Solid Tumor-derived Cell Lines Research Articles

Inhibition of γ-Secretase Activity Inhibits Tumor Progression in a Mouse Model of Pancreatic Ductal Adenocarcinoma

The Notch signaling pathway is required for the expansion of undifferentiated pancreatic progenitor cells during embryonic development and has been implicated in the progression of pancreatic ductal adenocarcinoma (PDAC). The interaction of Notch ligands with their receptors promotes a gamma-secretase-dependent cleavage of the Notch receptor and release of the Notch intracellular domain, which translocates to the nucleus and activates transcription. We investigated the role of this pathway in PDAC progression. We tested the effects of a gamma-secretase inhibitor (GSI) that blocks Notch signaling in PDAC cell lines and a genetically engineered mouse model of PDAC (Kras p53 L/+ mice). Notch signaling was activated in PDAC precursors and advanced tumors. The GSI inhibited the growth of premalignant pancreatic duct-derived cells in a Notch-dependent manner. Additionally, in a panel of over 400 human solid tumor-derived cell lines, PDAC cells, as a group, were more sensitive to the GSI than any other tumor type. Finally, the GSI completely inhibited tumor development in the genetically engineered model of invasive PDAC (P < .005, chi2 test; compared with mice exposed to vehicle). These results suggest that Notch signaling is required for PDAC progression. Pharmacologic targeting of this pathway offers therapeutic potential in this treatment-refractory malignancy.

Synthesis and evaluation of dihydroartemisinin and dihydroartemisitene acetal dimers showing anticancer and antiprotozoal activity

Twelve artemisinin acetal dimers were synthesized and tested for antitumor activity in the National Cancer Institute (NCI) in vitro human tumor 60 cell line assay, producing a mean GI 50 concentration between 8.7 (least active) and 0.019 μM (most active). The significant activity of the compounds in this preliminary screen led to additional in vitro antitumor and antiangiogenesis studies. Several active dimers were also evaluated in the in vivo NCI hollow fiber assay followed by a preliminary xenograft study. The title compounds were found to be active against solid tumor-derived cell lines and showed good correlation with other artemisinin-based molecules in the NCI database. The dimers were also evaluated for their antimalarial and antileishmanial activities. The antimalarial activity ranged from 0.3 to 32 nM (IC 50), compared to 9.9 nM for artemisinin.

Chemotherapy and zoledronate sensitize solid tumour cells to Vgamma9Vdelta2 T cell cytotoxicity.

Combinations of cellular immune-based therapies with chemotherapy and other antitumour agents may be of significant clinical benefit in the treatment of many forms of cancer. Gamma delta (gammadelta) T cells are of particular interest for use in such combined therapies due to their potent antitumour cytotoxicity and relative ease of generation in vitro. Here, we demonstrate high levels of cytotoxicity against solid tumour-derived cell lines with combination treatment utilizing Vgamma9Vdelta2 T cells, chemotherapeutic agents and the bisphosphonate, zoledronate. Pre-treatment with low concentrations of chemotherapeutic agents or zoledronate sensitized tumour cells to rapid killing by Vgamma9Vdelta2 T cells with levels of cytotoxicity approaching 90%. In addition, zoledronate enhanced the chemotherapy-induced sensitization of tumour cells to Vgamma9Vdelta2 T cell cytotoxicity resulting in almost 100% lysis of tumour targets in some cases. Vgamma9Vdelta2 T cell cytotoxicity was mediated by perforin following TCR-dependent and isoprenoid-mediated recognition of tumour cells. Production of IFN-gamma by Vgamma9Vdelta2 T cells was also induced after exposure to sensitized targets. We conclude that administration of Vgamma9Vdelta2 T cells at suitable intervals after chemotherapy and zoledronate may substantially increase antitumour activities in a range of malignancies.

