Targeted therapies for glioblastoma treatment.

Glioblastoma, the most common and aggressive type of brain tumor in adults, poses significant challenges in terms of treatment. Conventional approaches including surgery, chemotherapy, and radiotherapy have yielded limited success, with a median survival of approximately 15 months. However, extensive research into the biology of glioblastoma has identified molecular targets that can be exploited by newly developed drugs, leading to the emergence of precise personalized therapies. Several innovative treatment strategies are currently under development, aiming to enhance effectiveness while minimizing side effects. Clinical trials are underway to evaluate the efficacy of monoclonal antibodies that target glioblastoma cells, either by blocking specific receptors or by modifying molecular interactions that impede cell proliferation. Another promising avenue involves the use of oncolytic viruses designed to selectively infect glioblastoma cells. Additionally, the review explores the utilization of nanocarriers capable of surmounting the formidable obstacle of the blood-brain barrier, enabling efficient drug delivery. Cell therapies represent another promising approach, with dendritic cells, chimeric antigen receptor-T cells, and macrophages emerging as potential treatment modalities. By summarizing recent advances in targeted therapies against glioblastoma, this review aims to provide a comprehensive overview of ongoing efforts to discover effective and safe methods for treating glioblastoma patients. The ultimate goal is to improve patient outcomes and transform the landscape of glioblastoma treatment.

Effect of dietary canthaxanthin and iodine on the production performance and egg quality of laying hens.

In this study, we aimed to evaluate the effect of canthaxanthin (CX) and iodine (I) on the production of laying hens, on counteracting debilitation of the vitelline membrane, and on inhibiting Salmonella growth in eggs stored at 30°C. Three hundred hens were reared in cages. Birds were divided into six feeding groups (10 hens × 5 repetitions) that were administered 0, 3 or 6 ppm of CX and 1 or 10 ppm of I with their diets. Laying rate, egg weights, and feed conversion ratios were controlled. The quality of fresh eggs was assessed in wks 25-26, 48-50 and 62-63 of hens lives. An additional batch of eggs was incubated at the temperature of 30°C, and egg quality changes were monitored on days 3, 6 and 9 of storage. Additionally, eggs collected from four experimental groups of hens whose diets had been iodated with 1 or 10 ppm of I and supplemented with 0 or 6 ppm of CX were infected under laboratory conditions with Salmonella, and incubated for 5 and 10 d. The laying rate, egg weights, and feed conversion ratio were significantly improved. Dietary inclusion of CX contributed to a higher resistance of the vitelline membrane of egg yolks, but only for fresh eggs. Vitelline membrane degradation during egg storage at 30°C was significantly counteracted by dietary inclusion of I at a dose of 10 ppm. The same I dose resulted in the complete inhibition of Salmonella growth until day 10 of incubation, but exclusively for eggs collected from 40-week-old hens. Dietary supplementation with 10 ppm of I was found to impart high level of resistance to the vitelline membrane against the growth of Salmonella in case of eggs stored at 30°C; therefore, I was found to be more beneficial by ensuring longer preservation than that of CX. However, dietary supplementation with CX was found to increase the resistance of vitelline membrane in fresh eggs.

Rapid identification of Helicobacter pylori by capillary electrophoresis: an overview.

Helicobacter pylori can cause gastritis and peptic ulcers and is directly associated with the development of gastric cancer. There are many types of diagnostic methods used to identification H. pylori (invasive and non-invasive), but these methods usually require time-consuming and laborious procedures and therefore are not capable of fast diagnosis in cases of emergency. This contribution describes the new achievements, interdisciplinary significance and some future directions in the application of capillary electrophoresis for determination of H. pylori.

Preliminary study of volatile organic compounds from breath and stomach tissue by means of solid phase microextraction and gas chromatography-mass spectrometry.

