Hsc70 promotes anti-tumor immunity by targeting PD-L1 for lysosomal degradation.

Immune checkpoint inhibition targeting the PD-1/PD-L1 pathway has become a powerful clinical strategy for treating cancer, but its efficacy is complicated by various resistance mechanisms. One of the reasons for the resistance is the internalization and recycling of PD-L1 itself upon antibody binding. The inhibition of lysosome-mediated degradation of PD-L1 is critical for preserving the amount of PD-L1 recycling back to the cell membrane. In this study, we find that Hsc70 promotes PD-L1 degradation through the endosome-lysosome pathway and reduces PD-L1 recycling to the cell membrane. This effect is dependent on Hsc70-PD-L1 binding which inhibits the CMTM6-PD-L1 interaction. We further identify an Hsp90α/ß inhibitor, AUY-922, which induces Hsc70 expression and PD-L1 lysosomal degradation. Either Hsc70 overexpression or AUY-922 treatment can reduce PD-L1 expression, inhibit tumor growth and promote anti-tumor immunity in female mice; AUY-922 can further enhance the anti-tumor efficacy of anti-PD-L1 and anti-CTLA4 treatment. Our study elucidates a molecular mechanism of Hsc70-mediated PD-L1 lysosomal degradation and provides a target and therapeutic strategies for tumor immunotherapy.

Predictive and prognostic biomarkers of bone metastasis in breast cancer: current status and future directions.

The most common site of metastasis in breast cancer is the bone, where the balance between osteoclast-mediated bone resorption and osteoblast-mediated bone formation is disrupted. This imbalance causes osteolytic bone metastasis in breast cancer, which leads to bone pain, pathological fractures, spinal cord compression, and other skeletal-related events (SREs). These complications reduce patients' quality of life significantly and have a profound impact on prognosis. In this review, we begin by providing a brief overview of the epidemiology of bone metastasis in breast cancer, including current diagnostic tools, treatment approaches, and existing challenges. Then, we will introduce the pathophysiology of breast cancer bone metastasis (BCBM) and the animal models involved in the study of BCBM. We then come to the focus of this paper: a discussion of several biomarkers that have the potential to provide predictive and prognostic value in the context of BCBM-some of which may be particularly compatible with more comprehensive liquid biopsies. Beyond that, we briefly explore the potential of new technologies such as single-cell sequencing and organoid models, which will improve our understanding of tumor heterogeneity and aid in the development of improved biomarkers. The emerging biomarkers discussed hold promise for future clinical application, aiding in the prevention of BCBM, improving the prognosis of patients, and guiding the implementation of personalized medicine.

Incidence and risk factors of lymph node metastasis in breast cancer patients without preoperative chemoradiotherapy and neoadjuvant therapy: analysis of SEER data.

Background: Breast cancer (BC) is the leading cause of death in the female reproductive system, often linked to lymph node involvement, indicating poor prognosis. This study investigated lymph node metastasis incidence and risk factors in M0 stage BC patients who hadn't received preoperative chemoradiotherapy or neoadjuvant therapy. We explored the influence of various factors on lymph node metastasis. Methods: We conducted a retrospective analysis using Surveillance, Epidemiology, and End Results data from BC patients diagnosed between 2010 and 2015. Binary logistic regression and propensity score matching (PSM) assessed significant factors in BC patients without preoperative treatment. We developed predictive nomograms and evaluated model performance using the concordance index, calibration curve, area under the curve, and decision curve analysis. Results: Among 256,504 eligible BC patients, 25.57% had lymph node metastasis. Multivariate logistic regression revealed associations between lymph node metastasis and younger age, African-American ethnicity, central/nipple location, lobular carcinoma, human epidermal growth factor receptor 2 (HER2)-positive status, grade III classification, and T3 stage. PSM confirmed these findings. Interactions were identified between age, race, primary site, histology, breast subtype, grade, and T stage, all influencing lymph node metastasis. Conclusions: This retrospective study identified lymph node metastasis in female BC patients with distinct clinicopathological characteristics who received no preoperative treatment. We constructed valuable nomograms, revealing that: (I) young age (<35 years), African-American race, central/nipple location, infiltrating duct carcinoma, HER2 positivity, high histological grade (grade III), and larger tumor size are risk factors for regional lymph node metastasis; (II) lymph node metastasis may not solely represent the invasive nature of triple-negative BC; (III) patients with different BC subtypes in T1c-T2 stages may benefit from individualized neoadjuvant treatment strategies.

