Intralesional delivery of dendritic cells engineered to express T-bet promotes protective type 1 immunity and the normalization of the tumor microenvironment.

T-bet (Tbx21), a T-box transcription factor, has been previously identified as a master regulator of type 1 T cell polarization. We have also recently shown that the genetic engineering of human dendritic cells (DCs) to express human T-bet cDNA yields type 1-polarizing APCs in vitro (1). In the present study, murine CD11c(+) DCs were transduced with a recombinant adenovirus encoding full-length murine T-bets (DC.mTbets) and analyzed for their immunomodulatory functions in vitro and in vivo. Within the range of markers analyzed, DC.mTbets exhibited a control DC phenotype and were indistinguishable from control DCs in their ability to promote allogenic T cell proliferation in MLR in vitro. However, DC.mTbets were superior to control DCs in promoting Th1 and Tc1 responses in vitro via a mechanism requiring DC-T cell interaction or the close proximity of these two cell types and that can only partially be explained by the action of DC-elaborated IL-12p70. When injected into day 7 s.c. CMS4 sarcoma lesions growing in syngenic BALB/c mice, DC.mTbets dramatically slowed tumor progression (versus control DCs) and extended overall survival via a mechanism dependent on both CD4(+) and CD8(+) T cells and, to a lesser extent, asialoGM1(+) NK cells. DC.mTbet-based therapy also promoted superior tumor-specific Tc1 responses in the spleens and tumor-draining lymph nodes of treated animals, and within the tumor microenvironment it inhibited the accumulation of CD11b(+)Gr1(+) myeloid-derived suppressor cells and normalized CD31(+) vascular structures. These findings support the potential translational utility of DC.Tbets as a therapeutic modality in the cancer setting.

Effects of combined radiotherapy and immunotherapy with the use of pyran copolymer on murine fibrosarcoma.

A weakly immunogenic, 3-methylcholanthrene-induced, subcutaneous fibrosarcoma syngeneic to inbred C3H/HeJ mice was used. Pyran copolymer was injected either directly into the tumor, ip, or iv as soon as tumors appeared or when tumors were 8 mm in diameter. One, three, or five doses of pyran copolymer at 10 or 20 mg/kg/dose were injected, with multiple doses being given every other day. Pyran copolymer injected intratumorally once, three times, or five times significantly retarded tumor growth and prolonged the survival times of the hosts. Of the other routes and doses, only pyran copolymer given three times iv significantly retarded tumor growth, but none of these significantly prolonged the survival times of the hosts. Pyran copolymer alone did not induce any complete regression of tumor. Local tumor irradiation with a single exposure to 2,000 rads of X-ray induced complete regressions in some mice, but a higher percentage of tumor cure was observed when tumor irradiation was followed by pyran copolymer treatment.

Role of whole-body lipids and nitrogen as limiting factors for survival in tumor-bearing mice with anorexia and cachexia.

This study was designed to ascertain whether the overall availability of whole-body lipids and nitrogen is a limiting factor for survival in tumor-bearing mice suffering from anorexia and cachexia. Three-month-old nongrowing mice (C57BL/6J) were given s.c. transplants of a methylcholanthrene-induced sarcoma. Freely fed, starved, and pair-fed animals were used. Body and lipid composition, tumor growth, and survival time were measured. Freely fed sarcoma-bearing mice died with profoundly altered body composition. This was not explained by the anorexia assessed in pair-feeding experiments. Starvation had caused a more severe depletion in body composition in both tumor-bearing and nontumor-bearing animals than the tumor alone did in freely fed tumor-bearing mice. Freely fed tumor-bearing animals had normal proportions of whole-body triglycerides, cholesterol, and polar lipids, but they lost palmitic acid quantitatively more than any other fatty acid. It is unlikely that any single fatty acid became limiting during tumor growth. The results show that the overall availability of lipids, nitrogen, and glucose precursors is not a limiting factor for survival in experimental tumor cachexia. Other factors considered to be more likely as determining factors for the death of tumor-bearing animals are discussed.

