There are data to suggest that some ductal carcinoma in situ (DCIS) may evolve through an evolutionary bottleneck, where minor clones susceptible to the imposed selective pressure drive disease progression. Here, we tested the hypothesis that an impact of the inflammatory environment on DCIS evolution is HER2-dependent, conferring proliferative dominance of HER2-negative cells. In tissue samples, density of tumour-infiltrating immune cells (TIICs) was associated only with high tumour nuclear grade, but in 9% of predominantly HER2-negative cases, the presence of tumoral foci ('hot-spots') of basal-like cells with HIF1-α activity adjacent to the areas of dense stromal infiltration was noted. Results of in vitro analyses further demonstrated that IL-1β and TNF-α as well as macrophage-conditioned medium triggered phosphorylation of NF-κB and subsequent upregulation of COX2 and HIF1-α, exclusively in HER2-negative cells. Treatment with both IL-1β and TNF-α resulted in growth stimulation and inhibition of HER2-negative and HER2-positive cells, respectively. Moreover, ectopic overexpression of HIF1-α rescued HER2-positive cells from the negative effect of IL-1β and TNF-α on cell growth. Our data provide novel insight into the molecular basis of HER2-dependent proliferation of DCIS cells and indicate the NF-κB/COX2 → HIF1-α signalling axis as a dominant mechanism of DCIS evolution induced by inflammatory microenvironment. Presented findings also highlight the clinical significance of heterogeneity of DCIS tumours and suggest that HIF1-α might be considered as a predictive marker of disease progression.
Promising results have been obtained using brachytherapy in the treatment of brain tumors. Between November 99 and August 2000, 28 patients with brain tumors (15 newly diagnosed gliomas, 11 recurrent gliomas, 2 metastases) underwent implantation of temporary iridium 192 sources with stereotactic technique. This group received external beam radiation therapy (45 Gy) following implantation. Patients were followed-up with CT scans every 3 months. Serious complications occurred in two patients (postradiation brain oedema). Median survival has not been assessed due to short follow-up period and small number of patients. Further clinical assessment is required especially long-term follow-up. Brachytherapy appears to be a useful technique for the treatment of selected brain tumors.
- NRAS is a member of the RAS family oncoproteins implicated in cancer. Gain-of-function NRAS mutations were reported in a subset of colorectal cancers. These mutations occur at codons 12, 13, and 61 and are detected by molecular genetic testing. Recently, an antibody (clone SP174) became available to immunohistochemically pinpoint NRAS Q61R mutant protein. In malignant melanoma, NRAS Q61R mutant-specific immunohistochemistry was shown to be a valuable supplement to traditional genetic testing.- To evaluate the significance of NRAS Q61R mutant-specific immunohistochemistry in a cohort of colorectal carcinomas.- A total of 1185 colorectal carcinomas were immunohistochemically evaluated with SP174 antibody. NRAS Q61R mutant-specific immunohistochemistry was validated by molecular genetic testing including Sanger sequencing, quantitative polymerase chain reaction (qPCR), and next-generation sequencing.- Twelve tumors showed strong SP174 immunoreactivity. Sanger sequencing detected an identical c.182A>G substitution, causing NRAS Q61R mutation at the protein level, only in 8 SP174-positive cases. These results were confirmed by qPCR study. Subsequently, NRAS wild-type tumors with strong SP174 staining were evaluated by next-generation sequencing and revealed KRAS c.182A>G substitutions predicted to cause KRAS Q61R mutation. Review of colorectal carcinomas with known KRAS and NRAS genotype revealed that none of 62 wild-type tumors or 47 mutants other than Q61R were SP174 positive.- SP174 immunohistochemistry allows sensitive detection of NRAS and KRAS Q61R mutants. However, molecular genetic testing is necessary to determine specifically which RAS gene is mutated.
FIGURE 1 A -clinical photograph of the patient before treatment, showing bilateral, rubbery, yellow eyelid lesions; B -clinical photograph of the patient after treatment
Recently BRAF V600E mutant-specific antibody (clone VE1) became available to immunohistochemically pinpoint the occurrence of these BRAF-mutant proteins in different tumors, such as colon carcinoma. Detection of BRAF mutations is important for the accurate application of targeted therapy against BRAF serine-threonine kinase activation. In this study, we evaluated 113 colon carcinomas including 95 primary and 27 metastatic tumors with the VE1 antibody using Leica Bond-Max automated immunohistochemistry. To ensure comprehensive BRAF V600E mutation detection, all cases were evaluated using 4 molecular methods (Sanger sequencing, the Cobas 4800 BRAF V600 Mutation Test, BRAF V600 allele-specific polymerase chain reaction, and BRAF V600 quantitative polymerase chain reaction) with nearly 100% concordance. Molecular and immunohistochemical studies were blinded. Furthermore, all cases were evaluated for KRAS and NRAS mutations as parameters mutually exclusive with BRAF mutations offering parallel evidence for BRAF mutation status. Strong to moderate VE1 positivity was seen in 34 tumors. Twelve colon carcinomas showed weak VE1 immunohistochemical staining, and 67 were entirely negative. An identical c.1799T>A single nucleotide substitution leading to the BRAF V600E mutation was identified in 27 of 113 (24%) colon carcinomas. A majority of BRAF-mutant tumors were located in the right side of the colon and had mismatch-repair deficiency. V600E mutation-negative carcinomas were more often sigmoid tumors and usually showed intact mismatch-repair proteins and KRAS or NRAS mutations. The sensitivity and specificity of positive results (strong to moderate staining) of VE1 immunohistochemistry were 85% and 68%, respectively. If any positivity would be considered, then the specificity declined to 51% with no significant improvement of sensitivity. Therefore, only strong positivity should be considered when using the VE1 antibody and Leica Bond-Max automated immunohistochemistry with these parameters. Although VE1 antibody can be useful in the screening of colon carcinomas for BRAF V600E-mutant proteins, molecular genetic confirmation is always necessary for mutation diagnosis.
