Intratumor Heterogeneity of KIT Gene Mutations in Acral Lentiginous Melanoma.

Melanoma is an aggressive skin malignancy, and the acral lentiginous melanoma (ALM) subtype affects non-sun-exposed sites such as the volar surface of the hands and feet and the subungual region and is most common in Asians, Hispanics, and Afro-descendants. The presence of different clones within the same tumor seems to influence the aggressiveness of tumors. Patients with mutations in the KIT gene have shown a good response to tyrosine kinase inhibitor therapy. We tested the hypothesis of intratumor heterogeneity through analysis of KIT gene mutations in ALM and determined the correlation between KIT mutations and demographic, clinical, and histopathological variables. Twenty-five ALM samples were examined. We selected up to four different regions per tumor for sequencing by the Sanger method for analysis of KIT gene exon 11 and exon 13 mutations. Advanced lesions were predominant, and the main histopathological characteristics of lesions were Breslow index >4.0 mm (17/25, 68%), Clark level IV/V (21/25, 84%), ulceration (16/25, 64%), and >3 mitoses/mm (8/25, 32%). KIT gene mutations were detected in 11/25 cases (44%), and all these 11 cases displayed intratumor heterogeneity, that is, at least 2 tumor regions had different mutational profiles. The predicted effect of most mutations detected was detrimental to protein function. No significant correlations between histopathological variables and either KIT mutations or intratumor heterogeneity were observed. The hypothesis of intratumor heterogeneity of KIT gene mutations in acral lentiginous melanoma was supported.

BRAFV600E and KIT immunoexpression in early-stage melanoma.

Melanoma is widely known as the most lethal skin cancer. Specific tumor-related mortality can be significantly reduced if diagnosis and treatment are properly performed during initial phases of the disease. The current search for biomarkers in early-stage melanomas is a high-priority challenge for physicians and researchers. We aimed to assess the immunoexpression of BRAFV600E and KIT in a case series consisting of 44 early-stage melanomas. Formalin-fixed paraffin-embedded samples were systematically evaluated using a semi-quantitative method based on scores of percentage and intensity for immunostained tumor cells. We observed significant concordance between BRAFV600E and KIT immunoexpression in thin invasive melanomas. Our findings corroborate previous evidence showing abnormal expression of proteins associated with MAPK intracellular signaling pathway in early-stage melanomas.

BRAFV600E and KIT immunoexpression in early-stage melanoma

Abstract: Melanoma is widely known as the most lethal skin cancer. Specific tumor-related mortality can be significantly reduced if diagnosis and treatment are properly performed during initial phases of the disease. The current search for biomarkers in early-stage melanomas is a high-priority challenge for physicians and researchers. We aimed to assess the immunoexpression of BRAFV600E and KIT in a case series consisting of 44 early-stage melanomas. Formalin-fixed paraffin-embedded samples were systematically evaluated using a semi-quantitative method based on scores of percentage and intensity for immunostained tumor cells. We observed significant concordance between BRAFV600E and KIT immunoexpression in thin invasive melanomas. Our findings corroborate previous evidence showing abnormal expression of proteins associated with MAPK intracellular signaling pathway in early-stage melanomas.

Acral Lentiginous Melanomas Harbour Intratumor Heterogeneity in BRAF Exon 15, With Mutations Distinct From V600E/V600K.

The choice of appropriate therapeutic strategies may be influenced by intratumor heterogeneity and makes cancer treatment considerably more challenging. We aimed to evaluate the heterogeneity of BRAF exon 15 mutations in different areas of acral lentiginous melanoma (ALM). The entire exon 15 was sequenced in 4 different areas of paraffin-embedded samples from 26 patients with ALM. A total of 26 of 49 cases of ≥1 mm in depth of ALM identified by clinical, anatomical, and pathological data fulfilled the inclusion and exclusion criteria for this study. Tumors had a mean Breslow depth of 7.2 mm and an average mitotic index of 3 mitosis/mm. Mutations distinct from the common V600E and V600K were detected in 31%, and intratumor heterogeneity was observed in 31% of samples. Interestingly, 63.5% of all mutations had been previously associated with cancer. Most (62.5%) of the missense BRAF exon 15 mutations found in the ALM samples examined here were deemed "detrimental" for protein function according to at least 2 functional prediction programs, and 3 mutations (37.5%) were predicted to be "neutral," with no effect on protein function. BRAF exon 15 mutations were detected frequently in ALM and displayed heterogeneity, a finding to be further investigated.

