Diagnosis of a neonatal ophthalmic discharge, Ophthalmia neonatorum, in the molecular age: investigation for a correct therapy.

An early double case of acute Ophthalmia neonatorum in 3-day-old twins is reported. Culture of eye swabs showed a wide bacterial polymorphism, in which common bacteria, such as Klebsiella pneumoniae, Streptococcus pneumoniae, Corynebacterium ulcerans and other Enterobacteriaceae, coexisted with atypical Mycoplasmataceae and Chlamydiaceae from resident cervical-vaginal maternal microbiota. The neonates were in an apparently healthy state, but showed red eyes with abundant greenish-yellow secretion, mild chemosis and lid edema. The maternal cervical-vaginal ecosystem resulted differently positive to the same common cultivable, atypical bacteria culturally and molecularly determined. This suggested a direct maternal-foetal transmission or a further foetal contamination before birth. An extended culture analysis for common bacteria to atypical ones was decisive to describe the involvement of Mycoplasmas (M. hominis and U. urealyticum) within the scenario of the Ophthalmia neonatorum in a Caucasian couple. The introduction of a routine PCR molecular analysis for Chlamydiaceae and N. gonorrhoeae allowed to establish which of these were present at birth, and contributed to determine the correct laboratory diagnosis and to define an adequate therapeutic protocol obtaining a complete resolution after one year for culture and atypical bacteria controls. This study suggests to improve the quality of laboratory diagnosis as unavoidable support to a correct clinical diagnosis and therapy, in a standardized modality both for swabbing and scraping, to check the new-born microbial programming starting in uterus, overtaking the cultural age to the molecular age, and to revise the WHO guidelines of SAFE Strategy for trachoma eye disease, transforming it into SAFES Strategy where the S letter is the acronym of Sexual ecosystem and behavioural valuation/education.

Early deep anterior lamellar keratoplasty for fungal keratitis poorly responsive to medical treatment.

PurposeTo investigate the efficacy of early therapeutic deep anterior lamellar keratoplasty (DALK) in eradicating fungal keratitis that is poorly responsive to medical treatment.Patients and methodsTwenty-three eyes (23 patients) underwent early therapeutic DALK within 15 to 50 days from the onset of symptoms. The adopted eligibility criteria for early DALK included the following: active fungal keratitis affecting the optical zone with ulcer confined in the 6.00 mm central cornea; deeper than 150 µm but not exceeding 300 µm; and poorly responsive to medical treatment.ResultsThe big bubble technique was accomplished in 74% (17) of eyes, whereas manual dissection was performed in the remaining 26% (6) of eyes. Histopathological examination did not show any sign of fungal colonization in the peripheral and deep stromal lamellae in any case. All grafts were transparent postoperatively, and no recurrence of infection occurred. Median best spectacle corrected visual acuity significantly improved from 2.0 (1.0 interquartile range) logMAR to 0.1 (0.1 interquartile range) logMAR (P<0.01). The mean follow-up was 32±10 months. Neither episode of rejection nor graft failure was noted during the follow-up period.ConclusionEarly DALK could represent a safe therapeutic approach to eradicate fungal keratitis that affects the optical zone and is poorly responsive to medical treatment.

The Role of Gene Therapy in the Treatment of Retinal Diseases: A Review.

BACKGROUND: Gene therapy represents the therapeutic delivery of nucleic acid polymers into patient cells with the aim of treating an underlying disease. Over the past 2 decades this new therapy has made substantial progress owing to better understanding of the pathobiologic basis of various diseases coupled with growth of gene transfer biotechnologies. The eye, in particular, represents a suitable target for such therapy due to the immune privilege provided by the blood-ocular barrier, the ability to directly visualize, access and locally treat the cells and the minimal amount of vector needed given the size of this organ. It is not surprising therefore that several clinical trials are now ongoing in this field. OBJECTIVE: The purpose of this review was to provide an update on gene therapy for retinal diseases, discussing differences in treatment strategies, vector designs and surgical techniques. METHOD: Research was performed on PubMed, ClinicalTrials.gov, and Home Genetic Reference. We additionally utilized the internet database for genetics of retinal diseases, the portal for rare diseases and orphan drugs and the NCBI database Online Mendelian Inheritance in Man. No restriction was applied on the language of publications. RESULTS: We present the available results of current active clinical trials for inherited retinal disease such as Leber's congenital amaurosis type 2, choroideremia, Stargardt disease, achromatopsia and juvenile X-linked retinoschisis. We also illustrate a new approach of this therapy for the treatment of much more common ocular diseases such as age-related macular degeneration and diabetic retinopathy. CONCLUSION: Gene therapy represents an emerging and promising therapeutic approach for the treatment not only of rare inherited retinal diseases but also much more common retinal pathologies.

