Longitudinal analysis of the effect of water hardness on atopic eczema: evidence for gene-environment interaction.

BACKGROUND: Several studies have identified an association between water hardness and atopic eczema (AE); however, there is a paucity of longitudinal data in early life. OBJECTIVES: To examine whether water hardness is associated with an increased risk of AE and skin barrier dysfunction in infants and to assess effect modification by filaggrin (FLG) loss-of-function variants. METHODS: We performed a longitudinal analysis of data from infants in the Enquiring About Tolerance (EAT) study, who were enrolled at 3 months and followed up until 36 months of age. RESULTS: Of 1303 infants enrolled in the EAT study, 91·3% (n = 1189) attended the final clinic visit and 94·0% (n = 1225) of participants' families completed the 36-month questionnaire. In total, 761 (58·4%) developed AE by 36 months. There was no overall association between exposure to harder (> 257 mg L-1 CaCO3 ) vs. softer (≤ 257 mg L-1 CaCO3 ) water: adjusted hazard ratio (HR) 1·07, 95% confidence interval (CI) 0·92-1·24. However, there was an increased incidence of AE in infants with FLG mutations exposed to hard water (adjusted HR 2·72, 95% CI 2·03-3·66), and statistically significant interactions between hard water plus FLG and both risk of AE (HR 1·80, 95% CI 1·17-2·78) and transepidermal water loss (0·0081 g m-2 h-1 per mg L-1 CaCO3 , 95% CI 0·00028-0·016). CONCLUSIONS: There is evidence of an interaction between water hardness and FLG mutations in the development of infantile AE.

Is there a correlation between molecular markers and response to neoadjuvant chemoradiotherapy in locally advanced squamous cell esophageal cancer?

PURPOSE: To evaluate the expression of epidermal growth factor receptor (EGFR), p53, p21 and thymidylate synthase (TS) in a pretherapy biopsy specimen of locally advanced squamous cell esophageal cancer and correlate these markers with response to neoadjuvant chemoradiotherapy. METHODS: Sixty-two patients with histopathologically proven locally advanced (T3 or greater) squamous cell esophageal cancer were enrolled. The expression of EGRF, p53, p21 and TS markers was assessed with immunohistochemistry. Semiquantitative assessment of expression of these markers was performed based on the percent of the stained cells. Radiotherapy (45-50.4 Gy) was delivered concomitantly with 5-fluorouracil (5-FU)/leucovorin (LV)/cisplatin (CIS) chemotherapy. Five to 6 weeks after chemoradiation, response to treatment was assessed. Medically fit and operable patients were operated. The resected material underwent histopathological evaluation of tumor expansion, histological classification after initial multimodality treatment (yp TNM), residual status and tumor regression grade (TRG). RESULTS: Out of 62 patients enrolled, 41 (66%) were evaluated for molecular markers. Clinical response rate was 43.9%. Out of 41 patients, 12 (29%) underwent surgery. TRG 1 was noted in 58% of the patients. In a pretherapy tumor specimen, positive expression was noted in 80, 90, 80 and 71% for EGFR, p53, p21 and TS, respectively. We noted no statistically significant difference neither between tumor marker expression and clinical response to chemoradiation, nor between tumor marker expression and TRG. CONCLUSION: We registered no difference in response to treatment between EGFR, TS, p21 and p53 positive and negative staining.

Systematic reviews of sublingual immunotherapy (SLIT).

Allergic rhinitis is common worldwide, with significant morbidity and impact on quality of life. In patients who don't respond adequately to anti-allergic drugs. Subcutaneous allergen immunotherapy is effective although requires specialist administration. Sublingual immunotherapy may represent an effective and safer alternative. This Cochrane systematic review is an update of one published in 2003. We searched Cochrane ENT Group Trials Register, Central, PubMed, EMBASE, CINAHL, Web of Science, Biosis Previews, Cambridge Scientific Abstarcts, mRCT and additional sources. We included randomised, double-blind, placebo- controlled trials of sublingual immunotherapy in adults and children. Two authors selected studies and assessed them for quality. Data were put into RevMan 5.0 for a statistical analysis. We used standardised mean difference (SMD), with a random effect model to combine data. Sixty studies were included, with 49 suitable for meta-analysis. We found significant reductions in symptoms (SMD -0.49; 95%CI (-0.64 to -0.34, P < 0.00001)) and medication requirements (SMD -0.32; 95%CI (-0.43 to -0.21, P < 0.00001)) compared with placebo. None of the trials reported severe systemic reactions, anaphylaxis or use of Adrenaline. This updated review reinforces the conclusion of the original 2003 Cochrane Review that sublingual immunotherapy is effective for allergic rhinitis and appears a safe route of administration.

