Nitin Gangane

mTOR as a Molecular Target in HPV-Associated Oral and Cervical Squamous Carcinomas

Purpose: The incidence of head and neck squamous cell carcinomas (HNSCC) associated with human papillomavirus (HPV) infection has increased over the past decades in the United States. We aimed at examining the global impact of HPV-associated HNSCC and whether the established key role of mTOR activation in HNSCC is also observed in HPV+ HNSCC lesions, thereby providing novel treatment options for HPV-associated HNSCC patients. Experimental Design: An international HNSCC tissue microarray (TMA) was used to analyze the expression of p16INK4A, a surrogate for HPV infection, and Akt-mTOR pathway activation. Results were confirmed in a large collection of HPV− and HPV+ HNSCC cases and in a cervical cancer (CCSCC) TMA. Observations were validated in HNSCC and CCSCC-derived cell lines, which were xenografted into immunodeficient mice for tumorigenesis assays. Results: Approximately 20% of all HNSCC lesions could be classified as HPV+, irrespective of their country of origin. mTOR pathway activation was observed in most HPV+ HNSCC and CCSCC lesions and cell lines. The preclinical efficacy of mTOR inhibition by rapamycin and RAD001 was explored in HPV+ HNSCC and CCSCC tumor xenografts. Both mTOR inhibitors effectively decreased mTOR activity in vivo and caused a remarkable decrease in tumor burden. These results emphasize the emerging global impact of HPV-related HNSCCs and indicate that the activation of the mTOR pathway is a widespread event in both HPV− and HPV-associated HNSCC and CCSCC lesions. Conclusions: The emerging results may provide a rationale for the clinical evaluation of mTOR inhibitors as a molecular targeted approach for the treatment of HPV-associated malignancies. Clin Cancer Res; 18(9); 2558–68. ©2012 AACR.

Association of Epstein Barr Virus Infection (EBV) with Breast Cancer in Rural Indian Women

Introduction Breast cancer is the most common malignancy affecting females worldwide but conventional risk factors are able to explain only a small proportion of these cases. A possible viral etiology for breast cancer has been proposed and Epstein-Barr Virus (EBV) is a widely researched candidate virus. The aim of the present study, first one of its kind from India, was to determine if there is a greater association of EBV infection with breast cancer patients as compared to patients with benign breast diseases. Methods We looked for expression of Epstein-Barr Virus Nuclear Antigen-1 (EBNA-1) in breast cancer tissue specimens by employing immunohistochemistry (IHC). We also measured levels of anti-EBNA-1 Immunoglobulin (IgG) antibodies in stored sera of these patients using commercial Enzyme linked Immunosorbent Assay (ELISA) kit. Patients with benign breast diseases were used as a comparison group for both immunohistochemical and serological analysis. Results 58 cases of malignant breast disease and 63 of benign breast disease (controls) were included in the study. Using manufacturer determined cut-off of 3 IU/ml, 50/55 tested (90.9%) cases and 27/33 tested (81.8%) controls were seropositive for anti-EBNA-1 IgG. Mean antibody levels were significantly higher for cases (54.22 IU/ml) as compared to controls (18.68 IU/ml). IHC for EBNA-1 was positive in 28/51 cases (54.9%). No IHC positivity was noted in the tested 30 controls. Our results show that EBNA-1 expression is seen in a significant proportion of breast cancer tissue specimens from rural India and as compared to patients with benign breast diseases these patients also have a higher immunological response against EBNA-1.

Prevalence of human papillomavirus types in cervical and oral cancers in central India.

Cervical cancer that is associated with high-risk human papillomaviruses (HPV) is the most common malignancy in Indian women. Therefore, the establishment of a prevention program is urgently required considering both vaccination and screening. However, relatively little is known about the prevalence of the different HPV types in cervical cancers in different regions of India, particularly central India. In this study, we have determined the HPV type distribution in 180 cervical cancers of women from Sevagram, a rural area from central India. In addition, we have analyzed other epithelial cancers that are known to be in part associated with high-risk HPV infection, i.e. oral (n=65), vulva and vagina (n=7) and penis (n=7). Approximately 93% of cervical cancers were positive for one or more of the high-risk HPV types. HPV16 was the predominant type being present in 81.7% of the cases. The remaining most predominant high-risk HPV types were: 18, 31, 35, 45, 56 and 59. Oral and vulva/vagina cancers were exclusively associated with HPV16, 27.7% and 70%, respectively. None of penile cancers was found positive for any of the high-risk HPV types. These data show that HPV16 vaccination in this geographical region will have considerable impact on the prevention of cervical and other epithelial cancers.

