How we treat locoregional melanoma.

Cutaneous melanoma is the most lethal form of skin cancer and its incidence has been increasing in the past 30 years. Although this is completely resectable in most cases, thicker melanoma and those with regional lymph-node involvement are at a high risk of relapse. In recent years, the management of locoregional disease has drastically changed. In particular, in the 8th Edition of the American Joint Committee on Cancer (AJCC), subgroup classification of TNM (tumor-node-metastasis) has been modified, with the addition of the IIID stage. Furthermore, in recent randomized trials, completion lymph node dissection in case of sentinel lymph node biopsy positivity has not been shown to offer any improvement in overall survival versus observation. Consequently, radical dissection has been recommended as the standard treatment, but only in patients with palpable nodal metastases. However, the major novelty in the treatment of locally advanced melanoma has been the introduction of drugs, already used for metastatic disease, that have also shown clinical efficacy in the adjuvant setting. In fact, immunotherapies and, in the case of BRAF V600E/K-mutated melanoma, combination treatment of BRAF and MEK inhibitors have improved recurrence-free survival in these patients. In this paper, we will describe the current management of a patient with radically resectable melanoma and discuss the key points in light of the latest scientific evidence.

Triple blockade of EGFR, MEK and PD-L1 has antitumor activity in colorectal cancer models with constitutive activation of MAPK signaling and PD-L1 overexpression.

BACKGROUND: Molecular mechanisms driving acquired resistance to anti-EGFR therapies in metastatic colorectal cancer (mCRC) are complex but generally involve the activation of the downstream RAS-RAF-MEK-MAPK pathway. Nevertheless, even if inhibition of EGFR and MEK could be a strategy for overcoming anti-EGFR resistance, its use is limited by the development of MEK inhibitor (MEKi) resistance. METHODS: We have generated in vitro and in vivo different CRC models in order to underline the mechanisms of MEKi resistance. RESULTS: The three different in vitro MEKi resistant models, two generated by human CRC cells quadruple wild type for KRAS, NRAS, BRAF, PI3KCA genes (SW48-MR and LIM1215-MR) and one by human CRC cells harboring KRAS mutation (HCT116-MR) showed features related to the gene signature of colorectal cancer CMS4 with up-regulation of immune pathway as confirmed by microarray and western blot analysis. In particular, the MEKi phenotype was associated with the loss of epithelial features and acquisition of mesenchymal markers and morphology. The change in morphology was accompanied by up-regulation of PD-L1 expression and activation of EGFR and its downstream pathway, independently to RAS mutation status. To extend these in vitro findings, we have obtained mouse colon cancer MC38- and CT26-MEKi resistant syngeneic models (MC38-MR and CT26-MR). Combined treatment with MEKi, EGFR inhibitor (EGFRi) and PD-L1 inhibitor (PD-L1i) resulted in a marked inhibition of tumor growth in both models. CONCLUSIONS: These results suggest a strategy to potentially improve the efficacy of MEK inhibition by co-treatment with EGFR and PD-L1 inhibitors via modulation of host immune responses.

XB130, a tissue-specific adaptor protein that couples the RET/PTC oncogenic kinase to PI 3-kinase pathway.

XB130 is a recently cloned 130 kDa-adaptor protein and Src kinase substrate, structurally similar to actin-filament-associated protein. Here we show that XB130 is predominantly expressed in the thyroid. Given that XB130 is a thyroid-specific tyrosine kinase substrate, we asked whether it is targeted by RET/PTC, a genetically rearranged, constitutively active, thyroid-specific tyrosine kinase that plays a pathogenic role in papillary thyroid cancer. RET/PTC induced robust tyrosine phosphorylation of XB130, which promoted its subsequent association with the p85alpha subunit of phosphatidylinositol 3-kinase (PI 3-kinase). We identified tyrosine 54 of XB130 as the major target of RET/PTC-mediated phosphorylation and a critical binding site for the SH2 domains of p85alpha. Importantly, downregulation of XB130 in TPC1 papillary thyroid cancer cells, harboring the RET/PTC1 kinase, strongly reduced Akt activity without altering ERK1/2 phosphorylation, and concomitantly inhibited cell-cycle progression and survival in suspension. In conclusion, XB130 is a novel substrate of the RET/PTC kinase that links RET/PTC signaling to PI 3-kinase activation, and thereby plays an important role in sustaining proliferation and survival of thyroid tumor cells.