TP53 gene status and human papilloma virus infection in response to platinum plus taxane-based chemotherapy of epithelial ovarian carcinomas.

PURPOSE: Lack of symptoms in early stages of disease and resistance to chemotherapy make epithelial ovarian carcinomas one of the most lethal neoplasms among gynaecological malignancies. The aim of this study was to analyse the impact of TP53 mutations, codon 72 polymorphism and human papillomavirus (HPV) infection on the response to platinum-taxane combination chemotherapy in patients with epithelial ovarian carcinomas. METHODS: The study was conducted on 26 ovarian carcinoma patients who received carboplatin plus paclitaxel combination chemotherapy. DNA was isolated by salting-out procedure. Mutations in exons 4-8 of TP53 gene were detected by PCR-SSCP and confirmed by automatic DNA sequencing. Codon 72 polymorphism was assessed by the RFLP method. HPV infection was detected through amplification of one part of L1 viral gene. Genotyping was performed by DNA sequencing. Fisher's exact and log-rank tests were used for statistical analysis. RESULTS: TP53 mutations were present in 5/26 (19.2%) ovarian carcinomas. The distribution of codon 72 TP53 genotypes was: Arg/Arg 38.5%, Arg/Pro 50.0%, Pro/Pro 11.5%. HPV was present in 4/26 (15.4%) ovarian carcinomas. All HPV-positive tumors were HPV16 type. Patients with mutations in TP53 gene, Arg/Arg genotype of codon 72 and absence of HPV infection experienced the highest tumor response rate to platinum-taxane chemotherapy. However, no significant correlation between progression free interval (PFI) and the examined biomarkers was observed. CONCLUSION: Our results indicate that, based on the TP53 gene status and the presence/absence of HPV infection, the subgroups of patients having better initial response to platinum-taxane therapy could be distinguished. This might contribute to more adequate treatment and individual therapeutic approach.

Expression of ERCC1 protein in biopsy specimen predicts survival in advanced ovarian cancer patients treated with platinum-based chemotherapy.

PURPOSE: The purpose of this study was to investigate whether the expression of excision repair cross complementing 1 (ERCC1) protein I in tumor tissue was associated with resistance to standard carboplatin and paclitaxel (PC) combination chemotherapy in patients newly diagnosed with advanced epithelial ovarian carcinoma (EOC). METHODS: Fresh frozen tumor tissue was obtained from EOC patients. The protein expression levels of ERCC1 in tumor tissue were determined by Western blot analysis in 55 samples with advanced and metastatic EOC with different histologic subtypes; then these patients were treated with PC. RESULTS: The results showed that the clinical objective responses were significantly different in different categories of ERCC1 protein expression levels in patients with EOC. Time to progression (TTP) and overall survival (OS) in EOC patients previously treated with platinum-based chemotherapy were significantly longer in those with low expression compared with patients showing high expression of ERCC1 protein. CONCLUSION: Our results revealed that ERCC1 protein expression could potentially be used to customize chemotherapy by defining subsets of patients who would benefit the least from platinum-based chemotherapy.

Latest Developments in the Management of Nut Allergies.