Chemotherapy pretreatment sensitizes solid tumor‐derived cell lines to Vα24+ NKT cell‐mediated cytotoxicity

There is an increasing awareness of the therapeutic potential for combining immune-based therapies with chemotherapy in the treatment of malignant diseases, but few published studies evaluate possible cytotoxic synergies between chemotherapy and cytotoxic immune cells. Human V alpha 24+/V beta 11+ NKT cells are being evaluated for use in cell-based immunotherapy of malignancy because of their immune regulatory functions and potent cytotoxic potential. In this study, we evaluated the cytotoxicity of combinations of chemotherapy and NKT cells to determine whether there is a potential to combine these treatment modalities for human cancer therapy. The cytotoxicity of NKT cells was tested against solid-tumor derived cell lines NCI-H358, DLD-1, HT-29, DU-145, TSU-Pr1 and MDA-MB231, with or without prior treatment of these target cells, with a range of chemotherapy agents. Low concentrations of chemotherapeutic agents led to sensitization of cell lines to NKT-mediated cytotoxicity, with the greatest effect being observed for prostate cancer cells. Synergistic cytotoxicity occurred in an NKT cell in a dose-dependent manner. Chemotherapy agents induced upregulation of cell surface TRAIL-R2 (DR5) and Fas (CD95) expression, increasing the capacity for NKT cells to recognize and kill via TRAIL- and FasL-mediated pathways. We conclude that administration of cytotoxic immune cells after chemotherapy may increase antitumor activities in comparison with the use of either treatment alone.

Induction of Accelerated Senescence by γ Radiation in Human Solid Tumor-Derived Cell Lines Expressing Wild-Type TP53

Recent studies have demonstrated that p21WAF1 (now known as CDKN1A)-dependent and -independent accelerated senescence responses are a major determinant of the sensitivity of cancer cells to chemotherapeutic agents. The objective of the present study was to determine whether human solid tumor-derived cell lines that express wild-type TP53 can exhibit levels of CDKN1A induction after exposure to ionizing radiation that are sufficient to activate the accelerated senescence program. Exposure to 60Co gamma radiation (< or =8 Gy) triggered accelerated senescence in all five TP53 wild-type tumor cell lines examined, albeit to differing degrees. Three of the TP53 wild-type tumor cell lines, HCT116, A172 and SKNSH, activated the TP53 signaling pathway similarly to normal human fibroblasts, as judged by the nuclear accumulation of TP53, magnitude and duration of induction of CDKN1A mRNA and CDKN1A protein, and propensity to undergo accelerated senescence after radiation exposure. In the clonogenic survival assay, the degree of radiosensitivity of these three tumor cell lines was also in the range displayed by normal human fibroblasts. On the other hand, two other TP53 wild-type tumor cell lines, A498 and A375, did not maintain high levels of CDKN1A mRNA and CDKN1A protein at late times postirradiation and exhibited only low levels of accelerated senescence after radiation exposure. Studies with a CDKN1A knockout cell line (HCT116CDKN1A-/-) confirmed that the radiation-triggered accelerated senescence is dependent on CDKN1A function. We conclude that (1) clinically achievable doses of ionizing radiation can trigger CDKN1A-dependent accelerated senescence in some human tumor cell lines that express wild-type TP53; and (2) as previously documented for normal human fibroblasts, some TP53 wild-type tumor cell lines (e.g. HCT116, A172 and SKNSH) may lose their clonogenic potential in response to radiation-inflicted injury primarily through undergoing accelerated senescence.

Synthesis and biological evaluation of benzo[ d]isothiazole, benzothiazole and thiazole Schiff bases