Biochemical and molecular characterization of the novel BRAF(V599Ins) mutation detected in a classic papillary thyroid carcinoma.

Activating mutations of the BRAF gene are the most common genetic alterations in papillary thyroid carcinomas (PTCs) and the T1799A transversion, resulting in BRAFV600E, appeared virtually unique in this cancer type. Here, we report on the identification in a classic PTC of a novel BRAF mutation, namely a 1795GTT insertion, resulting in BRAFV599Ins, and describe its biochemical and molecular characterization. Kinase assays carried out on BRAFV599Ins and BRAFV600E revealed a three- to five-fold increase in the enzymatic activity of both mutants with respect to BRAFWT. Similarly, evaluation of BRAF-induced phosphorylation of MEK, MAPK and RSK revealed a significant MAPK cascade activation in cells expressing BRAFV599Ins or BRAFV600E, but not in cells expressing BRAFWT. Molecular dynamic simulations showed a destabilization of the inactive conformation of the enzyme in both BRAFV599Ins and BRAFV600E mutants, but not in BRAFWT. The analysis of the interaction energies inside the catalytic site allowed to demonstrate the presence of repulsive electrostatic forces acting on the activation loop and moving from inward to outward of the mutant enzymes. Finally, focus assays in NIH-3T3 cells confirmed a high transformation rate in the cells transfected either with BRAFV599Ins or BRAFV600E. In conclusion, this study demonstrated that BRAFV599Ins, as BRAFV600E, is a 'gain of function' mutation, characterized by a constitutive catalytic activation, which accounts for its causative role in the studied PTC.

A new germline RET mutation apparently devoid of transforming activity serendipitously discovered in a patient with atrophic autoimmune thyroiditis and primary ovarian failure.

Gain-of-function RET mutations are responsible for multiple endocrine neoplasia syndromes (MEN) 2A and 2B and familial medullary thyroid carcinoma (FMTC), whereas loss-of-function mutations are found in Hirschsprung disease. We report a new RET point mutation [R694Q (CGG-->CAG)], serendipitously found in a 23-yr-old woman with hypothyroidism due to atrophic Hashimoto's thyroiditis and primary ovarian failure, without altered calcitonin secretion. Familial history and clinical and biochemical evaluation of first-degree relatives were negative for FMTC, MEN 2A and 2B, and Hirschsprung disease. Genetic analysis showed that the mutation was inherited from the mother, who was submitted 2 yr before to thyroidectomy for goitrous Hashimoto's thyroiditis. Histological revision and immunohistochemical studies documented normal C cell number and morphology. We cloned the mutation in an expression vector encoding a full-length RET protein. The construct was transiently expressed in 293T cells in parallel with a wild-type RET and a C634R MEN 2A-associated RET mutant. Proteins were harvested from transfected cells, and tyrosine phosphorylation levels were assayed. The mutation did not exert significant potentiating effects on RET kinase. A focus assay was also performed on NIH3T3 fibroblasts; the mutant did not exert significant transforming activity. In conclusion, a new RET mutation was found in two subjects without any evidence of MEN and FMTC. In keeping with clinical data, transfection studies confirmed lack of activating activity. This serendipitous discovery, apparently devoid of oncogenic potential, underscores the problems that may be encountered in genomic studies on RET.