PURPOSE OF REVIEW: In this review, we sought to describe the most recent advances in the dietary and medical management of peanut and tree nut allergy, including selective introduction and immunotherapy. RECENT FINDINGS: Dietary updates include changes to labeling laws, improved information sources, and new apps for buying foods in shops and overseas to better protect individuals with nut allergies. There are still issues in the management of nut allergies in schools, such as parents having to resort to packed lunches instead of school meals and patients experiencing bullying. Air travel also poses concern, but additional resources are now available to travelers, and recent evidence suggest limited airborne exposure to nuts. The medical management of anaphylaxis is use of epinephrine; however, this remains underutilized. Needle length and administration devices have been recently debated considering the risk of bone penetration vs subcutaneous administration, and autoinjectors seem to deliver higher peak concentrations than syringes. Selective nut introduction has gained momentum in the last 5 years, demonstrating improved quality of life but with the need for motivated parents for continued consumption and available resources for challenges. Immunotherapy to nuts is also a rapidly developing field, with the balance of efficacy and safety being important considerations in the differing modes of administration. SUMMARY: The management of nut allergies is a rapidly developing field, and dietary and medical management have progressed significantly in the last 5 years. Future research directions include improving safety and efficacy of food immunotherapy and examining patients' goals for therapy and treatment outcomes.

Filaggrin loss-of-function mutations are associated with early-onset eczema, eczema severity and transepidermal water loss at 3 months of age.

BACKGROUND: Filaggrin loss-of-function (FLG) mutations are associated with eczema and skin barrier impairment, but it is unclear whether skin barrier impairment precedes phenotypic eczema in FLG mutation carriers. OBJECTIVES: To study the association between FLG mutations, skin barrier impairment and clinical eczema at 3 months of age. METHODS: A total of 88 infants were examined for eczema. Disease severity was determined by the SCORAD eczema severity score. Transepidermal water loss (TEWL) was measured on unaffected forearm skin. Venous blood samples were screened for the four most common FLG mutations found in the U.K. white population (R501X, 2282del4, R2447X and S3247X). Median SCORAD and TEWL measurements in children with and without eczema and FLG mutations were compared. RESULTS: Thirty-three per cent (29/88) of children had clinical eczema. Median SCORAD was 10·6 (range 3·5-31·0). TEWL (g m⁻² h⁻¹) was higher in children with eczema compared with unaffected infants (median TEWL 14·24 vs. 11·24, P < 0·001). Higher TEWL was associated with more severe disease (r = 0·59, P < 0·001, median TEWL, SCORAD < 15, 13·1 vs. 29·6, SCORAD ≥ 15, P = 0·029). Clinically dry skin was associated with higher TEWL, even in the absence of eczema (median TEWL 17·55 vs. 11·08, P = 0·008). Seventeen per cent (15/88) of children carried at least one FLG mutation. FLG mutation carriers were significantly more likely to have clinically dry skin, even in the absence of eczema [odds ratio (OR) 8·50, 95% confidence interval (CI) 1·09-66·58, P = 0·042]. FLG mutation carriers were also more likely to have eczema by 3 months of age (OR 4·26, 95% CI 1·34-13·57, P = 0·014). FLG mutations were significantly associated with higher median TEWL (all children, FLG 'yes' 21·59 vs. FLG 'no' 11·24, P < 0·001), even without clinical eczema (FLG 'yes' 15·99 vs. FLG 'no' 10·82, P = 0·01). CONCLUSIONS: By the age of 3 months, FLG mutations are associated with an eczema phenotype, dry skin and TEWL. The observation that TEWL is elevated in unaffected FLG mutation carriers suggests that skin barrier impairment precedes clinical eczema.

p53 gene mutations and codon 72 polymorphism in ovarian carcinoma patients from Serbia.

PURPOSE: Ovarian cancer is the leading cause of death from gynecological malignancies. The early stages of this disease are asymptomatic and more than 75% of the cases are diagnosed with regional or distant metastases. p53 gene is frequently mutated in some histological subtypes of ovarian carcinomas. The role of p53 mutations and polymorphic variant of codon 72 in the prognosis of disease is still unclear. The aim of this study was to determine the frequency of p53 mutations and polymorphic variants of codon 72 among ovarian carcinoma patients and to correlate them with clinicopathological characteristics of disease. METHODS: 54 ovarian carcinoma patients were included in the study. DNA was isolated from tumor tissue by the salting- out method. p53 mutations in exons 4-8 were detected by PCR-SSCP (polymerase chain reaction - single-stranded conformational polymorphism) electrophoresis. Codon 72 polymorphism was assessed by RFLP (restriction fragment-length polymorphism) method. RESULTS: p53 mutations were present in 11 out of 54 patients (20.4%). Twenty-four patients (44.4%) exhibited Arg/ Arg, 24 patients (44.4%) Arg/Pro and 6 patients (11.2%) Pro/ Pro genotype of 72 codon polymorphism. Correlations between p53 mutations and various clinicopathological characteristics were not found. However, we observed that the frequency of Pro/Pro genotype was increasing with higher histological grade as well as in advanced compared to localized disease, but without statistical significance. Distribution of p53 gene mutations between Pro/Pro genotype and Arg/Pro plus Arg/Arg genotypes was not statistically significant. CONCLUSION: Our study suggests that Pro/Pro genotype of 72 codon polymorphism could be an independent prognostic marker in ovarian carcinomas.

