Fertility preservation methods in young women with systemic lupus erythematosus prior to cytotoxic therapy: experiences from the FertiPROTEKT network

Despite new treatment options, some patients with systemic lupus erythematosus (SLE) need to be treated with the cytotoxic agent cyclophosphamide (CYC). Unlike malignant disease, there are no recommendations for ovarian protection in SLE. The clinical experience of the FertiPROTEKT network as well as recommendations after literature review will be presented in this paper.

Cryopreservation and autotransplantation of human ovarian tissue prior to cytotoxic therapy--a technique in its infancy but already successful in fertility preservation

Increasing survival rates in young cancer patients, new reproductive techniques and the growing interest in quality of life after gonadotoxic cancer therapies have placed fertility preservation as an important issue to oncologists, fertility specialists and patients. Several techniques are now available for fertility preservation in these patients. A new promising method is cryopreservation and transplantation of ovarian cortex. Ovarian tissue can be extracted by laparoscopy without any significant delay of gonadotoxic therapy. The tissue can be cryopreserved by specialised centres of reproductive medicine and transplanted in case the women experience premature ovarian failure (POF). This review summarises the European expertise on cryopreservation and transplantation of ovarian tissue, following around 30 reported transplantations globally, resulting in six live births and several ongoing pregnancies. It emphasises that fertility preservation by the cryopreservation of ovarian tissue is a new but already a successful clinical option, which can be considered for selected cancer patients.

Progress on BIG 1-02/IBCSG 27-02/NSABP B-37, a prospective randomized trial evaluating chemotherapy after local therapy for isolated locoregional recurrences of breast cancer

BACKGROUND: The utility of chemotherapy for women who experience a locoregional recurrence after primary treatment of early breast cancer remains an open question. An international collaborative trial is being conducted by the Breast International Group (BIG), the International Breast Cancer Study Group (IBCSG), and the National Surgical Adjuvant Breast and Bowel Project (NSABP) to determine the effectiveness of cytotoxic therapy for these patients, either alone or in addition to selective use of hormonal therapy and trastuzumab. METHODS: The trial population includes women who have had a previous diagnosis of invasive breast cancer treated by mastectomy or breast-conserving surgery, but subsequently develop an isolated local and/or regional ipsilateral invasive recurrence. Excision of all macroscopic tumor without evidence of systemic disease is required for study entry. Patients are randomized to receive chemotherapy or no chemotherapy; type of chemotherapy is not protocol-specified. Radiation, hormonal therapy, and trastuzumab are given as appropriate. The primary endpoint is disease-free survival (DFS). Quality-of-life measurements are collected at baseline, and then at 9 and 12 months. The accrual goal is 977 patients. RESULTS: This report describes the characteristics of the first 99 patients. Sites of recurrence at study entry were: breast (56%), mastectomy scar/chest wall (35%), and regional lymph nodes (9%). Two-thirds of patients have estrogen-receptor-positive recurrences. CONCLUSION: This is the only trial actively investigating the question of "adjuvant" chemotherapy in locally recurrent breast cancer. The case mix of accrual to date indicates a broad representation of this patient population.