A challenge to the striking genotypic heterogeneity of retinitis pigmentosa: a better understanding of the pathophysiology using the newest genetic strategies.

Retinitis pigmentosa (RP) is a group of inherited retinal disorders characterized by a complex association between tremendous genotypic multiplicity and great phenotypic heterogeneity. The severity of the clinical manifestation depends on penetrance and expressivity of the disease-gene. Also, various interactions between gene expression and environmental factors have been hypothesized. More than 250 genes with ~4500 causative mutations have been reported to be involved in different RP-related mechanisms. Nowadays, not more than the 50% of RPs are attributable to identified genes, whereas the rest of molecular defects are still undetectable, especially in populations where few genetic screenings have been performed. Therefore, new genetic strategies can be a remarkably useful tool to aid clinical diagnosis, potentially modifying treatment options, and family counseling. Genome-wide analytical techniques (array comparative genomic hybridization and single-nucleotide polymorphism genotyping) and DNA sequencing strategies (arrayed primer extension, Sanger sequencing, and ultra high-throughput sequencing) are successfully used to early make molecular diagnosis detecting single or multiple mutations in the huge heterogeneity of RPs. To date, further research needs to be carried out to better investigate the genotype/phenotype correlation, putting together genetic and clinical findings to provide detailed information concerning the risk of RP development and novel effective treatments.

Estrogen Receptors and Chronic Venous Disease.

OBJECTIVE/BACKGROUND: Chronic venous disease (CVD) is a common and relevant problem affecting Western people. The role of estrogens and their receptors in the venous wall seems to support the major prevalence of CVD in women. The effects of the estrogens are mediated by three estrogen receptors (ERs): ERα, ERß, and G protein-coupled ER (GPER). The expression of ERs in the vessel walls of varicose veins is evaluated. METHODS: In this prospective study, patients of both sexes, with CVD and varicose veins undergoing open venous surgery procedures, were enrolled in order to obtain vein samples. To obtain control samples of healthy veins, patients of both sexes without CVD undergoing coronary artery bypass grafting with autologous saphenous vein were recruited (control group). Samples were processed in order to evaluate gene expression. RESULTS: Forty patients with CVD (10 men [25%], 30 women [75%], mean age 54.3 years [median 52 years, range 33-74 years]) were enrolled. Five patients without CVD (three men, two women [aged 61-73 years]) were enrolled as the control group. A significant increase of tissue expression of ERα, ERß and GPER in patients with CVD was recorded (p < .01), which was also related to the severity of venous disease. CONCLUSION: ERs seem to play a role in CVD; in this study, the expression of ERs correlated with the severity of the disease, and their expression was correlated with the clinical stage.

Treatment of exophthalmos and strabismus surgery in thyroid-associated orbitopathy.

Endocrine orbitopathy (EO) can have important consequences, such as exophthalmos and restrictive strabismus. A retrospective study was performed of 35 patients with EO who underwent orbital decompression surgery and restrictive strabismus correction. Two surgical techniques for orbital decompression were analyzed: fat decompression by Olivari technique and three-wall bony expansion with fat decompression. Strabismus surgery was performed using adjustable or non-adjustable sutures under topical anaesthesia. Patients were divided into two groups according to the type of intra-orbital decompression performed, and the postoperative values resulting from the different fat decompression techniques were recorded. The preoperative and postoperative mean degrees of exophthalmos were 22.3 and 19.9mm, respectively, for the fat decompression group, and 24.3 and 19.8mm, respectively, for the bony expansion with transpalpebral fat decompression (combined form) group. The difference in residual prism dioptres between adjustable and non-adjustable suture techniques in patients who had previously undergone combined decompression was statistically significant. The management of patients with EO requires a multidisciplinary approach based on the collaboration of maxillofacial surgeons, ophthalmologists, and orthoptists. These results will allow the development of a more adequate strategy for the surgical treatment of restrictive strabismus in EO patients.

sTRAIL coupled to liposomes improves its pharmacokinetic profile and overcomes neuroblastoma tumour resistance in combination with Bortezomib.