siRNA and miRNA for the treatment of cancer.

In the process of RNA interference (RNAi), small RNAs pair with complementary messenger RNAs preventing their expression. The discovery of RNAi has revolutionized our understanding of gene regulation. Since cancer is a disease of altered genes, RNAi may have tremendous potential as a therapeutic strategy by downregulating altered genes. Just one decade after its discovery, this process is already being used in clinical trials and new technical achievements in delivering small RNAs to the cells are constantly improving the efficiency of this specific cancer treatment.

Therapeutic cancer vaccines in cervical cancer: phase I study of Lovaxin-C.

Producing effective therapeutic vaccines has proved much more difficult and challenging than developing cancer preventive vaccines. Despite huge research in the area of cancer immunology, FDA/EMEA have not approved any type of cancer treatment vaccine so far. More than 99% of cervical cancers have detectable amounts of human papillomavirus (HPV) DNA. Integration of high-risk HPV into the host cell genome is followed by continual expression of HPV E6 and E7 oncoproteins, making them excellent targets for developing vaccines which could be used in high grade precancerous (CIN) lesions or invasive cancer or in the prevention of cancer recurrence. Therapeutic cervical cancer vaccines have been extensively studied. Strategies used were vaccination with HPV peptides or proteins, alone or in pulsed dendritic cells, DNA vaccines, virus-like particles or viral and bacterial vectors. Lovaxin-C is a recombinant live-attenuated Listeria monocytogenes (Lm) that secretes the antigen HPV-16 E7 fused to a non-hemolytic listeriolysin O protein. In a phase I study Lovaxin-C was administered to advanced cervical cancer patients refractory to existing therapies. The dose-limiting toxicity was hypotension and flue-like syndrome. There were no serious adverse events. Specific T-cell response was detected as well as clinical response to Lovaxin-C. Several other therapeutic HPV vaccines are in clinical development and in most of the studies specific immunological and clinical responses were seen. Efficacious therapeutic vaccine for the treatment of cervical cancer should be expected in the near future.

Antitumor activity of Ru(III) complexes carrying beta-diketonato ligands in vitro and in vivo.

PURPOSE: To investigate the antitumor activity of two newly synthesized ruthenium(III) [Ru(III)] compounds carrying bidentate ligands: (acac)-acetylacetonate, [Ru(acac)3), and (tfac)-trifluoroacetylacetonate [Ru(tfac)3]. MATERIALS AND METHODS: The activity of ruthenium(III) analogues was evaluated on HeLa, B16, and Femx cell lines for cytotoxicity in vitro using MTT assay, and inhibition on tumor invading ability in vitro using cell migration and invasion assays, whereas inhibition of tumor growth in vivo was estimated on advanced B16 murine melanoma model. Both compounds were also investigated in combinations with cisplatin, oxaliplatin, or poly ADP-ribose polymerase- 1 (PARP-1) inhibitor, in order to determine the pattern of mutual interactions. RESULTS: Applied as single drugs, Ru(tfac)3 showed high cytotoxic activity against HeLa and Femx cell lines, while Ru(acac)3 did not reach the IC50 on any of the cell lines tested. In combinations, Ru(acac)3 with cisplatin gained synergistic interaction, antagonistic with oxaliplatin, and of different kind with (PARP-1) inhibitor in concentration-and cell line-dependent manner. Ru(acac)3 exhibited inhibition of HeLa cell migration and gelatinolytic activity of MMP-2 and MMP-9. Ru(tfac)3 complexes did not induce significant reduction of melanoma growth in vivo, whereas Ru(acac)3 did, but the latter failed to contribute in lifespan improvement. CONCLUSION: The investigated ruthenium complexes showed different levels of antitumor activity in vitro and in vivo, implicating on different mechanisms of their action as well as diverse perspectives in cancer treatment.