A retrospective comparative study on the diagnostic efficacy and the complications: between CassiII rotational core biopsy and core needle biopsy.

Accurate pathologic diagnosis and molecular classification of breast mass biopsy tissue is important for determining individualized therapy for (neo)adjuvant systemic therapies for invasive breast cancer. The CassiII rotational core biopsy system is a novel biopsy technique with a guide needle and a "stick-freeze" technology. The comprehensive assessments including the concordance rates of diagnosis and biomarker status between CassiII and core needle biopsy were evaluated in this study. Estrogen receptor (ER), progesterone receptor (PgR), human epidermal growth factor receptor 2 (HER2), and Ki67 were analyzed through immunohistochemistry. In total, 655 patients with breast cancer who underwent surgery after biopsy at Sir Run Run Shaw Hospital between January 2019 to December 2021 were evaluated. The concordance rates (CRs) of malignant surgical specimens with CassiII needle biopsy was significantly high compared with core needle biopsy. Moreover, CassiII needle biopsy had about 20% improvement in sensitivity and about 5% improvement in positive predictive value compared to Core needle biopsy. The characteristics including age and tumor size were identified the risk factors for pathological inconsistencies with core needle biopsies. However, CassiII needle biopsy was associated with tumor diameter only. The CRs of ER, PgR, HER2, and Ki67 using Cassi needle were 98.08% (kappa, 0.941; p<.001), 90.77% (kappa, 0.812; p<.001), 69.62% (kappa, 0.482; p<.001), and 86.92% (kappa, 0.552; p<.001), respectively. Post-biopsy complications with CassiII needle biopsy were also collected. The complications of CassiII needle biopsy including chest stuffiness, pain and subcutaneous ecchymosis are not rare. The underlying mechanism of subcutaneous congestion or hematoma after CassiII needle biopsy might be the larger needle diameter and the effect of temperature on coagulation function. In summary, CassiII needle biopsy is age-independent and has a better accuracy than CNB for distinguishing carcinoma in situ and invasive carcinoma.

ARIH1 activates STING-mediated T-cell activation and sensitizes tumors to immune checkpoint blockade.

Despite advances in cancer treatment, immune checkpoint blockade (ICB) only achieves complete response in some patients, illustrating the need to identify resistance mechanisms. Using an ICB-insensitive tumor model, here we discover cisplatin enhances the anti-tumor effect of PD-L1 blockade and upregulates the expression of Ariadne RBR E3 ubiquitin-protein ligase 1 (ARIH1) in tumors. Arih1 overexpression promotes cytotoxic T cell infiltration, inhibits tumor growth, and potentiates PD-L1 blockade. ARIH1 mediates ubiquitination and degradation of DNA-PKcs to trigger activation of the STING pathway, which is blocked by the phospho-mimetic mutant T68E/S213D of cGAS protein. Using a high-throughput drug screen, we further identify that ACY738, less cytotoxic than cisplatin, effectively upregulates ARIH1 and activates STING signaling, sensitizing tumors to PD-L1 blockade. Our findings delineate a mechanism that tumors mediate ICB resistance through the loss of ARIH1 and ARIH1-DNA-PKcs-STING signaling and indicate that activating ARIH1 is an effective strategy to improve the efficacy of cancer immunotherapy.

Patients achieved pCR during neoadjuvant chemotherapy had better outcome than adjuvant chemotherapy setting in breast cancer: A comparative study.

