Intravital and whole-organ imaging reveals capture of melanoma-derived antigen by lymph node subcapsular macrophages leading to widespread deposition on follicular dendritic cells.

Aberrant antigens expressed by tumor cells, such as in melanoma, are often associated with humoral immune responses, which may in turn influence tumor progression. Despite recent data showing the central role of adaptive immune responses on cancer spread or control, it remains poorly understood where and how tumor-derived antigen (TDA) induces a humoral immune response in tumor-bearing hosts. Based on our observation of TDA accumulation in B cell areas of lymph nodes (LNs) from melanoma patients, we developed a pre-metastatic B16.F10 melanoma model expressing a fluorescent fusion protein, tandem dimer tomato, as a surrogate TDA. Using intravital two-photon microscopy (2PM) and whole-mount 3D LN imaging of tumor-draining LNs in immunocompetent mice, we report an unexpectedly widespread accumulation of TDA on follicular dendritic cells (FDCs), which were dynamically scanned by circulating B cells. Furthermore, 2PM imaging identified macrophages located in the subcapsular sinus of tumor-draining LNs to capture subcellular TDA-containing particles arriving in afferent lymph. As a consequence, depletion of macrophages or genetic ablation of B cells and FDCs resulted in dramatically reduced TDA capture in tumor-draining LNs. In sum, we identified a major pathway for the induction of humoral responses in a melanoma model, which may be exploitable to manipulate anti-TDA antibody production during cancer immunotherapy.

Laparoscopic Indocyanine Green Sentinel Lymph Node Mapping in Pregnant Cervical Cancer Patients.

We present cases of 2 pregnant patients with early-stage cervical cancer who have undergone indocyanine green (ICG) sentinel lymph node (SLN) mapping followed by laparoscopic SLN biopsy, pelvic lymphadenectomy, and cervical conization. Eight milliliters of ICG were injected in the 4 quadrants of the cervix after having obtained an adequate pneumoperitoneum and having inspected the abdominal cavity. SLNs were identified in both hemipelvises in both patients. In the final pathologic analysis, both SLNs and non-SLNs were negative for metastatic disease. No adverse events from ICG injection were recorded. ICG SLN mapping seems to be feasible in pregnant cervical cancer patients.

A Comparison of Radiocolloid and Indocyanine Green Fluorescence Imaging, Sentinel Lymph Node Mapping in Patients with Cervical Cancer Undergoing Laparoscopic Surgery.

BACKGROUND AND PURPOSE (99)TC combined with blue-dye mapping is considered the best sentinel lymph node (SLN) mapping technique in cervical cancer. Indocyanine green (ICG) with near infrared fluorescence imaging has been introduced as a new methodology for SLN mapping. The aim of this study was to compare these two techniques in the laparoscopic treatment of cervical cancer. METHODS Medical records of patients undergoing laparoscopic SLN mapping for cervical cancer with either (99)Tc and patent blue dye (Group 1) or ICG (Group 2) from April 2008 until August 2012 were reviewed. Sensitivity, specificity, and overall and bilateral detection rates were calculated and compared. RESULTS Fifty-eight patients were included in the study-36 patients in Group 1 and 22 patients in Group 2. Median tumor diameter was 25 and 29 mm, and mean SLN count was 2.1 and 3.7, for Groups 1 and 2, respectively. Mean non-SLN (NSLN) count was 39 for both groups. SLNs were ninefold more likely to be affected by metastatic disease compared with NSLNs (p < 0.005). Sensitivity and specificity were both 100 %. Overall detection rates were 83 and 95.5 % (p = nonsignificant), and bilateral detection rates were 61 and 95.5 % (p < 0.005), for Groups 1 and 2, respectively. In 75 % of cases, SLNs were located along the external or internal iliac nodal basins. CONCLUSIONS ICG SLN mapping in cervical cancer provides high overall and bilateral detection rates that compare favorably with the current standard of care.