TP53 mutations in breast cancer: association with ductal histology and early relapse of disease.

PURPOSE: This study aimed to investigate the incidence of core domain TP53 mutations in Serbian breast cancer patients in view of their possible correlation with prognostic parameters, tumor characteristics and clinical disease course. METHODS: 145 breast cancer patients were included. Data on clinical disease course were available for 100 patients including 30 node-negative and 70 node-positive patients. After surgery, node-positive patients underwent adjuvant chemotherapy, mostly CMF. TP53 mutations were detected by PCR-SSCP. RESULTS: 31 mutations were found in 27/145 patients including 4/59 node-negative patients and 23/83 node-positive patients (4 double mutations). 26/31 TP53 mutations were found in patients with invasive ductal carcinoma and only 2 in patients with invasive lobular carcinoma. The presence of TP53 mutations was correlated with clinical disease course in premenopausal node-positive patients (n=70). 11/20 patients with TP53 mutations relapsed. Within the first 24 months of follow-up, significantly shorter disease-free intervals were observed in TP53-mutated patients. CONCLUSIONS: TP53 mutations correlated only with nodal status and ductal histology. The significance of the predominant distribution of TP53 mutations in tumors with a ductal histology for the aggressive behavior of these tumors has yet to be proved, since the favorable biological features of tumors with a lobular histology do not result in a better prognosis. Early relapse in mutated-TP53 carriers may support data on its predictive value with respect to adjuvant CMF.

p53 gene alterations and human papillomavirus type 16 infection in early stages of cervical carcinoma in Serbia.

PURPOSE: The incidence rate (age-standardized) of cervical carcinoma in Serbia is the highest in Europe. p53 is mainly inactivated at protein level in carcinomas associated with human papillomavirus (HPV) infection, such as cervical carcinomas. These tumors show low rate of p53 mutations. It is not clear if p53 mutations confer additional impact on disease prognosis. The role of polymorphic variant at codon 72 of p53 gene on patient's prognosis is controversial. The aim of this study was to determine the frequency of p53 mutations and to assess polymorphic variants of codon 72 among cervical carcinoma patients. PATIENTS AND METHODS: 53 patients, mainly FIGO stage I (n=50), with squamous cell carcinoma (n=49) were included. 30/32 (94%) patients who received adjuvant radiotherapy were followed-up (median 15 months, range 4-39). DNA was isolated by the salting out method from tumor tissue (n=53) and blood (42/53). p53 mutations were detected by PCR-SSCP (polymerase chain reaction - single-stranded conformational polymorphism) electrophoresis. Codon 72 polymorphism was assessed by the restriction fragment-length polymorphism method. RESULTS: Six p53 mutations were detected in 5/53 (9%) patients with FIGO stage I squamous cell carcinoma (one patient had double mutations). 25/42 (60%) patients exhibited Arg/Arg genotype. HPV16 type was detected in 29/51 (57%) cervical carcinoma samples. Relapse of disease occurred in only 2 patients- both with Arg/Arg genotype and HPV16 positive. One of them exhibited p53 mutation. CONCLUSION: Our results showed low incidence of p53 mutations and prevalence of Arg/Arg genotype polymorphic variant of codon 72 of p53 gene in early stages of cervical carcinoma.

Comparison between CoFactor and Leucovorin activity applied in combination with 5-fluorouracil against two human colon cancer cell lines.

