This dissertation mainly deals with some biochemical aspects regarding the efficacy of novel platinum anticancer compounds, as part of a broader study in which both chemistry and biochemistry are involved. Various novel diamine and N-S donor chelate compounds of platinum II and IV have been developed in which factors such as stereochemistry, ligand exchange rate and biocompatibility were considered as additional parameters. In the first order testing, each of these compounds was tested with reference to their “killing” potential by comparing their rate of killing, over a period of 48 hours with those of cisplatin and oxaliplatin. Some 80 compounds were tested in this way. Although only a few could be regarded as equal to or even better than cisplatin and oxaliplatin, the testing of these compounds on cancer cells provided useful knowledge for the further development of novel compounds. Four of the better compounds, namely Y9, Y14, Y16 and Lt16.2 were selected for further studies to obtain more detailed knowledge of their anticancer action, including some flow cytometric studies. In addition to the above, cisplatin resistant cells were produced for each of the three different cell lines tested, namely, HeLa, HT29 and MCF7 cancer cell lines, by intermittent and incremental exposure to cisplatin (all the cell lines tested became resistant to cisplatin). Each of the selected compounds were exposed to the cells in the same manner, in order to attempt the induction of resistance against these compounds in the three cell lines tested (i.e. whether these cells will become resistant to the various compounds). Each of these selected platinum containing compounds were subsequently tested against the “cisplatin resistant” cell lines in order to determine their efficacy against such cells. One such compound could be singled out, since cervical cancer cells (HeLa cells) do not become resistant to it. This behaviour is similar to that of oxaliplatin against cervical cancer and colon cancer (HT29) cells (oxaliplatin is the number one treatment for colon cancer at present). This compound also proved to be more active against cisplatin resistant cell lines. It was found that all the compounds induced apoptosis in the cell lines tested as well as inhibit the DNA cycle at one or more phase. Finally, an effort was made to evaluate the different compounds by comparing them with respect to their properties relating to anticancer action.

The progressive growth of tumors in human cancer patients and experimental animals has frequently been associated with a generalized depression of immunological responsiveness. Suppressor cells have been implicated as mediators of tumor-associated immunosuppression, but the identities of the cells causing suppression and the mechanisms by which they act have been unclear. The object of this thesis was thus to determine: if suppressor cells capable of non specifically suppressing immune responses were present in anergic mice bearing methylcholanthrene-induced sarcomas; the cell types responsible for suppression; and the mechanisms by which suppression occurs. The spleens of mice with large tumors were found to contain two distinct populations of non specific suppressor cells. One population inhibited the proliferative responses of normal lymphocytes to the T cell mitogen, Concanavalin A (Con A) and the B cell mitogen, lipopolysaccharide (LPS). These cells also inhibited the generation of antibody forming cells by normal lymphoid cells stimulated in vitro with the T cell dependent antigen, sheep red blood cells (SRBC) and the T cell independent antigen, dinitrophenylated-lipopolysaccharide (DNP-LPS). These suppressor cells appeared to be from the macrophage/monocyte line since they adhered to plastic and nylon wool, were removed by carbonyl iron and magnet, and were inactivated by carragheenan treatment, but were not removed by anti-Thy-1 or anti-mouse immunoglobulin sera plus complement. They were among the less dense spleen cells since they were retained in the light fraction after centrifugation on hypaque-ficoll of specific gravity 1.08 and did not appear to require cell division in order to suppress- since mitomycin C treatment did not inactivate them. Cell-cell contact appeared to be essential for suppression. The second population of suppressor cells, which pelleted to the bottom of a hypaque-ficoll gradient, inhibited only the generation of plaque forming cells to the T cell dependent antigen, SRBC. These cells appeared to be T cells since they were non adherent to plastic or nylon wool, were not removed by carbonyl iron and magnet, but were removed by anti-Thy-1 serum plus complement. Cell division was necessary since suppressive activity was totally removed by mitomycin C treatment. Suppression by this cell type appeared to be mediated by a soluble factor with a molecular weight of about 3,500 to 12,000. / Science, Faculty of / Microbiology and Immunology, Department of / Graduate