Regulation of bovine IL-12R beta 2 subunit mRNA expression in bovine lymph node cells

The beta 2 subunit of the interleukin (IL)-12 receptor (IL-12R beta 2) has been shown to play an essential role in differentiation of T helper 1 (Th1) cells in the murine and human system, and antibodies raised against IL-12R beta 2 recognized this molecule on human Th1 but not Th2 cells. However, while the cytokines secreted by clones of murine cells allowed the definition of distinct T helper cell subsets, bovine clones with polarized Th1 and Th2 cytokine profiles were rarely found. This raised important questions about the regulation of immune responses in cattle. We therefore cloned bovine IL-12R beta2 (boIL-12R beta 2) DNA complementary to RNA (cDNA) from the start codon to the 3' end of the mRNA. Comparison of boIL-12R beta 2 cDNA with human and murine IL-12R beta 2 cDNA sequences revealed homologies of 85 and 78%, respectively. The deduced protein sequence showed the hallmark motifs of the cytokine receptor superfamily including the four conserved cysteine residues, the WSXWS motif and fibronectin domains in the extracellular part as well as a STAT4 binding site in the intracellular part of the molecule. Using real-time reverse transcription-polymerase chain reaction, upregulation of mRNA expression of this molecule could be demonstrated in cultured bovine lymph node cells stimulated with phytohemagglutinin. Furthermore, cells with upregulated boIL-12R beta 2 mRNA responded with enhanced expression of interferon gamma to treatment with interleukin 12.

The role of SRC family kinase activation in prostate cancer growth and lymph node metastasis

Aberrant expression and/or activation of Src Family of non-receptor protein tyrosine kinases (SFKs) occur frequently during progressive stages of multiple types of human malignancies, including prostate cancer. Two SFKs, Src and Lyn, are expressed and implicated in prostate cancer progression. Work in this dissertation investigated the specific roles of Src and Lyn in the prostate tumor progression, and the effects of SFK inhibition on prostate tumor growth and lymph node metastasis in pre-clinical mouse models. ^ Firstly, using a pharmacological inhibitor of SFKs in clinical trials, dasatinib, I demonstrated that SFK inhibition affects both cellular migration and proliferation in vitro. Systemic administration of dasatinib reduced primary tumor growth, as well as development of lymph node metastases, in both androgen-sensitive and -resistant orthotopic prostate cancer mouse models. Immunohistochemical analysis of the primary tumors revealed that dasatinib treatment decreased SFK phosphorylation but not expression, resulting in decreased cellular proliferation and increased apoptosis. For this analysis of immunohistochemical stained tissues, I developed a novel method of quantifying immunohistochemical stain intensity that greatly reduced the inherent bias in analyzing staining intensity. ^ To determine if Src and Lyn played overlapping or distinct roles in prostate cancer tumor growth and progression, Src expression alone was inhibited by small-interfering RNA. The resulting stable cell lines were decreased in migration, but not substantially affected in proliferation rates. In contrast, an analogous strategy targeting Lyn led to stable cell lines in which proliferation rates were significantly reduced. ^ Lastly, I tested the efficacy of a novel SFK inhibitor (KX2-391) targeting peptide substrate-binding domain, on prostate cancer growth and lymph node metastasis in vivo. I demonstrated that KX2-391 has similar effects as dasatinib, an ATP-competitive small molecular inhibitor, on both the primary tumor growth and development of lymph node metastasis in vivo, work that contributed to the first-in-man Phase I clinical trial of KX2-391. ^ In summary, studies in this dissertation provide the first demonstration that Src and Lyn activities affect different cellular functions required for prostate tumor growth and metastasis, and SFK inhibitors effectively reduce primary tumor growth and lymph node metastasis. Therefore, I conclude that SFKs are promising therapeutic targets for treatment of human prostate cancer. ^

Disrupted splenic architecture, but normal lymph node development in mice expressing a soluble lymphotoxin-β receptor–IgG1 fusion protein

