Clinical Characteristics and prognostic analysis of Triple-negative Breast Cancer: Single institute experience.

BACKGROUND: Triple-negative breast cancer (TNBC) is a poor prognostic subset of breast cancer that lacks the benefit of specific targeted therapy. MATERIALS AND METHODS: A prospective study of the clinical profile of triple negative breast cancer cases at a tertiary referral centre. The duration of the study period was 26 months and the median follow up period was ten months. A total of 111 invasive breast cancer patients were evaluated from 1st August 2009 to 31st October 2011. We examined TNBC patients with respect to clinicopathological parameters, adjuvant chemotherapy regimens and relapse free survival. RESULTS: In our study, patients were young (median age at presentation, 47yrs), premenopausal (54%), tumour size was discordant with lymph node positivity, the histology was predominantly intraductal carcinoma (90%), histological grade higher than two (90%). Relapses were early and preferential visceral (32%) and CNS metastasises (11.7%). 91% of patients were eligible for adjuvant therapy but only 80% of the patients could complete full course of adjuvant chemotherapy. Anthracycline-based regimens (43%), sequential anthracycline and taxane-based regimen (24%) and other regimes like CMF (13%) were used as adjuvant chemotherapy in eligible TNBC patients. Median relapse free survival in patients following adjuvant chemotherapy was around 10 months at last follow-up. CONCLUSIONS: Patients with TNBC have aggressive clinicopathological characteristics with early and higher rate of disease relapse and therefore derive inadequate benefit from current adjuvant chemotherapy. So, new treatment strategies in adjuvant chemotherapy for TNBC are needed.

Platinum-based chemotherapy in metastatic triple negative breast cancer: Experience of a tertiary referral centre in India.

UNLABELLED: Experimental data suggest that triple-negative breast cancer (TNBC) may have increased sensitivity to platinum-based chemotherapy but there is lack of relevant clinical data. Clinical outcomes in patients with metastatic TNBC treated with Platinum-based chemotherapy were evaluated in this prospective study. METHODS: 21 selected patients with metastatic TNBC presenting at GCRI during the study period from 1st August 2009 to 31st October 2011 formed the study group with median follow up period of 10 months. They were given palliative chemotherapy based upon prior adjuvant chemotherapy along with an additional platinum compound. Response rates, response duration and toxicities of platinum-based chemotherapy were recorded and analyzed. RESULTS: In evaluable TNBC patients, overall response rate and complete clinical response were 72% and 38% with median response duration of four months. Response could not be assessed in three patients due to patient refusal for evaluation, lost to follow up and toxicities. In three TNBC patients after completion of platinum based chemotherapy have early isolated CNS relapse with systemic disease in remission. Haematological adverse effects were febrile neutropenia in 19% of patients, and grade 3­4 neutropenia (9%) thrombocytopenia and anaemia (7%). The main non-hematological adverse effects reported in the present study were peripheral neuropathy (14%) and severe emesis (9%). The most common Platinum-based chemotherapy combination was carboplatin and paclitaxel in 11 patients (52%) of evaluable patients. Patients who received this regime have complete response rate, overall response rate and toxicity was 45%, 65% and 10%. CONCLUSIONS: TNBC patients with platinum-based chemotherapy have better overall response rates, higher complete clinical response rates, prolonged response duration and acceptable safety profile. The results of the present study need to be confirmed with a larger randomized study with a longer follow up.

A comparative therapeutic management of anoestrus in buffaloes using insulin and GnRH.

