The anti-proliferative effects of a frankincense extract in a window of opportunity phase ia clinical trial for patients with breast cancer.

PURPOSE: Boswellic acids, active components of frankincense, suppress tumor proliferation in vitro with a strong clinical trial safety profile in patients with inflammatory diseases. We performed a Phase Ia window of opportunity trial of Boswellia serrata (B. serrata) in patients with breast cancer to evaluate its biologic activity and safety. METHODS: Patients with invasive breast cancer were treated pre-operatively with B. Serrata (2400 mg/day PO) until the night before surgery for a median of 11 days (SD 6 days; range: 5-23 days). Paraffin-embedded sections from pretreatment diagnostic core biopsies and post-treatment surgical excisions were evaluated using a tunnel assay and immunohistochemistry staining with Ki-67 antibodies. A non-intervention retrospective control arm consisting of core and surgical tissue specimens from untreated patients was used to compare patients treated with B. Serrata. The change in proliferation and apoptosis between diagnostic core specimens and surgical specimens was compared between the control and treatment groups using a two-tailed paired t-test. RESULTS: Twenty-two patients were enrolled, of which 20 received treatment, and 18 had sufficient tissue for IHC. There was an increase in percent change in proliferation from core biopsy to surgical excision in the control group (n = 18) of 54.6 ± 21.4%. In the B. serrata-treated group there was a reduction in proliferation between core biopsy and excision (n = 18) of 13.8 ± 11.7%. This difference was statistically significant between the control and B. serrata-treated groups (p = 0.008). There was no difference in change in apoptosis. There were no serious adverse events related to the drug. CONCLUSION: Boswellia serrata inhibited breast cancer proliferation and was well-tolerated in a Phase Ia window of opportunity trial.

Secreted frizzled related-protein 2 is prognostic for human pancreatic cancer patient survival and is associated with fibrosis.

Pancreatic adenocarcinoma (PDAC) is one of the deadliest cancers, with five-year survival rates of 9%. We hypothesized that secreted frizzled-related protein 2 (SFRP2) may influence stromal growth in pancreatic cancer, since it increases fibrosis and collagen production in non-neoplastic pathologies. We assessed SFRP2 value as a biomarker and assessed its function in PDAC. SFRP2 gene expression in patients with PDAC was analyzed using TCGA data. Disease free survival (DFS) was analyzed using Kaplan Meier test. The effect of KRAS inhibition on SFRP2 expression in PDAC cells was assessed. The associations of stromal content with SFPR2 mRNA and protein with fibrosis were analyzed. The role of SFRP2 in mesenchymal transformation was assessed by western blot in fibroblasts. Of all cancers in TCGA, SFRP2 levels were highest in PDAC, and higher in PDAC than normal tissues (n= 234, p= 0.0003). High SFRP2 levels correlated with decreased DFS (p= 0.0097). KRAS inhibition reduced SFRP2 levels. Spearman correlation was 0.81 between stromal RNA and SFRP2 in human PDAC, and 0.75 between fibrosis and SFRP2 levels in PDAC tumors. SFRP2-treated fibroblasts displayed mesenchymal characteristics. SFRP2 is prognostic for PDAC survival, regulated by KRAS, and associated with PDAC fibrosis.

The impact of nurse navigation on timeliness to treatment for benign high-risk breast pathology.

PURPOSE: High-risk breast pathology is a breast cancer risk factor for which timely treatment is crucial. Nurse navigation programs have been implemented to minimize delays in patient care. This study evaluated nurse navigation in terms of timeliness to surgery for patients with high-risk breast pathology. METHODS: This was a single-institution, retrospective review of patients with identified high-risk breast pathology undergoing lumpectomy between January 2017 and June 2019. Patients were stratified into cohorts based on periods with and without nurse navigation. Preoperative and postoperative time to care as well as demographic and tumor characteristics were compared using univariate and multivariate analysis. RESULTS: 100 patients had assigned nurse navigators and 29 patients did not. Nurse navigation was associated with reduced time from referral to date of surgery (DOS) by 16.9 days (p = 0.003). Patients > 75 years had a shorter time to first appointment (p = 0.03), and patients with Medicare insurance had a reduced time from referral to DOS (p = 0.005). 20% of all patients were upstaged to cancer on final surgical pathology. CONCLUSION: Nurse navigation was significantly associated with decreased time to care for patients with high-risk breast pathology undergoing lumpectomy. We recommend nurse navigation programs as part of a comprehensive approach for patients with high-risk breast pathology.

