Breast cancer cell-derived microRNA-155 suppresses tumor progression via enhancing immune cell recruitment and antitumor function.

Evidence suggests that increased microRNA-155 (miR-155) expression in immune cells enhances antitumor immune responses. However, given the reported association of miR-155 with tumorigenesis in various cancers, a debate is provoked on whether miR-155 is oncogenic or tumor suppressive. We aimed to interrogate the impact of tumor miR-155 expression, particularly that of cancer cell-derived miR-155, on antitumor immunity in breast cancer. We performed bioinformatic analysis of human breast cancer databases, murine experiments, and human specimen examination. We revealed that higher tumor miR-155 levels correlate with a favorable antitumor immune profile and better patient outcomes. Murine experiments demonstrated that miR-155 overexpression in breast cancer cells enhanced T cell influx, delayed tumor growth, and sensitized the tumors to immune checkpoint blockade (ICB) therapy. Mechanistically, miR-155 overexpression in breast cancer cells upregulated their CXCL9/10/11 production, which was mediated by SOCS1 inhibition and increased phosphorylated STAT1 (p-STAT1)/p-STAT3 ratios. We further found that serum miR-155 levels in breast cancer patients correlated with tumor miR-155 levels and tumor immune status. Our findings suggest that high serum and tumor miR-155 levels may be a favorable prognostic marker for breast cancer patients and that therapeutic elevation of miR-155 in breast tumors may improve the efficacy of ICB therapy via remodeling the antitumor immune landscape.

Optimizing Tumor Microenvironment for Cancer Immunotherapy: ß-Glucan-Based Nanoparticles.

Immunotherapy is revolutionizing cancer treatment. Recent clinical success with immune checkpoint inhibitors, chimeric antigen receptor T-cell therapy, and adoptive immune cellular therapies has generated excitement and new hopes for patients and investigators. However, clinically efficacious responses to cancer immunotherapy occur only in a minority of patients. One reason is the tumor microenvironment (TME), which potently inhibits the generation and delivery of optimal antitumor immune responses. As our understanding of TME continues to grow, strategies are being developed to change the TME toward one that augments the emergence of strong antitumor immunity. These strategies include eliminating tumor bulk to provoke the release of tumor antigens, using adjuvants to enhance antigen-presenting cell function, and employ agents that enhance immune cell effector activity. This article reviews the development of ß-glucan and ß-glucan-based nanoparticles as immune modulators of TME, as well as their potential benefit and future therapeutic applications. Cell-wall ß-glucans from natural sources including plant, fungi, and bacteria are molecules that adopt pathogen-associated molecular pattern (PAMP) known to target specific receptors on immune cell subsets. Emerging data suggest that the TME can be actively manipulated by ß-glucans and their related nanoparticles. In this review, we discuss the mechanisms of conditioning TME using ß-glucan and ß-glucan-based nanoparticles, and how this strategy enables future design of optimal combination cancer immunotherapies.

Identification of breast cancer in patients with pathologic nipple discharge: does ductoscopy predict malignancy?

OBJECTIVE: The purpose of the current study was to review characteristics of patients with nipple discharge who underwent ductoscopy-assisted excisional biopsy who had a final diagnosis of carcinoma. METHODS: A retrospective review was performed of patients presenting with pathologic nipple discharge (PND) who underwent ductoscopy-assisted excisional biopsy and had a final diagnosis of carcinoma. RESULTS: A total of 14 (7%) of 188 patients who underwent ductoscopy-assisted excision had a final pathology of ductal carcinoma-in-situ (DCIS) (12/14, 86%) or invasive breast cancer with DCIS (2/14, 14%). Duct wall irregularities or intraluminal growths were visualized during ductoscopy in 8 of the 14 (57%) breast cancer patients. There were no visual abnormalities noted during ductoscopy that accurately predicted a final diagnosis of malignancy. CONCLUSIONS: Although occult malignancies can be identified in patients undergoing ductoscopy-assisted biopsy for PND, no clear morphologic changes visualized during ductoscopy definitively indicated the presence of malignancy.

