The non-vesicle cell-free DNA (cfDNA) induces cell transformation associated with horizontal DNA transfer.

BACKGROUND: Cell-free DNA (cfDNA) is a source for liquid biopsy used for cancer diagnosis, therapy selection, and disease monitoring due to its non-invasive nature and ease of extraction. However, cfDNA also participates in cancer development and progression by horizontal transfer. In humans, cfDNA circulates complexed with extracellular vesicles (EV) and macromolecular complexes such as nucleosomes, lipids, and serum proteins. The present study aimed to demonstrate whether cfDNA not associated with EV induces cell transformation and tumorigenesis. METHODS: Supernatant of the SW480 human colon cancer cell line was processed by ultracentrifugation to obtain a soluble fraction (SF) and a fraction associated with EV (EVF). Primary murine embryonic fibroblast cells (NIH3T3) underwent passive transfection with these fractions, and cell proliferation, cell cycle, apoptosis, cell transformation, and tumorigenic assays were performed. Next, cfDNA was analyzed by electronic microscopy, and horizontal transfer was assessed by human mutant KRAS in recipient cells via PCR and recipient cell internalization via fluorescence microscopy. RESULTS: The results showed that the SF but not the EVF of cfDNA induced proliferative and antiapoptotic effects, cell transformation, and tumorigenesis in nude mice, which were reduced by digestion with DNAse I and proteinase K. These effects were associated with horizontal DNA transfer and cfDNA internalization into recipient cells. CONCLUSIONS: The results suggest pro-tumorigenic effects of cfDNA in the SF that can be offset by enzyme treatment. Further exploration of the horizontal tumor progression phenomenon mediated by cfDNA is needed to determine whether its manipulation may play a role in cancer therapy.

Exploring disparities in incidence and mortality rates of breast and gynecologic cancers according to the Human Development Index in the Pan-American region.

OBJECTIVE: To evaluate whether a country's Human Development Index (HDI) can help explain the differences in the country's breast cancer and gynecological cancer incidence and mortality rates in the Pan-American region. STUDY DESIGN: Ecological analysis. METHODS: Pan-American region countries with publicly available data both in GLOBOCAN 2012 and the United Nations Development Report 2012 were included (n = 28). Incidence and mortality rates age-standardized per 100,000 were natural log-transformed for breast cancer, ovarian cancer, corpus uteri cancer, and cervical cancer. The mortality-to-incidence ratio (MIR) was calculated for each site. Pearson's correlation test and a simple linear regression were performed. RESULTS: The HDI showed a positive correlation with breast cancer and ovarian cancer incidence and mortality rates, respectively, and a negative correlation with cervical cancer incidence and mortality rates. The HDI and corpus uteri cancer showed no association. MIR and the HDI showed a negative correlation for all tumor types except ovarian cancer. An increment in 1 HDI unit leads to changes in cancer rates: in breast cancer incidence ß = 4.03 (95% confidence interval [CI] 2.61; 5.45) P < 0.001, breast cancer mortality ß = 1.76 (95% CI 0.32; 3.21) P = 0.019, and breast cancer-MIR ß = -0.705 (95% CI 0.704; 0.706) P < 0.001; in cervical cancer incidence ß = -3.28 (95% CI -4.78; -1.78) P < 0.001, cervical cancer mortality ß = -4.63 (95% CI -6.10; -3.17) P < 0.001, and cervical cancer-MIR ß = -1.35 (95% CI -1.83; -0.87) P < 0.001; in ovarian cancer incidence ß = 3.26 (95% CI 1.78; 4.75) P < 0.001, ovarian cancer mortality ß = 1.82 (95% CI 0.44; 3.20) P = 0.012, and ovarian cancer-MIR ß = 5.10 (95% CI 3.22; 6.97) P < 0.001; in corpus uteri cancer incidence ß = 2.37 (95% CI -0.33; 5.06) P = 0.83, corpus uteri cancer mortality ß = 0.68 (95% CI -2.68; 2.82) P = 0.96, and corpus uteri cancer-MIR ß = -2.30 (95% CI -3.19; -1.40) P < 0.001. CONCLUSIONS: A country's HDI should be considered to understand disparities in breast cancer and gynecological cancer in the Pan-American region.