Early in ontogeny, the secondary lymphoid organs become populated with numerous cells of mesodermal origin which forms both the lymphoid and stromal elements. The critical receptor/ligand interactions necessary for lymphoid organogenesis to occur are for the most part unknown. Although lymphotoxin-α (LTα) has been shown to be required for normal lymph node, Peyer’s patch, and splenic development, it is unclear if soluble LTα3, and/or cell-bound lymphotoxin-αβ (LTαβ) mediate these developmental events. Here we report that blocking LTαβ/lymphotoxin-β receptor (LTβR) interaction in vivo by generating mice which express a soluble LTβR–Fc fusion protein driven by the human cytomegalovirus promoter results in an array of anatomic abnormalities affecting both the spleen and Peyer’s patches, but not the lymph nodes. These results demonstrate that surface LTαβ ligand plays a critical role in normal lymphoid organ development.

Heparanase and COX-2 expression as predictors of lymph node metastasis in large, high-grade breast tumors

Background/Aim: Heparanase (HPA) contributes to breast cancer metastasis by facilitating the breakdown of the basement membrane and extracellular matrix. High expression of HPA is thought to be associated with increased nodal involvement and poor survival in patients with breast cancer. Overexpression of cyclooxygenase-2 (COX-2) in breast cancer is associated with indicators of poor prognosis such as lymph node metastasis, poor differentiation, and large tumor size. The underlying mechanism by which HPA and COX-2 overexpression increases the metastatic potential of breast cancer is not fully-understood. To enhance our understanding over these mechanisms, we aimed to investigate the relationship between the size of the tumor and HPA expression, tumor grade as well as lymph node status in patients with breast cancer. Materials and Methods: Immunohistochemical analysis of HPA and COX-2 expression was performed on 246 breast tumor samples. The expression of HPA was correlated with COX-2 expression, tumor grade, lymph node status, oestrogen receptor status. Results: The overexpression of HPA and COX-2 was associated with increased likelihood of lymph node positivity in large, high-grade tumors. High-grade tumors with size greater than 20 mm, that overexpressed HPA, were 4-times more likely to be associated with lymph node involvement (OR 4.71, CI 1.21-18.25). Whereas, tumors greater than 20 mm in size were 5-times more likely to metastasize to the regional lymph nodes, if associated with overexpression of COX-2 (OR 5.5, CI 1.2-24.8). Conclusion: Expression of HPA appears to be a key mechanism by which large, highgrade breast tumors metastasize to regional lymph nodes, while COX-2 overexpression may be an independent predictor of lymph node positivity.

Prognostic value of lymph node ratio and extramural vascular invasion on survival for patients undergoing curative colon cancer resection

Peer reviewed

Surgical treatment of papillary thyroid carcinoma without lymph nodal involvement

Background. In the treatment of differentiated thyroid cancer (DTC), in absence of enlarged lymph nodes, the role of routine central lymph node dissection (RCLD) remains controversial. The aim of this study is to analyze data resulting from total thyroidectomy (TT) not combined with RCLD in the treatment of DTC. Methods. We retrospectively evaluated the clinical records of 80 patients treated between January 1996 and December 2003 with TT without RCLND, in absence of suspected enlarged lymph nodes at preoperative ultrasonography and intraoperatively during neck exploration. In this series, 75 patients (93.7%) underwent radioiodine (RAI) ablation, followed by Thyroid Stimulating Hormone (TSH) suppression therapy. In case of locoregional lymph nodal recurrence, a central (VI) and ipsilateral (III-IV) selective lymph node dissection was performed. Results. Incidence of permanent hypoparathyroidism (iPTH < 10 pg/ml) and unilateral temporary vocal fold paralysis were respectively 2.55% and 2.55%. Locoregional recurrence, with positive cervical lymph nodes, after a 10.3 ± 4.7 years mean follow-up was observed in 3 patients (3.75%). They were submitted to a central (VI) and ipsilateral (III-IV) selective neck dissection without significant complications. Conclusions. In our series, TT not combined with RCLD was associated to a low locoregional recurrence rate, even if the lack of a control group treated with RCLD does not allow any generalized assumption. RCLD may be indicated in high risk patients, in whom lymph nodal recurrence is more frequent. More prospective randomized studies are needed to better define the role of RCLD and postoperative radioiodine ablation.