Surgery for Localized Pancreatic Cancer: The Trend Is Not Improving.

OBJECTIVES: The aim of this study was to examine the trend in the use of surgery for localized pancreatic adenocarcinoma for the past 2 decades using the Surveillance, Epidemiology, and End Results database. METHODS: We identified a cohort of patients who received a diagnosis of localized pancreatic adenocarcinoma between 1988 and 2010 in the United States. Univariate and multivariate methods were used to determine factors associated with not receiving surgery. Cox proportional hazards regression modeling was used to determine factors associated with survival. RESULTS: Of 6742 patients with a diagnosis of localized pancreatic adenocarcinoma, 1715 patients (25.4%) underwent surgery. There was no significant change in use of surgery over time. Patients were less likely to undergo surgery if they were older than 50 years, black, unmarried, and located outside the East and had pancreatic head or body lesions, higher tumor grades, or tumor size greater than 2 cm (P < 0.0001). Receiving surgery had the most significant impact on the hazard of disease-specific death (hazards ratio, 1.41; 95% confidence interval, 1.29-1.53; P < 0.0001). CONCLUSIONS: In contrast to recent studies that suggest an increasing use of surgery, the present study demonstrates that there has been no change in the rate of use of surgery in patients with localized pancreatic disease.

The Fourth International Symposium on the Intraductal Approach to Breast Cancer, Santa Barbara, California, 10-13 March 2005.

Intraductal approaches encompass procedures and technologies that are designed to access and interrogate the ductal-alveolar systems of the human breast, and include nipple aspiration, ductal lavage, random periareolar fine needle aspiration, and ductoscopy. These approaches are being used to collect and analyze fluids and cells to develop methods for breast cancer detection and risk assessment; to introduce imaging technologies to explore the mammary tree for abnormalities; to administer therapeutic and/or preventive agents directly to the breast tissue; and to explore the biology of the normal mammary gland. The latest research findings in these areas, presented at The 4th International Symposium on the Intraductal Approach to Breast Cancer in 2005, are summarized in this report.

Non-viral nanoparticle delivers small interfering RNA to macrophages in vitro and in vivo.

Macrophages are increasingly being viewed as therapeutic target for various cancers and many inflammatory diseases. Sequence specific gene reduction by siRNA represents an attractive approach to modulate macrophage function. However, delivery of the therapeutic siRNA into macrophages by non-viral nanoparticles has been a major technical challenge. In this study, we developed a glucan-based siRNA carrier system (BG34-10-Re-I) and demonstrated that the BG34-10-Re-I can effectively assemble siRNA into uniformly distributed nanoparticles of the novel core-shell structure. The BG34-10-Re-I/siRNA nanoparticles effectively reduced gene expression of macrophage migration inhibitory factor (MIF) in primary macrophages at both protein and mRNA level. The nanoparticles also mediated a sustained reduction of MIF within primary macrophages. Moreover, systemic injection of the nanoparticles into the Balb/c mice bearing 4T1 mammary tumors resulted in the MIF reduction in tumor-associated macrophages. Mechanistic studies demonstrated that the glucan-shell and the siRNA-core structure contribute to the effective delivery of MIF siRNA to macrophages both in vitro and in vivo. This study represents the first development of the primary macrophage MIF gene targeted non-viral nanoparticle system for both in vitro and in vivo applications.

T Cells Derived From Human Melanoma Draining Lymph Nodes Mediate Melanoma-specific Antitumor Responses In Vitro and In Vivo in Human Melanoma Xenograft Model.