The influence of the ambient temperature on tumour growth, metastasis and survival in mice.

The effect of ambient temperature was analysed on the behaviour of MO4 cells which are malignant, invasive and metastasizing C3H mouse fibroblastic cells when implanted in the tail of the syngeneic animal. Raising the ambient temperature to 28 degrees C increases growth of the primary tumour, incidence of metastases and shortens survival. Lowering the ambient temperature to 13 degrees C prevents all these phenomena. Measurements at the implantation site show that the temperature of the tail remains in a range +/- 2-3 degrees C in accordance with the environment. From these results we conclude that room temperature may be important when experiments are performed in vivo.

Experimental studies on the carcinogenicity of five nitrogen containing polycyclic aromatic compounds directly injected into rat lungs.

Using a beeswax/tricaprylin mixture as vehicle, three doses each of acridine, benz[a]acridine (BaAC), benz[c]acridine (BcAC), dibenz[a,h]-acridine (DBa,hAC) and dibenz[a,j]acridine (DBa,jAC) were injected into the lungs of 35 female Osborne-Mendel rats per group. To compare the carcinogenic potency of the heterocycles, benzo[a]pyrene (BaP) was taken as reference substance. Dose-response relationships were obtained for DBa,hAC as well as for BaP. DBa,jAC and BcAC exhibited no carcinogenic effects in the dose range from 0.1 mg to 1 mg tested in the lung implantation model. From the results presented here, it cannot be excluded that tumor development may occur when testing higher doses. BaP, however, must be considered much more potent since it revealed, by far, higher tumor incidences and diminished life times.

Bone sarcoma characteristics and distribution in beagles fed strontium-90.

A total of 66 primary bone sarcomas were diagnosed in 47 beagles; 43 of these dogs were part of the 403 beagles fed 90Sr and 4 were part of the 162 controls. Multiple primary bone sarcomas were found in 15 of the 47 beagles (32%). The incidence of multiple primary bone sarcoma was restricted to the two highest dose groups, except for a single control dog which developed two bone sarcomas. A threshold-like radiation dose response was observed; no sarcomas were observed in the lowest three dose groups, but the number of primary bone sarcomas increased rapidly in the higher dose groups. Of the 66 primary sarcomas, 49 were osteosarcomas (74%). As the dose increased, the proportion of osteosarcomas increased sharply, 4/10 (40%), 26/29 (90%), and 16/18 (89%), in the three highest dose groups. Thirteen of the bone sarcomas of other types occurred in males, and 4 in females, whereas 21 osteosarcomas occurred in males, and 28 in females. The ratio of bone sarcomas of the appendicular skeleton to those in the axial skeleton was 40:26, with osteosarcomas occurring more often in the appendicular than the axial skeleton (32:17), whereas nonosteogenic tumors showed no predilection (8:9). A statistical study of the distribution of bone sarcomas among 16 separate bone groups showed a correlation only with the distribution of cancellous bone volume-to-surface ratio and not with either skeletal mass distribution or dose distribution. The highest occurrence of sarcomas was in the humeri, femora, and mandible, and no occurrence in the coccygeal vertebrae, paws, or sternum. It is postulated that the distribution of bone sarcomas reflects a critical combination of the osteosarcoma precursor cell population, their cell division rate, and the radiation dose absorbed by these cells.

Cholera toxin pretreatment protects against tumor necrosis factor lethality without compromising tumor response to therapy.