Omission of axillary clearance in patients with early breast cancer and positive sentinel node biopsy treated with conservative surgery: application of the ACOSOG Z011 trial in clinical practice

Background: Axillary lymph node dissection (ALND) in presence of sentinel lymph node (SLN) metastases has been the standard in breast cancer (BC) patients for many years. Today, after the publication of the ACOSOG Z0011 trial, ALND is a procedure restricted to a dwindling group of patients with a clearly metastatic axilla. Material and methods: This was a prospective observational trial involving two Italian Breast Units: Policlinico di Sant’Orsola and San Raffaele hospital. Objective was to evaluate that the omission of ALND in patients with cT1-2 cN0 BC undergoing breast conserving surgery (BCS) and histological finding of metastases in 1 or 2 SLN is not associated with a worse prognostic outcome. Primary endpoint was overall survival (OS). Secondary endpoints were disease free survival (DFS) and locoregional recurrence. All BC patients treated between the 1st of November 2020 and 31st of July 2023 with cT1-2 cN0 BC, preoperative negative axillary ultrasound and 1 or 2 metastatic SLN treated with sentinel node biopsy (SLNB) alone entered the study. Results: 795 cT1-2 cN0 BC patients underwent BCS and SLNB. Ninety patients were included. Median age was 60 (52-68) years. Seventy-five patients (83%) had T1 tumor and 15 (17%) T2. Median tumor size was 16 mm (11-19). The median SLN removed was 2 (1-3). Eighty-one patients had 1 positive SLN (90%), while 9 had 2 SLN metastasis (10%). 39 (43%) micrometastases were identified and 51 macrometastasis (57%). All patients underwent radiotherapy. Seventeen (19%) performed adjuvant chemotherapy. Two received immunotherapy with trastuzumab and pertuzumab. Endocrine therapy was given to 84 (93%). At a median follow-up of 19 months (IQR 13-23) OS and DFS were 100%. No loco-regional recurrence was seen. Conclusion: The preliminary results of our study confirm that omitting ALND in patients meeting Z011 criteria is oncologically safe and should be the standard of care.

Skip metaskses in medullary thyroid carcinoma: A single-center experience

Purpose. Total thyroidectomy (TT) with level VI and VII central neck dissection is the initial treatment for medullary thyroid carcinoma (MTC) without identifiable neck metastasis. Level II to V lateral neck dissection is performed if neck metastasis is present or suspected. We conducted this study to identify the frequency and clinical determinants of skip neck metastasis in MTC. Methods. We reviewed the medical records of 32 patients who underwent TT and bilateral neck dissection for MTC. The clinical features were correlated with pN status in the central versus lateral compartments of the neck. Results. Neck lymph node metastasis (pN+) was found in 20 patients (62.5%) and skip metastases were found in 7 (35%) patients. The sensitivity of the pN status of the central compartment of the neck to predict the pN status of the lateral compartment of the neck was 53.8% and specificity was 63.2%. We found pN+ in 90% of the patients with lymph nodes > 15mm in diameter versus 50% in those with lymph nodes < 1.5mm in diameter. Conclusions. There is skip metastasis in MTC. It is unsafe to use the lymph node status of the central compartment of the neck to define the pN status of the lateral neck. A lymph node greater than 15 mm in diameter is related to pN status.