AIM: Anoestrus is one of the most common functional disorders of the reproductive cycle in buffaloes. In spite of technical advancement, there is no single cure for the management of anoestrus. Therefore, the aim of this study was to find out the efficacy of gonadotropic releasing hormone (GnRH) and metabolic hormone for the management of true anoestrus in buffaloes. MATERIALS AND METHODS: The experimental animals were selected on the basis of history, gyneco-clinical examinations and progesterone estimation. Deworming was done with Fenbendazole and thereafter mineral mixture was given @ 50 g per animal per day for 10 days in all the selected buffaloes before the start of treatment. The selected buffaloes were randomly divided into four groups (n=25). In Group I, buffaloes were administered 20 µg of buserelin intramuscularly. Buffaloes of Group II were administered long-acting insulin @ 0.25 IU/Kg body weight subcutaneously for 5 consecutive days. In Group III, buffaloes were treated with a combination of insulin and buserelin in the above-mentioned doses whereas buffaloes of Group IV were kept as untreated control. RESULTS: The higher oestrus induction (64% vs. 28%) was found in Group III and differed significantly (p<0.05) as compared to control group. The conception rate (69.23% vs. 66.66%) was also found higher in Group III but did not differ significantly among the treated groups. The mean time taken for the onset of oestrus was recorded significantly shorter in insulin (8.80±0.69) and GnRH (7.60±0.92 days) alone and as compared to other (Group III, 14.43±0.83 and Group IV, 20.57±1.69 days) groups. CONCLUSION: The results of this study indicated better fertility response using Insulin plus Buserelin in true anoestrus buffaloes under field conditions.

Fertility response in postpartum anoestrus buffaloes (Bubalus bubalis) using modified Ovsynch based timed insemination protocols.

BACKGROUND AND AIM: Suboestrus constitutes the largest factor responsible for poor reproductive efficiency in buffaloes. Therefore, oestrus synchronization (OS) and fixed-time artificial insemination (AI) is considered an alternative approach to enhance reproductive efficiency in buffaloes. Thus, the present study was carried out to study the efficacy of modified Ovsynch protocol with fixed time insemination in post-partum anoestrus buffaloes. MATERIALS AND METHODS: Total 50 post-partum anoestrus dairy buffaloes were selected and randomly divided into 5 Groups, each comprising ten animals (n=10). Animals of Group I received buserelin acetate 10 µg(GnRH) at day 0 and 9, and prostaglandin F2α (PGF2α) at day 7; in Group II similar to Group I except double dose of Gn RH (20 µg) at day 0; in Group III, similar to Group I plus additional administration of insulin on day 0, 1 and 2; in Group IV, similar to Group II plus additional administration of insulin on day 0,1 and 2; in Group V similar to Group I except replacement of first Gn RH with insulin on day 0,1 and 2. Animal of all groups were inseminated at fixed time using frozen semen at 60 h and 72 h after PGF2α administration and confirmed for pregnancy at day 60 post-insemination. RESULTS: The results revealed satisfactory and comparable synchronization of oestrus (60-80%) and conception rate (20-40%) among the various treatment groups in the present study. The better synchronization was observed in modified protocols. However, non-significantly higher conception rate was recorded in modified Ovsynch protocol (40% each in Group II-V) as compared to 20% in Group-I (p>0.05). CONCLUSION: In this study comparatively higher OS and conception following insulin modified Ovsynch based timed AI protocols in post-partum anoestrus buffaloes was found satisfactory and comparable.

Gene polymorphisms, tobacco exposure and oral cancer susceptibility: a study from Gujarat, West India.

Polymorphic variability in the enzymes involved in biotransformation of tobacco-related pro-carcinogens plays an important role in modulating oral cancer susceptibility. CYP1A1*2A, CYP1A1*2C, GSTM1 and GSTT1 polymorphisms were determined in 122 oral carcinoma cases and 127 controls from Gujarat, West India using PCR-based methods. The results revealed that the polymorphic variants of CYP1A1 gene did not show association towards oral cancer risk. The GSTM1 and GSTT1 null genotypes were found to be over-represented in patients than controls, suggesting a moderate increase in risk of oral cancer. The oral cancer risk was significantly increased in the patients having either alone or concurrent deletion of GSTM1 and GSTT1. The results also suggested significant association between tobacco habits, especially chewing, variant genotypes of CYP1A1, GSTM1 and GSTT1 and oral cancer risk. Our data have provided evidence that GST polymorphism modified the susceptibility to oral cancer and individuals with variant genotypes of the three genes with tobacco habits are at significant risk of developing oral cancer.