It has been established in murine models that lymph nodes draining a progressively growing tumor contain antigen-specific T cells capable of mediating protective immune responses upon adoptive transfer. However, naturally occurring human tumor-draining lymph nodes (TDLNs) have yet to be fully investigated. In this study, we analyzed TDLNs from patients with stage III melanoma who were undergoing routine lymph node dissection. Following short-term (14 d) culture activation with anti-CD3/anti-CD28 microbeads and expansion in low concentrations of IL-2, the melanoma-draining lymph node (MDLN) cells were ∼ 60% CD4-activated and ∼ 40% CD8-activated T cells. The activated MDLN cells demonstrated reactivity in response to overlapping peptides spanning the sequence of 4 different known melanoma antigens MAGEA1, Melan-A/MART-1, NY-ESO-1, and Prame/OIP4, suggesting the presence of melanoma-specific T cells. Coculture of activated MDLN T cells with cancer cells in vitro resulted in preferential apoptosis of human cancer cell lines that were cocultured with T cells with high degree of MHC matching. Adoptive transfer of MDLN T cells with high degree of MHC matching to A375 to mice-bearing human A375 melanoma xenografts resulted in dose-dependent improvement in survival. Although prior human studies have demonstrated the immune responses within melanoma vaccine-draining lymph nodes, this study presents evidence for the first time that naturally occurring human MDLN samples contain melanoma-experienced CD4 and CD8 T cells that can be readily cultured and expanded to mediate protective immune responses both in vitro and in vivo in a human melanoma xenograft model.

Adoptive immunotherapy of cancer with polyclonal, 108-fold hyperexpanded, CD4+ and CD8+ T cells.

T cell-mediated cancer immunotherapy is dose dependent and optimally requires participation of antigen-specific CD4+ and CD8+ T cells. Here, we isolated tumor-sensitized T cells and activated them in vitro using conditions that led to greater than 108-fold numerical hyperexpansion of either the CD4+ or CD8+ subset while retaining their capacity for in vivo therapeutic efficacy. Murine tumor-draining lymph node (TDLN) cells were segregated to purify the CD62Llow subset, or the CD4+ subset thereof. Cells were then propagated through multiple cycles of anti-CD3 activation with IL-2 + IL-7 for the CD8+ subset, or IL-7 + IL-23 for the CD4+ subset. A broad repertoire of TCR Vbeta families was maintained throughout hyperexpansion, which was similar to the starting population. Adoptive transfer of hyper-expanded CD8+ T cells eliminated established pulmonary metastases, in an immunologically specific fashion without the requirement for adjunct IL-2. Hyper-expanded CD4+ T cells cured established tumors in intracranial or subcutaneous sites that were not susceptible to CD8+ T cells alone. Because accessibility and antigen presentation within metastases varies according to anatomic site, maintenance of a broad repertoire of both CD4+ and CD8+ T effector cells will augment the overall systemic efficacy of adoptive immunotherapy.

CDR3 spectratyping identifies clonal expansion within T-cell subpopulations that demonstrate therapeutic antitumor activity.

BACKGROUND: Antigen-specific T cells undergoing clonal expansion share common rearrangements of the variable complementary determining region 3 (CDR3) of the T-cell receptor (TCR), which can be identified using polymerase chain reaction-based V beta (VB) spectratyping. The purpose of this study was to determine whether CDR3 spectratyping identifies clonal expansion within tumor-draining lymph node (TDLN) subpopulations with antitumor therapeutic activity. METHODS: Recently sensitized T cells from 4T1 murine mammary carcinoma TDLN were fractionated based on CD62L (L-selectin) surface expression before RNA isolation and culture. L-selectinlow and L-selectinhigh TDLN were analyzed for T-cell receptor usage by immunophenotyping and CDR3 spectratyping, and then culture activated with anti-CD3/IL-2 to assess therapeutic efficacy after adoptive transfer. RESULTS: Adoptive transfer experiments confirmed that mice treated with culture-activated L-selectinlow TDLN cells exhibited delayed subcutaneous tumor growth and prolonged survival as compared to control or L-selectinhigh-treated mice (P < .01). CDR3 spectratyping demonstrated oligoclonal skewing of the CDR3 regions within several VB families including VB3, VB5.2, and VB17 in L-selectinlow but not in L-selectinhigh TDLN. Although fluorescence-activated cell sorter analysis demonstrated the highest percentage of cells expressing VB13 usage in both populations, CDR3 spectratyping did not identify the presence of clonal expansion. CONCLUSIONS: These data suggest that CDR3 spectratyping may be useful in identifying T cells undergoing clonal expansion that demonstrate antitumor therapeutic activity.