Genetic selection of volunteers and concomitant dose adjustment leads to comparable hydralazine/valproate exposure.

WHAT IS KNOWN AND OBJECTIVE: Hydralazine is an inhibitor of DNA methyltransferases, whereas valproate interferes with histone deacetylation. In combination, they show a marked synergism in reducing tumour growth as well as development of metastasis and inducing cell differentiation. Hydralazine is metabolized by the highly polymorphic N-acetyltransferase 2. The current pilot study was performed to analyse the pharmacokinetic parameters of a single dose of hydralazine in 24 h (one tablet with 83 mg for slow acetylators and one tablet with 182 mg for fast acetylators) and three fixed doses of valproate (one tablet of controlled liberation with 700 mg every 8 h) in healthy genetically selected volunteers. Selection was performed based on their NAT2 activity as deduced from their genotype. METHODS: An open label non-randomized single arm study was conducted in two groups of six healthy volunteers of both genders aged 20-45 years with a body mass index 22·2-26·9 which were classified as fast or slow acetylators after genotyping 3 SNPs that cover 99·9% of the NAT2 variants in the Mexican population. Blood samples were collected predose and serially post-dose in an interval of 48 h. Hydralazine and valproate concentrations were determined by ultra-high performance liquid chromatography (UPLC) coupled to tandem mass spectroscopy (MS/MS). RESULTS AND DISCUSSION: The AUC0-48 h and Cmax of hydralazine were almost identical (1410 ± 560 vs. 1446 ± 509 ng h/mL and 93·4 ± 16·7 vs. 112·5 ± 42·1 ng/mL) in both groups with NAT2 genotype-adjusted doses, whereas the multidose parameters of valproate were not significantly affected neither by the selection of the NAT2 genotype (AUC0-48 h 2064 ± 455 vs. 1896 ± 185 µg h/mL; Cmax 96·4 ± 21·1 vs. 88·8 ± 7·2 µg/mL, for the fast and slow acetylators, respectively) nor the co-administration of 83 or 182 mg of hydralazine. WHAT IS NEW AND CONCLUSION: Comparable hydralazine exposures (differences in AUC0-inf of only 7%) were observed in this study with genetic selection of volunteers and concomitant dose adjustment. However, the conclusions have yet to be confirmed with a full-powered 2 × 2 crossover study.

Radiosensitization of cervical cancer cells with epigenetic drugs hydralazine and valproate.

PURPOSE: To evaluate the radiosensitizing effects of the DNA methylation inhibitor hydralazine in combination with valproic acid, a histone deacetylase inhibitor in cervical cancer cells. MATERIALS AND METHODS: Cell viability assays were performed in the SiHa cervical cancer cell line treated with hydralazine and valproic acid for five days with and without cisplatin. Cell irradiation was performed using teletherapy (1.25 MV). RESULTS: Neither hydralazine, valproic acid or cisplatin as single agents increased the cytotoxicity from radiation, however, the combination of hydralazine with valproic acid at ten microM and one mM, respectively, did induce radiosensitization (p = 0.046). Interestingly, this effect was further increased with the triple combination of hydralazine, valproic acid, and cisplatin (p = 0.041), where cell viability decreased more than 50% as compared to radiation alone. CONCLUSIONS: The present results demonstrate that epigenetic drugs increase the efficacy of cisplatin chemoradiation in cervical cancer cells.

Brachytherapy versus radical hysterectomy after external beam chemoradiation with gemcitabine plus cisplatin: a randomized, phase III study in IB2-IIB cervical cancer patients.

