Unusual cerebrospinal fluid finding of intracytoplasmic granules in metaplastic carcinoma of the breast with acinar differentiation.

Cerebrospinal fluid (CSF) evaluation for total and differential cell count is a common practice in pathology for evaluation of various disease conditions. Although rare, these CSF samples yield interesting and unusual morphological findings, which are not only of academic interest, but also may play key roles in diagnosis. For diagnosing metastatic carcinoma in brain and meninges, CSF examination is one of the important tools along with imaging studies. Metaplastic breast carcinoma (MBC) encompasses a rare (<1% of all breast cancers), aggressive and highly heterogeneous group of tumors. MBC is almost always estrogen receptor, progesterone receptor and Her2 negative (triple negative) and shows frequent early distant metastases as well as sub-optimal response to systemic therapies. The involvement of leptomeninges is most commonly associated with these triple- negative subtypes. In this report, we present an unusual case of malignant cells with prominent intracytoplasmic granules in CSF smears of a 46-year-old female with metastatic MBC with acinar differentiation. An extensive review of literature in English language did not return any other reports of a similar finding.

Prozone phenomenon observed in indirect immunofluorescence assay by antibodies against neuronal antigens.

A marked prozone effect was observed in indirect immunofluorescence with human sera and human cerebrospinal fluid in two clinical cases involving breast carcinoma with paraneoplastic neuronal antibodies, and anti- N-methyl-D-aspartic acid (NMDA) receptor antibodies. Anti-Yo antibodies and anti-NMDA antibodies were not detectable under high concentrations (1:10 serum dilution and neat CSF respectively) but showed a true effect when sufficiently diluted at 1:80 and 1:5 respectively. This paper demonstrates that prozone effects have their occurrences in indirect immunofluorescence, and clinicians and laboratory technicians should be wary of its implications during screening of autoantibody markers in neurological diseases.

Intra-CSF topotecan in treatment of breast cancer patients with leptomeningeal metastases.

BACKGROUND: There are few treatment options for patients with leptomeningeal metastases (LM). METHODS: We report a case series of patients with breast cancer and LM treated with intra-CSF topotecan (TOPO). Outcome was assessed by clinical exam and MRI at baseline, at end of induction (4-5 weeks), then every 3 months; CSF cytology was determined at baseline and with each treatment. RESULTS: Thirty-one women [median age, 58 (37-81); median KPS 60 (40-100)] received treatment. At baseline, 68% had positive CSF cytology, and 90%, leptomeningeal enhancement on MRI. 84% of patients also received focal RT (not during TOPO) and 77% received concomitant systemic hormonal or chemotherapy. Median number of TOPO treatments was 14.5 (range, 3-71); median duration of treatment, 11 weeks (1-176); and median OS, 6.9 months (range, 0.9-48.8). Patients remaining progression-free during 4-6 weeks of induction (81%) had a median OS of 11.5 months (range, 1.8-48.8). Overall neurologic PFS at 6, 12, and 24 months was 39%, 26%, and 6%, respectively. Clearing of CSF malignant cells for >3 consecutive samples occurred in 10/21 (48%) patients with positive CSF cytology at baseline, remaining clear for a median duration of 15.9 months (range, 1.4-34.5). Grade 3 adverse events included headache or vomiting (3pts), T2 hyperintensity surrounding the ventricular catheter (2 pts), and meningitis (2 pts). CONCLUSIONS: Intra-CSF TOPO, with focal RT as needed for symptomatic areas of enhancement produced durable clearing of CSF malignant cells in 48% of patients positive at baseline, with promising median PFS and OS.

Cerebrospinal fluid circulating tumor cells as a quantifiable measurement of leptomeningeal metastases in patients with HER2 positive cancer.