PURPOSE: The reduced folate leucovorin (LV) has been used for over 30 years as an enhancer of 5-fluorouracil (5-FU) antitumor activity. CoFactor (CoF) is a calcium salt of LV's active metabolite. Thus, administration of CoF can prevent decrease of intratumoral folate, which was recognized as the main reason for resistance of tumors to 5-FU treatment. The aim of this study was to investigate the cytotoxic activity and type of interactions for 5-FU and CoF applied in combination, in comparison to combination of 5-FU and LV, against human colon cancer cell lines LS- 174 and HT-29. METHODS: The single agent activity was estimated in concentration range from 0.1 to 300 microM for 5-FU, CoF, and LV. In combined treatment each concentration of 5-FU (0.1, 1 and 10 microM) was matched with each concentration of CoF or LV (1, 10 and 100 microM). In both experiments the sulforodamine B cytotoxic test was used to assess drugs' activity after 72 h of incubation with cells. The type of interactions in combinations was calculated with the isobole method. RESULTS: 5-FU showed similar activity against LS-174 and HT-29 cell lines (IC50=30.8 and 21.6 microM, respectively). While LV affected both cell lines in a slightly proliferative manner, CoF showed cytotoxic effect on both cell lines, being more active against HT-29 cell (IC50=25.7 microM) and less active on LS-174 cells (IC50=72.6 microM). Addition of LV did not change the cytotoxicity of 5-FU, especially against LS-174 cells, and synergism was found only in HT-29 cells when LV was applied in high concentrations (10 and 100 microM). However, the combination of 5-FU with CoF revealed synergistic and additive interactions in LS-174 and HT-29 cells, respectively. CONCLUSION: These results should be taken into consideration when planning or analyzing the results of clinical trials using combination of CoF and 5-FU.

Dose-finding study of capecitabine in combination with weekly paclitaxel for patients with anthracycline-pretreated metastatic breast cancer.

PURPOSE: Capecitabine and paclitaxel show high efficacy, non-overlapping toxicity profiles and preclinical synergism, providing the rationale for their combination in metastatic breast cancer (MBC). This dose-escalation study aimed at determining the maximum tolerated dose (MTD) of capecitabine plus paclitaxel in anthracycline-pretreated MBC patients. PATIENTS AND METHODS: Patients with MBC received fl at-dose of oral capecitabine (1,000 mg/m(2) twice daily, days 1-14) plus weekly paclitaxel 60, 75, or 90 mg/m(2), i.v., days 1, 8 and 15, every 3 weeks. RESULTS: All 11 patients enrolled onto study were evaluable for toxicity and response. Two patients receiving paclitaxel 75 mg/m(2) experienced grade 3 nail toxicity, with grade 3 hand-foot syndrome (HFS) in one patient and grade 2 dermatitis in the other. Although not life-threatening, these were considered unacceptable and the preceding dose level was selected. Eight of 11 patients achieved objective responses. CONCLUSION: The recommended regimen is capecitabine 1,000 mg/m(2) twice daily, days 1-14, plus paclitaxel 60 mg/m(2)/week. Escalation of the paclitaxel dose above 60 mg/m(2)/week is not feasible due to severe skin toxicity.

Proliferative characteristics of Philadelphia-negative myeloproliferative neoplasms - clinical implications.

INTRODUCTION: Philadelphia-negative myeloproliferative neoplasms (Ph- MPN) are characterized by overproduction of one or more blood cell lines. METHODS: We studied the proliferative characteristics of 91 patients with de novo Ph- MPN. Colony-forming cells (CFC) and endogenous colonies (EC), from bone marrow (BM) and/or peripheral blood (PB), were analyzed by colony assay based on methylcellulose. The level of circulating CD34+ cells was determined by flow cytometry. RESULTS: The total number of PB CFC in primary myelofibrosis (PMF) was increased compared to the control sample (P < 0.01) and essential thrombocythemia (ET) (P < 0.05). The highest number of BM and PB EC was observed in polycythemia vera (PV) (P < 0.01). Increased levels of CD34+ cells characterized early-prefibrotic (57%) and advanced-fibrotic PMF (90%) as compared to PV (34%) and ET (32%) (P < 0.01). In the whole Ph- MPN group, the total number of PB CFC (P < 0.01), PB EC (P < 0.05), and CD34+ cells (P < 0.01) correlated with the degree of BM fibrosis. Higher levels of circulating CD34+ cells in PMF correlated with the total number of PB EC (P < 0.05) and degree of BM fibrosis (P < 0.01). CONCLUSIONS: Exploration of the PB proliferative characteristics of Ph- MPN on diagnosis may be helpful in revealing early-prefibrotic PMF. Monitoring the levels of circulating CD34+ cells may provide a sensitive indicator of fibrotic evolution in PV and PMF.