PURPOSE: Pathological complete response(pCR) during neoadjuvant chemotherapy(NAC) has been proposed as a predictor for better prognosis in breast cancer. However, few studies compare the outcomes of patients receiving NAC and adjuvant chemotherapy(AC). METHODS: We retrospectively matched the patients who received NAC(N = 462) and AC(N = 462) by age, time of diagnosis, and primary clinical stage using the propensity score match in breast cancer patients treated in Sir Run Run Shaw Hospital with the median follow up of 67 months. Death from breast cancer and recurrence were used as endpoints. A multivariable Cox models were used to estimate the hazard ratios for breast-cancer specific survival (BCSS) and DFS. A multivariable logistic regression model was simulated to predict pCR. RESULTS: In patients who received NAC, 18.0%(83/462) patients achieved pCR, while the rest of the patients did not. pCR subgroup demonstrated significant better BCSS and DFS than patients receiving AC(BCSS: HR = 0.39, 95% CI:0.12-0.93, P = 0.03; DFS: HR = 0.16, 95%CI 0.009-0.73, P = 0.013) and non-pCR patients(BCSS: HR = 0.32, 95%CI 0.10-0.77, P = 0.008; DFS: HR = 0.12, 95%CI 0.007-0.55, P = 0.002). Patients who received AC demonstrated insignificant survival compared to non-pCR patients(BCSS: HR= 0.82, 95%CI 0.62-1.10, P = 0.19; DFS: HR = 0.75, 95%CI 0.53-1.07, P = 0.12). Patients with AC had significant better DFS than non-pCR patients(HR = 0.33, 95% CI 0.10-0.94, P = 0.04) in luminal B Her2+ patients. More NAC cycles(>2), TNBC, lower cT stage, and mixed histology indicate higher possibility of pCR(AUC = 0.89). CONCLUSION: pCR patients with NAC indicated better prognosis than patients receiving AC or non-pCR patients from NAC. The timing of chemotherapy may need carefully pondering in luminal B Her2+ patients.

Everolimus combined with PD-1 blockade inhibits progression of triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer. Due to rapid progression and a lack of targetable receptors, TNBC is exceptionally difficult to treat. Available treatment options are nonspecific cytotoxic agents, which have had modest success; thus, there is a need for novel therapies for TNBC. The mammalian/mechanistic target of rapamycin (mTOR) signaling pathway is aberrantly activated in TNBC, and this pathway has been shown to promote cancer cell survival and chemoresistance. As such, mTOR inhibition has been considered a potential therapeutic strategy for TNBC. The mTOR inhibitor everolimus (EVE) has been approved for the treatment of estrogen positive breast cancer; however, its efficacy in TNBC is still undetermined. In this study, we evaluated the effects of EVE monotherapy and the mechanism of EVE resistance in the 4T1 model of TNBC. Whereas EVE monotherapy inhibited mTOR signaling activity, it did not attenuate tumor progression. Additionally, tumors from EVE-treated mice had abnormal vasculature characterized by disorganized architecture and hyperpermeability. We also found that treatment with EVE increased PD-L1 expression in intratumoral vascular endothelial cells, and this increase in endothelial cell-associated PD-L1 corresponded to reduced CD8 + T cell tumor infiltration. Importantly, combination treatment with anti-PD-1 antibody and EVE normalized the tumor vasculature, rescued CD8 + T cell tumor infiltration, and reduced tumor growth. Taken together, our findings improve our current understanding of mechanisms underlying mTOR inhibition resistance in TNBC and identify a novel combination treatment strategy in the treatment of mTOR resistant tumors.

Targeting pyroptosis in breast cancer: biological functions and therapeutic potentials on It.

Pyroptosis is a lytic and inflammatory type of programmed cell death that is mediated by Gasdermin proteins (GSDMs). Attractively, recent evidence indicates that pyroptosis involves in the development of tumors and can serve as a new strategy for cancer treatment. Here, we present a basic knowledge of pyroptosis, and an overview of the expression patterns and roles of GSDMs in breast cancer. In addition, we further summarize the available evidence of pyroptosis in breast cancer progression and give insight into the clinical potential of applying pyroptosis in anticancer strategies for breast cancer. This review will deepen our understanding of the relationship between pyroptosis and breast cancer, and provide a novel potential therapeutic avenue for breast cancer.

Modification of PLAC8 by UFM1 affects tumorous proliferation and immune response by impacting PD-L1 levels in triple-negative breast cancer.