Immune evasion is an emerging hallmark of cancer. Dysregulation of the major histocompatibility complex class I (MHC-I) is a frequent mechanism of immune evasion utilized by tumor cells and is particularly relevant to the pathogenesis of B-cell lymphomas, including diffuse large B-cell lymphoma (DLBCL) and classical Hodgkin lymphoma (cHL). A better understanding of MHC-I dysregulation in B-cell lymphomas is necessary to identify factors related to the risk, development, and progression of these tumors. In this thesis, we investigate the role of MHC-I dysregulation in DLBCL and cHL through the application of computational approaches to study genomic data. First, we introduce some background information about the normal function of MHC-I in the immune response to cancer and viral infection as well as the phenomenon of MHC-I dysregulation in the context of cancer. We provide an overview of how factors such as germline zygosity of HLA class I (HLA-I) genes and somatic alterations in the genes B2M and HLA-I that encode the protein subunits of MHC-I contribute to the development of DLBCL and cHL. Second, we present a study of the effects of HLA-I allele zygosity on survival in a cohort of 519 DLBCL patients treated with R-CHOP immunochemotherapy stratified by molecular subtype. Homozygosity in HLA-I was associated with a worse overall survival in patients whose tumors were classified in the “EZB” subtype, associated with somatic mutation in the epigenetic regulator EZH2. We find an association between the zygosity of the genes HLA-B and -C specifically and overall survival in EZB-DLBCL. These findings indicate that HLA-I zygosity may be a risk factor for worse clinical prognosis in patients with the EZB subtype of DLBCL. Third, we present a study of the genetic landscape of cHL tumors that are associated with infection with Epstein-Barr virus (EBV). We analyze inherited HLA-I allele types, somatic mutations, copy number changes, and mutational signatures in a cohort of 57 cHL patients (15 EBV-positive). We find that EBV-positive cHL is genetically distinct from EBV-negative cHL and is characterized by lower somatic mutation load and different activities of mutation signatures. Further, we find that cHL tumors are characterized by different patterns of MHC-I dysregulation depending on the EBV infection status. Germline homozygosity in HLA-I was associated with the EBV-positive subtype of cHL, while somatic alterations in HLA-I were associated with the EBV-negative subtype of cHL. These results suggest that inherited HLA-I homozygosity may be a risk factor for the EBV-positive subtype of cHL. Fourth, we expand our study of HLA-I in virus-associated cHL to perform a comparative analysis of virus-positive and virus-negative tumors in nine cancers linked to five viruses. We find that virus-positive tumors occur more frequently in males and show geographical disparities in incidence. Genomic analysis of 1,658 tumors reveals virus-positive tumors exhibit distinct mutation signatures, recurrent mutations in the RNA helicases DDX3X and EIF4A1, and a lower somatic mutation burden compared to virus-negative tumors of the same cancer type. We find that germline homozygosity in HLA-I is a potential risk factor for the development of EBV-positive cHL, but not other common virus-associated solid or hematological malignancies. Finally, we present in the Appendix a study of the genomic characterization of plasmablastic lymphoma (PBL), a rare EBV-associated B-cell lymphoma that occurs in the context of immunodeficiency caused by human immunodeficiency virus (HIV) infection. We find that PBL is characterized by mutations leading to constitutive activation of the JAK-STAT pathway. We additionally identify recurrent mutations in immune-related genes, such as B2M. These findings indicate a potential role for MHC-I and immune dysregulation in the pathogenesis of other B-cell lymphomas.

The human pancreatic cancer cell line T3M4, is known to produce transforming growth factor-alpha (TGF-alpha); as well as overexpress the receptor for this ligand, epidermal growth factor (EGF) receptor. TGF-alpha messenger RNA (mRNA) levels were assayed using northern blot, after addition of epidermal growth factor or TGF-alpha. The level of TGF-alpha mRNA was found to increase 2-fold at 2 hours and then return to near basal levels at 10 hours, after treatment with either ligand. Both ligands were also equipotent in a 2 hour dose response assay, with half maximal stimulation seen at 1 nM and maximal stimulation reached at 4 nM. Furthermore, there appeared to be a 2-fold increase in TGF-alpha transcription as determined by nuclear runoff experiments. Induction of TGF-alpha mRNA coupled with the overexpression of the EGF receptor, may result in a potent autocrine cycle; which may be found in other cancers.