Prognostic factors for residual occult disease in shave margins during partial mastectomy.

PURPOSE: Shave margins have been shown to decrease positive final margins in partial mastectomy. We investigated prognostic factors associated with residual disease in shave margins. METHODS: Patients with invasive breast carcinoma and ductal carcinoma in situ (DCIS) who had circumferential shave margins excised during lumpectomy were abstracted from a retrospective database from 2015 to 2018. We defined residual occult disease (ROD) as either (1) residual disease in a shave margin when the initial lumpectomy specimen had negative margins or (2) residual disease in a shave margin that did not correspond with the positive lumpectomy margin. We identified the frequency of ROD and conducted logistic regression analysis to identify associated prognostic factors. RESULTS: 166 Patients (139 invasive carcinoma, 27 DCIS) were included with median follow-up of 28 months (9-50 months). Residual occult disease existed in 34 (24.5%) with invasive carcinoma and 8 (29.6%) with DCIS. In univariate analyses of the invasive group, invasive lobular carcinoma and a positive initial, non-corresponding lumpectomy margin were predictive of ROD (OR 3.63, p = 0.04, OR 3.48, p = 0.003 respectively). In multivariate analysis, a positive lumpectomy margin remained significant, p = 0.007. No variables were associated with ROD in DCIS. CONCLUSION: Residual occult disease was shown to be a frequent event in this analysis of lumpectomy with circumferential shave margins. Having a positive initial lumpectomy margin was predictive of ROD in a non-corresponding margin. Surgeons should consider not being selective in their shave margins or margin of re-excision if shave margins were not obtained in their initial surgery.

Overcoming PD-1 Inhibitor Resistance with a Monoclonal Antibody to Secreted Frizzled-Related Protein 2 in Metastatic Osteosarcoma.

Secreted frizzled-related protein 2 (SFRP2) promotes the migration/invasion of metastatic osteosarcoma (OS) cells and tube formation by endothelial cells. However, its function on T-cells is unknown. We hypothesized that blocking SFRP2 with a humanized monoclonal antibody (hSFRP2 mAb) can restore immunity by reducing CD38 and PD-1 levels, ultimately overcoming resistance to PD-1 inhibitors. Treating two metastatic murine OS cell lines in vivo, RF420 and RF577, with hSFRP2 mAb alone led to a significant reduction in the number of lung metastases, compared to IgG1 control treatment. While PD-1 mAb alone had minimal effect, hSFRP2 mAb combination with PD-1 mAb had an additive antimetastatic effect. This effect was accompanied by lower SFRP2 levels in serum, lower CD38 levels in tumor-infiltrating lymphocytes and T-cells, and lower PD-1 levels in T-cells. In vitro data confirmed that SFRP2 promotes NFATc3, CD38 and PD-1 expression in T-cells, while hSFRP2 mAb treatment counteracts these effects and increases NAD+ levels. hSFRP2 mAb treatment further rescued the suppression of T-cell proliferation by tumor cells in a co-culture model. Finally, hSFRP2 mAb induced apoptosis in RF420 and RF577 OS cells but not in T-cells. Thus, hSFRP2 mAb therapy could potentially overcome PD-1 inhibitor resistance in metastatic osteosarcoma.

A Multifaceted Approach to Opioid Education, Prescribing, and Disposal for Patients with Breast Cancer Undergoing Surgery.

BACKGROUND: Standardized prescribing practices are recommended to decrease opioid abuse, however, data regarding the handling and disposal of leftover narcotics are lacking. This quality improvement project and analysis evaluated implementation of standardized prescribing, opioid education, and a narcotic disposal system. METHODS: This initiative was implemented over a 1-y period among patients who underwent breast surgery. The project included the following: 1) implementation of standardized prescribing, 2) voluntary and anonymous survey analysis, and 3) preoperative education regarding risks of opioids, charcoal disposal bag distribution, and follow-up survey to assess use and use of intervention. RESULTS: Preintervention surveys were completed by 53 patients, and 60% (n = 32) underwent lumpectomy. Narcotic prescriptions were filled by 90%; median number of pills taken was 3 (range 0-24), however 93% felt that a non-narcotic was more effective. Eighty three percentage of patients had unused pills, and 58% kept these pills in an unlocked cabinet. Postintervention surveys were completed by 66 patients, and 48% (n = 32) underwent lumpectomy. Narcotic prescriptions were filled by 88%, median number of pills taken was 4 (range 0-40), and 89% of patients had pills leftover. Sixty seven percentage of patients found the education handout useful and charcoal bag use was reported by 37% (n = 17). The median postoperative pain control satisfaction score was 4.5 (5-point Likert scale, 1 = very dissatisfied, 5 = very satisfied) on both preintervention and postintervention surveys. CONCLUSIONS: This study, which included standardized prescribing parameters, opioid education, and implementation of a disposal method, was found to be feasible, beneficial, and did not compromise postoperative pain control.