Antitumor therapy with tumor necrosis factor is limited by systemic toxic effects. We studied whether cholera toxin, a bacterial exotoxin that adenosine diphosphate-ribosylates the alpha-subunit of Gs proteins, could separate the lethal from the antitumor effects of tumor necrosis factor. A single dose of intravenous cholera toxin protected non-tumor-bearing mice from a lethal dose of Escherichia coli endotoxin administered 6 or 24 hours later. On the basis of these results, tumor-bearing mice were randomized to receive either cholera toxin or saline, followed 6 hours later by either human tumor necrosis factor (400 micrograms/kg) or saline. Tumor-bearing mice pretreated with cholera toxin had (1) reduced treatment-related mortality (0/11 vs 5/11 for saline controls) and (2) tumor regression similar to that of controls. In a separate experiment in tumor-bearing mice, intravenous human tumor necrosis factor treatment induced an increase in serum levels of murine tumor necrosis factor to a peak of 500 pg/mL at 1 hour in saline-pretreated controls, while a similar increase could not be detected in those mice pretreated with cholera toxin. These results suggest that pretreatment with cholera toxin can reduce the endogenous tumor necrosis factor response to administered tumor necrosis factor and separate the lethal from the antitumor effects. Cholera toxin may prove to be a useful tool for investigating the mechanisms underlying the varied effects of tumor necrosis factor.

Effective regional therapy of experimental cancer with paralesional administration of tumour necrosis factor-alpha + interferon-gamma.

Systemically administered tumour necrosis factor (TNF) has anti-tumour effects in animal tumour models but its clinical application is limited by severe toxicity. Interferon-gamma(IFN-gamma) has been shown to augment the anti-tumour effect of TNF. We evaluated the effect of paralesional (p.I.) injections of TNF plus IFN-gamma in a murine tumour model and compared the toxicity and anti-tumour effect with that seen with systemic administration. C57BL6 mice with 10-day subcutaneous MCA sarcomas were treated with daily p.I. injections of recombinant huTNF +/- IFN-gamma for 5 days. Optimal mean survival and 30-day cure rate was seen with doses of 5 micrograms TNF-alpha + 5000 U IFN-gamma (P < 0.05 vs. control or IFN-gamma alone). Tumour response after a single i.v. injection of 0-15 micrograms TNF + 5000 U IFN-gamma was then compared with five daily p.I. injections of the same dose of TNF-alpha and IFN-gamma. All animals with p.I. injections of > 5 micrograms TNF had initial complete necrosis of tumour with a variable degree of surrounding tissue necrosis, with rapid regrowth of tumour seen in some animals. Although treatment-related mortality was similar between i.v. and p.I. therapy, there was a higher percentage of animals cured with p.I. injections with overall cure rates in treated animals at 30 days of 17% vs. 72% (i.v. vs. p.I., P < 0.01) and 13% vs. 67% (P < 0.04) in a repeat study. 2+ clinical applications.

Long-term effects of flavone acetic acid on the growth of a rat tumour.

A rat tumour (MC7 sarcoma), growing subcutaneously, has been shown to be sensitive to a single application of flavone acetic acid. Thirteen rats were still alive after 50 days and 8 of these were tumour free, as compared with control rats which survived for 15.7 +/- 2.53 days. The 8 tumour free animals were rechallenged with MC7 sarcoma 40 weeks later, without further FAA treatment. The tumour grew initially but in all cases the animals became tumour free within 24 days. After a further 30 days they were rechallenged with D23 hepatoma which grew as effectively as in the controls.

Isothiazolopyrimidines--new group of anticancer agents. II.

In reactions of 5-aminoisothiazolopyrimidines 1 with glycosilisothiocyanates and phenyl isocyano- or isocyanates, N-glycosilo-N-phenylo- and N-isothiazolopyrimidinothiocarbamides type II were obtained. The same compound--5-aminoisothiazolopyrimidine 1 in reaction with glucose, arabinose and ribose bromoderivatives was converted into N-glycosiloaminoisothiazolopyrimidies type 3. Some of the newly synthetized compounds appeared effective against L-1210 leukemia and Sa-180 sarcoma.

A virally induced osteosarcoma in rats. A model for immunological studies of human osteosarcoma.