Prevention of tumor growth driven by PIK3CA and HPV oncogenes by targeting mTOR signaling with metformin in oral squamous carcinomas expressing OCT3

Most squamous cell carcinomas of the head and neck (HNSCC) exhibit a persistent activation of the PI3K–mTOR signaling pathway. We have recently shown that metformin, an oral antidiabetic drug that is also used to treat lipodystrophy in HIV-infected (HIV+) individuals, diminishes mTOR activity and prevents the progression of chemically induced experimental HNSCC premalignant lesions. Here, we explored the preclinical activity of metformin in HNSCCs harboring PIK3CA mutations and HPV oncogenes, both representing frequent HNSCC alterations, aimed at developing effective targeted preventive strategies. The biochemical and biologic effects of metformin were evaluated in representative HNSCC cells expressing mutated PIK3CA or HPV oncogenes (HPV+). The oral delivery of metformin was optimized to achieve clinical relevant blood levels. Molecular determinants of metformin sensitivity were also investigated, and their expression levels were examined in a large collection of HNSCC cases. We found that metformin inhibits mTOR signaling and tumor growth in HNSCC cells expressing mutated PIK3CA and HPV oncogenes, and that these activities require the expression of organic cation transporter 3 (OCT3/SLC22A3), a metformin uptake transporter. Coexpression of OCT3 and the mTOR pathway activation marker pS6 were observed in most HNSCC cases, including those arising in HIV+ patients. Activation of the PI3K–mTOR pathway is a widespread event in HNSCC, including HPV− and HPV+ lesions arising in HIV+ patients, all of which coexpress OCT3. These observations may provide a rationale for the clinical evaluation of metformin to halt HNSCC development from precancerous lesions, including in HIV+ individuals at risk of developing HPV− associated cancers. Cancer Prev Res; 8(3); 197–207. ©2015 AACR.

Accuracy of a multispecies rapid diagnostic test kit for detection of malarial parasite at the point of care in a low endemicity region

Although highly accurate rapid diagnostic tests (RDT) for Plasmodium falciparum [based on identification of histidine-rich protein-2 (PfHRP2)] have been developed, the accuracy of non-falciparum tests is relatively poor. Recently, a Plasmodium vivax-specific RDT [based on identification of species-specific lactate dehydrogenase (PvLDH)] became available, which along with PfHRP2 may improve malaria diagnosis by identifying the species correctly. A cross-sectional hospital-based study was designed to evaluate the diagnostic accuracy of FalciVax, a commercially available PfHRP2- and PvLDH-based RDT (index test), using malaria microscopy as a reference standard. All consecutive inpatients who presented with fever underwent both the index test and the reference standard. The study sample included 657 patients and the overall sensitivity and specificity of the RDT for diagnosis of any malarial species were 92.9% and 98.4%, respectively. The diagnostic accuracy estimates for correct species identification were lower (sensitivity 91.8%, specificity 96.8%). The accuracy of the PvLDH test to detect P. vivax was low (sensitivity 76.6%, specificity 98.1%).

Osteochondroma of the Sacrum with a Correlative Radiographic and Histological Evaluation

Primary benign sacral tumors are rare representing only 1–4% of solitary exostoses and constituting 4% of all solitary spinal tumor. The tumor usually comes to attention in childhood or adolescence. The treatment of symptomatic lesions is resection. We present here a case of a sacral osteochondroma with characteristic histological and radiological features in an adolescent presenting with radicular pain.