Growth inhibition of estrogen-dependent and estrogen-independent MXT mammary cancers in mice by the bombesin and gastrin-releasing peptide antagonist RC-3095.

BACKGROUND: Many breast cancers are estrogen independent, and even in patients who initially respond to estrogen suppression therapy, the regression is often temporary. We have recently shown that antagonists of bombesin and gastrin-releasing peptide, including RC-3095, inhibit the growth of pancreatic, colonic, and prostatic cancers in experimental animals. This effect was associated with a substantial decrease in epidermal growth factor (EGF) receptor levels in pancreatic and colon cancers. PURPOSE: In view of these findings, we investigated the effects of our synthetic bombesin and gastrin-releasing peptide receptor antagonist D-Tpi6,Leu13 psi (CH2NH)-Leu14 bombesin(6-14) (RC-3095) on the growth of hormone-dependent and hormone-independent MXT mouse mammary cancers in vivo. METHODS: Female (C57BL x DBA/2)F1 mice bearing estrogen-dependent or estrogen-independent MXT mammary carcinomas were treated with small doses (20 micrograms/d) of RC-3095 administered from osmotic minipumps. Separate groups of mice with estrogen-independent tumors received RC-3095, bombesin, or gastrin-releasing peptide(14-27) at 20 micrograms/d. We determined tumor volume and weight, mitotic index, apoptosis (programmed cell death), and argyrophilic nucleolar organizer regions, an indicator of tumor cell proliferation. Levels of receptors for EGF and bombesin were measured in tumor membrane fractions. RESULTS: Growth of both estrogen-dependent and estrogen-independent MXT breast cancers was significantly inhibited by RC-3095. Bombesin or gastrin-releasing peptide had no effect on the growth of estrogen-independent tumors. Inhibition of tumor cell proliferation was indicated by a 45%-65% reduction in tumor volume, a 35%-58% reduction in tumor weight, and statistically significant decreases in argyrophilic nucleolar organizer region counts after treatment with RC-3095. In estrogen-independent cancers, tumor inhibition was associated with a decrease in the capacity of EGF receptors from 0.21 +/- 0.016 pmol/mg membrane protein in controls to 0.03 +/- 0.003 pmol/mg membrane protein in the RC-3095-treated group. CONCLUSIONS: This is the first demonstration of inhibitory effects of bombesin and gastrin-releasing peptide antagonists on the growth of breast cancers in vivo. IMPLICATIONS: These findings suggest that bombesin antagonists should be considered for breast cancer therapy.

Cellular sensitivity to beta-diketonato complexes of ruthenium(III), chromium(III) and rhodium(III).

The aim of this study was to investigate cellular response to several ruthenium(III), chromium(III) and rhodium(III) compounds carrying bidentate beta-diketonato ligands: [(acac)--acetylacetonate ligand, (tfac)--trifluoroacetylacetonate ligand]. Cell sensitivity studies were performed on several cell lines (A2780, cisplatin-sensitive and -resistant U2-OS and U2-OS/Pt, HeLa, B16) using growth-inhibition assay. Effect of intracellular GSH depletion on cell sensitivity to the agents was analyzed in A2780 cells. Flow cytometry was used to assess apoptosis by Annexin-V-FITC/PI staining, and to analyze induction of caspase-3 activity. Possible DNA binding/damaging affinity was investigated, by inductively coupled mass spectrometry, and by 14C-thymidine / 3H-uridine incorporation assay. Cell sensitivity studies showed that the pattern of sensitivity to Ru(tfac)3 complex of the two cisplatin-sensitive/-resistant osteosarcoma cell lines, U2-OS and U2-OS/Pt, was similar to that of A2780 cells (72 h exposure), with the IC50 being around 40 microM. The growth-inhibitory effect of Ru(acac)3 ranged over 100 microM, while Cr(III) and Rh(III) complexes were completely devoid of antitumor action in vitro. Ru(tfac)3 exhibited strong potential for apoptosis induction on A2780 cells (up to 40%) and caused cell cycle arrest in the S phase as well as decrease of the percent of G1 and G2 cells. Ru(acac)3-induced apoptosis was slightly higher than 10%, whereas activation of caspase-3 in HeLa cells was moderate. DNA binding study revealed that only Cr(acac)3 was capable of binding DNA, while Cr(III) and Ru(III) compounds possess potential to inhibit DNA/RNA synthesis. In conclusion, only Ru(III) complexes showed potential for antitumor action.

