Differential diagnosis of parkinsonism based on deep metabolic imaging indices.

The clinical presentations of early idiopathic Parkinson's disease (PD) substantially overlap with those of atypical parkinsonian syndromes like multiple system atrophy (MSA) and progressive supranuclear palsy (PSP). This study aimed to develop metabolic imaging indices based on deep learning to support the differential diagnosis of these conditions. Methods: A benchmark Huashan parkinsonian PET imaging (HPPI, China) database including 1275 parkinsonian patients and 863 non-parkinsonian subjects with 18F-FDG PET images was established to support artificial intelligence development. A 3D deep convolutional neural network was developed to extract deep metabolic imaging (DMI) indices, which was blindly evaluated in an independent cohort with longitudinal follow-up from the HPPI, and an external German cohort of 90 parkinsonian patients with different imaging acquisition protocols. Results: The proposed DMI indices had less ambiguity space in the differential diagnosis. They achieved sensitivities of 98.1%, 88.5%, and 84.5%, and specificities of 90.0%, 99.2%, and 97.8% for the diagnosis of PD, MSA, and PSP in the blind test cohort. In the German cohort, They resulted in sensitivities of 94.1%, 82.4%, 82.1%, and specificities of 84.0%, 99.9%, 94.1% respectively. Employing the PET scans independently achieved comparable performance to the integration of demographic and clinical information into the DMI indices. Conclusion: The DMI indices developed on the HPPI database show potential to provide an early and accurate differential diagnosis for parkinsonism and is robust when dealing with discrepancies between populations and imaging acquisitions

CD1d-Expressing Breast Cancer Cells Modulate NKT Cell-Mediated Antitumor Immunity in a Murine Model of Breast Cancer Metastasis

Tumor tolerance and immune suppression remain formidable obstacles to the efficacy of immunotherapies that harness the immune system to eradicate breast cancer. A novel syngeneic mouse model of breast cancer metastasis was developed in our lab to investigate mechanisms of immune regulation of breast cancer. Comparative analysis of low-metastatic vs. highly metastatic tumor cells isolated from these mice revealed several important genetic alterations related to immune control of cancer, including a significant downregulation of cd1d1 in the highly metastatic tumor cells. The cd1d1 gene in mice encodes the MHC class I-like molecule CD1d, which presents glycolipid antigens to a specialized subset of T cells known as natural killer T (NKT) cells. We hypothesize that breast cancer cells, through downregulation of CD1d and subsequent evasion of NKT-mediated antitumor immunity, gain increased potential for metastatic tumor progression.In this study, we demonstrate in a mouse model of breast cancer metastasis that tumor downregulation of CD1d inhibits iNKT-mediated antitumor immunity and promotes metastatic breast cancer progression in a CD1d-dependent manner in vitro and in vivo. Using NKT-deficient transgenic mouse models, we demonstrate important differences between type I and type II NKT cells in their ability to regulate antitumor immunity of CD1d-expressing breast tumors.The results of this study emphasize the importance of determining the CD1d expression status of the tumor when tailoring NKT-based immunotherapies for the prevention and treatment of metastatic breast cancer

New and Emerging Targeted Therapies for Advanced Breast Cancer

In the United States, breast cancer is among the most frequently diagnosed cancers in women. Breast cancer is classified into four major subtypes: human epidermal growth factor receptor 2 (HER2), Luminal-A, Luminal-B, and Basal-like or triple-negative, based on histopathological criteria including the expression of hormone receptors (estrogen receptor and/or progesterone receptor) and/or HER2. Primary breast cancer treatments can include surgery, radiation therapy, systemic chemotherapy, endocrine therapy, and/or targeted therapy. Endocrine therapy has been shown to be effective in hormone receptor-positive breast cancers and is a common choice for adjuvant therapy. However, due to the aggressive nature of triple-negative breast cancer, targeted therapy is becoming a noteworthy area of research in the search for non-endocrine-targets in breast cancer. In addition to HER2-targeted therapy, other emerging therapies include immunotherapy and targeted therapy against critical checkpoints and/or pathways in cell growth. This review summarizes novel targeted breast cancer treatments and explores the possible implications of combination therapy

Regulation of COX2 expression in mouse mammary tumor cells controls bone metastasis and PGE2-induction of regulatory T cell migration.