Development of a Novel Humanized Monoclonal Antibody to Secreted Frizzled-Related Protein-2 That Inhibits Triple-Negative Breast Cancer and Angiosarcoma Growth In Vivo.

BACKGROUND: We previously reported that secreted frizzled-related protein-2 (SFRP2) is expressed in a variety of tumors, including sarcoma and breast carcinoma, and stimulates angiogenesis and inhibits tumor apoptosis. Therefore, we hypothesized that a humanized SFRP2 monoclonal antibody (hSFRP2 mAb) would inhibit tumor growth. METHODS: The lead hSFRP2 antibody was tested against a cohort of 22 healthy donors using a time course T-cell assay to determine the relative risk of immunogenicity. To determine hSFRP2 mAb efficacy, nude mice were subcutaneously injected with SVR angiosarcoma cells and treated with hSFRP2 mAb 4 mg/kg intravenously every 3 days for 3 weeks. We then injected Hs578T triple-negative breast cells into the mammary fat pad of nude mice and treated for 40 days. Control mice received an immunoglobulin (Ig) G1 control. The SVR and Hs578T tumors were then stained using a TUNEL assay to detect apoptosis. RESULTS: Immunogenicity testing of hSFRP2 mAb did not induce proliferative responses using a simulation index (SI) ≥ 2.0 (p < 0.05) threshold in any of the healthy donors. SVR angiosarcoma tumor growth was inhibited in vivo, evidenced by significant tumor volume reduction in the hSFRP2 mAb-treated group, compared with controls (n = 10, p < 0.001). Likewise, Hs578T triple-negative breast tumors were smaller in the hSFRP2 mAb-treated group compared with controls (n = 10, p < 0.001). The hSFRP2 mAb treatment correlated with an increase in tumor cell apoptosis (n = 11, p < 0.05). Importantly, hSFRP2 mAb treatment was not associated with any weight loss or lethargy. CONCLUSION: We present a novel hSFRP2 mAb with therapeutic potential in breast cancer and sarcoma that has no effect on immunogenicity.

Thioredoxin-1 improves the immunometabolic phenotype of antitumor T cells.

Adoptive transfer of tumor epitope-reactive T cells has emerged as a promising strategy to control tumor growth. However, chronically-stimulated T cells expanded for adoptive cell transfer are susceptible to cell death in an oxidative tumor microenvironment. Because oxidation of cell-surface thiols also alters protein functionality, we hypothesized that increasing the levels of thioredoxin (Trx), an antioxidant molecule facilitating reduction of proteins through cysteine thiol-disulfide exchange, in T cells will promote their sustained antitumor function. Using pre-melanosome protein (Pmel)-Trx1 transgenic mouse-derived splenic T cells, flow cytometry, and gene expression analysis, we observed here that higher Trx expression inversely correlated with reactive oxygen species and susceptibility to T-cell receptor restimulation or oxidation-mediated cell death. These Trx1-overexpressing T cells exhibited a cluster of differentiation 62Lhi (CD62Lhi) central memory-like phenotype with reduced glucose uptake (2-NBDGlo) and decreased effector function (interferon γlo). Furthermore, culturing tumor-reactive T cells in the presence of recombinant Trx increased the dependence of T cells on mitochondrial metabolism and improved tumor control. We conclude that strategies for increasing the antioxidant capacity of antitumor T cells modulate their immunometabolic phenotype leading to improved immunotherapeutic control of established tumors.

Accelerated blood clearance of targeted ultrasound contrast reduced molecular imaging signal intensity: Secreted Frizzled Related Protein-2 signal remained significantly higher than signal from either Vascular Endothelial Growth Factor Receptor-2 or alphaVbeta3 integrin.