Pleomorphic lobular carcinoma of the breast: A case report

BACKGROUND Pleomorphic lobular carcinoma of the breast is associated with aggressive behavior. CASE Fine needle aspiration cytology was performed on a breast lump in a 55-year-old woman. The aspirates showed highly pleomorphic, large cells in a dyscohesive pattern, with a tendency of few cells to aggregate in small groups. A diagnosis of ductal carcinoma was made on cytology. On histology, the paraffin sections showed features of pleomorphic lobular carcinoma of the breast. CONCLUSION The cytologic features of pleomorphic lobular carcinoma overlap with those of infiltrating ductal carcinoma. It is very difficult to make a diagnosis of pleomorphic lobular carcinoma prospectively on cytology. However, if Indian file arrangement and cytoplasmic vacuolation are present, pleomorphic lobular carcinoma must at least be suggested for the differential diagnosis as it has different clinical implications.

Primary cutaneous diffuse large B cell lymphoma, leg type.

Das diffus-großzellige B-Zell-Lymphom des Beines (primary cutaneous diffuse large B-cell lymphoma, leg type, PCDLBL, LT) zeichnet sich durch das Auftreten von rötlichen bis bläulichen Knoten und Tumoren im Bereich der unteren Extremität bei älteren Patienten aus [1–4]. Aufgrund des häufig vorkommenden extrakutanen Befalls ist die Prognose mit einer 5-Jahres-Überlebensrate von 55% erheblich schlechter als bei den indolenten kutanen B-Zell-Lymphomen [5]. In der aktuellen WHO-EORTC-Lymphom-Klassifikation der kutanen Lymphome werden neben dem PCDLBL, LT, auch noch andere Formen des primär kutanen, diffus-großzelligen B-Zell-Lymphoms aufgeführt (PCDLBL, other) [1,2,6]. Dabei handelt es sich um diffuse B-Zell-Lymphome, welche sich klinisch, histologisch und phänotypisch nicht eindeutig dem PCDLBL, LT, bzw. dem primär kutanen Keimzentrumslymphom zuordnen lassen [1,2,6]. Anamnese !

Role of modified bleach method in staining of acid-fast bacilli in lymph node aspirates.

OBJECTIVE To correlate acid-fast bacilli (AFB) positivity with cytomorphologic patterns of tuberculous lymphadenitis and evaluate bleach concentration method in diagnosing lymph node tuberculosis compared to Ziehl-Neelsen (ZN) method. STUDY DESIGN One hundred cases of tuberculous lymphadenitis diagnosed by fine needle aspiration cytology (FNAC) were analyzed and classified into 6 cytomorphologic patterns and correlated with bacillary load using routine and modified bleach methods of ZN staining. Smears were graded for AFB positivity. Sensitivity of routine ZN and modified bleach concentration was compared. RESULTS The classic cytomorphologic pattern of tuberculosis showing epithelioid granulomas, Langerhans giant cells and caseous necrosis was seen in 23% of cases. Routine ZN staining detected AFB in 27% of cases and the modified bleach method in 72%. In 58 cases the modified bleach method had a higher grade of AFB positivity than the routine method. The modified bleach method did not miss any AFB positivity detected on routine ZN staining. CONCLUSION The modified bleach method demonstrated AFB positivity in 72% of cases. AFB positivity grade was much higher than with routine ZN staining, making bacilli easily visible, with shorter screening time. The modified bleach method is inexpensive, easily performed and more sensitive and safe than routine ZN staining.

Cytodiagnosis of eumycotic mycetoma: a case report.

BACKGROUND Mycetoma is a late-stage clinical manifestation of a subcutaneous infection produced by bacteria (actinomycetoma) or fungi (eumycetoma). Only a few articles have described the morphologic appearance of this uncommon pathology on cytology. The distinction between eumycetoma and actinomycetoma in fine needle aspiration cytology (FNAC) is as accurate as in histopathology, as demonstrated in the present case. CASE A 30-year-old man presented with a large swelling on his left foot with a discharging sinus. FNAC of the swelling yielded pus-like material. Initial Papanicolaou and Giemsa stains showed the presence of septate, branching fungal hyphae and black granules against the inflammatory background. The presence of fungus was confirmed by PAS stain. The species was identified as Exophiala jeanselmei on fungal culture. CONCLUSION Mycotic mycetoma can be accurately diagnosed by FNAC, which is a simple, inexpensive and rapid technique when there is a high index of suspicion. Special stains and culture studies are helpful in confirmation of diagnosis and species identification.