Three new series of benzo[ d]isothiazole, benzothiazole and thiazole Schiff bases were synthesized and tested in vitro with the aim of identifying novel lead compounds active against emergent and re-emergent human and cattle infectious diseases (AIDS, hepatitis B and C, tuberculosis, bovine viral diarrhoea) or against drug-resistant cancers (leukaemia, carcinoma, melanoma, MDR tumors) for which no definitive cure or efficacious vaccine is available at present. In particular, these compounds were evaluated in vitro against representatives of different virus classes, such as a HIV-1 (Retrovirus), a HBV (Hepadnavirus) and the single-stranded RNA + viruses Yellow fever virus (YFV) and Bovine viral diarrhoea virus (BVDV), both belonging to Flaviviridae. Title compounds were also tested against representatives of Gram-positive and Gram-negative bacteria ( Staphylococcus aureus, Salmonella spp.) , various atypic mycobacterial strains ( Mycobacterium fortuitum and Mycobacterium smegmatis), yeast ( Candida albicans) and mould ( Aspergillus fumigatus). None of the compounds showed antiviral or antimicrobial activity. The benzo[ d]isothiazole compounds showed a marked cytotoxicity (CC 50=4–9 μM) against the human CD4 + lymphocytes (MT-4) that were used to support HIV-1 growth. For this reason, the most cytotoxic compounds of this series were evaluated for their antiproliferative activity against a panel of human cell lines derived from haematological and solid tumors. The results highlighted that all the benzo[ d]isothiazole derivatives inhibited the growth of leukaemia cell lines, whereas only one of the above mentioned compounds ( 1e) showed antiproliferative activity against two solid tumor-derived cell lines.

Synthesis and antiproliferative activity of basic thioanalogues of merbarone

Three series of 5-substituted 1,3-diphenyl-6-(ω-dialkyl- and ω-cyclo-aminoalkyl)thio-2-thiobarbiturates ( 11– 13) were synthesized as polysubstituted thioanalogues of merbarone, a topoisomerase II inhibitor acting on the catalytic site. To better understand pharmacophore requirements, a forth series of conformationally constrained analogues 14 was also prepared. Derivatives 11b, e, 14b, e, h, i, j were active in the low micromolar concentration range (IC 50: 3.3–4.3 μM), whereas compounds 11a, c, d, f,h, j and 13a, b, d, g, j and 14a, d, f showed IC 50 values between 10 and 15.5 μM. In constrast, compounds 12a– c, g– j, 13e, f, h and 14k were inactive. Cytotoxicity data provided from N.C.I. on selected compounds provided evidence that 11b, d, 13d, g and 14b, d, f, h, i, j were endowed with potent antiproliferative activity against leukemia and prostate cell lines (GI 50 up to 0.01 μM). In general, bicyclic derivatives 14 were up to 10-fold more potent than monocyclic counterparts against solid tumor-derived cell lines. SAR studies indicated that, in general, a certain tolerability in length of the alkyl side chains and in shape of distal amines is allowed in the four series, but in the monocyclic derivatives ( 11– 13) antiproliferative activity was strongly affected by the nature of the 5-substituents (COOC 2H 5>COCH 3≫C 6H 5). Compounds 11b and 14b were also evaluated against KB cell subclones expressing altered levels of topoisomerases or the multidrug resistance phenotype (MDR). In both cases the above compounds showed a decrease in potency. In enzyme assays, 11b and 14b turned out to be inhibitors of topoisonerase II as merbaron.

Synthesis and evaluation of anticancer benzoxazoles and benzimidazoles related to UK-1

UK-1 is a structurally unique bis(benzoxazole) natural product isolated from a strain of Streptomyces. UK-1 has been reported to possess anticancer activity but no activity against bacteria, yeast, or fungi. Previous work has also demonstrated the ability of UK-1 to bind a variety of di- and tri-valent metal ions, particularly Mg 2+ ions, and to form complexes with double-stranded DNA in the presence of Mg 2+ ions. Here we report the activity of UK-1 against a wide range of human cancer cell lines. UK-1 displays a wide spectrum of potent anticancer activity against leukemia, lymphoma, and certain solid tumor-derived cell lines, with IC 50 values as low as 20 nM, but is inactive against Staphylococcus aureus, a methicillin-resistant strain of S. aureus, or Pseudomonas aeruginosa. A series of analogues of the bis(benzoxazole) natural product UK-1 in which the carbomethoxy-substituted benzoxazole ring of the natural product was modified were prepared and evaluated for their anticancer and antibacterial properties. An analogue of UK-1 in which the carbomethoxy-substituted benzoxazole ring was replaced with a carbomethoxy-substituted benzimidazole ring was inactive against human cancer cell lines and the two strains of S. aureus. In contrast, a simplified analogue in which the carbomethoxy-substituted benzoxazole ring was replaced with a carbomethoxy group was almost as active as UK-1 against the four cancer cell lines examined but lacked activity against S. aureus. Metal ion binding studies of these analogues demonstrate that they both bind Zn 2+ and Ca 2+ ions about as well as UK-1. The non-cytotoxic benzimidazole UK-1 analogue binds Mg 2+ ions 50-fold weaker than UK-1, whereas the simple benzoxazole analogue binds Mg 2+ ions nearly as well as UK-1. These results support a role of Mg 2+ ion binding in the selective cytotoxicity of UK-1 and provide a minimal pharmacophore for the selective cytotoxic activity of the natural product.