The determination of volatile organic compounds (VOCs) in exhaled air and stomach tissue emission for the detection of cancer has been investigated. Solid phase microextraction (SPME) was used for sample preconcentration. The method presented in this paper showed satisfactory precision (RSD below 11%), linearity in the range of 2.8-136 ppb and limit of detection ranging from 0.6 to 2.1 ppb. The breath and emission from cancer tissue were collected from three patients with stomach cancer. Acetone, carbon disulfide, 2-propanol, ethyl alcohol and ethyl acetate were identified in breath and tissue samples. These compounds have been assumed as endogenous. Acetone ratio (AR) was calculated for carbon disulfide, 2-propanol and n-butane. The AR for carbon disulfide was found to be higher for normal tissue (20.64-44.95) than for emission from cancer tissue (2.01-18.20). A limitation of this study is that only a few clinical samples were investigated. These results should be evaluated as preliminary because of the small number of patients examined.

Influence of dendritic cells on the in vitro allogeneic cytotoxic reaction of lymphoid cells derived from normal or Listeria innocua-infected BALB/c mice.

The role of lymphoid dendritic cells (DCs) in the development of an allogeneic cytotoxic reaction in vitro was examined. The T+B and T cell subsets originating from the spleens or lymph nodes of normal and Listeria innocua-infected BALB/c mice were used as the effector cells. Their cytotoxicity to 51Cr-labeled C3H fibroblasts was determined after removal of DCs and replacing them again. Moreover, the influence of exogenous mrIL-12 on the potency of DCs in the allogeneic reaction developed in vitro was checked. It was found that the DC-deprived T+B or T subsets of splenocytes, regardless of their origin, exhibited 27-38% lower cytotoxicity than those accompanied by natural DCs. The cytotoxicity of these subsets from normal lymph nodes decreased by 22%, while the activity of bacteria-primed cells dropped by 38%. Replenishing effector cells with isolated DCs restored their cytotoxicity. Pulsation of normal DCs with IL-12 had no effect on the recovery of normal cell cytotoxicity. However, the IL-12-pulsed DCs were able to intensify the cytotoxicity of T+B subsets derived from the spleens or lymph nodes of L. innocua-infected mice. The results suggest that the alloantigen presentation by DCs to cytotoxic lymphocytes also takes place in the reaction developed in vitro, regardless of effector cell origin.

Quantitative reactions of anti 5,9-dimethylchrysene dihydrodiol epoxide with DNA and deoxyribonucleotides.

Native as well as denatured calf thymus DNA, deoxyguanylic and deoxyadenylic acid, respectively, were reacted with the racemic anti 5,9-dimethylchrysene dihydrodiol epoxide (5,9-DMCDE). The deoxyribonucleoside adducts were separated by HPLC and characterized by CD and NMR. Approximately 17% of the epoxide was trapped by native DNA and 76% of the adducts were derived from the RSSR enantiomer. The ratios of dAdo/dGuo modification in DNA were 14/86 and 19/81 for RSSR and SRRS enantiomers, respectively. By monitoring the product yields of anti 5,9-DMCDE with DNA and deoxyribonucleotides, we hoped to gain further insight into the factors responsible for deoxyguanosine adduct formation by 5-methylchrysene dihydrodiol epoxide (5-MCDE) compared to 5, 6-dimethylchrysene dihydrodiol epoxide (5,6-DMCDE). The adduct yields in deoxyribonucleotide reactions of 5,9-DMCDE were slightly higher than those from 5-MCDE. However, the reaction yields of 5, 9-DMCDE with DNA were lower than those with 5-MCDE in most cases, particularly for the cis and trans deoxyadenosine adducts. It seems that the 9-methyl group of 5,9-DMCDE significantly influences adduct formation with the deoxyadenosine residue in DNA in contrast to the 6-methyl group of 5,6-DMCDE. The 9-methyl group sterically decreases deoxyadenosine adduct yields more in reaction with native DNA than denatured DNA, but it has little effect on deoxyribonucleotide reactions. Adduct formation with deoxyguanosine residues in DNA by all three dihydrodiol epoxides correlate with their respective tumorigenic and mutagenic activities.

Reactions of dihydrodiol epoxides of 5-methylchrysene and 5, 6-dimethylchrysene with DNA and deoxyribonucleotides.