Inoculation of Moloney sarcoma virus into the medullary canal of the tibia in newborn Wistar-Lewis rats resulted in an initially localized osteosarcoma which usually metastasized to the lung and resulted in the death of the animal within four to five weeks. Tumor cells were grown in tissue culture and used as target cells in the assay of lymphocyte-mediated cytotoxicity using a microcytotoxicity and a radioisotope labeling method. Lymphocyte-mediated cytotoxicity was demonstrated throughout the course of the clinical disease as well as in a small number of animals which showed spontaneous regression of their tumors. Serum factors which could "block" or augment the cellular response were also identified. This model resembles the spontaneous osteosarcoma of humans in many respects and may be useful for studies of the human disease.

The potential for radiation sensitizers and possible strategy for use.

The search for radiation sensitzers for use in radiotherapy is based on the assumption that proper use of these compounds will provide a differential sensitization of tumor and normal tissue. Based on the evidence that hypoxic cells exist within human tumors but not within normal cell populations, there is reason to believe that specific sensitizers of hypoxic cells might provide useful therapeutic benefit. This paper discusses the possible advantages of such sensitizers, over reliance on the phenomenon of reoxygenation, and describes very briefly the current status of hypoxic cell sensitizers. Finally a proposal is made concerning the way in which such sensitizers might be tested for their efficacy in human radiotherapy.

Esters of chlorambucil with 2-substituted 1,4-dihydroxy-9,10-anthraquinones as multifunctional anticancer agents.

Novel twelve esters of chlorambucil with 2-(1-hydroxyalkyl)-1,4-dihydroxy-9,10-anthraquinone were synthesized and tested for their antitumor activity in mice bearing S-180 ascitic cells as well as cytotoxic activity against L1210 cells. Eight of them were highly cytotoxic on L1210 cells (ED(50), <6 microg mL(-1)) and derivatives 1 and 12 (T/C, 200 and 205%) appeared more active in vivo than chlorambucil (T/C, 168%).

Lack of carcinogenesis by the baked mushroom Agaricus bisporus in mice: different feeding regimen [corrected].

Agaricus bisporus, the cultivated mushroom of the western hemisphere, was baked at 220-230 degrees C for 10 minutes and subsequently fed to mice for 12 hours each day, five days each week throughout their life. After each feeding cycle, the animals received a well-balanced semisynthetic diet for 12 hours each day for five days plus the remaining two full days each week. The estimated average daily mushroom consumption per animal was 4.8 g for a female and 4.2 g for a male. Randomly bred Swiss mice, six weeks old at the start of the experiment, were used. In the baked mushroom-fed group, the incidences of tumors in the lungs, blood vessels, cecum, and colon increased when compared to the untreated controls. These increases were not, however, statistically significant. In another previous experiment, both the raw and the baked mushrooms, when used in different feeding regimens, induced statistically significant incidences of cancers in several organs of the mice. It is possible that the negative finding in the current study was due to insufficient mushroom consumption.

Adverse effect of splenectomy on growth and survival with murine lymphosarcoma.

Recent reports suggest that splenectomy may improve host resistance and inhibit solid tumor growth. The effect of splenectomy on lymphoid tumors in less clear. This study evaluates and compares the effect of splenectomy on tumor growth, therapy and survival in murine lymphosarcoma and mammary tumor. Gardner lymphosarcoma (5 x 10(5) cells) was implanted subcutaneously into 400 6C3HED mice (20 g). Two hundred mice underwent splenectomy 10 days previously. Animals were randomly placed into four groups. Group I (tumor alone) and Group II (splenectomy and tumor) received no further therapy. Group III (tumor) and Group IV (splenectomy and tumor) received cyclophosphamide (50 mg/kg/day x 3 days) beginning 10 days after implantation. The rate of implantation was similar in all groups (greater than 90%). Tumor growth was localized in controls, but was widespread in splenectomized mice. Survival analysis at 30 days showed an increased mortality in untreated mice (Group II) following splenectomy (p less than .02). Survival was (13/102) 12.75% Group I versus (8/103) 7.77% Group II. Survival was similar in mice receiving chemotherapy (36%) and was (p less than .001) greater than the untreated groups (12%). A similar protocol in mice with mammary tumor showed no differences between groups in tumor localization or survival postsplenectomy. These data suggest that splenectomy adversely affects localization and survival in murine lymphosarcoma but not in solid tumor. The variable effect of this operation on the natural history of lymphoid versus solid neoplasia questions the advisability of splenectomy in staging of patients with lymphosarcoma.