Neuroblastoma (NB), the most common and deadly extracranial solid tumour of childhood, represents a challenging in paediatric oncology. Soluble tumour necrosis factor (TNF)-related apoptosis-inducing ligand (sTRAIL) is a cancer cell-specific molecule exerting remarkable anti-tumour activities against paediatric malignancies both in vitro and in preclinical settings. However, due to its too fast elimination and to the undesired related side effects, the improvement of sTRAIL in vivo bioavailability and the specific delivery to the tumour is mandatory for increasing its therapeutic efficacy. In this manuscript, we developed an innovative pegylated liposomal formulation carrying the sTRAIL at the outer surface (sTRAIL-SL) with the intent to improve its serum half-life and increase its efficacy in vivo, while reducing side effects. Furthermore, the possibility to combine sTRAIL-SL with the proteasome inhibitor Bortezomib (BTZ) was investigated, being BTZ able to sensitize tumour cells toward TRAIL-induced apoptosis. We demonstrated that sTRAIL preserved and improved its anti-tumour activity when coupled to nanocarriers. Moreover, sTRAIL-SL ameliorated its PK profile in blood allowing sTRAIL to exert a more potent anti-tumour activity, which led, upon BTZ priming, to a statistically significant enhanced life spans in two models of sTRAIL-resistant NB-bearing mice. Finally, mechanistic studies indicated that the combination of sTRAIL with BTZ sensitized sTRAIL-resistant NB tumour cells to sTRAIL-induced cell death, both in vitro and in vivo, through the Akt/GSK3/ß-catenin axis-dependent mechanism. In conclusion, our results suggest that sTRAIL-SL might be an efficient vehicle for sTRAIL delivery and that its use in clinic, in combination with BTZ, might represent an adjuvant strategy for the treatment of stage IV, sTRAIL-resistant, NB patients.

Ruthenium-106 eye plaque brachytherapy in the conservative treatment of uveal melanoma: a mono-institutional experience.

BACKGROUND: Traditional treatment for uveal melanoma is the enucleation of the eye with outcomes cosmetically unacceptable and loss of useful vision. Plaque brachytherapy, compared to enucleation, had the advantage to preserve the eye with outcomes cosmetically acceptable and preservation of vision. PATIENTS AND METHODS: From July 1990 to December 2009 one hundred forty-two (142) patients (51 males and 91 females) with small to medium uveal melanoma were treated with 106Ru plaque brachytherapy. The patients underwent a complete staging before brachytherapy with indirect ophthalmoscopy and ultrasounds. Mean tumour thickness was 3.26 mm (1.6-6 mm). The dose scheduled was 80-100 Gy to the apex with a maximum dose of 800 Gy to the sclera. RESULTS: One hundred forty-two have been treated, nine patients had lost the follow-up and drop out; 133 patients were assessed. Mean follow-up was 7.7 years (6 months-18 years). The overall survival at 5, 10 and 15 years was 92%, 85% and 78% respectively. Cancer fee survival was 95%, 90% and 83%, respectively at 5, 10 and 15 year. Radiation-induced toxicity was represented in 47 patients with a 5 year actuarial survival rate free from complications of 54%. CONCLUSIONS: 106Ru plaque brachytherapy is a valid approach for treatment of uveal melanoma. This technique is efficacy and safe, with a low toxicity profile.

Prognostic factors and oncologic outcome in 146 patients with colorectal peritoneal carcinomatosis treated with cytoreductive surgery combined with hyperthermic intraperitoneal chemotherapy: Italian multicenter study S.I.T.I.L.O.

