cancer genetic immunotherapy.

The neutralization of interferons by antibody. I. Quantitative and theoretical analyses of the neutralization reaction in different bioassay systems.

The highly specific ability of antibodies to inhibit the biologic activity of cytokines or other therapeutic proteins is widely used in research and a subject of increasing clinical importance. The need exists for a standardized approach to the reporting of neutralizing antibody potency soundly based on theoretical and practical considerations and tested by experimental data. Pursuant to the original studies of Kawade on the theoretical and functional aspects of neutralization of interferons (IFN), experimental data were obtained by different laboratories employing varied methodology to address two hypotheses concerning the nature of IFN neutralization reactions, based on a derived formula that allows expression of neutralizing power as the reduction of 10 laboratory units (LU)/ml to 1 LU/ml, the end point of most bioassays. Two hypotheses are posed: (1) antibody acts to neutralize a fixed amount of biologically active IFN molecules, or (2) antibody reduces IFN activity in a set ratio of added/residual biologically active IFN. The first, or fixed amount, hypothesis relates to the reactivity of high-affinity antibodies neutralizing equimolar amounts of antigen, whereas the second, or constant proportion, hypothesis postulates a reduction in the ratio of total added IFN to residual active IFN molecules, such as a low-affinity antibody might exhibit. Analyses of data of the neutralization of IFN-alpha and IFN-beta are presented, employing human polyclonal antibodies and murine monoclonal antibodies (mAb). The theoretical constructs of Kawade are extended in the Appendix and correlated with new experimental data in the text. The data clearly indicate that the low-antibody affinity, constant proportion hypothesis, rather than the high-antibody affinity, fixed amount hypothesis, is applicable, if the bioassay is sensitive to IFN. The findings presented here and in the following paper (pp. 743-755, this issue) taken together provide the basis for a standardized method of expression of neutralizing potency and substantiate the earlier operational 10/1 LU/ml approach recommended by the World Health Organization. The accompanying paper relates neutralization results to the sensitivity of the bioassay to IFN and describes the rationale for a recommended unit of antibody neutralization.

Dendritic cells augment granulocyte-macrophage colony-stimulating factor (GM-CSF)/herpes simplex virus thymidine kinase-mediated gene therapy of lung cancer.

Lung cancer, the leading cause of cancer death in the United States, is resistant to most currently available therapies. To evaluate a multicomponent gene therapy approach that replaces tumor-bearing host immune deficits, we genetically modified Line 1 (L1C2), a weakly immunogenic alveolar cell carcinoma cell line. L1C2 was transduced ex vivo with a retroviral construct that contained two components: a cytokine gene (granulocyte-macrophage colony-stimulating factor) and a drug sensitivity gene (herpes simplex virus thymidine kinase). The third component of this therapy, in vitro-activated syngeneic bone marrow-derived dendritic cells, was included to augment antigen presentation. The addition of ganciclovir (GCV) caused the lysis of transduced tumor cells, resulting in the release of potential tumor antigens. Ex vivo-transduced tumor cells regressed in vivo following GCV therapy but were not effective in the treatment of established parental tumors. To treat established tumors, dendritic cells were administered in combination with transduced tumor cells and GCV. A total of 50% of these mice rejected the 5-day-old established tumors and were immune to rechallenge with parental L1C2 cells. Thus, this multicomponent gene therapy system leads to both the regression of established tumors and enhanced immunogenicity in this weakly immunogenic murine lung cancer model.

Metastases from fresh human non-small cell lung cancers propagated in nude mice.

Specimens from 69 freshly resected human non-small cell lung cancers (NSCLC) were transplanted into nude mice. Twelve mice died before the transplants were evaluable. There were 4 takes of 12 evaluable transplants into untreated athymic nude mice and 24 takes of 45 evaluable transplants into nude mice with decreased natural killer (NK) cell activity. Fourteen tumor lines were propagated into 2 or more successive transplant generations. Distant metastases occurred from the mid-dorsal transplant site in 8 of 81 (9.88%) recipients of 4 of those tumor lines, after 3-9 consecutive tumor growth cycles. Xenotransplantation of freshly resected human NSCLC provides a model with potential for serial assessment of cellular changes related to metastatic capability.

Immune reactivity in primary carcinoma of the lung and its relation to prognosis.

Detailed studies of immune reactivity were performed in 154 patients with primary lung cancer, 20 patients with benign thoracic lesions, and 109 healthy persons. Reactions to the 2,4-dinitrochlorobenzene (DNCB) skin test were postive in 73 per cent of patients with lung cancer and all (100 per cent) of the patients with benign disease (p less than 0.05). The incidence of DNCB reactions was 78 per cent for Stage I and II cancers (37 patinets), 73 per cent for resectable Stage III cancer (22 patients), and 66 per cent in patients with unresectable or inoperable Stage III cancer. DNCB reactivity showed a relationship to primary histology. The incidence of DNCB positive reactions was 80 per cent in patients with epidermold carcinoma versus 57 per cent in patients with adenocarcinoma, 64 per cent in patients with oat cell cancer, and 80 per cent in patients with terminal bronchiolar carcinoma. In vitro immune studeis correlated best with stage of disease. These included the absolute lymphocyte count and absolute T cell count and lymphoxyte stimulation witalen A (Com A). These values were in the normal range in patients with Stage I cancer but were significantly depressed in patients with Stage III cancer. Svrvival curves were plotted in patients with Stage III disease according to the responses to three immune parameters: DNCB, absolute lymphocyte count, and PHS stimulation. Although patients with normal reactions generally had better survival rates, PHA responses showed the most significant correlation to survival. These tests support the usefulness of immune testing as an additional parameter of assessing biological risk in patients with primary lung cancer.

Bronchioloalveolar carcinoma accompanied by severe bronchorrhea.

We report an unusual case of bronchioloalveolar carcinoma characterized by production of a large quantity of sputum accompanied by drastic electrolyte and fluid loss. The sputum contained a high level of gastrointestinal cancer-associated antigen (CA19-9) and carcinoembryonic antigen (CEA). An immunohistochemical study of tumor cells showed the specific distribution of gastrointestinal cancer-associated antigen and carcinoembryonic stains, which were localized in the apical region of tumor cells.

Bronchioloalveolar carcinoma in a child with congenital cystic adenomatoid malformation.

An 11-year-old girl was evaluated for chest pain, and chest radiographic findings of multiple nodules throughout both lungs. She underwent resection of several of the lesions from her left lung, which were found at pathologic examination to be bronchioloalveolar carcinoma. Her previous medical history included incomplete resection of a type I congenital cystic adenomatoid malformation in the neonatal period. To our knowledge, this girl is the youngest reported case of bronchioloalveolar carcinoma in a nonimmunocompromised patient, and one of several in which the association of congenital cystic adenomatoid malformation and bronchioloalveolar carcinoma has been observed.