Multiple doses of polyethylene glycol (PEG) decorated pharmaceuticals cause accelerated blood clearance (ABC) due to the generation of antibodies reactive to the PEG moiety. Using molecular imaging to monitor response to therapy could be complicated by the ABC effect due to PEG chains in microbubble lipid shells. Our objective was to measure the half-life of targeted contrast flowing through non-tumor tissue during longitudinal imaging studies, and to determine which targeted agent returned the highest signal intensity within tumors. The molecular imaging signals from contrast agents targeted to three distinct molecular targets, Secreted Frizzled-Related Protein-2 (SFRP2), Vascular Endothelial Growth Factor Receptor-2 (VEGFR2), AlphaVBeta3 Integrin (avb3) were all significantly correlated to contrast half-life. The molecular imaging signal from SFRP2 remained significantly higher than the signal returned by ultrasound contrast targeted to either VEGFR2 or avb3 before and after restricting analyses to imaging exams with similar half-lives. We hypothesize that increasing immune clearance rates during our longitudinal studies limited the amount of targeted contrast able to perfuse tumor vasculature, and that this resulted in a global dose-dependent decrease in molecular imaging signals. Molecular imaging may underestimate biomarker levels as longitudinal studies progress and as contrast half-lives decrease, unless contrast dosing is normalized by the amount of contrast able to reach the tumor and surrounding tissue rather than by the injected dosage.

Genetic Variants of VEGFA and FLT4 Are Determinants of Survival in Renal Cell Carcinoma Patients Treated with Sorafenib.

Molecular markers of sorafenib efficacy in patients with metastatic renal cell carcinoma (mRCC) are not available. The purpose of this study was to discover genetic markers of survival in patients with mRCC treated with sorafenib. Germline variants from 56 genes were genotyped in 295 patients with mRCC. Variant-overall survival (OS) associations were tested in multivariate regression models. Mechanistic studies were conducted to validate clinical associations. VEGFA rs1885657, ITGAV rs3816375, and WWOX rs8047917 (sorafenib arm), and FLT4 rs307826 and VEGFA rs3024987 (sorafenib and placebo arms combined) were associated with shorter OS. FLT4 rs307826 increased VEGFR-3 phosphorylation, membrane trafficking, and receptor activation. VEGFA rs1885657 and rs58159269 increased transcriptional activity of the constructs containing these variants in endothelial and RCC cell lines, and VEGFA rs58159269 increased endothelial cell proliferation and tube formation. FLT4 rs307826 and VEGFA rs58159269 led to reduced sorafenib cytotoxicity. Genetic variation in VEGFA and FLT4 could affect survival in sorafenib-treated patients with mRCC. These markers should be examined in additional malignancies treated with sorafenib and in other angiogenesis inhibitors used in mRCC. SIGNIFICANCE: Clinical and mechanistic data identify germline genetic variants in VEGFA and FLT4 as markers of survival in patients with metastatic renal cell carcinoma.

Molecular determinants of post-mastectomy breast cancer recurrence.

Breast cancer (BC) adjuvant therapy after mastectomy in the setting of 1-3 positive lymph nodes has been controversial. This retrospective Translational Breast Cancer Research Consortium study evaluated molecular aberrations in primary cancers associated with locoregional recurrence (LRR) or distant metastasis (DM) compared to non-recurrent controls. We identified 115 HER2 negative, therapy naïve, T 1-3 and N 0-1 BC patients treated with mastectomy but no post-mastectomy radiotherapy. This included 32 LRR, 34 DM, and 49 controls. RNAseq was performed on primary tumors in 110 patients; with no difference in RNA profiles between patients with LRR, DM, or controls. DNA analysis on 57 primary tumors (17 LRR, 15 DM, and 25 controls) identified significantly more NF1 mutations and mitogen-activated protein kinase (MAPK) pathway gene mutations in patients with LRR (24%, 47%) and DM (27%, 40%) compared to controls (0%, 0%; p < 0.0001 and p = 0.0070, respectively). Three patients had matched primary vs. LRR samples, one patient had a gain of a NF1 mutation in the LRR. There was no significant difference between the groups for PTEN loss or cleaved caspase 3 expression. The mean percentage Ki 67 labeling index was higher in patients with LRR (29.2%) and DM (26%) vs. controls (14%, p = 0.0045). In summary, mutations in the MAPK pathway, specifically NF1, were associated with both LRR and DM, suggesting that alterations in MAPK signaling are associated with a more aggressive tumor phenotype. Validation of these associations in tissues from randomized trials may support targeted therapy to reduce breast cancer recurrence.