Both syn and anti dihydrodiol epoxides from 5-methylchrysene (5-MCDE) and 5,6-dimethylchrysene (5,6-DMCDE) were reacted under the same conditions with native DNA, denatured DNA, and purine deoxyribonucleotides, and the products were quantified. The extents of reaction with the deoxyribonucleotides were consistently greater for 5,6-DMCDE than for 5-MCDE. The yield of adducts in the reaction with DNA ranged from being a few-fold to 50-fold greater than those found in the corresponding deoxyribonucleotide reactions for both 5-MCDE and 5,6-DMCDE. The DNA-dependent enhancement of product yield was greater for 5-MCDE than for 5,6-DMCDE with a few exceptions among cis and trans deoxyadenosine adducts. The most substantial differences in DNA-dependent enhancement were found for deoxyguanosine adducts; thus, steric hindrance between the 6-methyl group in the 5,6-DMCDE and the minor groove in the DNA double helix may account for the greater DNA-dependent enhancement found in the 5-MCDE reactions.

DNA reactions, mutagenic action and stealth properties of polycyclic aromatic hydrocarbon carcinogens (review).

A brief summary of recent research, primarily from the authors' laboratory, on polycyclic aromatic hydrocarbon carcinogens with respect to their DNA adduct formation, the mutational properties of these adducts and the effects of hydrocarbon dihydrodiol epoxide metabolites on the passage of cells through the cell cycle is presented. The concept of stealth properties of potent carcinogens, i.e. their ability to damage DNA without inducing a G1 arrest, is discussed. Also, mutation studies with dihydrodiol epoxide metabolites, the sequence-dependence of site-specific mutation, as well as the selectivity of hydrocarbon-DNA adduct formation are summarized.

Application of mixed mobile phases and a step gradient method in capillary electrochromatography for the separation of isomeric polycyclic aromatic hydrocarbon-deoxyribonucleoside adduct mixtures prepared in vitro.

Capillary electrochromatography (CEC) was used for the analysis of mixtures of neutral isomeric compounds derived from the reaction of carcinogenic hydrocarbon (benzo[g]chrysene and 5,6-dimethylchrysene) dihydrodiol epoxides with calf thymus deoxyribonucleic acid (DNA). The CEC analysis demonstrated higher resolution, greater speed and lower analyte consumption than high-performance liquid chromatography (HPLC) in the analysis of the same samples using the same type of stationary phase. Proper selection of the mixed mobile phases was critical for the separation of these complex mixtures with enhanced speed and selectivity. The use of a step gradient further improved the speed of the CEC analysis resulting in electrochromatograms that required only 25-70% of the corresponding HPLC analysis times.

Characterization of DNA adducts formed by the four configurationally isomeric 5,6-dimethylchrysene 1,2-dihydrodiol 3,4-epoxides.

The DNA adducts formed from the racemic syn and anti dihydrodiol epoxides of 5,6-dimethylchrysene were characterized through various spectroscopic methods. Substantial reaction with the amino groups of both deoxyadenosine and deoxyguanosine residues were detected with both the syn and anti derivatives. The chemical shifts and coupling constants for the cis and trans opened adducts from the syn dihydrodiol epoxide were distinctly different, whereas for the anti dihydrodiol epoxide these properties were fairly similar for cis and trans adducts. In the latter case, assignment of trans and cis configurations was less obvious, and the finding that trans adducts have always predominated over cis adducts for all dihydrodiol epoxides studied to date was helpful in making these assignments. The preferential formation of cis adducts in DNA by the syn dihydrodiol epoxide is more like the chemistry of the dihydrodiol epoxide of benzo[c]phenanthrene than of benzo[g]chrysene, although both of these, like 5,6-dimethylchrysene, are non-planar compounds.

Interleukin 12 and direct cytotoxicity of spleen lymphocytes to Listeria innocua-phagocyting syngeneic macrophages in C57BL/6 and BALB/c mice.