Directed duct excision by using mammary ductoscopy in patients with pathologic nipple discharge.

BACKGROUND: Duct excision for pathologic nipple discharge (PND) often requires "blind" surgical resection. Intraoperative mammary ductoscopy can allow for direct visualization of intraductal abnormalities during surgical resection. METHODS: We reviewed our experience with 119 patients with PND undergoing ductoscopy-directed duct excision. The variables that could interfere with a successful procedure were analyzed. RESULTS: Cannulation of the discharging duct was successful in 105 of 119 (88%) of patients, and ductoscopy-directed duct excision could be performed in 104 of 119 (87%). A preoperative ductogram was obtained in 70 patients and was positive in 53 of 70 (76%). In this same group, ductoscopy was positive in 63 of 70 (90%). The pathologic diagnoses were as follows: cancer (5), papilloma (84), and hyperplasia (16), for an abnormal pathology yield of 88%. Hyperplasia and cancer were significant predictors of unsuccessful cannulation. In 22 patients, ductoscopy visualized multiple lesions or abnormalities beyond 4 cm. CONCLUSIONS: Mammary ductoscopy for PND is a safe, effective procedure that offers advantages of a high lesion localization rate and intraoperative guidance, therefore negating the need for a preoperative ductogram. Lesions deep within the ductal system can be identified and removed, which would likely have been missed by blind duct excision.

Does core needle breast biopsy accurately reflect breast pathology?

BACKGROUND: Core needle breast biopsy (CB) has replaced excisional biopsy as the initial diagnostic biopsy procedure for many suspicious breast lesions; however, CB remains a sampling procedure. The purpose of this study was to determine the degree of agreement between histology obtained at CB and that obtained at a subsequent excisional procedure (EP). We hypothesized a high degree of agreement. METHODS: Data were collected prospectively for 3035 CBs performed by breast radiologists using either ultrasound or stereotactic guidance between January 1995 and July 2002, 1410 (46%) of which had a subsequent EP within 1 year. Histologic categories were defined as invasive breast cancer, duct carcinoma in-situ, atypia/lobular carcinoma in-situ, and benign. The principal histology (PH) from CB and EP was identified and compared. RESULTS: Overall, there was moderate agreement (kappa=0.669) between CB and EP histology. Complete agreement occurred in 1168 (83%) procedures. For the remaining 242, the PH was identified only at CB for 78 (5%) procedures, and only after EP for 164 (12%) procedures. CONCLUSIONS: Although the majority (83%) of CB and EP demonstrated exact histologic agreement, CB was diagnostic for 1246 (88%) procedures.

Nipple-sparing mastectomy: technique and results of 54 procedures.

HYPOTHESIS: The rationale for removal of the nipple-areolar complex (NAC) during total mastectomy centers on long-standing concerns about possible neoplastic involvement of the NAC and its postoperative viability. Nipple-sparing mastectomy (NSM) combines a skin-sparing mastectomy with preservation of the NAC, intraoperative pathological assessment of the nipple tissue core, and immediate reconstruction, thereby permitting better cosmesis for patients undergoing total mastectomy. Neoplastic involvement of the NAC can be predicted before surgery and assessed during the operation, and sustained postoperative viability of the NAC is likely with appropriate surgical technique. RESULTS: Fifty-four NSMs with immediate reconstruction were attempted among 44 patients. Six NAC core specimens revealed neoplastic involvement on frozen section analysis, resulting in conversion to total mastectomies. Forty-five of the 48 completed NSMs maintained postoperative viability of the NAC; 3 NACs had partial loss. CONCLUSION: Nipple-sparing mastectomy is a reasonable option for carefully screened patients.

Targeted therapies for the treatment of cancer.

