Pretreatment Circulating HPV16 DNA Viral Load Predicts Risk of Distant Metastasis in Patients with HPV16-Positive Oropharyngeal Cancer.

BACKGROUND: There are definite reasons to implement molecular diagnostics based on the measurement of human papillomavirus (HPV) DNA in liquid biopsy into clinical practice. It is a quick, repeatable, and health-safe test for cancer biomarkers in the blood. In this study, we investigated whether the circulating tumor-related HPV16 (ctHPV16) viral load (VL) in patients with oropharyngeal squamous cell carcinoma (OPSCC) was important for determining the risk of locoregional recurrence-free survival (LRFS), metastasis-free survival (MFS), and overall survival (OS). METHODS: This study included 91 patients with ctHPV16-positive OPSCC who had been treated with radical radiotherapy and chemotherapy. The VL was measured using quantitative PCR (qPCR) and a probe specific for HPV16. Based on 10 years of follow-up, the 2-, 3-, 5-, and 9-year LRFS, MFS, and OS were estimated. RESULTS: The 5-year actuarial LRFS, MFS, and OS rates of patients with ctHPV16-positive/OPSCC were 88%, 90%, and 81%, respectively. The VL was significantly higher in patients who subsequently developed distant metastases (DM) than in those who did not (VL 4.09 vs. 3.25; p = 0.009). In a Cox proportional hazards regression model for MFS, a higher ctHPV16 VL appeared to be a significant independent prognostic factor for the occurrence of DM (HR 2.22, p = 0.015). The ROC curve revealed a cutoff value of 3.556 for VL (p = 0.00001). CONCLUSIONS: A high VL before treatment indicates patients with a significant risk of DM, and should be used in OPSCC treatment stratification.

Determinants of the level of circulating-tumor HPV16 DNA in patients with HPV-associated oropharyngeal cancer at the time of diagnosis.

Circulating tumor HPV DNA (ctHPV16) assessed in liquid biopsy may be used as a marker of cancer in patients with HPV-associated oropharyngeal cancer (HPV + OPC). Factors influencing the initial ctHPV16 quantity are not well recognized. In this study we aimed to establish what factors are related to the level of ctHPV16 at the time of diagnosis. 51 patients (37 men and 14 women, median age of 57 years old) with HPV + OPC prior to definitive treatment were included. ctHPV16 was measured by qPCR. Tumor and nodal staging were assessed according to AJCC8. Blood derived factors included squamous cell carcinoma antigen (SCC-Ag), serum soluble fragment of cytokeratin 19 (CYFRA 21-1), C-reactive protein (CRP), albumin level (Alb), neutrophils (Neut), thrombocytes (Plt) and lymphocyte (Lym) count, Neut/Lym ratio were assessed. The volumes of the primary tumor (TV) and involved lymph nodes (NV) were calculated using MRI, CT or PET-CT scans. Data were analysed using parametric and nonparametric methods. Variables for multivariable linear regression analysis were chosen based on the results from univariable analysis (correlation, univariable regression and difference). There were 9 (18%), 10 (19%) and 32 (63%) patients who had TV and NV assessed in MRI, CT or PET respectively. Primary tumor neither as T-stage nor TV was related to ctHPV16 level. Significant differences in the ctHPV16 between patients with high vs low pain (P = 0.038), NV (P = 0.023), TV + NV (P = 0.018), CYFRA 21-1 (P = 0.002), CRP (P = 0.019), and N1 vs N3 (P = 0.044) were observed. ctHPV16 was significantly associated with CYFRA 21-1 (P = 0.017), N stage (P = 0.005), NV (P = 0.009), TV + NV (P = 0.002), CRP (P = 0.019), and pain (P = 0.038). In univariable linear regression analysis the same variables predicted ctHPV16 level. In multivariable analyses, CYFRA 21-1 and CRP (both as categorical variables) were predictors of ctHPV16 level even above NV. ctHPV16 at presentation is driven by tumor volume measured mostly by N. CYFRA 21-1 and CRP are additional factors related to ctHPV16 prior to the treatment.

Salivary Concentrations of Chemerin, α-Defensin 1, and TNF-α as Potential Biomarkers in the Early Diagnosis of Colorectal Cancer.