PURPOSE: The CellSearch® system has been used to identify circulating tumor cells (CTCs) in cerebrospinal fluid (CSF) to diagnose leptomeningeal metastasis (LM) in patients with epithelial cancers. Using this system, we prospectively explored sequential CSF CTC enumeration in patients with LM from HER2+ cancers receiving intrathecal (IT) trastuzumab to capture dynamic changes in CSF CTC enumeration. METHODS: CSF from patients enrolled in an IRB-approved phase I/II dose escalation trial of IT trastuzumab for LM in HER2+ cancer (NCT01325207) was obtained on day 1 of each cycle and was evaluated by the CellSearch® platform for CTC enumeration. The results were correlated with CSF cytology from the same sample, along with clinical and radiographic response. RESULTS: Fifteen out of 34 patients with HER2+ LM were enrolled in CSF CTC analysis; 14 were women. Radiographic LM was documented in 14 (93%) patients; CSF cytology was positive in 6 (40%) and CSF CTCs were identified in 13 (87%). Median CSF CTC was 22 CTCs (range 0-200 +) per 3 ml. HER2/neu expression analysis of CTCs was performed in 8 patients; 75% had confirmed expression of HER2/neu positivity in CSF and HER2/neu expression was absent in 25%. Four of 10 patients received 7 or more cycles of IT trastuzumab; in 3 of these patients, increase in CSF CTCs enumeration from baseline was detected 2-3 months prior to changes seen on MRI, and while CSF cytology remained negative. CONCLUSION: Our study demonstrates that enumeration of CSF CTCs may provide dynamic, quantitative assessment of tumor burden in the central nervous system compartment during treatment for LM and prior to changes on MRI or CSF cytology. TRIAL REGISTRATION: Clinicaltrials.gov: NCT01325207; registered March 29th, 2011.

Astrocytic IGFBP2 and CHI3L1 in cerebrospinal fluid drive cortical metastasis of HER2+breast cancer.

The brain is often reported as the first site of recurrence among breast cancer patients overexpressing human epidermal growth factor receptor 2 (HER2). Although most HER2+tumors metastasize to the subcortical region of the brain, a subset develops in the cortical region. We hypothesize that factors in cerebrospinal fluid (CSF) play a critical role in the adaptation, proliferation, and establishment of cortical metastases. We established novel cell lines using patient biopsies to model breast cancer cortical and subcortical metastases. We assessed the localization and growth of these cells in vivo and proliferation and apoptosis in vitro under various conditions. Proteomic analysis of human CSF identified astrocyte-derived factors that support the proliferation of cortical metastases, and we used neutralizing antibodies to test the effects of inhibiting these factors both in vivo and in vitro. The cortical breast cancer brain metastatic cells exhibited greater proliferation than subcortical breast cancer brain metastatic cells in CSF containing several growth factors that nourish both the CNS and tumor cells. Specifically, the astrocytic paracrine factors IGFBP2 and CHI3LI promoted the proliferation of cortical metastatic cells and the formation of metastatic lesions. Disruption of these factors suppressed astrocyte-tumor cell interactions in vitro and the growth of cortical tumors in vivo. Our findings suggest that inhibition of IGFBP2 and CHI3LI signaling, in addition to existing treatment modalities, may be an effective therapeutic strategy targeting breast cancer cortical metastasis.

CYFRA 21-1 levels in cerebrospinal fluid as a putative therapeutic monitoring biomarker for patients with leptomeningeal carcinomatosis: A pilot study.

BACKGROUND: To investigate the feasibility of cerebrospinal fluid (CSF) CYFRA 21-1 levels as a therapeutic monitoring biomarker in leptomeningeal carcinomatosis (LMC) patients undergoing ventriculo-lumbar perfusion (VLP) chemotherapy. METHODS: The levels of CYFRA 21-1 in 42 CSF samples from 15 LMC patients were analyzed using an electrochemiluminescence immunoassay. Samples were collected at individual time points during VLP chemotherapy. Therapeutic outcomes were measured as improvements in the Karnofsky Performance Status (KPS) score and decreasing intracranial pressure (ICP) as the main endpoint of VLP chemotherapy. Changes in CSF CYFRA 21-1 levels, protein levels, and cytology results were also investigated. We subsequently evaluated whether these changes were correlated with KPS score and ICP. RESULTS: The CSF CYFRA 21-1 levels at individual time points were associated with KPS score and ICP. The KPS scores (p= 0.007) and ICP (p= 0.018) of patients with high CSF CYFRA 21-1 levels were significantly different from those of patients with low CSF CYFRA 21-1 levels. By contrast, CSF protein levels and cytological responses were not significantly associated with KPS scores and ICP. CONCLUSIONS: CSF CYFRA 21-1 may have utility as a therapeutic monitoring biomarker to design personalized therapeutic strategies in LMC patients undergoing VLP chemotherapy.