Production of interleukin 12 (IL-12) during cytotoxic reaction of C57BL/6 and BALB/c spleen lymphocytes or their subpopulations: T+B, T, CD4+ and CD8+ cells to L. innocua-phagocyting syngeneic macrophages was examined. The effector cell donors were untreated or L. innocua-infected. The number of surviving bacteria in phagocytes was tested and IL-12 level in culture supernatants of reacting cells was determined. C57BL/6 mice, resistant to Listeria infection, were found to develop stronger cell cytotoxicity to bacteria-phagocyting syngeneic macrophages than BALB/c mice. The lymphocytes responsible for that phenomenon were of CD8+ phenotype. IL-12 was produced only during nonspecific cytotoxic reaction of CD4+ and CD8+ T cells. It is suggested that the innate resistance of mice to Listeria is dependent on their ability to develop a specific cytotoxic reaction and IL-12 production.

The host response to Listeria monocytogenes mutants defective in genes encoding phospholipases C (plcA, plcB) and actin assembly (actA).

Several genes involved in the determination of Listeria monocytogenes pathogenesis have been identified. Among them, plcA gene encodes phosphatidylinositol-specific phospholipase C (PI-PLC), plcB gene encodes a broad-range phospholipase C (PC-PLC), and actA encodes a protein contributing to actin assembly in infected cells. The interaction of L. monocytogenes wild type (LO 28) strain and two derivative mutants, plcA- (BUG 206) and actA-/plcB- (LUT 12), with macrophages and T lymphocytes was investigated in a mouse model of listeriosis. Both mutants showed evidence of attenuation. The plcA- mutant, but not the plcB- mutant, expressed an increase in susceptibility to the anti-listerial activity of macrophages. Both mutants showed a decreased ability to induce IL-12 production by bone marrow macrophages when co-stimulated with E. coli LPS or IFN-gamma. In vivo, L. monocytogenes plcA- mutant was found to be a more effective stimulator of T cells than the wild LO 28 strain.

Characterization of the adjuvant effect of IL-12 and efficacy of IL-12 inhibitors in type II collagen-induced arthritis.

A destructive joint disease can be induced in susceptible DBA/1 mice by immunization with type II collagen emulsified with oil and either killed Mycobacterium tuberculosis or IL-12 as adjuvant. Cellular and humoral anti-collagen immune mechanisms appear to be involved in the pathogenesis of arthritis. We have characterized the adjuvant effect or IL-12 in more detail and addressed the question whether mycobacteria might act via the induction of endogenous IL-12. Injections of IL-12 into collagen-immunized DBA/1 mice promoted the development of IFN-gamma-producing CD4+ T cells and strongly upregulated the production of complement-fixing IgG2a and IgG2b antibodies resulting in severe arthritis. Neutralization of IFN-gamma in vivo largely inhibited the increase in antibody synthesis and prevented joint disease in IL-12-treated mice. However, collagen-specific IFN-gamma synthesis by T cells was further enhanced in these animals. Furthermore, IL-12 treatment promoted the development of IFN-gamma-producing T cells but failed to enhance antibody synthesis and to induce arthritis in C57BL/6 or BALB/c mice immunized with collagen in oil. These results indicate that the induction (by IL-12) of a strong collagen-specific T-cell response alone is not sufficient to trigger arthritis. Attempts to show a role for endogenous IL-12 in DBA/1 mice immunized with collagen with mycobacteria as adjuvant gave no reliable results. Whereas anti-IL-12 treatment delayed the onset and ameliorated the disease in some experiments, it failed to do so in other experiments, or, control reagents also had some effect. A slight inhibition of collagen-specific IgG2a synthesis was observed in most experiments in the sera of anti-IL-12-treated mice. Taken together, the results show that exogenous IL-12 can promote arthritis via its direct effect on T cells and its effect on antibody production, which is at least in part IFN-gamma-dependent. On the other hand, whether or not endogenous IL-12 is involved in the adjuvant effect of mycobacteria needs further clarification.

IL-12 promotes cellular but not humoral type II collagen-specific Th 1-type responses in C57BL/6 and B10.Q mice and fails to induce arthritis.