Colorectal cancer is one of the most prevalent cancers worldwide. There is a great interest and need to find simple, inexpensive, and minimally invasive diagnostic tests. The aim of the study was to analyze the salivary concentrations of chemerin, α-defensin 1, and TNF-α in colorectal cancer (CRC) patients and in a healthy control group. The concentration of these proteins was simultaneously determined in the serum of subjects. We also aimed to assess the correlation of these results and selected clinicopathological features. This prospective study was comprised of 39 CRC patients and 40 control group patients. Salivary and serum concentrations were determined by enzyme immunoassays. The salivary and serum concentrations of chemerin, α-defensin 1, and TNF-α were significantly higher in cancer patients compared to the control group. No correlation was found between concentrations of the proteins and the clinical stage of cancer and tumor location. The ROC curve analysis showed that although salivary concentrations of all proteins showed 100% sensitivity and 100% specificity, serum concentrations of the analyzed proteins were characterized by 100% sensitivity and over 90% specificity. The assessment of chemerin, α-defensin 1, and TNF-α concentrations in saliva seem to have great potential as quick and useful biomarkers in the early diagnosis of CRC.

Analysis of Selected Nutritional Parameters in Patients with HPV-Related and Non-HPV-Related Oropharyngeal Cancer before and after Radiotherapy Alone or Combined with Chemotherapy.

Background: Radiotherapy plays an essential role in the treatment of oropharyngeal carcinoma (OPC). The aim of this study was to assess and compare the nutritional status (NS) of patients with HPV-related (HPV+) and non-HPV-related (HPV-) OPC before and after radiotherapy (RT) or chemoradiotherapy (CRT). Methods: The analysis included 127 patients with OPC who underwent radiotherapy (RT) alone, or in combination with chemotherapy (CRT), in the I Radiation and Clinical Oncology Department of Maria Sklodowska-Curie National Research Institute of Oncology, Gliwice Branch, Poland. Patients were divided according to HPV status. Confirmation of HPV etiology was obtained from FFPE (formalin-fixed, paraffin-embedded) tissue material and/or extracellular circulating HPV DNA. Basic anthropometric and biochemical parameters before and after RT/CRT were compared between the HPV- and HPV+ groups. The effect of NS on survival was also analyzed. Results: In both groups, a significant decrease in all analyzed nutritional parameters was noted after RT/CRT (p < 0.01). CRT caused significant weight loss and decreases in BMI, albumin, total lymphocyte count (TLC), and hemoglobin concentration, as well as an increase in the Nutritional Risk Score (NRS) 2002, in HPV- and HPV+ patients. A significant decrease in prealbumin levels after CRT was noted only in HPV+ patients. RT caused a significant decrease in hemoglobin concentration and TLC in HPV- patients. There were no significant differences regarding other nutritional parameters after RT in either group. RT did not have negative impact on body mass index (BMI), weight, NRS, CRP, Alb, Prealb, or PNI. Overall survival (OS) and disease-free survival (DFS) were significantly better in patients with a higher BMI in the HPV- group (OS, p = 0.011; DFS, p = 0.028); DFS was significantly better in patients with C-reactive protein (CRP) < 3.5 g/dL in the HPV- (p = 0.021) and HPV+ (p = 0.018) groups, and with total lymphocyte count (TLC) >1.28/mm3 in the HPV+ group (p = 0.014). Higher NRS 2002 was an independent adverse prognostic factor for OS and DFS in HPV-, but not in the HPV+ group. Kaplan−Meier analysis showed that both OS and DFS were significantly better in HPV- patients with lower NRS 2002 scores. However, this relationship was not observed in the HPV+ group. Conclusions: Regardless of HPV status, patients with OPC can develop malnutrition during RT/CRT. Therefore, nutritional support during RT/CRT is required in patients with HPV- and HPV+ OPC.

Prognostic value of human papillomavirus detection and the eighth edition of the TNM classification staging system in oropharyngeal squamous cell carcinoma: A single-center Polish study.