Hygrolidin induces p21 expression and abrogates cell cycle progression at G1 and S phases

Hygrolidin family antibiotics showed selective cytotoxicity against both cyclin E- and cyclin A-overexpressing cells. Among them, hygrolidin was the most potent and inhibited growth of solid tumor-derived cell lines such as DLD-1 human colon cancer cells efficiently more than that of hematopoietic tumor cells and normal fibroblasts. FACS analysis revealed that hygrolidin increased cells in G1 and S phases in DLD-1 cells. While hygrolidin decreased amounts of cyclin-dependent kinase (cdk) 4, cyclin D, and cyclin B, it increased cyclin E and p21 levels. Hygrolidin-induced p21 bound to and inhibit cyclin A–cdk2 complex more strongly than cyclin E–cdk2 complex. Furthermore, hygrolidin was found to increase p21 mRNA in DLD-1 cells, but not in normal fibroblasts. Thus, hygrolidin inhibited tumor cell growth through induction of p21. In respect to p21 induction, inhibition of vacuolar-type (H+)-ATPase by hygrolidin was suggested to be involved.

Immunocytochemical analysis of cell lines derived from solid tumors.

Antibodies recognizing tissue-specific antigens are widely used to identify the histological origin of tumors. Here we tested the fidelity of selected tissue markers on all 167 solid tumor-derived continuous cell lines in the DSMZ cell lines bank. Most lines had an intermediate filament content consistent with the tumor type from which they were derived. Thus, 93% of all carcinoma cell lines expressed keratin filaments. With certain antibodies, some subclassification was possible. For example, the CK7 keratin 7 antibody can differentiate between colon and pancreas-derived carcinoma cell lines. Cell lines derived from non-carcinomas, in general, did not express keratin but were vimentin-positive. Four of 10 glioma/astrocytoma cell lines expressed GFAP, five of six neuroblastoma cell lines expressed neurofilaments, and the TE-671 rhabdomyosarcoma cell line expressed desmin. When other tissue markers were tested, 12/16 melanoma-derived cell lines expressed HMB-45, while PSA, CA125, and thyroglobulin were less useful. These results demonstrate that cell lines retain some but not all markers typical of the original tumor type and identify certain markers useful in characterizing the histological origin of cell lines. Our data question the identity of some cell lines submitted to the bank in the past. The immunoprofiles of 167 solid tumor-derived and 131 hematopoetic cell lines can be found at www.dsmz.de.

Hematopoietic-specific expression of MEFV, the gene mutated in familial Mediterranean fever, and subcellular localization of its corresponding protein, pyrin