Overexpression of Bcl2 protein predicts chemoresistance in acute myeloid leukemia: its correlation with FLT3.

Potential prognostic biomarkers in acute myeloid leukemia (AML) can be identified by understanding the cellular pathway and molecular changes underlying leukemogenesis. Deregulation of apoptosis is one of the important features of AML and to understand the molecular mechanism underlying apoptosis and its contribution to tumor progression, this study aimed to evaluate anti-apoptotic Bcl2 protein expression in AML and correlate with FLT3 parameters for their role in prognosis of disease.Bcl2 and FLT3 protein expression was quantified by flow cytometry on leukemic blasts in total 174 de novo AML, myelodysplastic syndrome (MDS) and aplastic anemia patients. FLT3 internal tandem duplication (ITD), Tyrosine kinase domain (TKD) point mutations and quantification of mRNA level was carried out using PCR and RT-PCR methods. The incidence of Bcl2 positivity was 71% in AML patients. Bcl2 positivity was significantly associated with CD34+ and CD117+ AML. Bcl2 positivity tended to be associated with reduced DFS while Bcl2 positivity with FLT3 protein positivity was significantly associated with reduced DFS. In multivariate analysis, Bcl2+ and combined Bcl2+/FLT3 protein+ along with high WBC count emerged as poor prognostic factors for reduced DFS and high blast count for predicting reduced OS. In MDS patients, the incidence of Bcl2 expression was high while in aplastic anemia patients, incidence of Bcl2 expression was low.Patients with Bcl2 and FLT3 protein positivity showed significantly reduced DFS suggesting parallel role of these proteins in imparting chemoresistance to the leukemic cells.

Identification of site-specific prognostic biomarkers in patients with oral squamous cell carcinoma.

The purpose of the present study was to identify site-specific prognostic biomarkers in patients with oral squamous cell carcinoma (OSCC). For this purpose, Epidermal growth factor receptor (EGFR), Stat3, H-ras, c-myc, p53, cyclin D1, p16, Rb and Bcl-2 were localized immunohistochemically in buccal mucosa carcinoma (n=74) and tongue carcinoma (n=61) patients. Expression of markers was compared between buccal mucosa and tongue carcinoma and assessed for their prognostic value in site-specific manner. On comparison, only cyclin D1 showed significant difference in expression with higher accumulation in tongue tumors (r=+0.177, p=0.039). Moreover, univariate survival analysis showed that in buccal mucosa patients, loss of p16 and overexpression of H-ras were significant prognosticators for relapse-free survival (RFS) and overall survival (OS), respectively. However, in Cox multivariate analysis, they lost their significance after adjusting for significant clinicopathological parameters. On the other hand, in tongue cancer patients, Cox multivariate analysis showed that for RFS, Stat3 and c-myc, and for OS, Stat3, Bcl-2 and p53 were significant prognosticators after adjusting for significant confounding factors. Our findings indicated that buccal mucosa and tongue carcinoma exhibit different biological behavior which is reflected in prognosis. Therefore, this approach might be helpful to precisely identify patients for more effectively tailored treatment strategy.

Clinical significance of matrix metalloproteinase 2 and 9 in breast cancer.

BACKGROUND: Invasion and metastasis are the most strenuous problems in the management of breast cancer. These events require diverse proteolytic enzymes, among which MMP-2 and MMP-9 play a significant role in degradation of type IV collagen, the major component of the basement membrane. Therefore, the major objective of the study is to evaluate the clinical usefulness of MMP-2 and MMP-9 with respect to malignant tumor growth, invasion, and metastasis in breast cancer. MATERIALS AND METHODS: Gelatin zymography was performed on 157 tissue extracts of malignant and adjacent normal breast tissues as well as negative and positive lymph nodes from 49 breast cancer patients. Statistical analysis was carried out using SPSS statistical software (version 10). RESULTS: ProMMP-2 levels were significantly higher in adjacent normal tissues. Active MMP-2 and MMP-9 levels were higher in malignant breast tissues. Activation ratios of MMP-2 and MMP-9 were significantly higher in malignant breast tissues and in patients with lymph node metastasis. ProMMP-2, active MMP-2, and active MMP-9 could significantly discriminate between malignant and adjacent normal breast tissues. The MMP-2 activation ratio showed significant discriminatory efficacy between patients with and without lymph node metastasis and significant association with increased risk of lymph node metastasis in node-negative patients. CONCLUSION: The results indicate significant clinical utility of these proteolytic enzymes in malignant tumor growth, invasion, and metastasis in breast cancer.