Effect of 17á-estradiol or alendronate on the bone densitometry, bone histomorphometry and bone metabolism of ovariectomized rats

The objective of the present study was to evaluate the effect of 17á-estradiol or alendronate in preventing bone loss in 3-month-old ovariectomized Wistar rats. One group underwent sham ovariectomy (control, N = 10), and the remaining three underwent double ovariectomy. One ovariectomized group did not receive any treatment (OVX, N = 12). A second received subcutaneous 17á-estradiol at a dose of 30 æg/kg for 6 weeks (OVX-E, N = 11) and a third, subcutaneous alendronate at a dose of 0.1 mg/kg for 6 weeks (OVX-A, N = 8). Histomorphometry, densitometry, osteocalcin and deoxypyridinoline measurements were applied to all groups. After 6 weeks there was a significant decrease in bone mineral density (BMD) at the trabecular site (distal femur) in OVX rats. Both alendronate and 17á-estradiol increased the BMD of ovariectomized rats, with the BMD of the OVX-A group being higher than that of the OVX-E group. Histomorphometry of the distal femur showed a decrease in trabecular volume in the untreated group (OVX), and an increase in the two treated groups, principally in the alendronate group. In OVX-A there was a greater increase in trabecular number. An increase in trabecular thickness, however, was seen only in the OVX-E group. There was also a decrease in bone turnover in both OVX-E and OVX-A. The osteocalcin and deoxypyridinoline levels were decreased in both treated groups, mainly in OVX-A. Although both drugs were effective in inhibiting bone loss, alendronate proved to be more effective than estradiol at the doses used in increasing bone mass

Effect of impact load and active load on bone metabolism and body composition of adolescent athletes.

PURPOSE: It is unclear whether adolescents involved in nonweight-bearing activities experience a delay in bone growth acquisition and sexual maturation. The purpose of this study was to compare bone mineral density (BMD), body composition, hormonal profile, and bone biochemical markers of adolescent athletes active in sports involved in impact load sports with those participating in active load sports. METHODS: Forty-five male Caucasian athletes aged 12--18 yr were divided into two groups according to type of skeleton loading, impact (N = 18), or active (N = 27). Twenty-four male Caucasian adolescents (12--18 yr) served as controls and only performed the activities included in their physical education classes. All subjects were assessed for bone mass, body composition, and bone age by dual x-ray absorptiometry (DXA). Serum calcium (Ca), phosphorus (P), bone-specific alkaline phosphatase (BAP), total testosterone, FSH, LH, urinary calcium to creatinine ratio (Ca/Cr), and urinary deoxypyridinoline crosslinks to creatinine ratio (DPD/Cr) were measured. RESULTS: The impact load group presented the highest BMD among the three groups for all studied sites. Lean mass and absolute weight were correlated with all of the bone mass measurements. BAP levels were significantly higher and testosterone levels significantly lower in the active load group compared with the impact group. CONCLUSION: High-impact load exercises have a beneficial effect on bone mass in male adolescents. There is also a positive correlation of weight and body composition with BMD. However, further longitudinal studies are necessary to determine whether there is a delay in bone growth acquisition among adolescents involved in a nonweight-bearing exercise regimen and its association with sex hormones.

Effect of 17beta-estradiol or alendronate on the bone densitometry, bone histomorphometry and bone metabolism of ovariectomized rats.

The objective of the present study was to evaluate the effect of 17beta-estradiol or alendronate in preventing bone loss in 3-month-old ovariectomized Wistar rats. One group underwent sham ovariectomy (control, N = 10), and the remaining three underwent double ovariectomy. One ovariectomized group did not receive any treatment (OVX, N = 12). A second received subcutaneous 17beta-estradiol at a dose of 30 microg/kg for 6 weeks (OVX-E, N = 11) and a third, subcutaneous alendronate at a dose of 0.1 mg/kg for 6 weeks (OVX-A, N = 8). Histomorphometry, densitometry, osteocalcin and deoxypyridinoline measurements were applied to all groups. After 6 weeks there was a significant decrease in bone mineral density (BMD) at the trabecular site (distal femur) in OVX rats. Both alendronate and 17beta-estradiol increased the BMD of ovariectomized rats, with the BMD of the OVX-A group being higher than that of the OVX-E group. Histomorphometry of the distal femur showed a decrease in trabecular volume in the untreated group (OVX), and an increase in the two treated groups, principally in the alendronate group. In OVX-A there was a greater increase in trabecular number. An increase in trabecular thickness, however, was seen only in the OVX-E group. There was also a decrease in bone turnover in both OVX-E and OVX-A. The osteocalcin and deoxypyridinoline levels were decreased in both treated groups, mainly in OVX-A. Although both drugs were effective in inhibiting bone loss, alendronate proved to be more effective than estradiol at the doses used in increasing bone mass.