BACKGROUND: In contrast to conventional cytotoxic chemotherapy and radiation therapy, a new method of targeted cancer therapeutics is being directed towards molecular pathways that underlie the malignant phenotype. These therapies target specific tumor cell receptors or signaling events that are critical to tumor progression while reducing toxicity to normal cells. DATA SOURCES: The purpose of this review is to highlight several examples of novel targeted therapeutics that are currently approved by the FDA for treatment of patients with cancer. Rituxan is a humanized monoclonal antibody that binds to the CD20 antigen present on B cell lymphomas and is currently approved for the treatment of patients with relapsed or refractory low-grade CD20 positive follicular lymphoma. The humanized anti-HER-2/neu herceptin is approved for use in patients with metastatic breast cancer that demonstrates overexpression of HER-2/neu. Finally, Gleevec is a tyrosine kinase inhibitor that inhibits abl-specific phosphorylation and is approved for use in select patients with chronic myelogenous leukemia that is refractory to interferon therapy. CONCLUSIONS: The lessons learned from the use of these therapeutics will add to the growing knowledge of mechanistic approaches to the treatment of patients with cancer based upon targeted therapies, and herald a bright future that will improve the lives of patients with cancer.

Prospective study of intraoperative mammary ductoscopy in patients undergoing partial mastectomy for breast cancer.

BACKGROUND: The purpose of this study was to critically evaluate the added benefit of intraoperative mammary ductoscopy on margin assessment and identification of occult intraductal pathology in patients undergoing therapeutic partial mastectomy for in-situ and invasive breast carcinoma. METHODS: Eligible patients underwent intraoperative mammary ductoscopy before partial mastectomy. In patients where an intraductal abnormality was identified and the mammary ductoscope was not within the partial mastectomy cavity, an additional ductoscopy-directed margin of tissue was analyzed. RESULTS: Nineteen of 30 (63%) patients yielded nipple aspirate fluid and were able to undergo mammary ductoscopy; an intraductal abnormality was identified in 15 of 19 (79%) patients. Only 1 patient had an occult infiltrating carcinoma, which was outside of the resection cavity and identified by ductoscopy. CONCLUSIONS: Although mammary ductoscopy can identify intraductal abnormalities during partial mastectomy in a significant number of patients, many of these findings may be either benign or within the standard field of resection, thus adding no benefit to the patient.

Axillary lymph node metastases in patients with a final diagnosis of ductal carcinoma in situ.

BACKGROUND: Recent studies report the incidence of axillary metastases in patients with ductal carcinoma in-situ (DCIS) approaches 13%. The purpose of this study was to define the incidence of axillary micrometastases in patients with pure DCIS before and after the introduction of sentinel lymph node biopsy. METHODS: Patients with a final diagnosis of DCIS form the basis of this study. Data were entered prospectively into an Institutional Review Board approved Oracle database from January 1997 through July 2002. RESULTS: One hundred and thirty-four patients had lymph nodes evaluated. Ninety-eight percent of patients had no evidence of metastatic disease and 2% were found to have micrometastases. This was consistent in those who had level I or II lymph node sampling or both and those who had lymphatic mapping and a sentinel lymph node biopsy procedure. CONCLUSIONS: These data do not support axillary lymph node removal of any type in patients with pure DCIS.

Low-risk palpable breast masses removed using a vacuum-assisted hand-held device.

BACKGROUND: This study evaluates the safety, efficacy, and patient acceptance of a vacuum-assisted, hand-held biopsy device (Mammatome) in the percutaneous removal of breast masses using ultrasound guidance. METHODS: A multicenter, nonrandomized study evaluated 216 women with low-risk palpable lesions. Lesions 1.5 to 3.0 cm in size were removed using an 8-gauge probe. Those lesions <1.5 cm were removed with the 11-gauge probe. Follow-up evaluation was performed at 10 days and 6 months after biopsy. RESULTS: A total of 127 patients had biopsies using the 8-gauge probe, and 89 patients had biopsies using the 11-gauge probe. At 6-month follow-up, 98% of the lesions remained nonpalpable, 73% with no ultrasonographically visible evidence of the original lesion. Most complications were mild and anticipated. Most patients (98%) were satisfied with incision appearance, and 92% of patients would recommend the procedure to others. CONCLUSIONS: Percutaneous removal of palpable benign breast masses using the Mammotome system is feasible and safe, and yields high patient satisfaction. The results at 6 months after biopsy demonstrated the effectiveness of benign lesion removal, with correlative clinical data demonstrating lack of palpability and no need for additional procedures. Continuing evaluation of long-term efficacy is ongoing.