OBJECTIVES: This study aimed to determine human papillomavirus (HPV) status and genotypes, the HPV status-dependent survival, and the applicability of the eighth TNM classification in Polish patients diagnosed with oropharyngeal squamous cell carcinoma (OPSCC). STUDY DESIGN: All patients with primary OPSCC, diagnosed and treated from 2007 to 2017 at the National Research Institute of Oncology, Warsaw, Poland, who underwent radical radiotherapy were included. The Kaplan-Meier method was deployed to produce 3- and 5-year observed survival (OS) estimates. RESULTS: A total of 110 OPSCC cases were identified. Double positivity for HPV (IHC p16INK4a and HPV-DNA) was recorded in 70.9% of cases, with HPV16 being the most prevalent genotype (96.2%). The disease stage was significantly less advanced in the HPV-related group than in the HPV-negative group (P < .001). Three- and 5-year OS in HPV-related carcinoma was 80.7% and 74.0%, respectively; in the HPV-negative group, OS was 52.9% and 48.5%. OS rates were associated with HPV status, tumor stage, and disease stage according to the eighth edition TNM classification. CONCLUSIONS: The majority of Polish patients with OPSCC are HPV16-positive. In HPV-related OPSCC, survival rates are significantly higher than in HPV-negative OPSCC. The findings support the requirement of HPV testing in Polish patients with OPSCC because HPV-positive status influences tumor prognosis.

Comparison of Selected Immune and Hematological Parameters and Their Impact on Survival in Patients with HPV-Related and HPV-Unrelated Oropharyngeal Cancer.

Several immune and hematological parameters are associated with survival in patients with oropharyngeal cancer (OPC). The aim of the study was to analyze selected immune and hematological parameters of patients with HPV-related (HPV+) and HPV-unrelated (HPV-) OPC, before and after radiotherapy/chemoradiotherapy (RT/CRT) and to assess the impact of these parameters on survival. One hundred twenty seven patients with HPV+ and HPV- OPC, treated with RT alone or concurrent chemoradiotherapy (CRT), were included. Patients were divided according to HPV status. Confirmation of HPV etiology was obtained from FFPE (Formalin-Fixed, Paraffin-Embedded) tissue samples and/or extracellular circulating HPV DNA was determined. The pre-treatment and post-treatment laboratory blood parameters were compared in both groups. The neutrophil/lymphocyte ratio (NLR), platelet/lymphocyte ratio (PLR), monocyte/lymphocyte ratio (MLR), and systemic immune inflammation (SII) index were calculated. The impact of these parameters on overall (OS) and disease-free (DFS) survival was analyzed. In HPV+ patients, a high pre-treatment white blood cells (WBC) count (>8.33 /mm3), NLR (>2.13), SII (>448.60) significantly correlated with reduced OS, whereas high NLR (>2.29), SII (>462.58) significantly correlated with reduced DFS. A higher pre-treatment NLR and SII were significant poor prognostic factors for both OS and DFS in the HPV+ group. These associations were not apparent in HPV- patients. There are different pre-treatment and post-treatment immune and hematological prognostic factors for OS and DFS in HPV+ and HPV- patients. The immune ratios could be considered valuable biomarkers for risk stratification and differentiation for HPV- and HPV+ OPC patients.

Circulating HPV16 DNA may complement imaging assessment of early treatment efficacy in patients with HPV-positive oropharyngeal cancer.

BACKGROUND: Early detection of treatment failure may improve clinical outcome and overall survival in patients with head and neck cancer after first-line treatment. Circulating cell-free HPV16 DNA (cfHPV16 DNA) was evaluated as a possible complementary marker to radiological assessment of early response in patients with HPV-related oropharyngeal cancer (OPC) after radiotherapy alone or combined with chemotherapy. METHODS: The study included 66 patients with HPV-related OPC receiving radical radiotherapy alone or in combination with chemotherapy. cfHPV16 DNA was assessed in the blood of all patients before treatment using TaqMan-based qPCR. Subsequent analysis of cfHPV16 DNA was performed 12 weeks after treatment completion, along with radiological assessment of early treatment results. RESULTS: Complete (CRR) and incomplete radiological response (IRR) was found in 43 (65%) and 23 (35%) patients respectively. cfHPV16 DNA was present in 5 (28%) patients with IRR, while only in 1 (4%) with CRR. Three of five patients with IRR that were positive for cfHPV16 DNA exhibited histopathologically confirmed local or regional treatment failure, and other two developed distant metastases. None of the patients with negative cfHPV16 DNA presented disease failure. CONCLUSION: The post-treatment assessment of cfHPV16 DNA in patients with HPV-related OPC may be used as a complementary biomarker to conventional imaging-based examinations for early identification of treatment failure.