[The effect of sodium nitrite on rats infected with avian sarcoma virus]. / Vplyv Dusitanu sodného na krysy infikované vtácím sarkómovým vírusom.

A potential effect of sodium nitrite was studied as exerted on carcinogenesis elicited by the avian sarcoma virus in the body of rats. In the case of the infection of the rats with indefective avian sarcoma virus B77 (Bratislava strain; ID50 10-25/0.1 ml) and i. p. inoculation of sodium nitrite, great mortality occurred already on the 20th day p. i. On the 125th day p. i. the cumulative mortality in this group was 76.92%; on the other hand, in the control groups (rats inoculated with sodium nitrite or avian sarcoma virus) no deaths occurred before this period. The rats infected with a double dose of the virus were not observed to show pronounced changes in mortality. It was only on the 125th day that cumulative mortality in the group of rats infected with the avian sarcoma virus and rats inoculated s. c. with sodium nitrite was 87.5% and in the group of animals which had only been infected with the virus the mortality was 64.26%. The patho-morphological changes were in keeping with the hitherto described changes in rats infected with the avian sarcoma virus. In the trial no effect of sodium nitrite on the patho-morphological changes induced by the avian sarcoma virus was observed.

[Use of a BCG vaccine from a Soviet substrain for the immunotherapy of tumors caused by SA7 (C8) virus]. / Ispol'zovanie vaktsiny BTsZh iz otechestvennogo subshtamma dlia immunoterapii opukholei, vyzvannykh virusom SA7 (C8).

The oncolytic and immunotherapeutic effect of the national BCG vaccine with respect to undifferentiated sarcoma of hamster induced by the highly oncogenic adenovirus of green monkeys SA7(C8) was studied. Preliminary data on significant positive effect of the vaccine inoculated before tumor formation were obtained. A single inoculation of the vaccine into tumors did not result in tumor regression in any of the cases.

[Therapeutic effect from the combined action of pyrogenal and irradiation on sarcoma 45 in rats]. / Terapevticheskii éffekt pri kompleksnom vozdeistvii pirogenala i oblucheniia na sarkomu 45 krys.

The use of pyrogenal associated with radiotherapy in the experiment (rat sarcoma-45) contributes to a successful realization of radiotherapeutic effect, longer life-terms and non-recurrence period, the increased number of completely cured and survived animals. Pyrogenal would produce no adverse side-effects.

[The inhibitory effect of ascorbic acid on the estrogen-stimulated promotion of uterine sarcoma development in mice]. / Tormoziashchii éffekt askorbinovoi kisloty na stimulirovannuiu éstrogenom promotsiiu sarkomogeneza matki u myshei.

The administration of estradiol propionate after the discontinuation of 1,2-dimethylhydrazine (DMH) treatment increased the incidence of uterine sarcoma in CBA mice from 32.5 (DMH alone) to 62.5%. The addition of the ascorbic acid (0.3% solution in drinking water) to estradiol propionate was followed by a decrease in the tumor incidence to 35%, i. e. the acid levelled the promoting effect of estradiol propionate almost completely. Sodium ascorbate did not share that effect. Mechanisms underlying the antiestrogenic effect of the ascorbic acid are discussed.