Does ultrasound core breast biopsy predict histologic finding on excisional biopsy?

BACKGROUND: The purpose of this study was to determine whether ultrasound-guided core breast biopsy accurately predicts the histologic finding of a subsequent excisional procedure. METHODS: Data were collected prospectively from 1997 to 2001 for 832 ultrasound-guided core breast biopsies (USB) that were followed by excisional breast procedure (EP) within 1 year at our institution. The principal histologic finding obtained at USB and EP was identified for each procedure and the degree of agreement was assessed. RESULTS: The USB histology predicted EP histology in 90% (n = 746) of the procedures. The USB histology was more significant than EP histology in 3% (n = 22) of procedures; USB histology underdetermined EP histology in 7% (n = 64) of procedures. Overall, our results indicate moderate agreement between the principal histology identified at USB relative to that identified at EP. CONCLUSIONS: Ultrasound-guided core breast biopsy is an effective diagnostic method, but sampling limitations do exist.

Percutaneous removal of benign breast masses using a vacuum-assisted hand-held device with ultrasound guidance.

BACKGROUND: This study evaluates the safety, efficacy, and patient acceptance of a vacuum-assisted, hand-held biopsy device (Mammotome) in percutaneous removal of breast masses using ultrasound guidance. METHODS: A multicenter, nonrandomized study evaluated 124 women with low-risk palpable lesions. Lesions 1.5 cm but

Feasibility and Technical Considerations of Mammary Ductoscopy in Human Mastectomy Specimens.

Recent advances in endoscopic technology have made visualization of human mammary ducts possible. The purpose of this study was to assess the feasibility and technical factors influencing the ability to successfully visualize the epithelium of the human mammary ductal system. Lacrimal duct probes were used to dilate nipple orifices to 1.2 mm on 42 mastectomy specimens. The Depth of Field Imaging Micro-Minimally Invasive (DOFI(R) MMI) system consisting of a 1.2 mm rigid ductoscope with a 350 &mgr;m working channel was introduced into mammary ducts under air insufflation or saline irrigation. At least one major duct could be dilated and cannulated in all 42 specimens. Visualization of the proximal duct was accomplished in 34 of 42 (81%) specimens, whereas more extensive navigation through the distal subsegmental ducts was achieved in 22 of 42 (52%) specimens. Ductoscopy into the terminal ducts was accomplished in all patients with a previous history of nipple discharge or discharge at the time of the procedure (10 of 10). In three patients with no history of nipple discharge prior to ductoscopy, incidental papillomas were discovered and confirmed by the pathologist. In conclusion, mammary ductoscopy is technically feasible and may have an application as an additional diagnostic modality for patients with pathologic nipple discharge.

CD40 monoclonal antibody activation of antigen-presenting cells improves therapeutic efficacy of tumor-specific T cells.

OBJECTIVE: The goal of this study was to determine whether CD40 ligation of antigen presenting cells (APCs) enhances the anti-tumor effector function of tumor draining lymph node (TDLN) T lymphocytes in an adoptive immunotherapy model. STUDY DESIGN: MCA 205 TDLNs were culture activated both in the presence and absence of a stimulatory anti-CD40 monoclonal antibody (mAb) and effector cell phenotype, cytokine secretion in vitro and therapeutic efficacy in vivo were compared. RESULTS: Anti-CD40 mAb induced upregulation of APC cell surface activation markers that promoted generation of T cells that demonstrated an increase in tumor-specific IFN-gamma secretion and a statistically significant reduction in the number of pulmonary tumors (p< 0.01) after adoptive transfer. CONCLUSION: CD40 ligation of APCs in vitro results in the generation of T cells with enhanced effector function against established pulmonary tumors in vivo. SIGNIFICANCE: These findings have direct implications in the development of effective T cell-based immunotherapy of malignant conditions in human beings.