The Value of a Second Opinion for Breast Cancer Patients Referred to a National Cancer Institute (NCI)-Designated Cancer Center with a Multidisciplinary Breast Tumor Board.

BACKGROUND: This study aimed to investigate the changes in diagnosis after a second opinion for breast cancer patients from a multi-disciplinary tumor board (MTB) review at an National Cancer Institute (NCI)-designated cancer center. METHODS: A retrospective study analyzed patients with a breast cancer diagnosed at an outside institution who presented for a second opinion from August 2015 to March 2016 at the Medical University of South Carolina (MUSC). Radiology, pathology, and genetic testing reports from outside institutions were compared with reports generated after an MTB review and subsequent workup at MUSC. The second-opinion cases were categorized based on whether diagnostic variations were present or not. RESULTS: The review included 70 patients seeking second opinions, and 33 (47.1%) of these patients had additional radiologic images. A total of 30 additional biopsies were performed for 25 patients, with new cancers identified in 16 patients. Overall, 16 (22.8%) of the 70 of patients had additional cancers diagnosed. For 14 (20%) of the 70 patients, a second opinion led to a change in pathology interpretation. Genetic testing was performed for 11 patients (15.7%) who met the National Comprehensive Cancer Network (NCCN) guidelines for genetic testing, but none showed a mutation other than a variant of unknown significance. After a complete workup, 30 (42.8%) of the 70 patients had a change in diagnosis as a result of the MTB review. CONCLUSION: A review by an MTB at an NCI-designated cancer center changed the diagnosis for 43% of the patients who presented for a second opinion for breast cancer. The study findings support the conclusion that referral for a second opinion is beneficial and has a diagnostic impact for many patients.

TRAF3-interacting protein 3, a new oncotarget, promotes tumor growth in melanoma.

TRAF3-interacting protein 3 (TRAF3IP3) is expressed in the immune system and participates in cell maturation, tissue development, and immune response. In a previous study, we reported that TRAF3IP3 levels were substantially increased in the vasculature of breast cancer tissues, suggesting a proangiogenic role. In this study, we investigated TRAF3IP3 tumorigenic function. TRAF3IP3 protein was present in several cancer cell lines, with highest levels in melanoma. In addition, tumor microarray analysis on 23 primary melanoma and nine positive lymph nodes revealed that 70% of human primary melanoma and 66% of lymph node metastases were positive for TRAF3IP3. Importantly, TRAF3IP3 downregulation correlated with an 83% reduction of tumor growth in a subcutaneous xenograft mouse model (n=10, P=0.005). Immunohistochemistry analysis of the tumors revealed that TRAF3IP3-shRNA tumors had increased apoptosis (n=4, P<0.01) and reduced microvascular density (n=4, P<0.002). In addition, TRAF3IP3 downregulation in malignant endothelial cells reduced tube formation in a Matrigel tube formation assay. In melanoma cells, decreased levels of TRAF3IP3 were also associated with reduced viability (n=4, P=0.03) and proliferation (n=3, P=0.03), together with increased sensitivity to ultraviolet-induced apoptosis (n=4, P=0.0004). Furthermore, TRAF3IP3 downregulation correlated with increased amounts of interferon-γ. Interferon-γ inhibits tumor growth and angiogenesis, thus suggesting a new pathway for TRAF3IP3 in cancer. Collectively, the association of TRAF3IP3 with malignant properties of melanoma suggest a clinical potential for targeted therapy.

Frizzled-5: a high affinity receptor for secreted frizzled-related protein-2 activation of nuclear factor of activated T-cells c3 signaling to promote angiogenesis.