Characterization of exon 2 and intron 2 of leptin gene in Indian goats.

Leptin, the hormonal product of the leptin (obese) gene (LEP), has multiple physiological effects and plays a pivotal role in the control of body growth, immune function, and reproduction. LEP gene polymorphism has been analyzed in cattle, buffalo, and pigs, and the polymorphic pattern has been associated with energy balance, milk production, and live weight and fertility trait in different livestock species. The present study has been designed to analyze polymorphism in exon 2 and intron 2 region of leptin gene in Indian goats. Genotyping was carried out in 111 kids including 70 samples from Barbari and 41 samples from Jamunapari breeds. The amplified product of exon 2 and intron 2 region of leptin gene was 152 bp and 400 bp, respectively, in both breeds. Sequencing of the exon 2 and intron 2 region of leptin gene and restriction analysis were carried out to analyze the polymorphism in goats. Five major haplotypes were observed in exon 2 region and six major haplotypes observed in intron 2 region in both breeds. Restriction fragment analysis and sequence analysis confirmed the mutation at 60 bp position of exon 2 and 100 bp position in intron 2 in all the analyzed samples. Sequences of exon 2 region of goats were unique as compared to other livestock species in BLAST analysis.

E-Cadherin truncation and sialyl Lewis-X overexpression in oral squamous cell carcinoma and oral precancerous conditions.

The present study aimed to determine significance of E-cadherin, a cell adhesion molecule, and sialyl Lewis-X (sLeX), a cell surface antigen, in oral carcinogenesis. Expressions of E-cadherin and sLeX were detected using western blot analysis from oral malignant (n=25), and oral precancerous tissues (OPC, n=20) and their adjacent normal tissues. An altered expression of E-cadherin (E-cad) and sLeX was observed in malignant and precancerous tissues. E-cad western blot revealed presence of two bands, a 120 kDa (native, E-cad120) and a 97 kDa (known as truncated E-cad97). The accumulation of truncated E-cad97 and sLeX in malignant and OPC tissues compared to their adjacent normal tissues was observed. Receiver's Operating Characteristics (ROC) curve analysis showed good discriminatory efficacy of E-cad97, E-cad97:120 ratio and sLeX between the malignant and adjacent. normal tissues. Further, a positive correlation of E-cad97 and sLeX overexpression with advanced stage of the disease and lymphnode metastasis was observed. The data suggest that E-cadherin truncation and sLeX overexpression are early events which may facilitate the tumor cells to metastasize. Also, overexpression E-cad97 and sLeX in OPC tissues may be useful to predict metastatic potentials of tumors at an early stage of oral carcinogenesis. Key words: Oral cancer, oral precancerous conditions, E-cadherin, sialyl Lewis-X, metastasis.

A new recurring chromosome 13 abnormality in two older patients with de novo acute myeloid leukemia: An Indian experience.

We report here two cases of trisomy 13 in acute myeloid leukemia M1 subtype. short-term unstimulated bone marrow and peripheral blood lymphocyte culture showed 47, XY, +13 in all metaphase plates and trisomy 13 was confirmed with whole chromosome paint probes. Trisomy 13 in AML-M1 is a rare numerical abnormality. This is the first Indian report of sole trisomy 13 in AML-M1. Here, we present two cases of elder male patients, which may constitute a distinct subtype.