Cytotoxic activity of Fagonia cretica against human breast cancer cells

In many parts of the world, plants are directly utilised for their medicinal properties. Traditional medicine from Pakistan, India and the Far East is well documented and its history is embedded in folklore. It has been documented that an aqueous extract of the desert shrub, Fagonia cretica, is a popular treatment for breast cancer in Pakistan. The administration of an aqueous extract of Fagonia cretica is reported effective at reducing tumour size and improving the quality of life of breast cancer patients, is well tolerated and does not exhibit adverse effects like vomiting, diarrhoea or alopecia which are common side effects of standard cytotoxic therapy. In the past, many pharmacologically active and chemotherapeutic compounds have been isolated from plants which subsequently have proven to be successful in clinical trials and been used as primary compounds in therapeutic regimes. Fagonia cretica has historical use as a treatment for breast cancer, yet there is little scientific evidence which shows chemotherapeutic potential towards breast tumours. Preparation and analysis of an aqueous extract of Fagonia cretica may reveal novel chemotherapeutic agents that can be used to effectively target cancer cells. An understanding of the mechanism of any activity may improve our understanding of cancer cell biology and reveal novel therapeutic targets. This thesis describes for the first time that an aqueous extract of Fagonia cretica shows potent in vitro cytotoxic activity towards breast cancer epithelial cell lines which was not seen towards normal mammary epithelial cells. Elucidation and characterisation of the cytotoxic mechanism was undertaken by analysing DNA damage, cell cycle status, apoptosis, metabolic state and expression of transcription factors and their targets. Finally, methods for the isolation and identification of active compound(s) were developed using various chromatographic techniques. An aqueous extract of Fagonia cretica was able to reduce cell viability significantly in two phenotypically different breast cancer cell lines (MCF-7 and MDA-MB-231). This activity was markedly reduced in normal mammary epithelial cells (HMEpC). Further investigation into the mode of action revealed that extract treatment induced cell cycle arrest and apoptosis in both MCF-7 and MDA-MB-231 cell lines. This coincided with the formation of DNA double stranded breaks and the DNA repair marker ?-H2AX. In MCF-7 cells, ATM/ATR activation resulted in increased p53 expression and of its transcriptional targets p21 and bax, suggesting a role for a p53-mediated response. Furthermore, inhibition of extract-induced p53 expression with siRNA reduced the cytotoxic effect against MCF-7 cells. Extract treatment was also associated with increased FOXO3a expression in MCF-7 and MDA-MB-231 cells. In the absence of functional p53, siRNA knockdown of extract-induced FOXO3a expression was completely abrogated, suggesting that FOXO3a plays a vital role in extract-induced cytotoxicity. Isolation and characterisation of the active compound(s) within the extract was attempted using liquid chromatography and mass spectrometry in conjunction with a cell viability assay. Multiple fractionations generated an active fraction that contained four major compounds as detected by mass spectrometry. However, none of these compounds were identified structurally or chemically due to constraints within the methodology.

Cilengitide: an integrin-targeting arginine-glycine-aspartic acid peptide with promising activity for glioblastoma multiforme.

BACKGROUND: Glioblastoma multiforme (GBM), a highly invasive and vascular cancer, responds poorly to conventional cytotoxic therapy. Integrins, widely expressed in GBM and tumor vasculature, mediate cell survival, migration and angiogenesis. Cilengitide is a potent alphavbeta3 and alphavbeta5 integrin inhibitor. OBJECTIVE: To summarize the preclinical and clinical experience with cilengitide for GBM. METHODS: Preclinical studies and clinical trials evaluating cilengitide for GBM were reviewed. RESULTS/CONCLUSIONS: Cilengitide is active and synergizes with external beam radiotherapy in preclinical GBM models. In clinical trials for recurrent GBM, single-agent cilengitide has antitumor benefits and minimal toxicity. Among newly diagnosed GBM patients, single-arm studies incorporating cilengitide into standard external beam radiotherapy/temozolomide have shown encouraging activity with no increased toxicity and have led to a planned randomized Phase III trial.

A randomized clinical trial of adjuvant chemotherapy for radically resected locoregional relapse of breast cancer: IBCSG 27-02, BIG 1-02, and NSABP B-37

In this phase III, multinational, randomized trial, the International Breast Cancer Study Group, Breast International Group, and the National Surgical Adjuvant Breast and Bowel Project will attempt to define the effectiveness of cytotoxic therapy for patients with locoregional recurrence of breast cancer. We will evaluate whether chemotherapy prolongs disease-free survival and, secondarily, whether its use improves overall survival and systemic disease-free survival. Quality of life measurements will be monitored during the first 12 months of the study. Women who have had a previous diagnosis of invasive breast cancer treated by mastectomy or breast-conserving surgery and who have undergone complete surgical excision of all macroscopic disease but who subsequently develop isolated local and/or regional ipsilateral invasive recurrence are eligible. Patients are randomized to observation/no adjuvant chemotherapy or to adjuvant chemotherapy; all suitable patients receive radiation, hormonal, and trastuzumab therapy. Radiation therapy is recommended for patients who have not received previous adjuvant radiation therapy but is required for those with microscopically positive margins. The radiation field must encompass the tumor bed plus a surrounding margin to a dose of >or= 40 Gy. Radiation therapy will be administered before, during, or after chemotherapy. All women with estrogen receptor-positive and/or progesterone receptor-positive recurrence must receive hormonal therapy, with the agent and duration to be determined by the patient's investigator. Adjuvant trastuzumab therapy is permitted for those with HER2- positive tumors, provided that intent to treat is declared before randomization. Although multidrug regimens are preferred, the agents, doses, and use of supportive therapy are at the discretion of the investigator.