Primary intraosseous carcinoma of the mandible: a case report.

A case of primary intraosseous carcinoma of the mandible in a 75-year-old man is reported. This tumor is an uncommon lesion arising from odontogenic rests. This tumor affects men more than women and is more frequent in the sixth and seventh decades of age. Most tumors occur in the posterior mandible as painful, non-ulcerated lesions, even if several cases have shown complete absence of subjective symptoms in early phases; often they are detected first on routine radiographs. Radiographically the lesion usually shows a fully enclosed, irregular pattern of bone destruction, even if sometimes the margins are well defined. The accepted treatment is radical surgery; but metastases could occur in lymph nodes.

[Hormonal and biochemical parameters in postmenopausal osteoporosis]. / Parâmetros bioquímios e hormonais na osteoporose pós-menopausa.

In the recent years, many authors had studied the relationship between the calciotropic and gonadotropic hormones actions on postmenopausal bone loss, named calcium negative balance, with different results (Riggs et al 1983; Prince et al, 1995). We evaluated 187 female patients, aged 40 to 80 with the following distribution: 24 normals, 49 patients with osteopenia and 114 patients with osteoporosis, according to WHO classification. The aim of this study was to analyse the relationship between biochemical parameters (seric and urinary calcium), gonadotropic (seric FSH-foliculi stimulant-hormone and seric oestradiol) and calciotropic hormone (PTH) and postmenopausal bone loss. The results had shown the diminution on bone mineral density was related with elevated levels of FSH (p < 0.00001), lower levels of oestradiol (p < 0.00001) and, however, no differences on seric and urinary calcium (respectively, p > 0.70 and p > 0.52) or PTH (p > 0.70) were demonstrated.

[Adenolymphoma of the salivary glands. The anatomicoclinical, epidemiological and therapeutic aspects of 76 cases]. / Adenolinfoma della ghiandole salivari. Aspetti anatomo-clinici, epidemiologici e terapeutici di 76 casi.

A series of 76 cases of adenolymphoma (Warthin's tumor) of the salivary glands is presented with emphasis on its epidemiological, clinico-pathological and therapeutic features; together with the analysis of the clinical records and of the epidemiological data, this study has reevaluated the histopathological features of the 76 cases of Warthin's tumor on hematoxylin-eosin stained slides. On these bases, the authors underline the frequent multifocality and/or bilaterality of Warthin's tumor (11.3% of cases), its frequent association with tumors of other sites (11.3% of cases), the prevalence in males (M:F ratio = 24:1) and in adults (mean age: 57 years) of this type of tumor. Furthermore, this study confirms the parotid gland as the elective site for Warthin's tumor and the possibility to categorize this entity into four different histological types. The latter finding should be kept in mind especially when fine needle aspiration (FNA) is used as a preoperative diagnostic procedure for Warthin's tumor to avoid misleading diagnosis and overtreatment. From the analysis of the results the authors underline the need for a complex and multidisciplinary approach to the diagnosis of Warthin's tumor in order to allow conservative surgery with preservation of the facial nerve.

[Bilateral adenolymphoma (Warthin's tumor) of the parotid. The anatomicoclinical, diagnostic and therapeutic aspects of 2 cases]. / L'adenolinfoma (tumore di Warthin) bilaterale della parotide. Aspetti anatomo-clinici, diagnostici e terapeutici di due casi.