Impact of Surgical Staging in Locally Advanced Cervical Cancer and Subsequent Chemotherapy

Background: Surgical staging (SS) is the gold standard for determination of the true extent of a patient`s disease and is an important prognostic factor in cervical cancer. We investigated whether lymph node dissection (LND) prior to chemotherapy (CT) followed by radical surgery (RS) could modified overall (OS) and disease-free survival (DFS). Methods: We performed a cohort analysis of 98 patients with cervical carcinoma. The experimental group consisted of 36 patients who underwent SS followed by neoadjuvant chemotherapy, and then by RS (objective response) or chemo-radiation therapy (with or without subsequent surgery when not possible). The control group consisted of 62 similarly treated patients without pretreatment SS. The value of this procedure as a diagnostic tool in defining the extent of disease was evaluated. Furthermore, LND/CT-associated treatment complications and the impacts on OS and DFS were also evaluated. Results: Fourteen (38.9%) patients had pelvic LN metastases and three (8.3%) patients had pelvic and para-aortic LN metastases. The 39-month OS and DFS rates for the current study were 90.6% for the staged group and 52% for non-staged treatment (P < 0.001). Conclusion: SS in cervical cancer is a feasible and safe pretreatment procedure, and when associated with CT, it improves OS and DFS. J. Surg. Oncol. 2009;100:505-510. (C) 2009 Wiley-Liss, Inc.

Breast conserving surgery after neoadjuvant therapy for large primary breast cancer

Aim: The aim of this study was to evaluate the safety of breast conserving surgery ill patients with breast tumours satisfactorily downstaged after neoadjuvant therapy. Methods: A retrospective cohort study was undertaken to analyze the loco-regional recurrence (LRR) after breast conserving surgery. We enrolled 88 patients with breast cancer subjected to neoadjuvant therapy (NAT group) who achieved an objective response due to neoadjuvant treatment and compared them with 191 patients with early breast cancer (EBC group) who were submitted to primary conserving surgery. Lumpectomy or quadrantectomy with axillary lymph node dissection was performed in all patients who received adjuvant radiotherapy. Systemic adjuvant therapy was offered to all patients. The mean periods of observation were 61.3 months in the NAT group and 67.5 months in the EBC group. Results: The mean age was 53 years in the NAT group and 56 years in the EBC group (p = 0.04). There was no histological type and histological grade difference between groups. In the NAT group, the mean diameter of residual tumour was lower and the mean volume of breast tissue resection was higher than in the EBC group (p = 0.01 and p = 0.002, respectively). The ipsilateral recurrence rate was 7.9% in the NAT group and 7.8% in the EBC group (p = 0.9). The most important predictive factor of recurrence in the NAT group was the age of patient. Conclusion: Breast conserving therapy is a safe procedure in satisfactorily downstaged breast cancer after neoadjuvant therapy. (c) 2008 Elsevier Ltd. All rights reserved.

Peritoneal cytology: impact on disease-free survival in clinical stage I endometrioid adenocarcinoma of the uterus

The prognostic significance of positive peritoneal cytology in endometrial carcinoma has led to the incorporation of peritoneal cytology into the current FIGO staging system, While cytology was shown to be prognostically relevant in patients with stage II and III disease, conflicting data exists about its significance in patients who would have been stage I but were classified as stage III solely and exclusively on the basis of positive peritoneal cytology (clinical stage I). Analysis was based on the data of 369 consecutive patients with clinical stage I endometrioid adenocarcinoma of the endometrium. Standard treatment consisted of an abdominal total hysterectomy, bilateral salpingo-oophorectomy with or without pelvic lymph node dissection. Peritoneal cytology was obtained at laparotomy by peritoneal washing of the pouch of Douglas and was considered positive if malignant cells could be detected regardless of the number of malignant cells present. Disease-free survival (DFS) was considered the primary statistical endpoint. In 13/369 (3.5%) patients, positive peritoneal cytology was found. The median follow-up was 29 months and 15 recurrences occurred. Peritoneal cytology was independent of the depth of myometrial invasion and the grade of tumour differentiation, Patients with negative washings had a DFS of 96'7e at 36 months compared with 67% for patients with positive washings (log-rank P < 0.001). The presence of positive peritoneal cytology in patients with clinically stage I endometrioid adenocarcinoma of the endometrium is considered an adverse prognostic factor. (C) 2001 Elsevier Science Ireland Ltd. All rights reserved.