Abstract Despite the introduction of new molecular classifications, advanced colorectal cancer (CRC) is treated with chemotherapy supplemented with anti-EGFR and anti-VEGF targeted therapy. In this study, 552 CRC cases with different primary tumor locations (250 left side, 190 rectum, and 112 right side) were retrospectively analyzed by next generation sequencing for mutations in 50 genes. The most frequently mutated genes were TP53 in left-sided tumors and BRAF in right-sided tumors. Mutations in KRAS , NRAS , and BRAF were not detected in 28.6% of patients with right-sided tumors and in 45% of patients with left-sided tumors. Liver metastases were more common in patients with left-sided tumors. Tumors on the right side were larger at diagnosis and had a higher grade (G3) than tumors on the left. Tumors located in the rectum differed from those in other locations in biology, site of metastasis (lung), and mutation rates (e.g., BRAF, FBXW7, and TP53). KRAS , NRAS , and BRAF gene mutations were not detected in >47% of rectal tumors compared with 42.8% of left-sided and 28.6% of right-sided tumors. Primary tumor location has implications for the potential treatment of CRC with anti-EGFR therapy.
Aggressive fibromatosis (desmoid tumor) is a mesenchymal lesion originating from fascial, aponeurotic and muscular connective tissue. It rarely becomes histologically malignant. In this study we analyzed the cell cycle regulation proteins: pRb, p16, and proliferating antigens: Ki-67, PCNA, MCM5 with immunohistochemical method in archival material derived from 27 extra-abdominal (E-AD), 18 abdominal (AD) and 5 intra-abdominal (I-AD) cases of desmoid tumor. None of the examined cases (n=50) of aggressive fibromatosis was pRb-immunonegative. Heterogeneous expression of pRb was observed in 51.85% (14/27) of Group AD cases and in 5.56% (1/18) of Group E-AD cases; positive expression in 48,15% (13/27) of Group AD cases, in 94.44% (17/18) of Group E-AD cases, and in 100% (5/5) of Group I-AD cases. There were no negative cases for p16 staining in any of the examined groups. The number of heterogeneous cases in individual groups was: 33.33% (9/27) in Group AD, 50% (9/18) in Group E-AD and 40% (2/5) in Group I-AD, and positive cases: 66.67% (18/27), 50% (9/18) and 60% (3/5), respectively. Overexpression of PCNA was noted in 98% (49/50) of cases. The positive staining for Ki-67 protein was noted in 25.93% (7/27) in Group AD, in 16.67% (3/18) in Group E-AD and in 60% (3/5) in Group I-AD. None of the examined cases was immunopositive for MCM5 protein. The noted levels of pRb and p16 expression in desmoid cells reflect their function in cell cycle regulation. Probably the unsettled cell cycle progression, especially in G1 phase, is not the cause of aggressive fibromatosis pathogenesis.
The 5 cases of salivary duct carcinoma (SDC); very rare, but distinct group of highly malignant salivary gland tumor are presented, and difficulties with pathological and clinical diagnosis is discussed. The SDC developed in single cases in parotid salivary gland, submandibular salivary and in mucosa of maxillary sinus, pyriform fossa and oral cavity (check). In 3 cases the second malignant tumor was present--synchronously (SDC + pleomorphic adenoma in parotid gland; SDC + squamous cell carcinoma in hypopharynx) or metachroneously (squamous cell carcinoma of upper lip followed by SDC). In one case the high levels of PSA suggesting of metastases from unknown primary within the prostate gland, or PSA expression related to SDC was observed. The four patients received radical treatment - surgical resection followed by radiotherapy; in one case only palliative treatment was applied, due to patient's poor general condition and high advancement of the primary disease. The observation ranged from 10 to 77 months (average time--31 months). The one patient died 13 months after diagnosis and palliative treatment. The three patients are alive with distant metastases to the lung and bones (77, 38 and 18 months after primary treatment was completed). Only one patient with 10 months observation after treatment is living without symptoms of recurrence or metastases.
Aggressive fibromatosis, usually called desmoid tumor develops from muscle connective tissue, fasciae and aponeuroses. This neoplasm is composed of spindle (fibrocyte-like) cells. As regards the site, aggressive fibromatoses can be divided into: extra-abdominal in the area of the shoulder and pelvic girdle or chest and neck wall; abdominal in abdominal wall muscles; intra-abdominal concerning pelvis, mesentery connective tissue or retroperitoneal space. Desmoid tumor is a neoplasm which rarely turns malignant and is non-metastasizing but demonstrates ability to local infiltration into tissue and is characterized by high risk of recurrence (25-65%) after surgical treatment. Desmoid tumor etiology is uncertain. This neoplasm occurs in sporadic (idiopathic) form and is also associated with some familial neoplastic syndromes. Most sporadic cases of aggressive fibromatosis contain a somatic mutation in either the adenomatous polyposis coli (APC) or beta-catenin genes. Sporadic tumors are more frequent in women than in men from 2 : 1 to 5 : 1. In about 10-15 per cent of patients with familial adenomatous polyposis (FAP), aggressive fibromatosis is a parenteral manifestation of this familial syndrome conditioned by APC gene mutation. Abdomen injury--most frequently due to surgery is said to play an important role in the initiation of fibrous tissue proliferative process in the cases of abdominal and intra abdominal forms. High cells growth potential with relatively high local malignancy is observed in about 10% of cases with sporadic tumors as well as in those FAP-associated.