Aquaporin-4 antibody positive short transverse myelitis associated with breast cancer.

Neuromyelitis optica spectrum disorder (NMOSD) is an inflammatory demyelinating disease of the central nervous system (CNS). A typical finding on spinal magnetic resonance imaging (MRI) of NMOSD is longitudinally extensive transverse myelitis (LETM). However, patients with NMOSD presenting with short-segment transverse myelitis (STM) during myelitis attacks associated with breast cancer are uncommon. We report a case of a 35-year-old woman with STM and left eye optic neuritis. The patient was positive for serum aquaporin-4 antibodies (AQP4-IgG), and a biopsy of the left breast showed invasive ductal carcinoma. The patient was diagnosed with NMOSD and breast malignancy. This is the first report of a patient with NMOSD whose spinal MRI showed STM and serum test showed that the patient's AQP4-IgG was positive and complicated by breast cancer. This case improves our understanding of the association between NMOSD and cancer and raises the question of whether it was a coincidental occurrence. It is important to search for extensive malignancies in patients presenting with atypical MRI or no reaction to traditional therapies.

Therapeutic targeting of HER2-CB2R heteromers in HER2-positive breast cancer.

Although human epidermal growth factor receptor 2 (HER2)-targeted therapies have dramatically improved the clinical outcome of HER2-positive breast cancer patients, innate and acquired resistance remains an important clinical challenge. New therapeutic approaches and diagnostic tools for identification, stratification, and treatment of patients at higher risk of resistance and recurrence are therefore warranted. Here, we unveil a mechanism controlling the oncogenic activity of HER2: heteromerization with the cannabinoid receptor CB2R. We show that HER2 physically interacts with CB2R in breast cancer cells, and that the expression of these heteromers correlates with poor patient prognosis. The cannabinoid Δ9-tetrahydrocannabinol (THC) disrupts HER2-CB2R complexes by selectively binding to CB2R, which leads to (i) the inactivation of HER2 through disruption of HER2-HER2 homodimers, and (ii) the subsequent degradation of HER2 by the proteasome via the E3 ligase c-CBL. This in turn triggers antitumor responses in vitro and in vivo. Selective targeting of CB2R transmembrane region 5 mimicked THC effects. Together, these findings define HER2-CB2R heteromers as new potential targets for antitumor therapies and biomarkers with prognostic value in HER2-positive breast cancer.

[Cerebrospinal fluid examination after liposomal cytarabine intrathecal injection]. / Examen cytologique d'un liquide céphalorachidien après administration intrathécale de cytarabine liposomale.

The patient is a 36 year old female who presented breast cancer with leptomeningeal involvement. A systematic lumbar puncture was performed and sent to the laboratory for CSF analysis. CSF examination using wet mount preparation showed a large number of round spherules. After discussion with the ordering physician, we learnt that the patient had received intrathecal liposomal cytarabine injection 19 days earlier. Cytarabine liposomes are spherules with a granular interior and range in size from 10-30 µm. It can be confused with leukocytes and lead to spurious elevation of CSF leukocytes count. Care needs to be taken in interpreting CSF results in patients who have received intrathecal liposomal cytarabine.

Lower motor neuron syndrome in a patient with HER2-positive metastatic breast cancer: A case report and review of the literature.

Paraneoplastic neurological syndromes are very rare and often associated to breast, ovarian and small cells lung cancers. Paraneoplastic motor neuron diseases (MNDs) are even rarer, and frequently described in patients with breast cancer. We presented the first case of patient affected by HER2-positive breast tumor and possible paraneoplastic lower motor neuron disease. In literature, few cases are reported but no one highlights the tumor receptors' profile. Instead, HER2-positive breast cancers are prone to be related to anti-Yo-associated paraneoplastic cerebellar disorders. Anti-onconeural antibodies positivity can be rarely found, confirming that paraneoplastic MND have no specific biomarkers. The presence of CSF oligoclonal bands (OBs) suggests the presence of immune-mediated mechanism, in absence of other possible OBs causes.

