Diptendra Kumar Sarkar

Curcumin reverses T cell-mediated adaptive immune dysfunctions in tumor-bearing hosts

Immune dysfunction is well documented during tumor progression and likely contributes to tumor immune evasion. CD8+ cytotoxic T lymphocytes (CTLs) are involved in antigen-specific tumor destruction and CD4+ T cells are essential for helping this CD8+ T cell-dependent tumor eradication. Tumors often target and inhibit T-cell function to escape from immune surveillance. This dysfunction includes loss of effector and memory T cells, bias towards type 2 cytokines and expansion of T regulatory (Treg) cells. Curcumin has previously been shown to have antitumor activity and some research has addressed the immunoprotective potential of this plant-derived polyphenol in tumor-bearing hosts. Here we examined the role of curcumin in the prevention of tumor-induced dysfunction of T cell-based immune responses. We observed severe loss of both effector and memory T-cell populations, downregulation of type 1 and upregulation of type 2 immune responses and decreased proliferation of effector T cells in the presence of tumors. Curcumin, in turn, prevented this loss of T cells, expanded central memory T cell (TCM)/effector memory T cell (TEM) populations, reversed the type 2 immune bias and attenuated the tumor-induced inhibition of T-cell proliferation in tumor-bearing hosts. Further investigation revealed that tumor burden upregulated Treg cell populations and stimulated the production of the immunosuppressive cytokines transforming growth factor (TGF)-β and IL-10 in these cells. Curcumin, however, inhibited the suppressive activity of Treg cells by downregulating the production of TGF-β and IL-10 in these cells. More importantly, curcumin treatment enhanced the ability of effector T cells to kill cancer cells. Overall, our observations suggest that the unique properties of curcumin may be exploited for successful attenuation of tumor-induced suppression of cell-mediated immune responses.

Aspirin Suppresses the Acquisition of Chemoresistance in Breast Cancer by Disrupting an NFκB-IL6 Signaling Axis Responsible for the Generation of Cancer Stem Cells.

Acquired chemoresistance has curtailed cancer survival since the dawn of chemotherapy. Accumulating evidence suggests a major role for cancer stem cells (CSC) in chemoresistance, although their involvement in acquired resistance is still unknown. The use of aspirin has been associated with reduced cancer risk and recurrence, suggesting that the anti-inflammatory drug may exert effects on CSCs. In this study, we investigated the contribution of CSCs to acquired chemoresistance of breast cancer and the avenues for reversing such effects with aspirin. We observed that the residual risk of recurrence was higher in breast cancer patients who had acquired chemoresistance. Treatment of preexisting CSCs with a genotoxic drug combination (5-fluorouracil, doxorubicin, and cyclophosphamide) generated an NFκB-IL6-dependent inflammatory environment that imparted stemness to nonstem cancer cells, induced multidrug resistance, and enhanced the migration potential of CSCs. Treatment with aspirin prior to chemotherapy suppressed the acquisition of chemoresistance by perturbing the nuclear translocation of NFκB in preexisting CSCs. Therefore, disruptions to the NFκB-IL6 feedback loop prevented CSC induction and sensitized preexisting CSCs to chemotherapy. Collectively, our findings suggest that combining aspirin and conventional chemotherapy may offer a new treatment strategy to improve recurrence-free survival of breast cancer patients. Cancer Res; 76(7); 2000-12. ©2016 AACR.

Mithramycin A sensitizes therapy-resistant breast cancer stem cells toward genotoxic drug doxorubicin

Chemotherapy resistance is a major clinical challenge for the management of locally advanced breast cancer. Accumulating evidence suggests a major role of cancer stem cells (CSCs) in chemoresistance evoking the requirement of drugs that selectively target CSCs in combination with chemotherapy. Here, we report that mithramycin A, a known specificity protein (Sp)1 inhibitor, sensitizes breast CSCs (bCSCs) by perturbing the expression of drug efflux transporters, ATP-binding cassette sub-family G, member 2 (ABCG2) and ATP-binding cassette sub-family C, member 1 (ABCC1), survival factors, B-cell lymphoma 2 (Bcl-2) and X-linked inhibitor of apoptosis (XIAP), and, stemness regulators, octamer-binding transcription factor 4 (Oct4) and Nanog, which are inherently upregulated in these cells compared with the rest of the tumor population. In-depth analysis revealed that aberrant overexpression of Sp1 in bCSCs transcriptionally upregulates (1) resistance-promoting genes to protect these cells from genotoxic therapy, and (2) stemness regulators to sustain self-renewal potential of these cells. However, mithramycin A causes transcriptional suppression of these chemoresistant and self-renewal genes by inhibiting Sp1 recruitment to their promoters. Under such antisurvival microenvironment, chemotherapeutic agent doxorubicin induces apoptosis in bCSCs via DNA damage-induced reactive oxygen species generation. Cumulatively, our findings raise the possibility that mithramycin A might emerge as a promising drug in combinatorial therapy with the existing chemotherapeutic agents that fail to eliminate CSCs. This will consequently lead to the improvement of therapeutic outcome for the treatment-resistant breast carcinomas.