BACKGROUND: The aim of the present study was to demonstrate that radical hysterectomy (RH) leads to improved survival outcomes in FIGO stage IB2-IIB cervical cancer when compared with standard brachytherapy (BCT) after identical external beam chemoradiation (EBRT-CT). PATIENTS AND METHODS: EBRT-CT treatment consisted of six courses of cisplatin at 40 mg/m² and gemcitabine at 125 mg/m² per week concurrent with 50.4 Gy of radiation. In the BCT arm, EBRT-CT was followed by BCT to reach a point A dose of 85 Gy, whereas in the experimental arm, a type III RH with bilateral pelvic lymph node dissection and para-aortic lymph node sampling (RH) was carried out within 4-6 weeks after EBRT-CT. RESULTS: Between May 2004 and June 2009, 211 patients were enrolled (BCT, 100 and RH, 111). At a median follow-up time of 36 months (3-80), progression-free survival (PFS) and overall survival (OS) rates were similar in both the arms. PFS rates were 74.8% and 71.7% in the BCT and RH arms [HR 0.6516 (95% confidence interval (CI) 0.3504-1.2116)], P = 0.186. OS rates were 76.3% in the BCT versus 74.5% in the surgical arm [HR 0.6981 (95% CI 0.3106-1.3439)], P = 0.236. No differences were observed in the pattern of local and systemic failures. CONCLUSIONS: This study failed to demonstrate that RH after EBRT-CT is superior to standard BCT.

Efficacy in high burden locally advanced cervical cancer with concurrent gemcitabine and cisplatin chemoradiotherapy plus adjuvant gemcitabine and cisplatin: prognostic and predictive factors and the impact of disease stage on outcomes from a prospective randomized phase III trial.

OBJECTIVE: We recently published results of a phase III trial demonstrating superior outcomes in patients with locally advanced cervical cancer (LACC) when concurrent cisplatin chemoradiotherapy is supplemented with concurrent gemcitabine and adjuvant gemcitabine/cisplatin. We present prognostic and predictive factors identified in that study, along with analyses of the effect of disease stage and post-study therapy. PATIENTS AND METHODS: In that trial, 515 patients with stage IIB-IVA LACC were administered concurrent cisplatin chemoradiotherapy with or without gemcitabine and adjuvant gemcitabine/cisplatin. Cox models were used to identify prognostic and predictive factors. Survival was estimated using the Kaplan-Meier method. RESULTS: Advanced (stage III-IVA) disease, squamous histology, low hemoglobin, the presence of ≥1 enlarged para-aortic lymph nodes, and large tumor size, were associated with poorer prognosis, regardless of treatment assigned. Tumor size and histology were predictive of treatment efficacy. Chemoradiotherapy supplemented with gemcitabine produced relatively greater benefit in patients with stage III/IVA disease. Post-study therapy did not appear to affect the overall survival outcome. CONCLUSION: Prognostic factors identified in this study are consistent with other reports. The finding of relatively greater benefit in advanced-stage patients makes for an important factor in consideration of treatment for these patients and the design of future studies.

Arylamine N-acetyltransferase 2 genotypes in a Mexican population.

The acetylating activity of N-acetyltransferase 2 (NAT2) has critical implications for therapeutics and disease susceptibility. To date, several polymorphisms that alter the enzymatic activity and/or protein stability of NAT2 have been identified. We examined the distribution and frequency of NAT2 genotypes in the Mexican population. Among 250 samples amplified and sequenced for the NAT2 gene, we found seven different SNPs; the most frequent allele was 803 A>G (35.8%), followed by 282 C>T, 341 T>C, and 481 C>T. There were no differences in the distribution of SNPs between healthy subjects and cancer patients. These eight polymorphisms defined 26 diplotypes; 11.6% were wild type (NAT2*4/NAT2*4), while the most common diplotype was NAT2*4/NAT2*5B, present in 17.2%. We did not identify other common polymorphisms. The results were compared with the NAT2 SNPs reported from other populations. All but the Turkish population was significantly different from ours. We conclude that the mixed-race Mexican population requires special attention because NAT2 genotype frequencies differ from those in other regions of the world.

Late recurrence of cervical cancer: a report of 16 cases.