Familial Mediterranean fever (FMF) is a recessively inherited disorder characterized by recurrent, self-limited attacks of fever and serositis and by infiltration of affected tissues by large numbers of neutrophils. A candidate gene for FMF was identified by positional cloning and named "MEFV." The corresponding protein was named "pyrin." To elucidate the currently unknown function of pyrin, we characterized its tissue distribution, regulation of expression during hematopoietic differentiation, and subcellular localization. Reverse transcription-polymerase chain reaction analysis, followed by hybridization with an internal oligonucleotide, demonstrated expression of MEFV in different populations of peripheral blood cells. Among hematopoietic cell lines, MEFV was almost exclusively expressed in cells of the myeloid lineage. Furthermore, MEFV messenger RNA was strongly expressed within 24 hours of dimethyl sulfoxide-induced granulocytic differentiation of HL-60 cells. Analysis of complementary DNA from human solid tumor-derived cell lines revealed expression of MEFV in several cell lines derived from colon and prostate cancers. Expression of MEFV fused to enhanced green fluorescent protein showed that pyrin localized in distinct patches in the cytoplasm, forming a perinuclear cap. Taken together, MEFV is predominantly expressed in myeloid cells and upregulated during myeloid differentiation, and the corresponding protein, pyrin, is expressed in the cytoplasm. (Blood. 2000;95:1451-1455)

Indolo[3,2-c]cinnolines with antiproliferative, antifungal, and antibacterial activity

A series of indolo[3,2-c]cinnoline derivatives was prepared and tested to evaluate their biological activity. Most of them inhibited the proliferation of leukemia, lymphoma and solid tumor-derived cell lines at micromolar concentrations, whereas none of the compounds were active against HIV-1. With the exception of 7g Table 1Antiproliferative activity of indolo[3,2-c]cinnolinesIC50 [μM]aCell lines7a7b7c7d7e7f7g7h7iDoxoLeukemia/lymphomaL1210121.03.5305024>1000.3ND0.23Wil2-NS4.50.51.0201018890.21.10.02CCRF-SB50.72.8213.227>1000.21.10.01Raji6.80.817>1012>10>1001.21.80.08CCRF-CEM50.51.9107.35.0540.082.20.10MOLT-43.30.41.6123.212>1000.080.20.03MT-42.20.94.7252.350>1000.30.50.09CarcinomaHT-29505.20.82.8>502.1941.94.50.05HeLa8.91.01048.133>10>1007.71.00.25ACHN>5.03.11.241.515.5>10>1003.03.00.4456374.00.32.11019>1018.90.72.70.02NeuroblastomaIMR-325.80.6NDND7.0NDNDNDND<0.01aCompound concentration required to reduce cell multiplication by 50% under conditions allowing untreated controls to undergo at least three consecutive rounds of multiplication. L1210, mouse leukemia; Wil2-NS, human splenic B-lymphoblastoid cells; CCRF-SB, human acute B-lymphoblastoic leukemia; Raij, human Burkitt lymphoma; CCRF-CEM and MOLT-4, human acute T-lymphobastic leukemia; MT-4, human CD4+ T-cells containing an integrated HTLV-1 genome; HT-29, human colon adenocarcinoma; HeLa, human cervix carcinoma; ACHN, human renal adenocarcinoma; 5637, human bladder carcinoma; IMR-32, human neuroblastoma. ND: not determined.Table 3Antifungal activity of indolo[3,2-c]cinnolinesMIC [μM]aSpecies7a7b7c7d7e7f7g7h7iMiconazoleC. albicans1509.412>200>200>200>2001.6667.5C. parapsilosis1.82.3100>200>200>200>2001.6220.9C. paratropicalis371950>200>200>200>2001.6667.5C. neoformans0.80.16.2>200>200>200>2001.67.40.9T. mentagrophytes1.875NDND>200NDNDNDND0.9A. fumigatus5.04.7200>200>200>200>2003.12001.9aMinimum inhibitory concentration. ND: not determined.Table 4Antibacterial activity indolo[3,2c]cinnolinesMIC [μM]aSpecies7a7b7c7d7e7f7g7h7iStreptomycinStreptococcus0.20.10.46.23.16.2>2000.050.32.1Staphlyococcus0.10.070.46.26.26.21000.020.094.2Salmonella>200>200>200>200>200>200>2003.12008.6Shigella>200>200>200>200>200>200>2001.6662.1aMinimum inhibitory concentration., all title compounds showed antibacterial activity against gram-positive bacteria, being up to 200 times more potent than the reference drug streptomycin. Some of the indolo[3,2-c]cinnolines were also endowed with good antifungal activity, particularly against Cryptococcus neoformans.