MEK inhibition prevents tumour-shed transforming growth factor-β-induced T-regulatory cell augmentation in tumour milieu.

Tumour progression is associated with immune‐suppressive conditions that facilitate the escape of tumour cells from the regimen of immune cells, subsequently paralysing the host defence mechanisms. Induction of CD4+ CD25+ FoxP3+ T regulatory (Treg) cells has been implicated in the tumour immune escape mechanism, although the novel anti‐cancer treatment strategies targeting Treg cells remain unknown. The focus of this study is to define the interaction between tumour and immune system, i.e. how immune tolerance starts and gradually leads to the induction of adaptive Treg cells in the tumour microenvironment. Our study identified hyperactivated mitogen‐activated protein kinase kinase (MEK)/extracellular signal‐regulated kinase (ERK) ‐signalling as a potential target for reversing Treg cell augmentation in breast cancer patients. In more mechanistic detail, pharmacological inhibitors of MEK/ERK signalling inhibited transforming growth factor‐β (TGF‐β) production in tumour cells that essentially blocked TGF‐β‐SMAD3/SMAD4‐mediated induction of CD25/interleukin‐2 receptor α on CD4+ T‐cell surface. As a result high‐affinity binding of interleukin‐2 on those cells was prohibited, causing lack of Janus kinase 1 (JAK1)/JAK3‐mediated signal transducer and activator of transcription 3 (STAT3)/STAT5 activation required for FoxP3 expression. Finally, for a more radical approach towards a safe MEK inhibitor, we validate the potential of multi‐kinase inhibitor curcumin, especially the nano‐curcumin made out of pure curcumin with greater bioavailability; in repealing tumour‐shed TGF‐β‐induced Treg cell augmentation.

Role of Nuclear Factor-κB in female Breast Cancer: A Study in Indian Patients

INTRODUCTION The nuclear factor κB (NF-κB) is a super family of transcription factors which plays important roles in development and progression of cancer. The present investigation concerns NF-κB /p65 activity in human breast cancers with overexpression of ER, PR, HER-2/neu, as well as the significance of p65 expression with regard to menopausal status, stage, grade, tumor size, nodal status, and NPI of invasive ductal carcinomas in Eastern India. MATERIALS AND METHODS In this hospital based study 57 breast cancer patients attending a Breast Clinic of a reputed institute of Eastern India were assessed for p65 protein expression in breast tumor tissue samples by Western blotting. ER, PR and HER-2/neu expression was determined by immunohistochemistry. RESULTS NF-κB/p65 was significantly associated with advanced stage, large tumor size (≥5 cm), high grade, negative ER, negative PR, and positive HER-2/neu. High NF-κB/p65 expression was more frequent in patients with a high NPI (NPI≥5.4, 84.6%) compared with low NPI (<5.4, 44.4%) and this association was statistically significant (p=0.002). CONCLUSION NF-κB/p65 overexpression was associated with advanced stage, large tumor size, high grade, and high NPI which are poor prognostic factors linked to enhanced aggressiveness of the disease. NF-κB/p65 expression implies aggressive biological behavior of breast cancer and this study validates significant association of NF-κB /p65 overexpression with negative estrogen and progesterone receptor status and overexpression of HER-2/neu oncoprotein. In our good clinical practice, patients with NF-κB positive tumors need to be treated aggressively.

Prognostic Significance of HER-2/neu and Survival of Breast Cancer Patients Attending a Specialized Breast Clinic in Kolkata, Eastern India

INTRODUCTION The worldwide incidence of breast cancer has increased rapidly in recent years. The scenario of Eastern India is also showing the same trend. It is necessary to study the utility of HER-2/neu as a prognostic factor in breast cancer survival. However, there have not been detailed studies in this respect with the breast cancer patients of Eastern India. Thus this study was conducted. MATERIALS AND METHODS In this hospital-based study 86 breast cancer patients attending a breast clinic of a reputed institute of Eastern India and having invasive ductal carcinomas were observed for a period of 5 years after surgery. Associations between 5 years observed survival and status of ER, PR and HER-2/neu of the patients were critically evaluated. RESULTS There was statistically significant association between survival pattern for 5 years and the HER-2/neu status (p=0.00001). Better survival was observed for the patients with HER-2/neu negative tumors 67(100%) compared to HER-2/neu positive tumors 7(36.8%). CONCLUSION There is strong interaction between survival and HER-2/neu expression of breast cancer patients. Thus the patients with HER-2/neu positive tumors need to be treated aggressively.