PURPOSE: To analyze the clinical characteristics and outcome of cervical cancer patients presenting late recurrence. MATERIALS AND METHODS: The medical records of 16 patients who were treated between 1974 and 1999 at the Institution and whose cancer recurred after a five-year disease-free interval were reviewed. RESULTS: Mean time from initial therapy to recurrence was 162.5 months (60-360 mean). Smear abnormalities, atypical genital bleeding, abdominal and lumbar pain, and respiratory findings were the most common symptoms and signs associated with late recurrence. Fourteen patients were diagnosed by physical examination. Three of the six patients with local recurrence who were re-irradiated developed a vesico-vaginal fistula. At a median follow-up time of 12.5 months (4-38 mean), 12 patients were alive and the median survival time was 30 months. CONCLUSIONS: Cervical cancer patients surviving free of disease after the fifth year post-treatment are still at risk for relapse and in most of them, the recurrence is suspected by clinical examination alone.

Hereditary diffuse gastric cancer (HDGC). An overview.

It is estimated that up to 10% of gastric carcinomas show familial aggregation. In contrast, around 1-3 % (approximately 33,000 yearly) are genuinely hereditary. Hereditary diffuse gastric cancer (HDGC) is a rare malignancy characterized by autosomal dominant inheritance of pathological variants of the CDH1 and CTNNA1 genes encoding the adhesion molecules E-cadherin and α-catenin, respectively. The multifocal nature of the disease and the difficulty of visualizing precursor lesions by endoscopy underscore the need to be aware of this malignancy as surgical prevention can be fully protective. Here, we provide an overview of the main epidemiological, clinical, genetic, and pathological features of HDGC, as well as updated guidelines for its diagnosis, genetic testing, counseling, surveillance, and management. We conclude that HDGC is a rare, highly penetrant disease that is difficult to diagnose and manage, so it is necessary to correctly identify it to offer patients and their families' adequate management following the recommendations of the IGCL. A critical point is identifying a mutation in HDGC families to determine whether unaffected relatives are at risk for cancer.

Pharmacokinetics of hydralazine, an antihypertensive and DNA-demethylating agent, using controlled-release formulations designed for use in dosing schedules based on the acetylator phenotype.

PURPOSE: The antihypertensive hydralazine has recently been repositioned as DNA demethylating for the epigenetic therapy of cancer. As the acetylator phenotype is the key determinant of its plasma levels, the dose of hydralazine needs to be adjusted for the acetylation status of patients. METHODS: The pharmacokinetics of orally administered hydralazine was evaluated in 26 healthy volunteers (13 slow and 13 fast acetylators) after a single dose of 182 mg administered as a controlled-release tablet. Plasma levels of hydralazine were analyzed in 85 cancer patients treated with this formulation at a dose of 83 mg/day and 182 mg/day for slow and fast acetylators, respectively. RESULTS: The C(max) and t(max) of hydralazine for fast acetylators were 208.4 ± 56.9 SD ng/ml and 2.8 ± 2.5 h, respectively. The corresponding results for slow acetylators were 470.4 ± 162.8 ng/ml, and 4.4 ± 3.1 h. Healthy volunteers who were fast acetylators had no clinically significant changes in blood pressure and heart rate or any other side-effect, however, slow acetylators had transient episodes of headache, tachycardia and faintness. Among 85 cancer patients that received either 182 mg or 83 mg of hydralazine daily, according to their acetylator status, the mean concentrations of hydralazine in plasma were 239.1 ng/ml and 259.2 ng/ml for fast and slow acetylators, respectively. These differences were not significantly different, p = 0.3868. CONCLUSIONS: The administration of dose-adjusted controlled-release hydralazine according to the acetylation status of cancer patients yields similar levels of hydralazine.

Epigenetic therapy and cisplatin chemoradiation in FIGO stage IIIB cervical cancer.