BACKGROUND: Triple-negative breast cancer is characterized by a poor prognosis and lack of targeted treatments, and thus, new targeting markers and therapeutic strategies are urgently needed. We previously indicated that PLAC8 promotes tumorigenesis and exerts multidrug resistance in breast cancer. Therefore, we aimed to characterize the PLAC8-regulated network in triple-negative breast cancer. METHODS: We measured the levels of PLAC8 in breast cancer cell lines and found that PLAC8 is post-translationally modified by ubiquitin-fold modifier 1 (UFM1). Then, we revealed a new regulatory system of PD-L1 by PLAC8 in triple-negative breast cancer. We also tested the molecular functions of PLAC8 in triple-negative breast cancer cell lines and measured the expression of PLAC8 and PD-L1 in breast cancer tissues. RESULTS: PLAC8 was generally highly expressed in triple-negative breast cancer and could be modified by UFM1, which maintains PLAC8 protein stability. Moreover, PLAC8 could promote cancer cell proliferation and affect the immune response by regulating the level of PD-L1 ubiquitination. Additionally, among patients with breast cancer, the expression of PLAC8 was higher in triple-negative breast cancer than in non-triple-negative breast cancer and positively correlated with the level of PD-L1. CONCLUSIONS: Our current study discoveries a new PLAC8-regulated network in triple-negative breast cancer and provides corresponding guidance for the clinical diagnosis and immunotherapy of triple-negative breast cancer.

Surgical management of monomorphic epitheliotropic intestinal T-cell lymphoma followed by chemotherapy and stem-cell transplant: A case report and review of the literature.

BACKGROUND: Monomorphic epitheliotropic intestinal T-cell lymphoma (MEITL) is a rare and rapidly progressive intestinal T-cell non-Hodgkin lymphoma associated with a very poor prognosis and a median survival of 7 mo. Advances in the identification of MEITL over the last two decades have led to its recognition as a separate entity. MEITL patients, predominantly male, typically present with vague and nonspecific symptoms and diagnosis is predominantly confirmed at laparotomy. Currently, there are no standardized treatment protocols, and the optimal therapy remains unclear. CASE SUMMARY: We report a case of MEITL that was initially considered to be gastrointestinal stromal tumor (GIST) and Imatinib was administered for one cycle. The 62-year-old man presented with abdominal pain, abdominal distension, and weight loss of 20 pounds. Within 2 wk, the size of the mass considerably increased on computed tomography scans. The patient underwent surgery followed by chemotherapy with CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisone) and stem-cell transplant. A correct diagnosis of MEITL was established based on postoperative pathology. Immunophenotypically, the neoplastic cells fulfilled the diagnostic criteria for MEITL as they were CD3+, CD4+, CD8+, CD56+, and TIA-1+. CONCLUSION: Given that MEITL has no predisposing factor and presents with vague symptoms with rapid progression, the concomitant presence of abdominal symptoms and B symptoms (weight loss, fever, and night sweats) with hypoalbuminemia, anemia, low lymphocytic count and endoscopic findings of diffuse infiltrating type lesions should alert physicians to this rare disease, especially when it comes to Asian patients. Immediate laparotomy should then be carried out followed by chemotherapy and stem-cell transplant.

The role of lncRNA H19 in tumorigenesis and drug resistance of human Cancers.

Systemic therapy is one of the most significant cancer treatments. However, drug resistance often appears and has become the primary cause of cancer therapy failure. Regulation of drug target, drug metabolism and drug efflux, cell death escape (apoptosis, autophagy, et al.), epigenetic changes, and many other variables are complicatedly involved in the mechanisms of drug resistance. In various types of cancers, long non-coding RNA H19 (lncRNA H19) has been shown to play critical roles in tumor development, proliferation, metastasis, and multiple drug resistance as well. The efficacy of chemotherapy, endocrine therapy, and targeted therapy are all influenced by the expression of H19, especially in breast cancer, liver cancer, lung cancer and colorectal cancer. Here, we summarize the relationship between lncRNA H19 and tumorigenesis, and illustrate the drug resistance mechanisms caused by lncRNA H19 as well. This review may provide more therapeutic potential targets for future cancer treatments.

Detection of circulating tumor cells: opportunities and challenges.

Circulating tumor cells (CTCs) are cells that shed from a primary tumor and travel through the bloodstream. Studying the functional and molecular characteristics of CTCs may provide in-depth knowledge regarding highly lethal tumor diseases. Researchers are working to design devices and develop analytical methods that can capture and detect CTCs in whole blood from cancer patients with improved sensitivity and specificity. Techniques using whole blood samples utilize physical prosperity, immunoaffinity or a combination of the above methods and positive and negative enrichment during separation. Further analysis of CTCs is helpful in cancer monitoring, efficacy evaluation and designing of targeted cancer treatment methods. Although many advances have been achieved in the detection and molecular characterization of CTCs, several challenges still exist that limit the current use of this burgeoning diagnostic approach. In this review, a brief summary of the biological characterization of CTCs is presented. We focus on the current existing CTC detection methods and the potential clinical implications and challenges of CTCs. We also put forward our own views regarding the future development direction of CTCs.