[Infiltrates of M2-Like Tumour-Associated Macrophages Are Adverse Prognostic Factor in Patients with Human Papillomavirus-Negative but Not in Human Papillomavirus-Positive Oropharyngeal Squamous Cell Carcinoma].

INTRODUCTION: Human papillomavirus with a high oncogenic potential (HR-HPV) is responsible for more than a half of squamous cell carcinomas of the oropharynx. The HR-HPV-dependent cases of this tumour have a better prognosis compared to the HR-HPV-negative cases, despite the usually more advanced disease at the time of diagnosis. In addition to genetic and epigenetic factors, the causes of this more favourable course of the disease are also seen in the participation of the tumour microenvironment, including the patient's immune system. Macrophages are one of the most important elements of the immunocompetent cells landscape that make up the tumour microenvironment. Traditionally, they are divided into 2 groups: inflammatory macrophages with the M1 phenotype and tumour-associated macrophages known as M2 phenotype macrophages. OBJECTIVE: The aim of this study was to investigate the impact of the macrophage infiltrates intensity of the M1/M2 and M2 phenotype separately on the clinical outcome of patients with squamous cell carcinoma of the oropharynx (OPSCC), taking into account the HR-HPV status of tumours. METHODS: The study involved 85 patients with OPSCC in which HR-HPV status in tumour tissue was determined using a double-check algorithm including the detection of viral DNA by RT-PCR method with subsequent confirmation of its biological activity by immunohistochemical demonstrating the P16INK4A protein overexpression. In each of the groups formed on the basis of HR-HPV status, macrophages were discriminated using CD68 and CD163 proteins as markers of pan-macrophage and M2 phenotype. The intensity of infiltrates was quantified by means of computer-assisted analysis in digital images of whole slides (virtual slides) separately in tumour tissue and stroma. RESULTS: In HPV-positive patients, significantly more intense infiltration of both M1/M2 and M2 macrophages was found in the tumour stroma compared to HPV-negative patients. The infiltrates from both types of macrophages in the tumour tissue were less intense and did not differ between these groups. Intensive infiltration of CD68+ macrophages in the tumour front was associated with higher rate of nodal failures and a shorter nodal control in both HR-HPV groups. In the group of HR-HPV-negative patients, heavy infiltration of CD163+ macrophages was associated with significantly shorter: loco-regional control (LRC), metastasis-free survival and overall survival (OS). These parameters and prognosis in patients with scanty CD163+ infiltration were similar to favourable outcomes in HR-HPV-positive patients. The relative risk of local-regional recurrence, distant metastases and disease-related death in HR-HPV-negative patients with intense CD163+ infiltrates was, respectively, 4.7, 5.4 and 5.7 compared to patients with scanty infiltrates. CONCLUSIONS: Tumours with a positive HR-HPV status demonstrate intense infiltrations of total pool M1/M2 and M2 macrophages. In the group of HPV-negative patients, intensive M1/M2 macrophage infiltrates correlate with higher risk of nodal failures, and intensive M2 infiltrates are an adverse prognostic factor for LRC, metastasis-free survival and OS.

Detection of circulating HPV16 DNA as a biomarker in the blood of patients with human papillomavirus-positive oropharyngeal squamous cell carcinoma.

BACKGROUND: Development of biomarker analysis using the circulating cell-free DNA (cfDNA) methodology is a challenge for noninvasive cancer diagnosis. In this study, a comparison between the plasma and tumor tissue HPV16 DNA viral loads (VLs) has been presented. METHODS: Real-time polymerase chain reaction was performed for quantitating of HPV16 DNA in the plasma and tumor samples of patients with oropharyngeal cancer. RESULTS: Among the tissues, HPV16-positive patients with oropharyngeal squamous cell carcinoma, nonsmoking patients, displayed significantly higher HPV16 DNA VLs in their tissue. No smoking and advanced N disease were the most important predictors for cHPV16 DNA (circulating HPV16 DNA) detection. The cHPV16-positive women displayed significantly higher VLs in their tumor tissues compared to the men, although without notable impact on the blood detection. CONCLUSIONS: Many factors were responsible for human papillomavirus DNA circulation in blood. As a result of the small size of the analyzed group, some observed discrepancies need to be proven on a larger cohort.