A case of acute myeloid leukemia-M2 with trisomy 4 in addition to t(8;21).

t(8;21)(q22;q22) is the most frequently observed karyotypic abnormality associated with acute myeloid leukemia (AML), specifically in FAB-M2. Short-term unstimulated bone marrow (BM) and peripheral blood lymphocyte culture showed 47,XX, +4,t(8;21) in all metaphase plates; and interphase and metaphase results of AML-ETO fusion was positive and trisomy of 4 was confirmed with WCP probes. Trisomy 4 in AML with t(8;21) is a rare numerical abnormality. Here we present such case of patient which may constitute a distinctive subtype.

Cytoplasmic localization of BAG-1 in leukoplakia and carcinoma of the tongue: correlation with p53 and c-erbB2 in carcinoma.

BACKGROUND: The present study evaluated the clinical significance of BAG-1, an antiapoptotic protein, in leukoplakia and carcinoma of the tongue. METHODS: BAG-1 expression was evaluated by immunohistochemistry in paraffin-embedded tissues of leukoplakia (n=25) and carcinoma of the tongue (n=61). RESULTS: Cytoplasmic expression was predominantly seen in 80% and 70% of patients with leukoplakia and carcinoma, respectively. BAG-1 expression was found to be significantly lower in tobacco users than in non-tobacco users. BAG-1 expression in tobacco-using leukoplakia and carcinoma patients was compared by grouping the carcinoma patients according to lymph node status and disease stage. Carcinoma patients with tumor-positive lymph nodes had significantly lower BAG-1 expression than patients with negative lymph nodes and leukoplakia. Further, a trend towards an inverse correlation was observed with p53 and c-erbB2. In univariate and multivariate survival analysis, patient subgroups with 2+ or 3+ marker positivity (BAG-1 negativity, p53 and c-erbB2 positivity) had a reduced overall survival compared with patient subgroups with 1+ marker positivity or negativity. CONCLUSION: BAG-1 negativity in association with p53 and c-erbB2 positivity identified a subgroup of tongue cancer patients with an aggressive phenotype. Hence, an antiapoptotic protein, BAG-1, was found to be down-regulated in chewing-tobacco-mediated tongue carcinogenesis.

Molecular alterations in oral carcinogenesis: significant risk predictors in malignant transformation and tumor progression.

In this study an attempt was made to establish the significance of a battery of molecular alterations and thereby identify risk predictors in oral carcinogenesis. For this purpose, EGFR, Stat3, H-ras, c-myc, p53, cyclin D1, p16, Rb, Ki-67 and Bcl-2 were localized immunohistochemically in normal mucosa (n=12), hyperplasia (n=35), dysplasia (n=25), early stage carcinoma (n=65) and advanced stage carcinoma (n=70). Deregulation occurred at an early stage and the number of alterations increased with disease progression. Using multivariate logistic regression analysis, the significant risk predictor for hyperplasia from normal mucosa was Ki-67 (OR=5.75, p=0.021); the significant risk predictors for dysplasia from hyperplasia were EGFR (OR=12.96, p=0.002), Stat3 (OR=17.16, p=0.0001), p16 (OR=5.50, p=0.039) and c-myc (OR=5.99, p=0.052); the significant risk predictors for early stage carcinoma from dysplasia were p53 (OR=6.63, p=0.0001) and Rb (OR=3.81, p=0.056); and the significant risk predictors for further progression were EGFR (OR=5.50, p=0.0001), Stat3 (OR=4.49, p=0.0001), H-ras (OR=4.05, p=0.001) and c-myc (OR=2.99, p=0.015). Cyclin D1 holds a key position linking upstream signaling pathways to cell cycle regulation. Gene products of the mitogenic signaling pathway play an equally significant role as cell cycle regulatory proteins in the hyperplasia-dysplasia-early-advanced-carcinoma sequence and together may provide a reference panel of markers for use in defining premalignant lesions and predicting the risk of malignant transformation and tumor progression.