Study of Immunohistochemistry in Prostatic Lesions with Special Reference to Proliferation and Invasiveness

Prostatic lesions on routine staining sometimes cause diagnostic dilemma especially in premalignant lesions like atypical adenomatous hyperplasia and prostatic intraepithelial neoplasia. Benign small acinar lesions also may be difficult to differentiate from small acinar adenocarcinoma. An important differentiating point is the loss of basal cell layer in adenocarcinoma and its presence in benign lesions. Basal cell markers (e.g. 34βE12 cytokeratin) & proliferative markers (e.g. AgNOR and PCNA) can help in this regard. Total 60 cases of different prostatic lesions studied. After history taking, clinical examination, radiological & other investigations were done. Routine H&E staining, immunohistochemical staining against 34βE12 cytokeratin & proliferative markers (AgNOR & PCNA) was performed. Statistically significant differences found in expression of 34βE12 cytokeratin and proliferative markers between benign, premalignant and malignant prostatic lesions. Basal cell markers and proliferative markers are important parameters to distinguish between different benign, premalignant and malignant prostatic lesions.

Is estrogen receptor study useful in prognostication of breast cancer patients in India

BackgroundTo assess the validity of estrogen receptor (ER) status of breast tumor as a prognostic marker in clinical practice in the Indian perspective.Materials and methodsSixty-three patients of breast cancer attending breast clinic IPGMER, Kolkata were included in this study. All patients underwent surgery. Metastatic status and tumor were done. Immunohistochemistry was also used for further analysis.ResultsStatistically we have proved that advancing course of the disease, there is a tendency of the tumors to become hormone insensitive.ConclusionsER has a strong prognostic importance in early breast cancer and can play a major role in optimizing treatment modalities in node negative early breast cancer.

Cytological and Morphometric Study of Urinary Epithelial Cells with Histopathological Correlation

This study has been done to asses the utility and accuracy of urinary cytology and morphometric study of exfoliated cells in early detection and follow-up of urothelial neoplasms and thereby help to reduce the disease-related mortality and morbidity. A total 100 patients with urinary symptoms were studied by cytological examination of urine along with morphometric analysis of suspicious epithelial cells. Immunostaining to detect CK-20 expression and p53 over expression was done in smears showing atypical cells. Histopathological confirmation was done in cases which were suspicious on cystoscopy. P value was determined by using unpaired t-test. Statistically significant difference was found between neoplastic and non-neoplastic lesions of the urinary bladder as far as morphometry is concerned. Urinary cytology along with morphometry is an important tool in early detection of urothelial neoplasms. It is also helpful to find out the recurrences during post-operative follow-up period. Overexpression of CK-20 and p53 immunostain in cytology can act as an adjunct to the cytological diagnosis.

Oncoplastic Breast Surgery - Our Experiences in the Breast Clinic, IPGME&R, Kolkata

Oncoplastic breast surgery is the fusion of oncological and surgical principles to gain successful breast tumour excision with good cosmesis. It is an widely accepted and popular method in the western world. However, the picture is different in India. The major issues here, like late detection and advanced tumours, poor socio-economic status overriding quality of life issue and shortage of dedicated oncoplastic surgeons result in a poor acceptance and practice of oncoplastic breast surgery. This article explores the use of various oncoplastic techniques in clinical practice and discusses future directions in this emerging field in an Indian perspective. In our institute (breast care unit, I.P.G.M.E&R,Kolkata), we performed a retrospective study over a period of five years (2005–2009). It included a sample size of 30 patients with diagnosed breast cancer or Phylloides Tumor (PT). The study focused on the indications, type of oncoplastic procedure used, cosmetic outcome (shape / volume replacement, ptosis correction & chest wall coverage), complications faced and response to subsequent therapy. The indications, for which oncoplastic techniques were applied, were DCIS (2), LCIS (1), IDC (19) and phylloides tumor (8). Of the 30 patients, RAT was used in 8, LDMF (of various types and volume) was used in 16, pedicled TRAM flap was used in 3 and reduction mastopexy in 3.The procedural indications of reconstruction were total glandular replacement by TRAM flap, mini-LDMF to fill volume loss after BCS or wide local excision, rotation advancement technique for reshaping / symmetry maintenance after BCS or wide local excision, LDMF for chest wall coverage after MRM and reduction mastopexy after wide local excision. From patient’s point of view the outcome of surgery was highly satisfactory (score 3 or more) in 19 out of 30 patients (63.33%). LDMF was the most commonly used (16 out of 30) oncoplastic procedure with least complication rates (0 out of 16). 4 out of 30 patients had complications related to the procedure. Chest wall coverage after MRM still forms the main indication of oncoplastic surgery in this country.