Detection of cerebrospinal fluid tumor cells and its clinical relevance in leptomeningeal metastasis of breast cancer.

Circulating tumor cells are commonly observed in the peripheral blood of advanced breast cancer patients. We tested the feasibility of tumor cell detection in the cerebrospinal fluid (CSF) and studied its clinical relevance in leptomeningeal metastasis (LM) of breast cancer. CSF samples were collected from 38 metastatic breast cancer patients known or suspected to have LM. Control CSF samples were collected from 14 individuals without solid tumor malignancy. We used a modified CellSearch™ assay and an alternative EPCAM-based method involving immunomagnetic enrichment followed by flow cytometry (IE/FC) to enumerate CSF tumor cells (CSFTCs). CSFTCs were assayed at time of LM diagnosis and over the course of LM-directed therapy. We analyzed a total of 102 CSF samples with modified CellSearch™. The CSFTC counts were strongly correlated with the corresponding IE/FC results (Pearson's r = 0.94). Twenty-eight out of 30 samples in which malignant cells were identified by CSF cytology were CSFTC-positive by modified CellSearch™. Baseline CSFTC levels from 21 patients eventually diagnosed with LM were significantly higher than the controls (p = 0.0202), whereas 13 patients deemed not to have LM showed CSFTC results indistinguishable from the controls. In patients with serial samples, it was possible to monitor CSFTC levels as a potential biomarker of treatment response. CSFTC detection using a modified CellSearch™ assay demonstrated high sensitivity in detecting malignant cells in CSF and may be a promising method for diagnosing LM and monitoring LM during treatment.

Systemic Therapy for HER2-Positive Central Nervous System Disease: Where We Are and Where Do We Go From Here?

Patients with human epidermal growth factor receptor 2 (HER2)-positive breast cancer are at an increased risk of developing brain metastases. The incidence and prevalence of central nervous system (CNS) disease are increasing due to improved survival, which can be attributed to better systemic therapies for extracranial disease. The current standard of care for brain metastases includes a combination of surgery and/or radiation. Systemic therapies are typically reserved for patients with intracranial progression following radiation, due to their limited ability to cross the blood-brain barrier. None of the available anti-HER2 agents (trastuzumab, lapatinib, pertuzumab, and ado-trastuzumab emtansine (T-DM1)) are currently approved for the treatment of brain metastases. Research is underway evaluating novel anti-HER2 agents, which have demonstrated CNS activity. This article discusses the current data on using anti-HER2 therapies to treat CNS disease as well as the newer anti-HER2 agents, which may overcome the current challenges faced in treating brain metastases in the HER2-positive patient population.

A pilot study of bevacizumab combined with etoposide and cisplatin in breast cancer patients with leptomeningeal carcinomatosis.

BACKGROUND: Elevated vascular endothelial growth factor (VEGF) was associated with poor prognosis in leptomeningeal carcinomatosis and anti-angiogenic therapy was found to prolong the survival of mice in preclinical studies. This prospective pilot study investigated the efficacy of anti-VEGF therapy plus chemotherapy in patients with leptomeningeal carcinomatosis originating from breast cancer. METHODS: Eligible patients were scheduled to receive bevacizumab combined with etoposide and cisplatin (BEEP) every 3 weeks for a maximum of 6 cycles or until unacceptable toxicity. The primary objective was the central nervous system (CNS)-specific response rate, which was defined as disappearance of cancer cells in the cerebrospinal fluid (CSF) and an improved or stabilized neurologic status. The impact of VEGF inhibition on etoposide penetration into the CSF was analyzed. RESULTS: Eight patients were enrolled. The CNS-specific response rate was 60% in 5 evaluable patients. According to intent-to-treat analysis, the median overall survival of the eight patients was 4.7 months (95% confidence interval, CI, 0.3-9.0) and the neurologic progression-free survival was 4.7 months (95% CI 0-10.5). The most common grade 3/4 adverse events were neutropenia (23.1%), leukopenia (23.1%), and hyponatremia (23.1%). The etoposide concentrations in the CSF were much lower than those in plasma, and bevacizumab did not increase etoposide delivery to the CSF. CONCLUSIONS: BEEP exhibited promising efficacy in breast cancer patients with leptomeningeal carcinomatosis. Additional studies are warranted to verify its efficacy and clarify the role of anti-angiogenic therapy in this disease. TRIAL REGISTRATION: ClinicalTrials.gov identifying number NCT01281696 .