HJURP regulates cell proliferation and chemo-resistance via YAP1/NDRG1 transcriptional axis in triple-negative breast cancer.

Triple-negative breast cancer is still a difficult point in clinical treatment at present, and a deep study of its pathogenesis has great clinical value. Therefore, our research mainly focuses on exploring the progression of triple-negative breast cancer and determines the important role of the HJURP/YAP1/NDRG1 transcriptional regulation axis in triple-negative breast cancer. We observed significantly increased HJURP expression levels in triple-negative breast cancer compared to other subtypes. HJURP could affect the level of ubiquitination modification of YAP1 protein and then regulate its downstream transcriptional activity. Mechanistically, we found that YAP1 positively regulates NDRG1 transcription by binding the promoter region of the NDRG1 gene. And HJURP/YAP1/NDRG1 axis could affect cell proliferation and chemotherapy sensitivity in triple-negative breast cancer. Taken together, these findings provide insights into the transcriptional regulation axis of HJURP/YAP1/NDRG1 in triple-negative breast cancer progression and therapeutic response.

Cancer-associated fibroblasts in breast cancer: Challenges and opportunities.

The tumor microenvironment is proposed to contribute substantially to the progression of cancers, including breast cancer. Cancer-associated fibroblasts (CAFs) are the most abundant components of the tumor microenvironment. Studies have revealed that CAFs in breast cancer originate from several types of cells and promote breast cancer malignancy by secreting factors, generating exosomes, releasing nutrients, reshaping the extracellular matrix, and suppressing the function of immune cells. CAFs are also becoming therapeutic targets for breast cancer due to their specific distribution in tumors and their unique biomarkers. Agents interrupting the effect of CAFs on surrounding cells have been developed and applied in clinical trials. Here, we reviewed studies examining the heterogeneity of CAFs in breast cancer and expression patterns of CAF markers in different subtypes of breast cancer. We hope that summarizing CAF-related studies from a historical perspective will help to accelerate the development of CAF-targeted therapeutic strategies for breast cancer.

Corrigendum: STAT5a Confers Doxorubicin Resistance to Breast Cancer by Regulating ABCB1.

[This corrects the article DOI: 10.3389/fonc.2021.697950.].

Fascin enhances the vulnerability of breast cancer to erastin-induced ferroptosis.

Ferroptosis, which is characterized by intracellular iron accumulation and lipid peroxidation, is a newly described form of regulated cell death that may play a key role in tumour suppression. In the present study, we investigated the expression profiles and biological effects of fascin actin-bundling protein 1 (Fascin, gene name FSCN1) in breast cancer. In addition, bioinformatics analysis of the TCGA cancer database and gain- and loss-of-function studies showed that Fascin enhances sensitivity to erastin-induced ferroptosis. Mechanistically, Fascin directly interacts with cysteine/glutamate transporter (xCT, gene name SLC7A11) and decreases its stability via the ubiquitin-mediated proteasome degradation pathway. Furthermore, we observed that Fascin is substantially upregulated in tamoxifen-resistant breast cancer cell lines, and drug-resistant cells were also more vulnerable to erastin-induced ferroptosis. Taken together, our findings reveal a previously unidentified role of Fascin in ferroptosis by regulating xCT. Thus, ferroptosis activation in breast cancer with high Fascin level may serve as a potential treatment.

Multifaced roles of PLAC8 in cancer.

The role of PLAC8 in tumorigenesis has been gradually elucidated with the development of research. Although there are common molecular mechanisms that enforce cell growth, the impact of PLAC8 is varied and can, in some instances, have opposite effects on tumorigenesis. To systematically understand the role of PLAC8 in tumors, the molecular functions of PLAC8 in cancer will be discussed by focusing on how PLAC8 impacts tumorigenesis when it arises within tumor cells and how these roles can change in different stages of cancer progression with the ultimate goal of suppressing PLAC8-relevant cancer behavior and related pathologies. In addition, we highlight the diversity of PLAC8 in different tumors and its functional output beyond cancer cell growth. The comprehension of PLAC8's molecular function might provide new target and lead to the development of novel anticancer therapies.