Assessment of PI3K/AKT/PTEN signaling pathway activity in colorectal cancer using quantum dot-conjugated antibodies.

In certain patients with advanced colorectal cancer, loss of phosphatase and tensin homolog deleted on chromosome 10 (PTEN) activity is observed. PTEN is a major gatekeeper gene of the AKT serine/threonine kinase (AKT) signaling pathway responsible for the proliferative activity of cells. The assessment of AKT activity may be a prognostic factor or a predictor of response to the targeted therapies against particular signaling proteins. To precisely identify the cause and the place of the pathway deregulation, it is necessary to identify phosphorylation states and concentrations of several proteins located at different levels of the regulatory cascade. In the present study, we propose the simultaneous use of specific antibodies conjugated with different quantum dots to highlight the nature of AKT/PKB cascade deregulation in patients with colorectal cancer and the loss of PTEN expression in tumor tissue. Fifty patients with colorectal cancer of no specific location were enrolled in the study. The expression of the PTEN protein, and concentrations of phosphorylated/activated forms of 3-Phosphoinositide-dependent kinase 1 (PDK1) and AKT were assessed using quantum dot-conjugated antibodies. In patients with a diminished or complete loss of the PTEN expression in the tumor tissue increased levels of activated/phosphorylated forms of PDK1 (Phospho-PDK1-Ser241) and AKT (Phospho-AKT-Thr308) proteins were found, which are responsible for the permanent activation of the phosphoinositide 3-kinase/AKT/PTEN signaling pathway in certain cases of colorectal cancer.

Oral and oropharyngeal papillomas are not associated with high-risk human papillomavirus infection.

The role of different types of human papillomavirus (HPV) in the development of oral and oropharyngeal papillomas remains unclear. High-risk HPV (HR-HPV) was shown to be involved in the pathogenesis of significant proportion of squamous cell carcinomas of the oropharynx. In this study, we hypothesized that in some oropharyngeal papillomas, low-risk HPV (LR-HPV) and HR-HPV infection could co-exist, similar to what is observed in genital warts, and thus contribute to the elevated risk of malignancy. To test this hypothesis, we used real-time PCR to assess the presence of HPV DNA of 16 types (2 LR-HPV and 14 HR-HPV), in 75 formalin-fixed and paraffin-embedded histopathological samples of oral and oropharyngeal papillomas and in 57 squamous cell carcinomas from the same regions. We investigated the biological activity of HPV by demonstrating accumulation of P16(INK4A) protein in the viral-infected tissue samples. The presence of the LR-HPV genome from the HPV6 or HPV11 types was confirmed in 42 (56%) papillomas and in no carcinomas. HPV6/HPV11 co-infection was detected in 17 (22.7%) of the papillomas. HR-HPV DNA presence and HR-HPV activity hallmarks were not observed in any of the investigated papillomas. Thus, a causative role for HR-HPV or its contribution to LR/HR-HPV co-infection in the pathogenesis of oral or oropharyngeal papillomas is unlikely. Additionally, HR-HPV and LR-HPV infections seem to be mutually exclusive in papillomas and squamous cell carcinomas of the oral cavity and oropharynx.

Tumor-Associated Macrophages and Regulatory T Cells Infiltration and the Clinical Outcome in Colorectal Cancer.