AIM: The present study was specifically designed to assess the major clinical and pathological variables of patients with colorectal peritoneal carcinomatosis in order to investigate whether currently used criteria appropriately select candidates for peritonectomy procedures (cytoreductive surgery) combined with hyperthermic intraperitoneal chemotherapy (HIPEC). PATIENTS AND METHODS: Preoperative, operative and follow-up data on 146 consecutive patients presenting with peritoneal carcinomatosis of colorectal origin and treated by surgical cytoreduction combined with HIPEC in 5 Italian Hospital and University Centers were prospectively entered in a common database. Univariate and multivariate analyses were used to assess the prognostic value of clinical and pathologic factors. RESULTS: Over a minimum 24-month follow-up, the overall morbidity rate was 27.4% (mortality rate: 2.7%) and was directly related to the extent of surgery. Peritoneal cancer index (PCI), unfavorable peritoneal sites, synchronous or previously resected liver metastasis and the completeness of cytoreduction, all emerged as independent prognostic factors correlated with survival. CONCLUSIONS: Until research provides more effective criteria for selecting patients based upon the biomolecular features of carcinomatosis, patients should be selected according to the existing independent prognostic variables.

Recent advances in targeted anti-vasculature therapy: the neuroblastoma model.

Novel anti-vasculature strategies that are emerging for the treatment of cancer and for the inhibition of angiogenesis may be a promising new tool for the adjuvant therapy of malignant tumours. Over the last fifteen years, several reports have been published concerning the relationship between tumour progression and angiogenesis in experimental models of neuroblastoma in vitro and in vivo. Moreover, a high vascular index in neuroblastoma correlates with poor prognosis, suggesting dependence of aggressive tumour growth on active angiogenesis. Here, we present an overview of the most recent advances in anti-vasculature therapy of neuroblastoma, and describe some preclinical results as well as future perspectives.

Hyperthermic isolated perfusion with tumor necrosis factor-alpha and doxorubicin for the treatment of limb-threatening soft tissue sarcoma: the experience of the Italian Society of Integrated Locoregional Treatment in Oncology (SITILO).

BACKGROUND: Tumor necrosis factor-alpha (TNFalpha)-based hyperthermic isolated limb perfusion (HILP) is routinely carried out at most oncological institutions in the treatment of locally advanced soft tissue limb sarcoma (STS), employing high TNFalpha dosages. After a phase I-II study, the SITILO (Italian Society of Integrated Locoregional Therapies in Oncology) centers began to employ the lower dose of 1 mg of TNFalpha. The aim of this paper is to report on the results obtained in 75 patients with limb-threatening STS treated with a low TNFalpha dose and doxorubicin (Dx). PATIENTS AND METHODS: HILP with TNFalpha (at a dosage of either 1 mg) and Dx was administered to 75 patients with limb-threatening STS: 37 males and 38 females; median age 50 years; tumor in the lower and upper limbs in 58 and 17 patients, respectively; primary and recurrent tumors in 45 and 30 patients, respectively. Most tumors (77%) were high grade. Tumor resection was carried out 6 to 8 weeks after HILP. RESULTS: The grade of limb toxicity was mild to moderate in the vast majority of patients (76%). Grades IV and V were observed, but only when high muscle temperatures were recorded and high TNFalpha dosages were employed. Systemic toxicity was also mild to moderate and there were no postoperative deaths. Complete and partial tumor responses were 34% and 48%, respectively, with an overall response of 82% . Limb sparing surgery was carried out in 85.3% of patients. At a median follow-up of 28 months, 16 recurrences (21.3%) were recorded, with a 5-year locoregional disease-free survival of 63% . The 5-year disease-free survival and overall survival were 36.7% and 61.6%, respectively. CONCLUSION: HILP with 1 mg of TNFalpha is an effective neoadjuvant therapy resulting in a high rate of limb sparing in limb-threatening STS, with acceptable local reactions and negligible systemic toxicity.

Prognostic factors influencing tumor response, locoregional control and survival, in melanoma patients with multiple limb in-transit metastases treated with TNFalpha-based isolated limb perfusion.