Cystadenoma lymphomatosum or Warthin's tumor of the parotid glands appears bilateral in 8-10% of cases, synchronous (rarely), metachronous (frequently) and sometimes with multicentric aspects. After the review of the epidemiological, pathogenetic, clinical, radiographic and histologic data, the authors present two cases of bilateral Warthin's tumor, metachronous (7 years), nodular-isolated in the first one and metachronous (5 years), nodular-multicentric in the second one. The patients after CAT scan and NMR examinations, and a fine needle aspiration biopsy were treated with superficial parotidectomy with preservation of the facial nerve. In our experience the management of patients affected by Warthin's tumors, needs the following steps: CAT scan and NMR examinations to outline multiple and bilateral lesions, fine needle aspiration biopsy, superficial parotidectomy with preservation of the facial nerve, an extended follow-up (10 years after the last surgery).

[Heterologous mesodermal adenosarcoma of the endocervix. Description of a clinical case]. / Adenosarcoma mesodermico eterologo dell' endocervice. Descrizione di un caso clinico.

We describe a case of an endocervical heterologous mesodermal adenosarcoma, found in a 43 year old woman. Among mesodermal neoplasms, various histological types are distinguished: the carcinosarcoma, the embryonal rhabdomyosarcoma or botryoid sarcoma, and the adenosarcoma; the last is formed by a benign epithelial component and by a malignant stromal component, that may contain heterologous tissues, such as cartilage, skeletal muscle, etc. Adenosarcoma is a tumor of the uterine corpus and seems to be most common among menopausal women. A primitive adenosarcoma of the uterine cervix is very rare; in fact the overall percentage of the uterine cervical sarcoma is 0.2-0.4%. The patient, age 43 years, with regular menstrual bleeding came to an outpatient clinic referring a post coital metrorrhagia. After a control examination, a polyp from the uterine cervix was removed; the histopathologic diagnosis was: fibroangioadenomatous polyp of the isthmus with cartilaginous metaplastic areas. Two months later, the patient was referred to our clinic and another cervical polyp was removed. The histological diagnosis was adenosarcoma with chondrosarcomatous heterologous mesodermal component. Then the patient was operated and the postoperative histological examination confirmed the preoperative diagnosis. A literature review about the uterine adenosarcoma etiopathogenesis is reported, and a suitable diagnostic iter, is discussed.

[Intraepithelial cervical lesions: colpo-microcolposcopic diagnosis]. / Lesioni intraepiteliali cervicali: diagnosi integrata colpo-microcolposcopica.

Integration of colposcopy and microcolposcopy allows to perform a complete study of eso- endo-cervix, for a diagnostic and topographic evaluation of cervical intraepithelial lesion. We have endoscopically studied 172 patients with histologically demonstrated CIN. In 218 cases (74.4%) the microcolposcopical evaluation corresponded to the histological diagnosis. These data confirm the methodological effectiveness of microcolposcopy and recommend it as helpful mean for cervical pathology investigation. However we believe that histological evaluation is necessary for a correct staging of the lesion. According to our experience indications for microcolposcopy are: 1) identification of endocervical lesion and squamo-columnar junction, when colposcopically not evident; 2) possibility of performing a "tailored" conization; 3) study of endocervix after conization.

Mechanisms of thrombocytopenia in the acute phase of antigen-induced arthritis in rabbits.

Arthritis induced in hyperimmune rabbits by the intra-articular injection of the specific antigen was associated with a fall in circulating platelet number that lasted up to 60 days. Pretreatment of the animals with indomethacin and econazol at doses that significantly decreased thromboxane levels in the synovial fluids reduced the arthritis-related thrombocytopenia in the acute phase of arthritis. A similar inhibition was seen when L-655,240, a specific Thromboxane A2 antagonist, and BN 52021, a Platelet Activating Factor antagonist were used. The results suggest that both thromboxane and PAF are involved in the mechanisms leading to thrombocytopenia in this experimental model of arthritis.