STAT5a Confers Doxorubicin Resistance to Breast Cancer by Regulating ABCB1.

Chemoresistance is a daunting challenge to the prognosis of patients with breast cancer. Signal transducer and activator of transcription (STAT) 5a plays vital roles in the development of various cancers, but its function in breast cancer is controversial, and its role in chemoresistance in breast cancer remains unexplored. Here we identified STAT5a as a chemoresistance inducer that regulates the expression of ABCB1 in breast cancer and can be targeted by pimozide, an FDA-approved psychotropic drug. First, we found that STAT5a and ABCB1 were expressed at higher levels in doxorubicin-resistant cell lines and chemoresistant patients, and their expression was positively correlated. Then, we confirmed the essential roles of STAT5a and ABCB1 in doxorubicin resistance in breast cancer cells and the regulation of ABCB1 transcription by STAT5a. Subsequently, the efficacy of pimozide in inhibiting STAT5a and sensitizing doxorubicin-resistant breast cancer cells was tested. Finally, we verified the role of STAT5a in doxorubicin resistance in breast cancer and the efficacy of pimozide in reversing this resistance in vivo. Our study demonstrated the vital role of STAT5a in doxorubicin resistance in breast cancer. Targeting STAT5a might be a promising strategy for treating doxorubicin-resistant breast cancer. Moreover, repurposing pimozide for doxorubicin resensitization is attractive due to the safety profile of pimozide.

Metformin induces Ferroptosis by inhibiting UFMylation of SLC7A11 in breast cancer.

BACKGROUND: Ferroptosis is a newly defined form of regulated cell death characterized by the iron-dependent accumulation of lipid peroxidation and is involved in various pathophysiological conditions, including cancer. Targeting ferroptosis is considered to be a novel anti-cancer strategy. The identification of FDA-approved drugs as ferroptosis inducers is proposed to be a new promising approach for cancer treatment. Despite a growing body of evidence indicating the potential efficacy of the anti-diabetic metformin as an anti-cancer agent, the exact mechanism underlying this efficacy has not yet been fully elucidated. METHODS: The UFMylation of SLC7A11 is detected by immunoprecipitation and the expression of UFM1 and SLC7A11 in tumor tissues was detected by immunohistochemical staining. The level of ferroptosis is determined by the level of free iron, total/lipid Ros and GSH in the cells and the morphological changes of mitochondria are observed by transmission electron microscope. The mechanism in vivo was verified by in situ implantation tumor model in nude mice. RESULTS: Metformin induces ferroptosis in an AMPK-independent manner to suppress tumor growth. Mechanistically, we demonstrate that metformin increases the intracellular Fe2+ and lipid ROS levels. Specifically, metformin reduces the protein stability of SLC7A11, which is a critical ferroptosis regulator, by inhibiting its UFMylation process. Furthermore, metformin combined with sulfasalazine, the system xc- inhibitor, can work in a synergistic manner to induce ferroptosis and inhibit the proliferation of breast cancer cells. CONCLUSIONS: This study is the first to demonstrate that the ability of metformin to induce ferroptosis may be a novel mechanism underlying its anti-cancer effect. In addition, we identified SLC7A11 as a new UFMylation substrate and found that targeting the UFM1/SLC7A11 pathway could be a promising cancer treatment strategy.

Establishment and Characterization of a HER2-Positive Cell Line Derived From the Pleural Effusion of a Drug-Resistant Breast Cancer Patient.

Drug resistance is a daunting challenge in the treatment of breast cancer, making it an urgent problem to solve in studies. Cell lines are important tools in basic and preclinical studies; however, few breast cell lines from drug-resistant patients are available. Herein, we established a novel HER2-positive breast cancer cell line from the pleural effusion of a drug-resistant metastatic breast cancer patient. This cell line has potent proliferative capability and tumorigenicity in nude mice but weak invasive and colony-forming capability. The molecular subtype of the cell line and its sensitivity to chemotherapeutics and HER2-targeting agents are different from those of its origin, suggesting that the phenotype changes between the primary and metastatic forms of breast cancer.