INTRODUCTION: This trial aimed to evaluate the safety and efficacy of epigenetic therapy associated with cisplatin chemoradiation in FIGO Stage IIIB patients. METHODS: Hydralazine containing either 182 mg for rapid-, or 83 mg for slow acetylators and magnesium valproate were administered at 30 mg/kg tid. Both drugs were taken until intracavitary therapy was finished. Pelvic external beam radiation and low-dose rate brachytherapy were administered at a total cumulative dose to point A of at least 85 Gy. Weekly cisplatin at 40 mg/m2 was delivered for six cycles. RESULTS: Twenty-two patients were included and 18 (82%) patients completed treatment. Mean dose to point A was 84.6 + 2.2. Median number of cisplatin cycles was 5.5 (range, 1-6). Brachytherapy was delayed for technical reasons; the mean overall treatment time was 11.8 weeks. Grade 3 anemia, leucopenia, neutropenia, and thrombocytopenia were observed in 9%, 45%, 45%, and 9% of patients, respectively. CONCLUSIONS: Hydralazine and valproate are well-tolerated and safe when administered with cisplatin chemoradiation. Unfortunately, the suboptimal administration of brachytherapy for technical reasons in this study, precluded assessing the efficacy of epigenetic therapy. However, the tolerability of this regimen administered concurrent to radiation needs to be further tested.

Mutational analysis of BRCA1 and BRCA2 genes in Mexican breast cancer patients.

Germline mutations in the BRCA1 and BRCA2 genes predispose to breast and ovarian cancer. A variable incidence of mutations has been reported for these genes. The contribution of BRCA1 and BRCA2 mutations to Mexican women with breast and/or ovarian cancer is not known. Because of the increasing prevalence of breast cancer in this population, it is necessary to study the presence of mutations in both genes. We screened BRCA1 and BRCA2 genes in 40 patients: 29 patients with a history of breast and/or ovarian cancer, and 11 patients with early-onset breast cancer (< 40 years), through denaturing high performance liquid chromatography analyses. We found two frameshift mutations in BRCA1 and one missense mutation in each gene. Additionally we found several intronic variants as well as synonymous mutations. We found 5% of deleterious mutations in the BRCA genes. Larger studies are needed to establish the significance and prevalence of BRCA mutations among Mexican women.

Concurrent chemoradiation with carboplatin for elderly, diabetic and hypertensive patients with locally advanced cervical cancer.

INTRODUCTION: Chemoradiation based on cisplatin is the standard treatment of locally advanced cervical cancer, however, a subset of patients are either elderly and/or have comorbidities such as diabetes and hypertension. These conditions may compromise the administration of cisplatin. We report our Institution experience with weekly carboplatin as a radiosensitizer for the management of this subset of patients. PATIENTS AND METHODS: We reviewed the files of 59 patients with locally advanced cervical cancer who were treated with primary chemoradiation with weekly carboplatin. Response rate, toxicity and survival were analyzed. RESULTS: Mean age was 62 years (range, 36-83 years). The majority of cases were squamous cell carcinoma (88.14%), and distribution according to FIGO Stage was IB2 8.4%, IIA 13.5%, IIB 52.5%, IIIA 3.3% and IIIB 18.6%; Overall, 100% and 91% of patients completed external beam and intracavitary therapy. Seventy-nine percent received from five to six planned cycles of weekly carboplatin. Complete responses were achieved in 49 (83.05 %) patients, whereas ten patients (16.95%) had either persistent or progressive disease. The most common toxicities were grades 1 and 2 hematological and gastrointestinal. At median follow-up (20 months; range 2-48 months), 16 patients (32.65%) have relapsed. Estimated 30-month overall survival is 63%. CONCLUSIONS: Weekly carboplatin concurrent with pelvic radiation is well tolerated in patients with locally advanced carcinoma of the cervix who are older than 70 years and/or have diabetes mellitus and/or high blood pressure, however, the apparently slighty lower survival observed cautions against its routine use.

Ultrasound, histopathological, and genetic features of uveal melanoma in a Mexican-Mestizo population. / Características ultrasonográficas, histopatológicas y genéticas del melanoma uveal en población mexicana mestiza☆.