Glutamine as a regulator of DNA and protein biosynthesis in human solid tumor cell lines.

The transport of glutamine by six different human solid tumor-derived cell lines (e.g., breast, colon, liver) was characterized and the impact of glutamine deprivation on rates of tumor cell proliferation and DNA and protein synthesis was assayed. Glutamine is added routinely to cell culture media and its importance for cellular growth has been established. However, carrier-mediated glutamine transport by solid tumors has not been studied extensively, and the mechanisms by which glutamine contributes to cell growth regulation require further investigation. In a panel of different human solid tumor-derived cells, sodium-dependent glutamine transport was characterized in vitro and rates of cell proliferation, protein and DNA synthesis, as well as thymidine transport, were correlated with glutamine concentrations in the culture media. In all cells, regardless of tissue origin, sodium-dependent glutamine transport was mediated almost exclusively by a single carrier. There was a range of Michaelis constants (Km) and maximal transport velocities (Vmax) for the glutamine transporter in each cell type, but the amino acid inhibition profiles were nearly identical, consistent with uptake by the System ASC family of transporters. Rates of cell growth, DNA and protein synthesis, and thymidine transport correlated with the glutamine concentration in the culture media, indicating the central role of this amino acid in regulating cellular proliferation. These data indicate that glutamine transport by all solid tumors is mediated by the System ASC family of transporters. The variation in Km values suggests that some cancers may be better suited to survive in a low glutamine environment than others. The mechanism by which glutamine supports cell proliferation and regulates cell cycle kinetics involves its modulation of DNA and protein biosynthetic rates.

Retinoic acid receptor and retinoid X receptor expression in retinoic acid—resistant human tumor cell lines

Retinoic acid (RA) has profound effects on cell proliferation and differentiation both in vitro and in vivo. Many human cell lines are known to be sensitive to the growth-inhibitory action of RA. We analyzed established human solid tumor-derived cell lines for their RA sensitivity. Growth inhibition by RA in monolayer was examined by [3H]thymidine incorporation and cell proliferation. Here we report that 11 widely used human cell lines were RA resistant. The majority are carcinoma derived (A-431, BT-20, C-41, ACHN, HCT116, 293, A549, and PA-1); two are sarcoma derived (Saos-2 and A673); and one is a melanoma cell line (A-375). Since nuclear retinoid receptors are implicated in the biological effects of RA, we examined the expression of retinoic acid receptors (RARs) RAR alpha, RAR beta, RAR gamma, and the retinoid X receptors (RXRs) RXR alpha, RXR beta, and RXR gamma in the RA-resistant cell lines by northern blotting and by RNase protection analysis for RAR beta. RAR alpha transcripts were constitutively expressed in all cell lines. By contrast, RAR beta was expressed in only seven RA-resistant cell lines (Saos-2, ACHN, 293, A549, A-375, A673, and PA-1), and its level was enhanced by RA in some cases. In most cell lines, RAR gamma expression was low and was not affected by RA. The RXR genes showed a very distinct expression pattern in the group of selected cell lines. In general, RXR alpha was the most abundantly expressed subtype, RXR beta was expressed at low levels, and RXR gamma could not be detected. In none of the RA-resistant cell lines was RXR expression modulated by RA. The results presented here indicate that the resistance of these human tumor cell lines to RA cannot be simply correlated with expression of RAR or RXR or both.

Production of natural killer cell stimulatory factor (interleukin 12) by peripheral blood mononuclear cells.