A model for the study of the relationship of immunity to cancer is found in AKR mice which harbor Gross virus. This genetically transmitted virus is present in a latent form for months before it spontaneously induces leukemia. Many investigators have demonstrated near normal humoral responses, but abnormal cellular immunity in the preleukemic animal. With increasing age, pathology of the disease is expressed, reflecting diminished immunity. In this study, the ontogeny of humoral antibodies of AKR/J and SWR/J mice was assayed by microagglutination techniques in response to thymus-independent, thymus-dependent, and solubilized antigens. Simultaneous injections of thymusdependent and -independent antigens provided data suggesting an impaired humoral response in the AKR mouse.

Extracellular stimuli may influence the M1/M2 phenotypic polarization of macrophages. We examined M1/M2 biomarkers, phagocytic activity, and tumoricidal activity in RAW 264.7 mouse macrophages. Macrophages were treated with conditioned media (CM) from 4T1 breast cancer cells, curcumin, 22-oxacalcitriol, LPS, or a combination of the previously listed. Arginase activity, a M2 phenotypic biomarker, was upregulated by the treatment of macrophages with conditioned media. Curcumin, 22- oxacalcitriol, and LPS partially inhibited RAW 264.7 arginase activity in the presence of 4T1 breast cancer media. 22-oxacalcitriol increased the phagocytic ability of RAW 264.7 macrophages in the presence of M2 polarizing substances produced by the 4T1 breast cancer cells. Also, LPS increased RAW 264.7 phagocytic ability in the presence of 4T1 breast cancer CM. This study looked at the potential substances that would possibly reverse the M2 tumor promoting macrophage phenotype seen in the breast cancer tumor environment. / Includes bibliography. / Thesis (M.S.)--Florida Atlantic University, 2017. / FAU Electronic Theses and Dissertations Collection

Graduation date: 2006

A foundation for the control of tumors is presented, based upon the formulation of a realistic, knowledge-based mathematical model of the interaction between tumor cells and the immune system. The parametric control variables relevant to the latest experimental data, e.g., the sigmoidal dose-response relationship and Michaelis-Menten dynamics, are also considered. The model consists of 12 states, each composed of first-order, nonlinear differential equations based on cellular kinetics and each of which can be modeled bilinearly. In recent years a great deal of clinical progress has been achieved in the use of optimal controls to improve cancer therapy patient care. For this study, a cancer immunotherapy problem is investigated in which the aim is to minimize the tumor burden at the end of the treatment period, while penalizing excessive administration of interleukin-2 as a limit of toxicity. The optimal solution developed for this investigation is a mixture of an initially large dose of interleukin-2, followed by a gradually decreased dosage and a continuing infusion to maintain the tumor cell population at its allowable limit. Sensitivity analysis is applied to an investigation of the influences of system parameters. It has been found that the immune system is influenced greatly by several parameters such as macrophage level, tumor killing rate, tumor growth rate, and IL-2 level. The simulation results suggest that parametric control variables are important in the destruction of tumors and that the application of exacerbation theory is a good method of tumor control. / Graduation date: 1991