The aim of the study is the assessment of the intensity of the infiltration of tumor-associated macrophages (TAMs) CD68+/iNOS- and Tregs CD8+/FoxP3+ in colorectal cancer (CRC) patients as prognostic factors with respect to disease-free survival (DFS) and overall survival (OS). In this retrospective study, tissue samples were obtained from 89 patients undergoing resection for CRC (stage IIA, pT3N0M0 and stages IIIB and IIIC, pT3N1-2M0). Recurrence was observed in 45 patients at the time of the follow-up (10 local recurrences, 35 distant metastases). In patients with recurrence the following were present: a tendency to an older average age at the time of diagnosis (p = 0.07), higher nodal involvement (p = 0.002) and more advanced clinical disease (p = 0.01). The analysis of the clinical data and immunohistochemical studies were performed with the methodology of identification of TAM and Treg subsets in histological sections, with the aim to use it in routine clinical management. Both DSF and OS were the clinical parameters assessed in the study. The presence of intense infiltration of TAMs in the tumor stroma was related to shorter DFS (p = 0.005) and OS (p = 0.006). The opposite tendency was observed in the tumor front (p = 0.061). The relative risks of recurrence and cancer-related death were more than twice higher in the group of patients with intense infiltration of TAMs in the tumor stroma (RR 2.05, 95% CI 1.33-3.14; p = 0.001 and RR 2.08, 95% CI 1.28-3.39; p = 0.003, respectively). Intense infiltration of Tregs in the tumor stroma was related to shorter DFS and OS (p < 0.0001). The relative risks of recurrence and death in a group of patients with intense infiltration of Tregs in the tumor stroma were more than 12 times higher than in patients with less intense infiltration (RR 12.3, 95% CI 5.44-27.9; p < 0.0001 and RR 12.5, 95% CI 4.9-32.4; p < 0.0001, respectively). Infiltration of TAMs CD68+/iNOS- and Tregs CD8+/FoxP3+ in the tumor stroma are negative prognostic factors with a positive correlation between them. Tregs may constitute an independent prognostic factor in patients with CRC.

Vertically transmitted HPV-dependent squamous cell carcinoma of the external auditory canal : Case report of a child.

BACKGROUND: There is much evidence that high-risk human papillomavirus (HPV) plays a causative role in a subset of head and neck squamous cell cancer (HNSCC) in adults. HPV-positive tumors behave differently even in their response to treatment and are therefore a distinct subset. Both HPV-positive and HPV-negative tumors of the head and neck region are usually in the domain of adults and cases in children are rare; thus when a 2­year-old child was diagnosed with this cancer in the external auditory canal, an in-depth assessment of the tumor was considered necessary. CASE REPORT: A 2­year-old girl was born to a HPV-positive mother who was diagnosed with cervical cancer during pregnancy. The child was delivered by caesarean section and the mother died of her cancer 7 months after delivery. After the diagnosis of locally invasive HPV-positive squamous cell cancer of the external auditory canal, the child was treated surgically, and with chemotherapy and radiotherapy. Full remission was obtained lasting up to 325 weeks since treatment was started, resulting in over 6 years of disease-free survival. CONCLUSION: This is the first case of advanced, HPV-related HNSCC in a 2­year-old child, in whom the tumor was located in the external auditory canal and who made a dramatic recovery after treatment with nonradical surgery, chemotherapy and radiotherapy. The child has currently been disease free for 6 years. This case supports the observation that HPV-related HNSCC tumors appear to respond favorably to treatment despite the patient's young age and the clinically advanced stage of the tumor.

Clinical importance of follicular lesion of undetermined significance (diagnostic category III according to Bethesda System) diagnosed from Fine-Needle Aspiration Biopsy.

INTRODUCTION: Follicular Lesion of Undetermined Significance (FLUS) belongs to the most controversial category of the Bethesda System. The aim of the study was to specify the risk of malignancy in patients with FLUS diagnosis in the material from the Institute of Oncology in Gliwice. This is the first Polish study specifying the risk of malignant neoplasm presence when Fine-Needle Aspiration Biopsy (FNAB) results in a report of diagnostic category III (DC III). MATERIAL AND METHODS: Three hundred and ninety-five primary DC III diagnoses from FNABs of the thyroid gland performed from 2010 to 2015 were analysed. Correspondence of DC III with diagnoses from repeated FNABs and histopathology reports was evaluated. RESULTS: From 395 DC III patients, 27 were treated surgically for clinical indications, receiving six diagnoses of cancer. Repeat FNAB was performed in 180 cases, and primary diagnosis was confirmed in 41 cases. In the second FNAB there was one diagnosis of "Papillary Thyroid Carcinoma" and one "Suspicious for Papillary Thyroid Carcinoma". From eight patients treated surgically in these series prior cytological cancer diagnosis was confirmed in two cases. Forty-six patients were subjected to third and subsequent FNABs; in one case the diagnosis was "Suspicious for Malignancy". In the analysed material the risk of cancer in patients with FLUS is 2.78%. Taking into account all 56 subsequent FNABs in which the primary diagnosis was confirmed, the risk decreases to 2.43%. CONCLUSIONS: The diagnosis of FLUS in the absence of clinical indications is not a basis for surgical treatment.