The control of hematopoiesis and leukemia: from basic biology to the clinic.

Hematopoiesis gives rise to blood cells of different lineages throughout normal life. Abnormalities in this developmental program lead to blood cell diseases including leukemia. The establishment of a cell culture system for the clonal development of hematopoietic cells made it possible to discover proteins that regulate cell viability, multiplication and differentiation of different hematopoietic cell lineages, and the molecular basis of normal and abnormal blood cell development. These regulators include cytokines now called colony-stimulating factors (CSFs) and interleukins (ILs). There is a network of cytokine interactions, which has positive regulators such as CSFs and ILs and negative regulators such as transforming growth factor beta and tumor necrosis factor (TNF). This multigene cytokine network provides flexibility depending on which part of the network is activated and allows amplification of response to a particular stimulus. Malignancy can be suppressed in certain types of leukemic cells by inducing differentiation with cytokines that regulate normal hematopoiesis or with other compounds that use alternative differentiation pathways. This created the basis for the clinical use of differentiation therapy. The suppression of malignancy by inducing differentiation can bypass genetic abnormalities that give rise to malignancy. Different CSFs and ILs suppress programmed cell death (apoptosis) and induce cell multiplication and differentiation, and these processes of development are separately regulated. The same cytokines suppress apoptosis in normal and leukemic cells, including apoptosis induced by irradiation and cytotoxic cancer chemotherapeutic compounds. An excess of cytokines can increase leukemic cell resistance to cytotoxic therapy. The tumor suppressor gene wild-type p53 induces apoptosis that can also be suppressed by cytokines. The oncogene mutant p53 suppresses apoptosis. Hematopoietic cytokines such as granulocyte CSF are now used clinically to correct defects in hematopoiesis, including repair of chemotherapy-associated suppression of normal hematopoiesis in cancer patients, stimulation of normal granulocyte development in patients with infantile congenital agranulocytosis, and increase of hematopoietic precursors for blood cell transplantation. Treatments that decrease the level of apoptosis-suppressing cytokines and downregulate expression of mutant p53 and other apoptosis suppressing genes in cancer cells could improve cytotoxic cancer therapy. The basic studies on hematopoiesis and leukemia have thus provided new approaches to therapy.

Peptide-antibody conjugates for tumour therapy: a MHC-class-II-restricted tetanus toxin peptide coupled to an anti-Ig light chain antibody can induce cytotoxic lysis of a human B-cell lymphoma by specific CD4 T cells.

Anti-idiotype antibody therapy of B-cell lymphomas, despite numerous promising experimental and clinical studies, has so far met with limited success. Tailor-made monoclonal anti-idiotype antibodies have been injected into a large series of lymphoma patients, with a few impressive complete tumour remissions but a large majority of negative responses. The results presented here suggest that, by coupling to antilymphoma idiotype antibodies a few molecules of the tetanus toxin universal epitope peptide P2 (830-843), one could markedly increase the efficiency of this therapy. We show that after 2-hr incubation with conjugates consisting of the tetanus toxin peptide P2 coupled by an S-S bridge to monoclonal antibodies directed to the lambda light chain of human immunoglobulin, human B-lymphoma cells can be specifically lysed by a CD4 T-lymphocyte clone specific for the P2 peptide. Antibody without peptide did not induce B-cell killing by the CD4 T-lymphocyte clone. The free cysteine-peptide was also able to induce lysis of the B-lymphoma target by the T-lymphocyte clone, but at a molar concentration 500 to 1000 times higher than that of the coupled peptide. Proliferation assays confirmed that the antibody-peptide conjugate was antigenically active at a much lower concentration than the free peptide. They also showed that antibody-peptide conjugates required an intact processing function of the B cell for peptide presentation, which could be selectively inhibited by leupeptin and chloroquine.(ABSTRACT TRUNCATED AT 250 WORDS)