BACKGROUND: In isolated limb perfusion (ILP) with tumor necrosis factor-alpha (TNFalpha) and interferon (IFN)-gamma, pioneered by Lienard and Lejenne in 1988, TNFalpha was empirically employed at a dosage (3-4 mg) ten times higher than the systemic maximum tolerable dose (MTD). We previously conducted a phase I/II study in 20 patients with in-transit melanoma metastases, using a combination of melphalan and TNFalpha at dosages ranging from 0.5 to 3.3 mg. The dose of 1 mg of TNFalpha was identified as optimal in terms of both efficacy and toxicity. The aim of the present study was to describe our experience with 113 stage IIIA/IIIAB melanoma patients treated with a TNFalpha-based ILP and identify prognostic factors for response, locoregional control and survival. PATIENTS AND METHODS: Patients at stage IIIA-IIIAB (presence of in-transit metastases and/or regional node involvement) were considered eligible. The disease was bulky (>or=10 nodules3 cm) in 42.5% of the patients and unresectable in 33% . Forty patients were treated with a TNFalpha dosage of >1 mg and 73 with 1 mg. Patients with tumors in the upper and lower limbs were submitted to ILP via axillary and iliac vessels, respectively. TNFalpha was injected in the arterial line of an extracorporeal circuit at the pre-established dose, followed by melphalan (13 and 10 mg/l of limb volume for the upper and lower limbs, respectively) 30 minutes later. RESULTS: Complete responses (CR) and partial responses (PR) were 63% and 24.5%, respectively, with an objective response (OR) of 87.5%. No change (NC) was observed in only 12.5% of the patients. Upon multivariate analysis, only bulky disease maintained its independent value for tumor response with an odds ratio of 4.07 and a p-value of 0.02. The 5-year locoregional disease-free survival was 42.7%. Upon multivariate analysis, the only prognostic factors were stage, age and bulky disease. The 5-year overall survival was 49%. Multivariate analysis showed that only sex, stage and CR maintained their independent values. CONCLUSION: TNFalpha-based ILP was proven to be an effective treatment for melanoma patients with in-transit metastases. The TNFalpha dosage of 1 mg was as effective as 3-4 mg, with lower toxicity and cost. We propose that TNFalpha and melphalan-based ILP should be employed for bulky tumors or after failure of melphalan-based ILP.

PHOX2A and PHOX2B genes are highly co-expressed in human neuroblastoma.

The detection of PHOX2B mutations in a small proportion of patients affected with either familial or sporadic neuroblastoma (NB), has arisen interest on the possible pathogenic role of this gene in the disease determination. In this light, we have carried out a quantitative expression analysis of PHOX2B and its paralogue PHOX2A on a panel of NB cell lines and NB tumour samples to identify a possible differential expression between NB cells and their normal counterpart (adrenal medulla cells). Our results revealed that both PHOX2A and PHOX2B are over-expressed in tumour samples and NB cell lines. Particularly, the expression levels of the two genes in NB cell lines show a highly significant correlation, suggesting their possible synergistic role or a coordinated expression regulation. Furthermore, PHOX2 gene over-expression in NB tumours and cell lines suggests these genes may be widely involved in NB development through either a direct mechanism of up-regulation or a failure in maintaining proper transcript levels after embryonic development.

Docetaxel in the adjuvant therapy of HER-2 positive breast cancer patients.

Considering the clinical benefit of trastuzumab in advanced breast cancer, fi ve prospective adjuvant randomized trials have recently been completed and early results have been published. Two of them, (NSABP-B31 and NCCTG N9831), employed anthracycline-containing regimens with sequential paclitaxel, with or without trastuzumab. The third study, HERA trial, randomized patients after adjuvant chemotherapy into an observational arm, one or two years of trastuzumab. Results of these studies, after a median follow up of 2-3 years confirm a DFS and OS benefit for the experimental arms. The worst rate of cardiotoxicity, in terms of incidence of CHF, with the use of trastuzumab and anthracycline based regimens was 4.1% in the trastuzumab arm of the NSABP-B31 trial. Among the fi ve trastuzumab trials, two, BCIRG 006 and FinHer, employed docetaxel-based regimens. The innovative BCIRG 006 trial compared ACdocetaxel (T) with two trastuzumab-containing regimens, ACTH, and a non-anthracycline-containing regimens, TCH, with a clear advantage in DFS for both trastuzumab arms. Data from the second interim analysis indicate that, in the subgroup of patients without co-amplification of topoisomerase 2 (TOPO-2), the arm without trastuzumab (ACT) showed a DFS significantly poorer that in the other arms; moreover, if we consider the lower toxicity of TCH regimen in comparison with anthracycline-containing arms, the innovative statements offered by BCIRG 006 trial appear evident, and these findings opened an important question about the consolidated employment of anthracyclines in adjuvant setting.The FinHer trial was a small trial testing a short course of trastuzumab (9 weeks) concomitantly with a chemotherapy including docetaxel, and there was a significant advantage in DFS for the trastuzumab based arms, without relevant toxicity and without any cardiotoxicity. Although data from all trastuzumab adjuvant trials, and without particulary from BCIRG-006 and FinHer trials, appear very intriguing, further follow-up is required.