Inhibition of growth of HT-29 human colon cancer xenografts in nude mice by treatment with bombesin/gastrin releasing peptide antagonist (RC-3095).

Nude mice bearing xenografts of HT-29 human colon cancer cell line were treated for 4 weeks with a [D-Trp6] agonist of luteinizing hormone releasing hormone (LH-RH), somatostatin analogue (RC-160), and bombesin/gastrin releasing peptide antagonist (RC-3095). Some inhibitory effect of [D-Trp6] LH-RH microcapsules releasing 25 micrograms/day on tumor growth was observed that could be due to sex steroid deprivation, but the inhibition was not statistically significant. Microcapsules of RC-160, releasing 50 micrograms/day, significantly reduced tumor volume after 21 and 24 days of treatment, but at the end of the experiment the inhibition in tumor volume and weight was not significant. Bombesin/gastrin releasing peptide antagonist RC-3095, at a dose of 20 micrograms/day administered by daily s.c. injections or by continuous infusion using Alzet osmotic minipumps, had the greatest inhibitory effect on tumor growth. Tumor volume, percentage change in tumor volume, and tumor weights were significantly decreased in both groups treated with RC-3095. This is the first report on inhibition of human colon cancer growth in vivo by bombesin antagonists.

Inhibitory effect of bombesin/gastrin-releasing peptide antagonist RC-3095 and high dose of somatostatin analogue RC-160 on nitrosamine-induced pancreatic cancers in hamsters.

Female Syrian golden hamsters with N-nitrosobis(2-oxopropyl)amine-induced pancreatic cancers were treated for 2 months with new pseudononapeptide bombesin receptor antagonist [D-Tpi6,Leu13 psi (CH2NH)-Leu14]bombesin(6-14)(RC-3095), administered s.c. with implanted osmotic minipumps releasing 20 micrograms/day of the analogue. The results were compared to those obtained by treatment with somatostatin analogue RC-160 (35 micrograms/day and 150 micrograms/day) or [D-Trp6]luteinizing hormone-releasing hormone (25 micrograms/day), which inhibited the growth of pancreatic cancers in our previous studies. A new acetylated somatostatin analogue [formula: see text] (30 micrograms/day) also was used for comparison of therapeutic response. All peptide analogues induced tumor inhibition by at least one of the measured parameters. Bombesin antagonist RC-3095 and high dose of RC-160 (150 micrograms/day) had the greatest inhibitory effect on pancreatic cancers: A significant decrease in the number of animals with tumors, reduced pancreatic weight, 87-89% inhibition of tumorous pancreas weight, and a significant diminution in the number of tumor nodules and argyrophilic nucleolar organizer region count in tumor cell nuclei were observed in the groups treated with these regimens. We were able to detect receptors for bombesin in membranes of N-nitrosobis(2-oxopropyl)amine-induced pancreatic tumors and these receptors were not down-regulated after treatment with the bombesin antagonist. In hamsters treated with bombesin antagonists, tumor inhibition might be explained by a significant decrease in the binding capacity of epidermal growth factor receptors in pancreatic cancers. The acetylated somatostatin analogue RC-160-II had a similar inhibitory effect on the tumors as the original analogue RC-160. Our results suggest that the increase in the dose of RC-160 improves the therapeutic response, and this finding should be taken into account in clinical use of this somatostatin analogue. In view of its strong inhibitory effect on experimental pancreatic tumors, the bombesin antagonist RC-3095 might be considered as a possible new agent for the therapy of human exocrine pancreatic cancer.