Breast cancer leptomeningeal metastasis: the results of combined treatment and the comparison of methotrexate and liposomal cytarabine as intra-cerebrospinal fluid chemotherapy.

BACKGROUND: This was a prospective observational study to assess the results of the treatment of patients with breast cancer leptomeningeal metastasis (LM) and to compare the efficacy of methotrexate and liposomal cytarabine in patients treated intrathecally by lumbar puncture. PATIENTS AND METHODS: In this prospective observational study, 149 consecutive patients with breast cancer and LM treated between the years 1999 and 2011 were assessed. Multimodality treatment methods were used: systemic therapy in 77 patients, radiotherapy in 92 patients, intrathecal methotrexate in 81 patients, and intrathecal liposomal cytarabine in 15 patients. RESULTS: The median survival of all patients was 4.2 months. The median survival of patients in whom systemic intravenous/oral treatment was used was 6 months, in those who did not have systemic treatment, the median survival was 2 months (P < .001). The median survival of patients treated with intrathecal methotrexate was 4.2 months; in patients treated with intrathecal liposomal cytarabine, the median survival was 4.6 months, and in patients who did not receive intrathecal treatment, the median survival was 3.7 months (P = .717). Median survival after whole-brain radiotherapy was 4.6 months and with no radiotherapy, it was 3.2 months (P = .028). Multivariate analysis revealed a Karnofsky performance status (KPS) of > 70. Systemic intravenous/oral treatment and bone as a site of metastasis were factors prolonging survival from LM. CONCLUSION: Among treatment methods, only systemic therapy prolonged survival in patients with LM. Neither radiotherapy nor lumbar intrathecal therapy influenced survival in those patients; however, both methods alleviated signs and symptoms of LM. No difference in survival was observed in patients treated intrathecally with methotrexate and those treated with liposomal cytarabine. Treatment with both drugs was comparable.

COX-2 drives metastatic breast cells from brain lesions into the cerebrospinal fluid and systemic circulation.

Breast cancer is among the most common malignancies that metastasize to the brain, with 15% to 20% of patients with metastatic breast cancer eventually developing brain metastases. We previously reported a method to enumerate tumor cells in the cerebrospinal fluid (CSF) of patients with breast cancer with central nervous system (CNS) metastases, a setting that lacks sufficiently informative biomarkers. Here, we show that breast cancer cells can spontaneously disseminate into the CSF from brain lesions in mice in a COX-2-dependent manner and can escape from the CNS to systemic circulation. Enumeration of tumor cells in the peripheral blood (circulating tumor cells, CTC) and CSF (cerebrospinal fluid tumor cells, CSFTC) of nine breast cancer patients with brain metastases revealed dynamic changes in tumor cell burden in both the peripheral blood and CSF compartments that correlated with clinical disease progression. Interestingly, four of the enrolled patients exhibited rapid intercompartmental transitioning of the disease reflected in the CTC and CSFTC counts that preceded corresponding evidence by clinical imaging or neurologic symptoms. Two of these patients had systemic disease recurrence involving the primary malignant site. Intercompartmental cycling of tumor cells may represent an important mechanism for disease persistence and recurrence that may involve tumor self-seeding. Our findings demonstrate the involvement of COX-2 in the genesis of CSFTCs and suggest that COX-2 inhibitors should be investigated in patients with breast cancer with brain metastases for their ability to reduce CSFTC counts and prevent systemic recurrence.

Molecular profiling of tumor cells in cerebrospinal fluid and matched primary tumors from metastatic breast cancer patients with leptomeningeal carcinomatosis.