Stat3 expression in oral squamous cell carcinoma: association with clinicopathological parameters and survival.

The present study sought to explore the occurrence of signal transducer and activator of transcription 3 (Stat3) in patients with oral squamous cell carcinoma (n=135) and its potential relationship with clinicopathological parameters and survival. Stat3 expression was studied by immunohistochemistry. Cytoplasmic or nuclear localization of Stat3 was observed in 62% of patients, whereas only nuclear Stat3 expression was found in 44%. Stat3 positivity in early-stage patients was 45% compared to 79% in advanced-stage patients. However, early-stage Stat3-positive patients showed a gradual increase in staining intensity, with intense staining seen in 52% of the tumors compared to 18% in Stat3-positive advanced-stage patients, where a gradual decrease in intensity expression was observed (p=0.001). Stat3 showed a significant positive correlation with disease stage (p=0.001), nodal status (p=0.033) and tumor size (p=0.001). Multivariate survival analysis using the Cox proportional hazard regression model showed that nuclear Stat3 was a significant independent prognosticator for both relapse-free survival (p=0.014) and overall survival (p=0.042) in early-stage patients. Our results indicated that Stat3 activation is an early event in oral squamous cell carcinoma and represents a potential risk factor for poor prognosis in early-stage patients.

p53 expression in leukoplakia and carcinoma of the tongue.

There is growing interest in assessing multistep carcinogenesis and predicting its course using different molecular markers. TP53 is a tumor suppressor gene and appears to be one of the molecular targets of tobacco-related carcinogens in oral cancer. The present study evaluated the role of p53 expression in patients with leukoplakia and carcinoma of the tongue. p53 expression was studied by immunohistochemistry. All patients with leukoplakia of the tongue were male tobacco users. Nuclear staining of p53 was observed in 79% of those patients. Fifty percent, 25% and 4% of the patients expressed 1+, 2+ and 3+ nuclear staining, respectively. When leukoplakia patients were graded according to histopathology, 67% had hyperplasia and 33% had dysplasia. Nuclear p53 accumulation was 88% in hyperplasia and 62% in dysplasia. In patients with tongue cancer, nuclear accumulation of p53 was seen in only 19% of the tumors, with a staining intensity of 1+ in 13%, 2+ in 2% and 3+ in 4% of the tumors. The prevalence of nuclear p53 positivity (79%) was significantly higher in patients with leukoplakia than in patients with tongue cancer (19%; chi2 = 34.32, r = -0.45, df = 1, p = 0.0001; odds ratio (OR) = 16.66, 95% CI, 5.25-52.86). Therefore, leukoplakia patients who show p53 expression have a higher risk of developing tongue cancer than those who do not show p53 expression. As the percentage of positivity of nuclear p53 was very low, none of the clinicopathological parameters or disease status showed any significant association with it. The interesting finding is that none of the female cancer patients showed nuclear p53 expression. Therefore, p53 accumulation is believed to be an early event in neoplastic progression of the tongue.

Usefulness of terminal deoxynucleotidyl transferase as prognosticator in leukemia patients.

Peripheral blood samples from 55 previously untreated leukemia patients (33 males, 22 females) were analysed for terminal deoxynucleotidyl transferase (TdT) activity. TdT was significantly higher in patients with acute myelogenous leukemia (AML; P < 0.001), chronic myelogenous leukemia (CML; P < 0.05) and acute lymphoblastic leukemia (ALL; P < 0.001) when compared with controls. One patient with chronic lymphocytic leukemia (CLL) had undetectable TdT. Among leukemic patients, ALL patients had higher concentration of TdT than CML and AML patients. Females had higher TdT activity than males, although the difference between the two groups was not statistically significant. 68% TdT+ and 32% TdT- patients were in blastic crisis. Patients with more than 10% of blasts in the circulation had significantly higher TdT than blast-negative patients (P < 0.001). No difference in survival was observed between TdT+ and TdT- group. From these results, we conclude that the absolute TdT concentration is of little prognostic value in leukemia patients.