Sentinel node biopsy in cutaneous melanoma: correlations between melanoma prognostic factors and sentinel node status.

This study aims to correlate the most important prognostic factors of primary melanoma with sentinel node (SN) positive for metastases. We have enrolled 84 patients subjected to sentinel node biopsies for cutaneous melanomas of Breslow's thickness > or = 0.75 mm by using an intra-operative gamma probe after lymphoscintigraphy, without blue dye support. SN metastases were reported in 27% of cases (14% by histology and 13% by immunohistochemistry). By chi-square test Breslow's thickness > 2mm (p= 0.004), IV and V Clark's level (p= 0.02), ulceration (p= 0.05) and high mitotic rate (p= 0.05) were statistically significant (p < 0.05) with reference to SN positive for metastases, unlike the site of cutaneous melanoma, vertical growth phase, tumour infiltrating lymphocytes, regression and vascular invasion. Breslow's thickness remains the first prognostic factor to be considered for sentinel node biopsy in cutaneous melanoma, but other markers must be carefully estimated.

120 peritoneal carcinomatoses from colorectal cancer treated with peritonectomy and intra-abdominal chemohyperthermia: a S.I.T.I.L.O. multicentric study.

A multicentric study has been carried out on 120 patients affected by peritoneal carcinomatosis from colorectal cancer. Patients have been treated by cytoreductive surgery and intra-operative hyperthermic chemoperfusion (HIPEC) with cisplatin (CDDP) and mitomycin-c (MMC). A small group of patients were treated with oxaliplatin (LOHP) following the Elias et al. scheme [intravenous 5-fluorouracil (400 mg/m2) and leucovorin (20 mg/m2) followed by intraperitoneal perfusion with LOHP (460 mg/m2) in 2 l/m2, during 30 min at 43 degrees C]. CC-0 cytoreduction was achieved in 85.2% of the patients. Major morbidity and mortality was 22.5% and 3.3%, respectively. No G4 toxicity was registered. The three-year survival was 25.8%. The difference in survival evaluating complete cytoreduction (CC-0) vs. incomplete (CC1-2; residual tumor nodules greater than 2.5 mm) was statistically significant (p < 0.0001). Evaluating only the patients that could be cytoreduced to CC-0, the 3-year survival was raised to 33.5%. In our experience the peritoneal cancer index (PCI) has been demonstrated to be a weak prognostic factor reaching a statistical significance only after the exclusion of patients with resected hepatic metastases. The patients treated with oxaliplatin were alive and free-of-disease after a 16-month median follow-up.

[Doxorubicin in antiblastic hyperthermic perfusion in the treatment of advanced soft-tissue sarcoma of the limbs]. / La doxorubicina nella perfusione ipertermico antiblastica nel trattamento dei sarcomi avanzati dei tessuti molli degli arti.

Different antineoplastic drugs have been associated to hyperthermia in the treatment of advanced soft tissue limb sarcoma with a good results in terms of conservative surgery (77%-97%), locoregional control (77%-87%) and overall survival (72%). Two different studies were performed: the first was carried out to analyze the doxorubicin-TNFalpha-hyperthermia association (three different trials) in terms of toxicity and efficacy of the treatment (tumor response, locoregional control, disease free and overall survival). The results showed that the trimodality association (doxorubicin TNFalpha and hyperthermia) is the best regimen able to obtain a 77% of objective response and 77% of limb sparing in patients candidate to amputation but may result in high local toxicity if high temperatures (>41.5 degrees C) were maintained during perfusion. The second study describes the employment of liposomal doxorubicin in hyperthermic antiblastic perfusion (HAP) in terms of tumor response, conservative surgery and toxicity; the maximum tolerable dose (MTD) of doxorubicin was 16 mg/l of perfused limb volume at the temperature of 41.5 degrees C; the conservative surgery was possible in 91% of the cases and mild (grade I and II) toxicity was observed in the perfused limb with high temperature (>41.5 degrees C).