Natural killer cell stimulatory factor (NKSF), or interleukin 12 (IL-12), is a 70-kD heterodimeric cytokine composed of two covalently linked chains, p40 and p35. NKSF/IL-12 has multiple effects on T and NK cells and was originally identified and purified from the supernatant fluid of Epstein-Barr virus (EBV)-transformed human B lymphoblastoid cell lines. We have produced a panel of monoclonal antibodies against both chains of NKSF/IL-12. Some of these antibodies have neutralizing activity, and several combinations of them have been used to establish sensitive radioimmunoassays detecting the free p40 chain, the free p35 chain, or the p70 heterodimer. Using these reagents, we have determined that most EBV-transformed human B lymphoblastoid cell lines constitutively produce low levels of the p70 heterodimer and an excess of the free p40 chain, whereas Burkitt lymphoma-derived, T, myeloid, and many solid tumor-derived cell lines produce neither. Production of both p40 and p70 is increased several-fold upon stimulation of the EBV-transformed cell lines with phorbol diesters. The ability of supernatant fluids from unstimulated and phorbol diester-stimulated cell lines to induce interferon gamma (IFN-gamma) production from T and NK cells, one of the effects of NKSF/IL-12, parallels the levels of production of the p70 heterodimer, known to be the biologically active form of NKSF/IL-12. Staphylococcus aureus Cowan I strain (SAC) and other stimuli induce accumulation of p40 mRNA and production of both p40 and p70 by peripheral blood mononuclear cells (PBMC). The producer cells appear to include both adherent cells and nonadherent lymphocytes, possibly B cells. The supernatant fluids from SAC-stimulated PBMC mediate the typical functions of NKSF/IL-12 (i.e., IFN-gamma induction, mitogenic effects on T/NK blasts, enhancement of NK cell cytotoxicity) at concentrations of p70 similar to those at which recombinant NKSF/IL-12 mediates the same functions. Moreover, these activities are significantly inhibited by anti-NKSF/IL-12 antibodies. The neutralizing anti-NKSF/IL-12 antibodies also inhibit 85% of the IFN-gamma production in response to SAC, an NKSF/IL-12 inducer, and approximately 50% of the IFN-gamma production in response to non-NKSF/IL-12-inducers such as IL-2, phytohemagglutinin, and anti-CD3 antibodies. These results indicate that induced or constitutively produced NKSF/IL-12 has a major role in facilitating IFN-gamma production by peripheral blood lymphocytes.(ABSTRACT TRUNCATED AT 400 WORDS)

Development of Monoclonal Antibodies to the Human Breast Carcinoma Cell Line PMC42&lt;xref ref-type="fn" rid="FN1"&gt;2&lt;/xref&gt;

In an attempt to identify antigens expressed during breast differentiation, three murine monoclonal antibodies, CIBr2, CIBr7, and CIBr18, were produced against the human pleomorphic breast carcinoma cell line PMC42. All three monoclonal antibodies reacted with previously undescribed antigenic determinants on the PMC42 cell line. Antibody CIBr18 reacted only with the immunizing cell line PMC42, whereas antibodies CIBr2 and CIBr7 showed minimal reactivity toward a panel of 34 human leukemia- and solid tumor-derived cell lines. The antigenic determinants detected by the three antibodies were distinct, and each showed variable expression in PMC42 monolayer and organoid cultures. The heterogeneity of staining seen on PMC42 cultures may reflect the fact that this cell line contains up to eight morphologically distinct cell types. Antigen expression correlated with cell type in some instances, whereas in other instances phenotypic subdivision within a cell type was apparent. Antigens recognized by antibodies CIBr7 and CIBr18 were characterized biochemically. In Western blotting, antibody CIBr7 identified a single band of an apparent molecular weight of 38,000 within PMC42 cell lysates. Sodium dodecyl sulfate-polyacrylamide gel analysis of polypeptides immunoprecipitated by antibody CIBr18 from [35S]methionine-labeled PMC42 cell lysates identified two glycoproteins of apparent molecular weights of 115,000 and 120,000, respectively. No biochemical data for the CIBr2 antigen are yet available. All three antigens were detected in human mammary epithelium and some non-breast tissues. The expression of these antigens in normal and neoplastic mammary epithelia is discussed in terms of antigen heterogeneity and changes in antigen expression upon conversion to the malignant state.