Although leptomeningeal carcinomatosis is a well-established clinical syndrome, virtually nothing is known about the tumor cells responsible for this particularly aggressive metastatic process. To isolate cerebrospinal fluid-derived tumor cells (CSFTC) from 15 patients with metastatic breast cancer diagnosed with leptomeningeal carcinomatosis, CSF samples were subjected to a two-step method involving immunomagnetic enrichment and fluorescence-activated cell sorting (IE/FACS), a technique previously used for isolating circulating tumor cells (CTC) from blood. CSFTCs were subjected to genome-wide copy number analysis by array comparative genomic hybridization. Genomic profiling was successfully performed for 13 of 15 patients (87%). Copy number analysis in CSFTCs revealed genomic alterations commonly observed in primary breast cancer and CTCs, indicating their malignant origin. Interestingly, 12 (92%) harbored high-level gains on the 8q24 locus, which includes the MYC oncogene. Comparison of CSFTCs against corresponding archival primary tumors in six patients revealed clonal relationships with some divergence. Good concordance among serial samples attested to the reproducibility of the assay. Our approach for isolation and molecular analysis of CSFTCs yielded new insights into the molecular nature of these cells. Further genomic and functional analyses may help elucidate mechanisms by which tumor cells metastasize to the central nervous system.

Salvage intracerebrospinal fluid thiotepa in breast cancer-related leptomeningeal metastases: a retrospective case series.

There is currently a paucity of data on salvage intracerebrospinal fluid (intra-CSF) chemotherapy in leptomeningeal metastases (LM). This report is a single-institution experience with salvage treatment in patients with breast cancer (BC) and LM. This retrospective cohort describes 24 consecutive patients with BC selected for a second-line of treatment for LM. The first line of LM treatment consisted of intra-CSF liposomal cytarabine in all patients combined with systemic therapy in 18 cases and radiotherapy in four cases. Second-line (salvage) treatment utilized intra-CSF thiotepa in all and systemic chemotherapy in nine patients. No patient received CNS-directed radiotherapy. The median Eastern Cooperative Oncology Group performance status at initiation of intra-CSF thiotepa treatment was 3 (range 1-4). The median progression-free survival and median survival following intra-CSF thiotepa was 3.1 months (range 3 days-2 years) and 4.0 months (range 6 days-2.5 years), respectively. The median overall survival from LM diagnosis was 9.5 months (range 1.3 months-2.7 years). No grade 3 or higher toxicity was observed. Recognizing the limits of a retrospective study, intra-CSF thiotepa has an acceptable toxicity profile and appears to be a reasonable option for selected BC patients.

Clinical features and outcome of leptomeningeal metastasis in patients with breast cancer: a single center experience.

BACKGROUND: Leptomeningeal metastasis (LM) is one of the major problems in the management of metastatic breast cancer; typically, LM has a devastating prognosis and often represents a terminal event. The present study analyzed the clinical features and outcome of LM in patients with breast cancer. METHODS: The medical records of patients diagnosed with LM from breast cancer at Asan Medical Center, between 2002 and 2012, were reviewed retrospectively. RESULTS: Of 95 LM patients, 38 (40 %) had an ECOG performance status (PS) ≤ 2, and the median age was 47 years (range 26-72 years). At the time of LM diagnosis, 46 patients (48.4 %) presented with coincidental failure of systemic disease control. Seventy-eight patients (82.1 %) underwent intrathecal (IT) chemotherapy, resulting in cytologic negative conversion in 26 patients, and 46 patients (48.4 %) received systemic chemotherapy. The median overall survival (OS) time was 3.3 months, and 7.8 % of the patients survived for more than 1 year. OS tended to be higher in patients who achieved cytologic negative conversion from IT chemotherapy than in those who did not (4.5 vs. 2.4 months, P = 0.088). Multivariate analysis demonstrated that ECOG PS ≤ 2, controlled extracranial disease at the time of LM diagnosis, and systemic chemotherapy after LM diagnosis were independent factors associated with survival. CONCLUSIONS: The prognosis of patients with LM from breast cancer is poor. Systemic chemotherapy, in addition to intrathecal chemotherapy, might confer a survival benefit, even after the detection of LM.