BACKGROUND: The targeting of the immune system through immunotherapies to prevent tumor tolerance and immune suppression are at the front lines of breast cancer treatment and research. Human and laboratory studies have attributed breast cancer progression and metastasis to secondary organs such as the bone, to a number of factors, including elevated levels of prostaglandin E2 (PGE2) and the enzyme responsible for its production, cyclooxygenase 2 (COX2). Due to the strong connection of COX2 with immune function, we focused on understanding how variance in COX2 expression manipulates the immune profile in a syngeneic, and immune-competent, mouse model of breast cancer. Though there have been correlative findings linking elevated levels of COX2 and Tregs in other cancer models, we sought to elucidate the mechanisms by which these immuno-suppressive cells are recruited to breast tumor and the means by which they promote tumor tolerance. METHODOLOGY/PRINCIPAL FINDINGS: To elucidate the mechanisms by which exacerbated COX2 expression potentiates metastasis we genetically manipulated non-metastatic mammary tumor cells (TM40D) to over-express COX2 (TM40D-COX2). Over-expression of COX2 in this mouse breast cancer model resulted in an increase in bone metastasis (an observation that was ablated following suppression of COX2 expression) in addition to an exacerbated Treg recruitment in the primary tumor. Interestingly, other immune-suppressive leukocytes, such as myeloid derived suppressor cells, were not altered in the primary tumor or the circulation. Elevated levels of PGE2 by tumor cells can directly recruit CD4+CD25+ cells through interactions with their EP2 and/or EP4 receptors, an effect that was blocked using anti-PGE2 antibody. Furthermore, increased Treg recruitment to the primary tumor contributed to the greater levels of apoptotic CD8+ T cells in the TM40D-COX2 tumors. CONCLUSION/SIGNIFICANCE: Due to the systemic effects of COX2 inhibitors, we propose targeting specific EP receptors as therapeutic interventions to breast cancer progression

PGE2 directly involved in inducing Treg migration.

<p>CD4+ CD25+ Tregs were enriched by magnetic column (MACS) from naïve thymus and lymph nodes (A). Treg migration was measured using media alone, cell-free supernatant collected from TM40D, TM40D-COX2, or TM40D-MB cells (B). Additionally, the role of PGE2 was assessed by anti-PGE2 blocking antibody added to TM40D-COX2 media. Purified GFP-FoxP3+ cells were isolated from spleen following CD4+ T cell enrichment (C) and analyzed for expression of the PGE2 receptors EP1, EP2, EP3, or EP4 via RT-PCR with GAPDH as a control (D). #p<0.05, *p<0.01 compared to Media alone, N = 3 samples per group.</p

Breast cancer rate of metastasis to bone is not correlated to <i>in vitro</i> or <i>in vivo</i> growth rate.

<p>A. COX2 expression, measured by quantitative RT-PCR, by TM40D, TM40D-MB, TM40D-COX2, and TM40D-MB-shCOX2 cells. B. Metastasis to bone from primary TM40D, TM40D-COX2, highly aggressive TM40D-MB, or TM40D-MB-shCOX2 cells which also over-express COX2 was assessed at maximum tumor size by homogenizing both hind leg bones followed by in vitro growth under selection media and complemented with genomic DNA PCR for GFP. C. MTT cell proliferation assay performed on TM40D and TM40D-COX2 cells to measure <i>in vitro</i> growth. D. Mice were implanted with either low tumorigenic/COX2 expressing TM40D cells or COX2 over-expressing TM40D-COX2 cells into fourth mammary fat pad and monitored for <i>in vivo</i> tumor growth rates. N = 1/10, 4/8, 5/9, and 0/5 for TM40D, TM40D-COX2, TM40D-MB, and TM40D-MB-shCOX2, respectively. *p<0.05 compared to TM40D and TM40D-MB-shCOX2.</p

Tumor immune profile reveals elevated Tregs levels.

<p>TM40D (grey bar) or COX2 over-expressing TM40D-COX2 (black bar) cells were implanted in the fourth mammary pad of Balb/c mice. At maximum tumor volume tumors were assessed for (A) CD4+ and CD8+ T-cells, (B) immature monocytes (CD11b+ F4/80+ Ly6g+), tumor associate macrophages (CD11b+ F4/80+ Ly6g−), G MDSCs (CD11b+ Ly6c^low Ly6g+) and M MDSCs (CD11b+ Ly6C^hi Ly6g−). Tregs (FoxP3+CD4+ CD25+) were assessed via flow cytometry of spleens from TM40D and TM40D-COX2 challenged mice (C). Quantitative analysis of Tregs observed in the tumor of tumor challenged mice (D). Representative Treg levels in the primary tumor of mice challenged with TM40D or TM40D-COX2 mammary tumor cells (E). Quantitative analysis of Tregs observed in the spleen of TM40D versus TM40D-COX2 challenged mice (F). p<0.05 compared to TM40D group, n = 4–5 animals per group. *p<0.05 compared to TM40D.</p

TM40D-COX2 tumors have increased apoptotic CD8+ T cells.

<p>Histological sections of TM40D and TM40D-COX2 tumors were stained with AlexaFluor 594 for CD8 (RED) AlexaFluor 488 for Cleaved Caspase 3 (ClCasp3+) (GREEN), and Dapi for nucleus (BLUE) (A and B, respectively). C. Quantitative analysis of panels A and B measuring the percent of CD8+ cells that have cleaved Caspase 3 co-localized to the cell. N = 3 samples per group, 8–10 sections per sample. *p<0.01 compared to TM40D.</p