DBA/1 (H-2q) and C57BL/6 (H-2b) mice develop an intermediate immune responses when immunized with chicken type II collagen (CII) emulsified with incomplete Freund's adjuvant (IFA). Only a few animals develop a mild form of arthritis. As reported before and confirmed herein, administration of IL-12 to DBA/1 mice immunized with CII in IFA strongly enhances the cellular and humoral (auto)immune response to CII and induces severe destructive joint disease with an incidence of 80-100%. In contrast, the same treatment did not promote joint disease in C57BL/6 mice. Characterization of the IL-12 effect on the CII-specific immune response of C57BL/6 mice revealed that IL-12 promoted the development of CII-specific T cells producing IFN-gamma in DBA/1 and C57BL/6 mice equally well. However, whereas treatment with IL-12 in DBA/1 mice strongly up-regulated the synthesis of CII-specific antibodies, especially of the IgG2a and IgG2b subclasses, it rather slightly down-regulated the CII-specific IgG2a and IgG2b synthesis in C57BL/6 mice. This may indicate that the effect of IL-12 on the CII-specific antibody synthesis is of crucial importance in the pathogenesis of type II collagen-induced arthritis (CIA). The failure of IL-12 to up-regulate IgG2a and IgG2b synthesis in C57BL/6 mice is specific for CII as antigen and not a general property of this strain because the keyhole limpet hemacyanin-specific antibody response is up-regulated by IL-12 in C57BL/6 mice. Furthermore, it is not the H-2b haplotype of C57BL/6 mice but rather the genetic background (DBA/1 versus BL/6 or BL/10) that limits the effect of IL-12 on the CII-specific antibody response because IL-12 treatment of CII-immunized B10.Q (H-2q) mice also failed to induce arthritis and to enhance CII-specific IgG2a and IgG2b synthesis. However, as in the two other strains, injection of IL-12 promoted the development of splenic T cells producing IFN-gamma upon activation with CII. These results indicate that an enhancement of the cellular and humoral anti-CII response by IL-12 is required for inducing arthritis.

High doses of interleukin-12 inhibit the development of joint disease in DBA/1 mice immunized with type II collagen in complete Freund's adjuvant.

Collagen-induced arthritis (CIA) is an (autoimmune) joint disease readily elicited in DBA/1 mice by immunization with type II collagen (CII) emulsified with complete Freund's adjuvant. It is a destructive arthritis involving about 50% of the limbs and occurs with an incidence of 70% to 100%. In this study we evaluated the effect of mouse recombinant interleukin-12 (mrIL-12) on CIA. Administration of mrIL-12 at high doses (1 micrograms/mouse, daily) for 2 or 3 weeks delayed the onset and reduced the incidence of CIA. Furthermore, the severity of CIA was much milder and in most cases restricted to single digits of the paws. Short-term administration of high doses of IL-12 exerted some, but less pronounced, disease-suppressing effect. In contrast, 10-fold lower doses of IL-12 given during the first 3 weeks, or high doses of IL-12 administered therapeutically proved to be ineffective. Only those regimens of IL-12 treatment that ameliorated CIA were associated with a down-regulation of the CII-specific antibody response. A strong inhibition of CII-specific IgG1 antibodies (10- to 20-fold) and a moderately (2- to 6-fold) suppressed IgG2b response was observed, whereas the level of CII-specific IgG2a antibodies remained high. Taken together, the results indicate that some initial events in the induction of CIA in DBA/1 mice injected with CII emulsified with CFA are suppressed by treatment with high doses of IL-12.

Characterization of DNA adducts formed by anti-benzo[g]chrysene 11,12-dihydrodiol 13,14-epoxide.