Predictors of response of AIDS-associated Kaposi-Sarcoma to standard chemotherapy Overview: Kaposi Sarcoma is the most common HIV-associated cancer. Its etiology and pathogenesis is not fully understood. Little is known about what predicts prognosis, survival and therapeutic response in HIV-KS. In South Africa given the high seroprevalence rates of HIV-l and human herpes virus 8 (HHV 8), Kaposi's sarcoma is a significant problem. The majority of patients have been treated solely with palliation due to the poor outcome associated with a diagnosis of HIV-KS, more so in the absence of highly active antiretroviral therapy (HAART). Since the national ARV rollout programme and the availability and accessibility of HAART to all patients with a diagnosis of HIV-KS, a new strategy has to be established to enable adequate patient selection for chemotherapy. There have been a few published studies addressing the predictors of response to chemotherapy in the first world. However, this is the first study of these factors in HIV-l infected African patients with Kaposi's sarcoma. Aim: To identify and assess the potential value of several parameters predictive of outcome, survival and therapeutic response in HIV- infected patients with KS. Clinical, hematological, biochemical, immunological and virological variables were evaluated. Methods: We collected data from 25 patients with AIDS-KS who were enrolled in a phase III randomized controlled trial comparing HAART alone with the combination of HAART and chemotherapy. All patients were from the combination therapy arm. The following variables were evaluated as predictors of prognosis and therapeutic response: age, gender, ethnic origin, Haemoglobin (Hb), white blood cells (WBCs), lymphocytes, neutrophils, platelets, S.albumin, ALP, GGT, CD4 count, HIV viral load. These variables were assessed in patients at baseline and month 6 of therapy. Patients were staged into good risk and poor risk according to the AIDS clinical trial group (ACTG) criteria. The outcomes assessed were response to treatment and mortality. Results: A total of 25 patients participated to the study. Of these 16(64%) were males and 9(36%) were females, with male: female ratio of 2.7:1. Median age was 34 years (24-47); all patients were of Black African origin. Of the 21 patients, 15 (71.4%) were of good prognosis and 6(28.6%) were of poor prognosis. At baseline the median values of the different variables were as follows: Hb 10.9 g/dl, WBCs 5.95x109/L, lymphocytes 1.7 x109/L, neutrophils 3 x10 9 /L, platelets 272 x10 9 /L, S.albumin 30 gil, total protein 88 gil, ALP 64 U/L, and GTT 21 U/L, CD4 count was 255 cells/mm 3 , HIV-RNA viral load was 42000( 4.610gs). At month 6, 22 patients remained alive, their median values were: Hb 12.2 g/dl, WBCs 4.65 x109/L, lymphocytes 1.5 x109/L, neutrophils 3 x10 9 /L, platelets 301 x109/L, S.albumin 36.5 gil, total protein 84.5 gil, ALP 78.5 U/L, GTT 44.5 U/L, CD4 count 288 cells/mm3 , HIV-RNA viral load was 50500( 4.6910gs). The baseline median CD4 and HIV-RNA viral load counts for the 3 patients who died before month 6 were 47 cells/mm3 and 31000(4.610gs); respectively. Response to therapy was evaluated in 21(84%) patients as 4(16%) patients were missing, of the 21 patients 3 (14.3%) had complete response and 18(85.7%) had partial response. With respect to sex 2(14.3%) males had complete response and 12(85.7%) had partial response, 1(14.3%) female had complete response and 6 (85.7%) had partial response. Non-parametric statistics were used because of the small sample size and the skewness of the data. Variables were described using medians and ranges, and compared between two independent groups using Mann-Whitney tests. Baseline and month 6 comparisons were done using Wilcoxon signed ranks tests. Receiver Operating Characteristic (ROC) curves were used to analyze cut points to optimize sensitivity and specificity of a quantitative variable for a dichotomous outcome. Discussion In the univariate analysis age and sex didn't influence prognosis and therapeutic response, the influence of ethnic origin couldn't be assessed as all patients were of the same ethnic origin. Baseline WBCs (P= 0.004) and lymphocytes (P=0.026) were significantly associated with complete response. Higher values of GGT (p=O.OOl); ALP (P=0.006) were associated with more deaths. Baseline CD4 count and HIV viral load were not of predictive value, lthough change CD4 (P=002) and VL (p=.OOO) over time was significant and most likely attributed to response to therapy. 90.9 % of patients reached undetectable HIV-l Viral loads at month 6. CONCLUSION: Neither CD4 count nor HIV viral load at baseline predicted prognosis or survival; however there was a borderline significance of CD4 (P=0.058) towards a better survival. / Thesis (M.Med.)-University of KwaZulu-Natal, Durban, 2006.

Doctorat en Sciences / info:eu-repo/semantics/nonPublished