Secreted frizzled-related protein 2 (SFRP2) is a pro-angiogenic factor expressed in the vasculature of a wide variety of human tumors, and modulates angiogenesis via the calcineurin-dependent nuclear factor of activated T-cells cytoplasmic 3 (NFATc3) pathway in endothelial cells. However, until now, SFRP2 receptor for this pathway was unknown. In the present study, we first used amino acid alignments and molecular modeling to demonstrate that SFRP2 interaction with frizzled-5 (FZD5) is typical of Wnt/FZD family members. To confirm this interaction, we performed co-immunofluorescence, co-immunoprecipitation, and ELISA binding assays, which demonstrated SFRP2/FZD5 binding. Functional knock-down studies further revealed that FZD5 is necessary for SFRP2-induced tube formation and intracellular calcium flux in endothelial cells. Using protein analysis on endothelial cell nuclear extracts, we also discovered that FZD5 is required for SFRP2-induced activation of NFATc3. Our novel findings reveal that FZD5 is a receptor for SFRP2 and mediates SFRP2-induced angiogenesis via calcineurin/NFATc3 pathway in endothelial cells.

Optimizing ultrasound molecular imaging of secreted frizzled related protein 2 expression in angiosarcoma.

Secreted frizzled related protein 2 (SFRP2) is a tumor endothelial marker expressed in angiosarcoma. Previously, we showed ultrasound molecular imaging with SFRP2-targeted contrast increased average video pixel intensity (VI) of angiosarcoma vessels by 2.2 ± 0.6 VI versus streptavidin contrast. We hypothesized that redesigning our contrast agents would increase imaging performance. Improved molecular imaging reagents were created by combining NeutrAvidin™-functionalized microbubbles with biotinylated SFRP2 or IgY control antibodies. When angiosarcoma tumors in nude mice reached 8 mm, time-intensity, antibody loading, and microbubble dose experiments optimized molecular imaging. 10 minutes after injection, the control-subtracted time-intensity curve (TIC) for SFRP2-targeted contrast reached a maximum, after subtracting the contribution of free-flowing contrast. SFRP2 antibody-targeted VI was greater when contrast was formulated with 10-fold molar excess of maleimide-activated NeutrAvidin™ versus 3-fold (4.5 ± 0.18 vs. 0.32 ± 0.15, VI ± SEM, 5 x 106 dose, p < 0.001). Tumor vasculature returned greater average video pixel intensity using 5 x 107 versus 5 x 106 microbubbles (21.2 ± 2.5 vs. 4.5 ± 0.18, p = 0.0011). Specificity for tumor vasculature was confirmed by low VI for SFRP2-targeted, and control contrast in peri-tumoral vasculature (3.2 ± 0.52 vs. 1.6 ± 0.71, p = 0.92). After optimization, average video pixel intensity of tumor vasculature was 14.2 ± 3.0 VI units higher with SFRP2-targeted contrast versus IgY-targeted control (22.1 ± 2.5 vs. 7.9 ± 1.6, p < 0.001). After log decompression, 14.2 ΔVI was equal to ~70% higher signal, in arbitray acoustic units (AU), for SFRP2 versus IgY. This provided ~18- fold higher acoustic signal enhancement than provided previously by 2.2 ΔVI. Basing our targeted contrast on NeutrAvidin™-functionalized microbubbles, using IgY antibodies for our control contrast, and optimizing our imaging protocol significantly increased the SFRP2-specific signal returned from angiosarcoma vasculature, and may provide new opportunities for targeted molecular imaging.

Dysregulated connexin 43 in HER2-positive drug resistant breast cancer cells enhances proliferation and migration.

Connexin 43 (Cx43) is a gap junction protein whose function in the development of breast cancer and in breast cancer progression remains unclear. Evidence suggests that Cx43 (GJA1) mRNA and protein expression is altered in breast tumors. However, reports indicate both increased and decreased Cx43 levels in human breast cancer samples. Studies also suggest that loss of Cx43 regulated gap junction intercellular communication is a common feature of breast malignancies that potentially correlates with histological stage. Further evidence suggests that Cx43 (GJA1) mRNA expression is negatively correlated with HER2 positivity but a relationship between Cx43 and HER2 in breast cancer is not well defined. Therefore, in this study, we sought to evaluate the relationship between Cx43 activity, HER2, and drug resistance. Using HER2+ breast cancer cell lines that are sensitive or resistant to HER2 inhibitor, we evaluated Cx43 gap junction function. We found that Cx43 gap junction activity is completely lost in drug resistant HER2-positive (HER2+) breast cancer cells, whereas Cx43 gap junction activity can be restored by Cx43 overexpression in drug sensitive HER2+ cells. Moreover, the dysregulation of Cx43 resulted in increased tumorigenic and migratory capacity of the HER2+ drug resistant breast cancer cells.