Suspicious for follicular neoplasm or follicular neoplasm? The dilemma of a pathologist and a surgeon.

INTRODUCTION: Cytological material obtained from Fine Needle Aspiration Biopsy (FNAB) does not permit us to distinguish between follicular carcinomas, adenomas, and hyperplastic nodules. The limitations of the method are: lack of possibility to assess the presence of tumour capsule, eventual capsular invasion, and angioinvasion. An unequivocal conclusion of whether what we have to deal with is a neoplastic or benign lesion is possible only after histopathological examination. The aim of the study was to confirm justification for using the term "Suspicious for Follicular Neoplasm" (SFN) in cytological diagnostics of thyroid carcinoma. MATERIAL AND METHODS: Three hundred and fifty-two primary SFN FNAB diagnoses (diagnostic category IV [DC IV] - according to Bethesda System) obtained from 2010 to 2015 in the Institute of Oncology in Gliwice were analysed, and their correlation with histopathological diagnoses was verified. RESULTS: In the Institute of Oncology in Gliwice, 352 primary SFN diagnoses (diagnostic category IV [DC IV] - according to Bethesda System) were established. Surgical treatment was undertaken after first FNAB in six cases, giving confirmation of a neoplasm in five cases, one of which was a follicular carcinoma. Second FNAB performed in 90 patients confirmed DC IV diagnosis in 53 cases. Third FNAB concerned 26 patients, providing another 14 diagnoses of DC IV. 26 out of 352 patients were subjected to surgery, and then histopathological examination confirmed a neoplasm in 19 cases (which comprises 73%), five of which were carcinomas. CONCLUSIONS: High positive predictive value PPV = 73% of SFN diagnosis justifies undertaking surgical treatment in any case of this diagnosis.

Human papillomavirus DNA in pharyngeal scrapes as a marker of HPV-related squamous cell cancer of the oropharynx.

BACKGROUND: Recent studies have indicated that human papillomavirus is an etiologic agent for a subset of head and neck cancers associated with better prognosis, therefore, prompt confirmation of such etiology seems to be crucial for choosing the optimal therapeutic option. Standard HPV diagnosis is currently based on histopathological material. In the present study, the novel diagnostic method based on pharyngeal brush biopsy is proposed. OBJECTIVES: The aim of this study was to evaluate the usefulness of the Real-Time PCR-based (RT-PCR) test in detecting HPV-related cancer of the oropharynx using superficial scraps taken from the oropharyngeal region. STUDY DESIGN: Ninety patients with head and neck squamous cell cancer were enrolled in the study. The presence of HPV DNA in pharyngeal superficial scrapes assessed by RT-PCR was compared to the HPV status in the tumor tissue samples determined by a combined RT-PCR/P16(INK4A) expression algorithm. Analytical sensitivity and specificity were calculated and the clinical outcome was analyzed in correlation to the HPV status. RESULTS: HR-HPV DNA in pharyngeal swabs was revealed in 25 cases (28.4%) and simultaneously confirmed in all corresponding tissue samples. Sensitivity and specificity of the viral status assessment in the brush biopsies in respect to the RT-PCR/P16(INK4A) 20 were 100% and 96.2%, respectively. HR-HPV positive status was associated with an excellent clinical outcome and reduced hazard ratio of recurrence and disease-related death. CONCLUSIONS: The proposed novel method of HPV status assessment using RT-PCR and superficial scraps appeared to be highly sensitive, specific, and useful in predicting the clinical outcome.

The prognostic significance of indoleamine-2,3-dioxygenase and the receptors for transforming growth factor ß and interferon γ in metastatic lymph nodes in malignant melanoma.