Cytotoxic activity of a dichloromethane extract and fractions obtained from Eudistoma vannamei (Tunicata: Ascidiacea)

This study consists of the bioassay-guided fractionation of the dichloromethane extract from Eudistoma vannamei and the pharmacological characterization of the active fractions. The dried hydromethanolic extract dissolved in aqueous methanol was partitioned with dichloromethane and chromatographed on a silica gel flash column. The anti-proliferative effect was monitored by the MTT assay. Four of the latest fractions, numbered 14 to 17, which held many chemical similarities amongst each other, were found to be the most active. The selected fractions were tested for viability, proliferation and death induction on cultures of HL-60 promycloblastic leukemia cells. The results suggested that the observed cytotoxicity is related to apoptosis induction. (C) 2007 Elsevier Inc. All rights reserved.

Lentiviral vector-mediated tyro sinase-related protein 2 gene transfer to dendritic cells for the therapy of melanoma

Dendritic cells (DCs) are the most potent professional antigen-presenting cells (APCs), which play a vital role in primary immune responses. Introducing genes into DCs will allow constitutive expression of the encoded proteins and thus prolong the presentation of the antigens derived therefrom. In addition, multiple and unidentified epitopes encoded by the entire tumor-associated antigen (TAA) gene may enhance T cell activation. This study demonstrated that an HIV-1-based lentiviral vector conferred efficient gene transfer to DCs. The transgene, murine tyrosinase-related protein 2 (mTRP-2), encodes a clinically relevant melanoma-associated antigen (MAA), which has been found to be a tumor rejection antigen for B16 melanoma. The transfer and proper processing of mTRP-2 in DCs, in terms of RNA transcription activity and protein expression, were verified by RT-PCR and specific antibody, respectively. Administration of mTRP-2 gene-modified DCs (DC-HR'CmT2) to C57BL/6 mice evoked strong protection against tumor challenge, for which the presence of CD4(+) and CD8(+) cells during both the priming and challenge phase was essential. In a therapy model, our results showed that four of seven mice with preestablished tumor remained tumor free for 80 days after therapeutic vaccination. Given the results shown in this study, mTRP-2 gene transfer to DCs provides a potential therapeutic strategy for the management of melanoma, especially in the early stage of the disease.

Photophysical and photobiological studies of a silicon tribenzonaphthoporphyrazinato incorporated into liposomes for photodynamic therapy use

Nanostructured drug delivery systems (NDDS), such as liposomes, represent a growing area in biomedical research. These microheterogeneous media can be used in many biological systems to provide appropriate drug levels with a specific biodistribution. The photophysical properties of a silicon derivative of tribenzonaphthoporphyrazinato (Si-tri-PcNc) incorporated into liposome were studied by steady-state techniques, time-resolved fluorescence and laser flash photolysis. All the spectroscopy measurements performed allowed us to conclude that Si-tri-PcNc in liposome is a promising NDDS for PDT The in vitro experiments with liposomal NDDS showed that the system is not cytotoxic in darkness, but exhibits a substantial phototoxicity at 1 mu M of photosensitizer concentration and 10.0 J/cm(2) of light. These conditions are sufficient to kill about 80% of the cells.

Interaction of 10-(octyloxy) decyl-2-(trimethylammonium) ethyl phosphate with mimetic membranes and cytotoxic effect on leukemic cells