OBJECTIVE: To describe the ultrasound, histopathological and genetic characteristics of uveal melanoma in a Mexican-Mestizo population. MATERIAL AND METHODS: A total of 39 enucleated eyes with a histopathological diagnosis of uveal melanoma were assessed by describing the clinical findings, and ultrasound, histopathological and genetic features. RESULTS: A high correlation was observed between tumour height measurement using ultrasound and histopathology. In our cases, tumour size and reflectivity were higher compared with those reported in the literature. The preliminary data on the molecular assessment of the tumours show the presence of an unreported polymorphism (T>C IVS5+34) and one sample with GNAQ mutation (A>C CAA>CCA Gln 209 Pro). CONCLUSION: Ultrasound is a reliable method to identify the size of the tumour. Furthermore, knowledge of the molecular mechanisms promises new perspectives for the development of new targeted therapeutics. Fortunately this leads to progress in the treatment of patients with metastatic disease or prevents it in those at high risk.

Computational Study of the Binding Modes of Diverse DPN Analogues on Estrogen Receptors (ER) and the Biological Evaluation of a New Potential Antiestrogenic Ligand.

Estrogen (17ß-estradiol) is essential for normal growth and differentiation in the mammary gland. In the last three decades, previous investigations have revealed that Estrogen Receptor Alpha (ERα) plays a critical role in breast cancer. More recently, observations regarding the widespread expression of ERß-like proteins in normal and neoplastic mammary tissues have suggested that ERß is also involved in the mentioned pathology. Design of new drugs both steroidal and nonsteroidal that target any of these receptors represents a promise to treat breast cancer although it remains a challenge due to the sequence similarity between their catalytic domains. In this work, we propose a new set of compounds that could effectively target the estrogen receptors ERα and ERß. These ligands were designed based on the chemical structure of the ERß-selective agonist Diarylpropionitrile (DPN). The designed ligands were submitted to in silico ADMET studies, yielding in a filtered list of ligands that showed better drug-like properties. Molecular dynamics simulations of both estrogen receptors and docking analysis were carried-out employing the designed compounds, from which two were chosen due to their promising characteristics retrieved from theoretical results (docking analysis or targeting receptor predictions). They were chemically synthetized and during the process, two precursor ligands were also obtained. These four ligands were subjected to biological studies from which it could be detected that compound mol60b dislplayed inhibitory activity and its ability to activate the transcription via an estrogenic mechanism of action was also determined. Interestinly, this observation can be related to theoretical binding free energy calculations, where the complex: ERß-mol60b showed the highest energy ΔGbind value in comparison to others.

Nicotinamide sensitizes human breast cancer cells to the cytotoxic effects of radiation and cisplatin.

Poly(ADP-ribose) polymerase (PARP) inhibitors enhance the effect of DNA alkylating agents on BRCA1­ and BRCA2-deficient cell lines. The aim of this study was to analyze the effect of the PARP inhibitor nicotinamide (NAM) on breast cancer cells with different BRCA1 expression or function, such as BRCA1­deficient MDA-MB-436 cells, low expression BRCA1 MCF-7 cells, and the BRCA1 wild­type MDA-MB-231 cells, to demonstrate its effects as a chemo­ or radiosensitizing agent. PARP activity was analyzed in MDA-MB-436, MCF-7 and MDA-MB-231 breast cancer cells subjected or not to NAM. Inhibition of PARP by NAM in the presence of DNA damage was examined by Alexa Fluor 488 immunofluorescence. Crystal violet assays were used to test growth inhibition and the chemo­ and radiosensitization effects of NAM were investigated using clonogenic assays. Significant differences among data sets were determined using two-tailed ANOVA and Bonferroni tests. We demonstrated that NAM reduces PARP activity in vitro, and in cells subjected or not to DNA damage, it also reduces the viability of breast cancer cell lines and synergyzes the cytotoxicity of cisplatin in MDA-MB-436 and MCF-7 cells. Downregulation of PARP1 with siRNA led to modest growth inhibition, which was further increased by cisplatin. Nicotinamide also induced radiosensitization in MDA-MB-436 and MDA-MB-231 cells. In conclusion, NAM may be used as a chemo­ or radiosensitizing agent regardless of the BRCA1 status in breast cancer.

Analysis of nm23-H1 expression in breast cancer. Correlation with p53 expression and clinicopathologic findings.