We analyzed the prognostic significance of indoleamine-2,3-dioxygenase (IDO) and type 1 receptors for transforming growth factor beta (TGF-ßR1) and interferon gamma (IFN-γR1) in resected nodal metastases of 48 malignant melanoma patients. In 32 cases the corresponding skin tumors were available. We used immunohistochemical (IHC) staining which was assessed by pathologists and by a computer-aided algorithm that yielded quantitative results, both absolute and relative. We correlated the results with the patient outcome. We identified absolute computer-assessed IDO levels as positively correlated with increased risk of death in a multivariate model (HR = 1.02; 95% CI: 1.002-1.04; p = 0.03). In univariate analysis, patients with IDO levels below the median had a better overall survival time (30.3 vs. 17.5 months; p = 0.03). TGF-ßR1 and IFN-γR1 expression was modestly correlated (R = 0.34; p lt; 0.05) and TGF-ßR1 expression was lower in lymph nodes than in matched primary skin tumors (Z = 2.87; p = 0.004). The pathologists' and computer-aided IHC assessment demonstrated high correlation levels (R = 0.61, R = 0.74 and R = 0.88 for IDO, TGF-ßR1 and IFN-γR1, respectively). Indoleamine-2,3-dioxygenase is prognostic for the patient outcome in melanoma with nodal involvement and should be investigated prospectively for its predictive significance. IHC assessment by computer-aided methods is recommended as its gives IHC more objectivity and reproducibility. ecting mismatch repair deficiency. Association of CDX2 and PMS2 in the present study is necessary to conduct further genetic and pathological studies focusing on these two markers together.

The presence and clinical implications of α-2,6-galactose-linked sialic acids in non-small-cell lung cancer brain metastases--preliminary study.

Brain metastases (BM) in non-small-cell lung cancer (NSCLC) patients present an increasing clinical challenge. Identifying biomarkers which specifically identify patients at high risk of BM may improve their early diagnosis, which is crucial for surgical and radiotherapeutic treatment outcome. Alpha-2,6-sialyltransferase (α-2,6-ST) and the primary product of its activity, alpha-2,6-galactose-linked sialic acids (α-2,6-GalSA) have been found responsible for the adhesion of tumor cells to the brain vessels' endothelium and enabling their transmigration through the blood-brain barrier in brain metastatic tumors. The aim of the study was to investigate by histochemical method the presence and possible role of α-2,6-GalSA in the formation of brain metastasis in NSCLC. In the screening phase 76 metastatic brain tumors were stained for α-2,6-GalSA and the second phase involved an identical staining of 20 primary tumors of patients who had their primary tumors treated with surgery or definite radiochemotherapy yet who later developed BM. The results were compared to a control group of 22 patients treated with surgery for NSCLC and who survived 5 years without the recurrence of disease. Alpha-2,6-GalSA presence was found to be down-regulated in poorly differentiated tumor types, whereas majority of differentiated tumors overexpressed it. This was statistically significant for both BM and the primary tumors. The expression of α-2,6-GalSA remained stable in primary and metastatic tumor pairs, however, no statistically significant differences were observed between study and control groups. Within the study group, a higher α-2,6-GalSA expression was associated with better overall survival, but not all statistical models found this result significant. Further studies are recommended to validate these findings.

Potential role of human papilloma virus in the pathogenesis of gastric cancer.

AIM: To demonstrate the presence and biological activity of human papilloma virus (HPV) in gastric cancer (GAC) tissues. METHODS: The study involved 84 surgically treated patients with gastric adenocarcinoma, regardless of the clinical stage of the disease. The presence of HPV DNA of high oncogenic risk types in formalin-fixed, paraffin-embedded tumor samples was determined using quantitative polymerase chain reaction analysis. A stringent protocol of prevention of cross- and environmental contamination was applied during DNA isolation, and amplification, as well as confirmation of the biological activity of the virus in tumor cells, was implemented. The study utilized the Real-time High Risk HPV test, which detects the DNA of 14 HPV subtypes that are considered to have high oncogenic potential. The overexpression of the p16(INK4a) protein assessed immunohistochemically was considered confirmation of the HPV infection. RESULTS: Among the 89 patients initially included in the study group, diagnostic results were obtained for 84 individuals. In five cases, either the histopathological material was too scant to isolate the necessary amount of DNA, or the isolated DNA was significantly degraded, resulting in the failure of internal control amplification within the predefined number of 35 cycles. Those patients were excluded from further analysis. The amplification of HPV DNA was demonstrated in none of the 84 tissue samples; thus, all cases were considered to have a negative DNA status of highly oncogenic HPV subtypes. Immunohistochemical staining provided diagnostic results for all of the examined tissue samples, and excluded the accumulation of the p16(INK4a) protein in tumor cells, thus confirming the lack of active HPV infection in all of the individuals. CONCLUSION: The study does not confirm the presence or biological activity of HPV in tumor tissues. Thus, the relationship between GAC and HPV infection, in the Central European population seems doubtful.