10-(Octyloxy) decyl-2-(trimethylammonium) ethyl phosphate (ODPC) is an alkylphospholipid that can interact with cell membranes because of its amphiphilic character. We describe here the interaction of ODPC with liposomes and its toxicity to leukemic cells with an ED-50 of 5.4, 5.6 and 2.9 pM for 72 h of treatment for inhibition of proliferation of NB4, U937 and K562 cell lines, respectively, and lack of toxicity to normal hematopoietic progenitor cells at concentrations up to 25 pM. The ED-50 for the non-malignant HEK-293 and primary human umbilical vein endothelial cells (HUVEC) was 63.4 and 60.7 mu M, respectively. The critical micellar concentration (CMC) of ODPC was 200 mu M. Dynamic light scattering indicated that dipalmitoylphosphatidylcholine (DPPC) liposome size was affected only above the CMC of ODPC. Differential calorimetric scanning (DCS) of liposomes indicated a critical transition temperature (T(c)) of 41.5 degrees C and an enthalpy (Delta H) variation of 7.3 kcal mol(-1). The presence of 25 mu M ODPC decreased T(c) and Delta H to 393 degrees C and 4.7 kcal mol(-1), respectively. ODPC at 250 mu M destabilized the liposomes (36.3 degrees C. 0.46 kcal mol(-1)). Kinetics of 5(6)-carboxyfluorescein (CF) leakage from different liposome systems indicated that the rate and extent of CF release depended on liposome composition and ODPC concentration and that above the CMC it was instantaneous. Overall, the data indicate that ODPC acts on in vitro membrane systems and leukemia cell lines at concentrations below its CMC, suggesting that it does not act as a detergent and that this effect is dependent on membrane composition. (C) 2010 Elsevier B.V. All rights reserved.

Endostatin gene therapy enhances the efficacy of IL-2 in suppressing metastatic renal cell carcinoma in mice

We investigated whether the administration of IL-2 combined with endostatin gene therapy was able to produce additive or even synergistic immunomodulatory activity in a mouse model of metastatic renal carcinoma. Renca cells were injected into the tail vein of BALB/c mice. After 24 h, the animals were randomly divided into four groups (5 mice/group). One group of mice was the control, the second group received treatment with 100,000 UI of Recombinant IL-2 (Proleukin, Chiron) twice a day, 1 day per week during 2 weeks (IL-2), the third group received treatment with a subcutaneous inoculation of 3.6 x 10(6) endostatin-producing cells, and the fourth group received both therapies (IL-2 + ES). Mice were treated for 2 weeks. In the survival studies, 10 mice/group daily, mice were monitored daily until they died. The presence of metastases led to a twofold increase in endostatin levels. Subcutaneous inoculation of NIH/3T3-LendSN cells resulted in a 2.75 and 2.78-fold increase in endostatin levels in the ES and IL-2 + ES group, respectively. At the end of the study, there was a significant decrease in lung wet weight, lung nodules area, and microvascular area (MVA) in all treated groups compared with the control group (P < 0.001). The significant difference in lung wet weight and lung nodules area between groups IL-2 and IL-2 + ES revealed a synergistic antitumor effect of the combined treatment (P < 0.05). The IL-2 + ES therapy Kaplan-Meier survival curves showed that the probability of survival was significantly higher for mice treated with the combined therapy (log-rank test, P = 0.0028). Conjugated therapy caused an increase in the infiltration of CD4, CD8 and CD49b lymphocytes. An increase in the amount of CD8 cells (P < 0.01) was observed when animals received both ES and IL-2, suggesting an additive effect of ES over IL-2 treatment. A synergistic effect of ES on the infiltration of CD4 (P < 0.001) and CD49b cells (P < 0.01) was also observed over the effect of IL-2. Here, we show that ES led to an increase in CD4 T helper cells as well as cytotoxic lymphocytes, such as NK cells and CD8 cells, within tumors of IL-2 treated mice. This means that ES plays a role in supporting the actions of T cells.

Cytotoxic effects of butanolic extract from Pfaffia paniculata (Brazilian Ginseng) on cultured human breast cancer cell line MCF-7

Roots of Pfaffia paniculata have been well documented for multifarious therapeutic values and have also been used for cancer therapy in folk medicine. This study has been performed in a human breast tumor cell line, the MCF-7 cells. These are the most commonly used model of estrogen-positive breast cancer, and it has been originally established in 1973 at the Michigan Cancer Foundation from a pleural effusion taken from a woman with metastatic breast cancer. Butanolic extract of the roots of P. paniculata showed cytotoxic effect MCF-7 cell line. as determined with crystal violet assay, cellular death with acridine orange/ethidium bromide staining, and cell proliferation with immunocytochemistry of bromodeoxyuridine (BrdU). Subcellular alterations were evaluated by electron microscopy. Cells treated With butanolic extract showed degeneration of cytoplasmic components and profound morphological and nuclear alterations. The results show that this butanolic extract indeed presents cytotoxic substances, and its fractions merit further investigations. (C) 2008 Elsevier GmbH. All rights reserved.