Metastasis is the most frequent cause of death in patients with breast cancer. The nm23-H1 and p53 genes have been involved in the development of breast cancer metastasis. We have analyzed the correlation between the expression of nm23 protein and several established clinicopathologic factors. Our results show that the antimetastatic role of nm23-H1 is not related to the cell proliferative status or tumor grade and that it is not associated with the expression of p53. We also demonstrate a strong inverse relationship between the expression of nm23-H1 protein, lymph node metastasis and vascular invasion. These data support the antimetastatic role of the nm23-H1 gene and suggest that nm23-H1 and p53 genes may be involved in different steps of the metastatic process.

Misoprostol prophylaxis for high-dose chemotherapy-induced mucositis: a randomized double-blind study.

From April 1993 to September 1993, 15 patients with lymphoid or solid neoplasms underwent 16 non-cryopreserved peripheral stem cell transplantation courses using the ICE (ifosfamide, carboplatin, etoposide) program. They were randomized in a double-blind clinical trial to received oral misoprostol or placebo for mucositis prophylaxis. The active drug or placebo administration began jointly with chemotherapy at day -4 and was continued until day 16. The mucositis incidence and severity was significantly higher in patients who received misoprostol. We found no differences regarding myelosuppression, infections or other chemotherapy complications. Our results do not support the use of oral misoprostol as administered in this study, for high-dose chemotherapy-induced mucositis prophylaxis.

Gemcitabine activity in cervical cancer cell lines.

PURPOSE: Gemcitabine (2',2'-difluorodeoxycytidine) is an antineoplastic agent with activity against a variety of solid tumors. To investigate its in vitro activity toward cervical cancer, we exposed six cervical cancer cell lines to gemcitabine. METHODS: Combinational cytotoxic studies using viability tests and clonogenicity assays. RESULTS: Gemcitabine was cytostatic and cytotoxic in some of the lines at peak plasma concentrations similar to those achieved in clinical trials. Gemcitabine was also found to effectively synergize with cisplatin and showed a radiosensitizing effect in these cells. The cytotoxicity observed in sensitive cell lines was due to apoptosis, as demonstrated by DNA fragmentation assays. CONCLUSIONS: We recommend performing additional in vitro experimentation so that these results can be confirmed to support clinical trials of gemcitabine in cervical cancer patients both as first-line therapy and with concomitant radiation.

A pilot study of perilymphatic leukocyte cytokine mixture (IRX-2) as neoadjuvant treatment for early stage cervical carcinoma.

Clinical and experimental data demonstrate that local cytokines are able to induce tumor regression and in some cases antitumor systemic immune response. IRX-2 is a cell-free mixture of cytokines obtained from unrelated donor lymphocytes with demonstrated ability to induce immune mediated regression of squamous cell carcinomas of head and neck. The objective of this study was to evaluate the antitumor activity and toxicity of IRX-2 in untreated early stage cervical cancer patients. Ten consecutive patients clinically staged IB1, IB2 and IIA were treated with a neoadjuvant immunotherapy regimen that consisted in a single IV dose of cyclophosphamide at 300 mg/m2 on day 1, oral indomethacin or ibuprofen and zinc sulfate were administered from days I to 21 and 10 regional perilymphatic injections of IRX-2 on days 3 to 14. All patients were scheduled for radical hysterectomy on day 21. The clinical and pathological responses, toxicity and survival were evaluated. Clinical response was seen in 50% of patients (three partial responses, two minor responses). Seven patients underwent surgery and pathological tumor reduction associated with tumor fragmentation was found in five cases. Histological studies demonstrated a rather heterogeneous cell type infiltrating pattern in the tumor which included lymphocytes, plasma cells, neutrophils, macrophages and eosinophils. Immunohistochemical analysis of the surgical specimens demonstrated an increase of tumor infiltrating CD8+ cells. The treatment was well tolerated except for mild pain and minor bleeding during injections and gastric intolerance to indomethacin. At 31 months of maximum follow-up (median 29), eight patients are disease-free. Our results suggest that the immunotherapy approach used induces tumor responses in cervical cancer patients. Further studies are needed to confirm these results as well as to elucidate the mechanisms underlying these effects.