Long term safety of targeted internalization of cell penetrating peptide crotamine into renal proximal tubular epithelial cells in vivo.

Activated proximal tubular epithelial cells (PTECs) play a crucial role in progressive tubulo-interstitial fibrosis in native and transplanted kidneys. Targeting PTECs by non-viral delivery vectors might be useful to influence the expression of important genes and/or proteins in order to slow down renal function loss. However, no clinical therapies that specifically target PTECs are available at present. We earlier showed that a cationic cell penetrating peptide isolated from South American rattlesnake venom, named crotamine, recognizes cell surface heparan sulfate proteoglycans and accumulates in cells. In healthy mice, crotamine accumulates mainly in kidneys after intraperitoneal (ip) injection. Herein we demonstrate for the first time, the overall safety of acute or long-term treatment with daily ip administrated crotamine for kidneys functions. Accumulation of ip injected crotamine in the kidney brush border zone of PTECs, and its presence inside these cells were observed. In addition, significant lower in vitro crotamine binding, uptake and reporter gene transport and expression could be observed in syndecan-1 deficient HK-2 PTECs compared to wild-type cells, indicating that the absence of syndecan-1 impairs crotamine uptake into PTECs. Taken together, our present data show the safety of in vivo long-term treatment with crotamine, and its preferential uptake into PTECs, which are especially rich in HSPGs such as syndecan-1. In addition to the demonstrated in vitro gene delivery mediated by crotamine in HK-2 cells, the potential applicability of crotamine as prototypic non-viral (gene) delivery nanocarrier to modulate PTEC gene and/or protein expression was confirmed.

Oral treatment with a rattlesnake native polypeptide crotamine efficiently inhibits the tumor growth with no potential toxicity for the host animal and with suggestive positive effects on animal metabolic profile.

The efficacy of crotamine as antitumoral was first demonstrated by daily intraperitoneal (IP) injections of low doses of this toxin in an animal model bearing melanoma tumors. Significant inhibition of tumor growth and increased lifespan of mice bearing tumor was also noticed after 21 consecutive days of this daily IP administration of crotamine. However, due to the limited acceptance of treatments by IP route in clinical conditions, herein, we evaluated the antitumor effect of this native polypeptide employing the oral route. The efficacy of crotamine in inhibiting the melanoma growth in vivo, even after passing through the gastrointestinal tract of the animal, was confirmed here. In addition, biochemical biomarkers and also histopathological analysis showed both the absence of any potential toxic effects in tissues or organs of the animal in which the highest accumulation of crotamine is expected. Interestingly, a reduction of weight gain was observed mainly in animals with tumor treated with crotamine by IP route, but not by oral administration. Albeit, oral administered crotamine was able to significantly decrease the body weight gain of healthy animals without tumor. Taking advantage of this same experimental animal models receiving crotamine by oral route, it was possible to show metabolic changes as the increased capacity of glucose clearance, which was accompanied by a reduction of the total cholesterol, and by increased high-density lipoprotein levels, both observed mainly in the absence of tumor. Triglycerides and low-density lipoprotein were also significantly decreased, but only in the absence of tumor. Taken together, these data suggest a clear trend for metabolic positive effects and mischaracterize unhealthy condition of animals, with or without tumors, treated with crotamine for 21 days. In addition, this study confirmed the efficacy of crotamine administered by oral route as antitumor agent, which besides the additional advantage of administration convenience and decreased risk of toxic effects, allowed the serendipitous observation of several positive metabolic effects on treated animals.

Expression of Galectins 1, 3 and 9 in normal oral epithelium, oral squamous papilloma, and oral squamous cell carcinoma.

BACKGROUND: The aim of this study was to characterize the immunohistochemical expression of galectin 1, 3, and 9 in normal oral epithelium, oral squamous papilloma, and oral squamous cell carcinoma. MATERIALS AND METHODS: Immunohistochemical staining for galectins 1, 3, and 9 was evaluated in 8 samples of normal oral squamous epithelium, 15 samples of oral squamous papilloma, and 41 samples of oral squamous cell carcinoma. Immunohistochemical data were assessed by Kruskal-Wallis non-parametric test followed by Dunn's test. For all analyzes, it was adopted the value of P <0.05 for statistical significance. RESULTS: Significant differences were found in galectin- 3 expression when comparing ordinary mucosa and oral squamous papilloma with the oral squamous cell carcinoma samples. CONCLUSION: These findings indicate that galectin-3 is closely involved in malignant transformation of oral mucosa cells.