G Protein Coupled Receptor Kinase 3 Regulates Breast Cancer Migration, Invasion, and Metastasis.

Triple negative breast cancer (TNBC) is a heterogeneous disease that has a poor prognosis and limited treatment options. Chemokine receptor interactions are important modulators of breast cancer metastasis; however, it is now recognized that quantitative surface expression of one important chemokine receptor, CXCR4, may not directly correlate with metastasis and that its functional activity in breast cancer may better inform tumor pathogenicity. G protein coupled receptor kinase 3 (GRK3) is a negative regulator of CXCR4 activity, and we show that GRK expression correlates with tumorigenicity, molecular subtype, and metastatic potential in human tumor microarray analysis. Using established human breast cancer cell lines and an immunocompetent in vivo mouse model, we further demonstrate that alterations in GRK3 expression levels in tumor cells directly affect migration and invasion in vitro and the establishment of distant metastasis in vivo. The effects of GRK3 modulation appear to be specific to chemokine-mediated migration behaviors without influencing tumor cell proliferation or survival. These data demonstrate that GRK3 dysregulation may play an important part in TNBC metastasis.

Immune Checkpoint Inhibition for Hypermutant Glioblastoma Multiforme Resulting From Germline Biallelic Mismatch Repair Deficiency.

PURPOSE: Recurrent glioblastoma multiforme (GBM) is incurable with current therapies. Biallelic mismatch repair deficiency (bMMRD) is a highly penetrant childhood cancer syndrome often resulting in GBM characterized by a high mutational burden. Evidence suggests that high mutation and neoantigen loads are associated with response to immune checkpoint inhibition. PATIENTS AND METHODS: We performed exome sequencing and neoantigen prediction on 37 bMMRD cancers and compared them with childhood and adult brain neoplasms. Neoantigen prediction bMMRD GBM was compared with responsive adult cancers from multiple tissues. Two siblings with recurrent multifocal bMMRD GBM were treated with the immune checkpoint inhibitor nivolumab. RESULTS: All malignant tumors (n = 32) were hypermutant. Although bMMRD brain tumors had the highest mutational load because of secondary polymerase mutations (mean, 17,740 ± standard deviation, 7,703), all other high-grade tumors were hypermutant (mean, 1,589 ± standard deviation, 1,043), similar to other cancers that responded favorably to immune checkpoint inhibitors. bMMRD GBM had a significantly higher mutational load than sporadic pediatric and adult gliomas and all other brain tumors (P < .001). bMMRD GBM harbored mean neoantigen loads seven to 16 times higher than those in immunoresponsive melanomas, lung cancers, or microsatellite-unstable GI cancers (P < .001). On the basis of these preclinical data, we treated two bMMRD siblings with recurrent multifocal GBM with the anti-programmed death-1 inhibitor nivolumab, which resulted in clinically significant responses and a profound radiologic response. CONCLUSION: This report of initial and durable responses of recurrent GBM to immune checkpoint inhibition may have implications for GBM in general and other hypermutant cancers arising from primary (genetic predisposition) or secondary MMRD.

Bilateral syringomatous adenomas of the nipple: Case report with immunohistochemical characterization of a rare tumor mimicking malignancy.

OBJECTIVES: Syringomatous adenoma of the nipple (SAN) is a rare benign infiltrative neoplasm that may be confused with low-grade breast carcinomas. We report the case of 71-year-old woman with a fungating left breast mass and subareolar calcifications in the contralateral breast. METHODS: Histomorphologic features were evaluated with standard H&E-stained sections. Both lesions were also evaluated by immunohistochemistry for further characterization. A literature review was performed to place the current case in context relative to previous observations. RESULTS: Both lesions were infiltrating syringomatous adenomas, confirmed by detailed immunohistochemical analyses, which also provided new evidence for the putative sweat duct origin for these tumors. The debated presence of myoepithelial cells in these lesions was newly and convincingly supported in this study by strong diffuse immunostaining of the outer cell layer of the tubules for smooth muscle myosin, cytokeratin 34ßE12, cytokeratin 5/6, and p63. Previous reports describing similar detailed immunophenotypical characterization of these uncommon tumors are lacking. CONCLUSIONS: To our knowledge, this case represents only the second reported patient with bilateral SAN and the first such case to be reported in the pathology literature.