The anti-11,12-dihydrodiol 13,14-epoxide of benzo[g]chrysene, a fjord-region-containing hydrocarbon, was found to react with DNA in vitro to yield, as the major product, an adduct in which the epoxide of the 11R, 12S, 13S, 14R enantiomer was opened trans by the amino group of deoxyadenosine. The structures of this adduct and other deoxyadenosine and deoxyguanosine adducts were established by spectroscopic methods. In reactions with deoxyguanylic acid, a product tentatively identified as a 7-substituted guanine was also detected. The mutagenic properties of this dihydrodiol epoxide in shuttle vector pSP189 showed that mutation at AT pairs accounted for 39% of base change mutations whereas chemical findings indicated that about 60% of adducts formed in calf thymus DNA involved adenines. Since calf thymus DNA is 56% AT and the target supF gene is 41% AT, the findings represent a fairly close relationship between adduct formation and mutagenic response. Overall, the chemical and mutagenic selectivities for the two purine bases in DNA were similar, though not identical, to those for the only other fjord-region-containing hydrocarbon studied in depth, i.e., benzo[c]phenanthrene. A major difference for these two hydrocarbon derivatives, however, is that benzo[c]phenanthrene dihydrodiol epoxides react to much higher extents (approximately 4-fold) with DNA than did the benzo[g]chrysene derivative.

Administration of interleukin 12 in combination with type II collagen induces severe arthritis in DBA/1 mice.

The induction of arthritis in DBA/1 mice usually requires immunization with the antigen type II collagen emulsified with Mycobacterium tuberculosis in oil. Here we describe that interleukin 12 (IL-12) can replace mycobacteria and cause severe arthritis of DBA/1 mice when administered in combination with type II collagen. Immunization of DBA/1 mice with type II collagen emulsified in oil alone resulted in a weak immune response, and only a few animals (10-30%) developed arthritis. Administration of IL-12 for 5 days simultaneously with each immunization strongly enhanced the anti-type II collagen immune response. Collagen-specific interferon gamma (IFN-gamma) synthesis by ex vivo activated spleen cells was enhanced 3- to 10-fold. IFN-gamma was almost completely produced by CD4+ T cells. Furthermore, the production of collagen-specific IgG2a and IgG2b antibodies was upregulated 10- to 100-fold. As a consequence, the incidence of arthritis in the group of mice immunized with collagen plus IL-12 was very high (80-100%). The developing arthritis was severe, involving approximately 50% of all limbs with strongly increased footpad thickness in most cases. Furthermore, histological examination revealed massive, mainly polymorphonuclear cell infiltration, synovial hyperplasia, cartilage and bone destruction, as well as new bone formation. In many cases, this resulted in the complete loss of joint structure. Neutralization of IFN-gamma in vivo prevented the development of arthritis in collagen-immunized and IL-12-treated mice. In conclusion, our data show that in vivo administered IL-12 can profoundly upregulate a T helper I-type autoimmune response, resulting in severe joint disease in DBA/1 mice.

Mutational spectra for 5,6-dimethylchrysene 1,2-dihydrodiol 3,4-epoxides in the supF gene of pSP189.

Dihydrodiol epoxides from 5,6-dimethylchrysene exhibit properties similar to those of fjord region-containing hydrocarbon derivatives in that they react extensively with deoxyadenosine residues in DNA and consequently generate substantial numbers of mutations at AT pairs as well as GC pairs. The syn-dihydrodiol epoxide favors reaction with deoxyadenosine (68% of adducts) to a greater extent than does the anti-dihydrodiol epoxide (52% of adducts), and point mutations at AT pairs (72% for syn- and 45% for anti-dihydrodiol epoxide) follow the same trend. A novel feature of the mutagenicity of the 5,6-dimethylchrysene derivatives is that they exhibit a higher fraction of AT-->GC transitions (28% and 26% for syn and anti, respectively) than has been seen for other hydrocarbon derivatives to date.

Participation of IL-2 and direct cell cytotoxicity in the modulation of GvH reaction in mice infected with Listeria innocua.

Listeria innocua can intensify the development of local GvH reaction in a semiallogeneic system in mice. This phenomenon is observed in (BALB/c x AKR)F1 mice intraperitoneally injected with the live bacteria on the 7th day before the transfer of BALB/c spleen cells. The local GvH reaction develops as strongly as in normal hybrid recipients given the parental cell graft in the presence of exogenous interleukin 2 (IL-2). The development of the reaction is associated with an intensive increase of the direct cytotoxicity of lymph node cells which is also markedly enhanced after the bacteria injection itself. IL-2 seems to play a relevant role in the development of local GvH reaction in L. innocua-injected mice.