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OBJECTIVES To evaluate the diagnostic performance of ultrasound (US)-guided fine-needle aspiration with optional core needle biopsy of head and neck lymph nodes and masses, with attention to differences between biopsy of treated squamous cell carcinoma (SCC) and biopsy of other lesions. METHODS Institutional Review Board approval was obtained, and the need for consent was waived for this retrospective study. All 861 US-guided biopsies of head and neck lymph nodes and masses performed between March 1, 2012, and May 16, 2016, were reviewed. RESULTS Of the 861 biopsies, 53 targeted SCC with residual masses after treatment. The biopsy procedures yielded benign or malignant pathologic results in 71.7% (38 of 53) of treated SCC and 90.7% (733 of 808) of all other lesions (P

Renal cell carcinoma (RCC) is a common and devastating disease characterized by a hypoxic microenvironment, epithelial-mesenchymal transition and potent resistance to therapy evidencing the presence of cancer stem cells (CSCs). Various CSC markers have been studied in RCC, but overall there is limited data on their role and most markers studied have been relatively nonspecific. Doublecortin-like kinase 1 (DCLK1) is a validated CSC marker in the gastrointestinal tract and evidence for an equivalent role in other cancers is accumulating. We used bioinformatics, immunohistochemistry, flow cytometry, spheroid self-renewal and chemoresistance assays in combination with overexpression and siRNA-knockdown to study the stem cell-supportive role of DCLK1 alternative splice variants (DCLK1 ASVs) in RCC. To target tumor cells expressing DCLK1 ASVs directly, we developed a novel monoclonal antibody (CBT-15) and delivered it systemically to RCC tumor xenografts. DCLK1 ASVs were overexpressed, enriched together with CSC markers and predictive of overall and recurrence-free survival in RCC patients. In vitro, DCLK1 ASVs were able to directly stimulate essential molecular and functional characteristics of renal CSCs including expression of aldehyde dehydrogenase, self-renewal and resistance to FDA-approved receptor tyrosine kinase and mTOR inhibitors, while targeted downregulation of DCLK1 reversed these characteristics. Finally, targeting DCLK1 ASV-positive cells with the novel CBT-15 monoclonal antibody blocked RCC tumorigenesis in vivo. These findings establish DCLK1 as a CSC marker with implications for therapy, disease progression and survival in RCC and demonstrate the therapeutic value of DCLK1-targeted monoclonal antibodies against renal CSCs.

Objectives Preferential use of fine-needle aspiration (FNA) versus core needle biopsy (CNB) for distinguishing benign from malignant major salivary gland tumors is highly debated. The main disadvantage of FNA is lower sensitivity, whereas arguments against CNB include use of a larger bore needle and greater risk of complications. The aim of this study is to evaluate our experience performing ultrasound-guided (UG) FNA with selective use of CNB based on preliminary cytopathology, and to determine whether our preoperative diagnostic approach is more sensitive and specific than FNA alone—and at least as sensitive and specific as CNB alone. Study Design Retrospective review of UG needle biopsy sampling of lesions arising in or around parotid and submandibular glands. Methods Ultrasounds of 141 needle biopsies were identified. Patient/lesion/needle biopsy characteristics, preliminary cytopathology, final pathology, imaging studies, and subsequent clinical course and treatment were documented. Results Needle biopsies performed according to our protocol provided results that guided clinical decision making in 125 of 135 cases, 92.6% (95% confidence interval [CI], 86.8%–96.4%) of the time. Using 41 cases that had histologic verification, sensitivity was 100% (95% CI, 79.6%–100%), and specificity was 92.3% (95% CI, 75.9%–97.9%) for detecting malignancy. We definitively characterized 120 lesions as benign (84) or malignant (36). Conclusion Preoperative needle biopsy diagnoses allowed clinical management to progress 92.6% of the time. The protocol of FNA with selective use of CNB may potentially reduce patient exposure to risks associated with CNB without the tradeoff of lower sensitivity seen with FNA. Level of Evidence 4. Laryngoscope, 2017

RET fusions have been found in lung adenocarcinoma, of which KIF5B–RET is the most prevalent. We established inducible KIF5B-RET transgenic mice and KIF5B–RET-dependent cell lines for preclinical modeling of KIF5B–RET-associated lung adenocarcinoma. Doxycycline-induced CCSP-rtTA/tetO-KIF5B-RET transgenic mice developed invasive lung adenocarcinoma with desmoplastic reaction. Tumors regressed upon suppression of KIF5B–RET expression. By culturing KIF5B–RET-dependent BaF3 (B/KR) cells with increasing concentrations of cabozantinib or vandetanib, we identified cabozantinib-resistant RETV804L mutation and vandetanib-resistant-RETG810A mutation. Among cabozantinib, lenvatinib, ponatinib, and vandetanib, ponatinib was identified as the most potent inhibitor against KIF5B–RET and its drug-resistant mutants. Interestingly, the vandetanib-resistant KIF5B-RETG810A mutant displayed gain-of-sensitivity (GOS) to ponatinib and lenvatinib. Treatment of doxycycline-induced CCSP-rtTA/tetO-KIF5B-RET bitransgenic mice with ponatinib effectively induced tumor regression. These results indicate that KIF5B-RET–associated lung tumors are addicted to the fusion oncogene and ponatinib is the most effective inhibitor for targeting KIF5B–RET in lung adenocarcinoma. Moreover, this study finds a novel vandetanib-resistant RETG810A mutation and identifies lenvatinib and ponatinib as the secondary drugs to overcome this vandetanib resistance mechanism. Mol Cancer Ther; 15(10); 2521–9. ©2016 AACR.

// Janani Panneerselvam 1, 4 , Akhil Srivastava 1, 4 , Ranganayaki Muralidharan 1, 4 , Qi Wang 1, 4 , Wei Zheng 1 , Lichao Zhao 1, 4 , Alshine Chen 3, 4 , Yan D. Zhao 3, 4 , Anupama Munshi 2, 4 , Rajagopal Ramesh 1, 4, 5 1 Department of Pathology, The University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA 2 Department of Radiation Oncology, The University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA 3 Department of Biostatistics and Epidemiology, The University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA 4 Stephenson Cancer Center, The University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA 5 Graduate Program in Biomedical Sciences, The University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA Correspondence to: Rajagopal Ramesh, email: rajagopal-ramesh@ouhsc.edu Keywords: IL-24, lung cancer, HMGA1, miR222-3p, metastasis Received: April 21, 2016 Accepted: August 24, 2016 Published: September 02, 2016 ABSTRACT Interleukin (IL)-24, a novel tumor suppressor/cytokine exhibits antitumor activity against a broad-spectrum of human cancer cells. In a recent study, we showed that IL-24 inhibited AKT in lung cancer cells. However, the molecular mechanism of AKT inhibition by IL-24 remains elusive. The high mobility group (HMG) A1 a member of the non-histone chromosomal proteins and commonly referred to as architectural transcription factor, regulates transcription of various genes involved in cell growth and survival. Overexpression of HMGA1 has been shown to be associated with tumor progression and metastasis in several cancers, including human lung cancer. A recent study demonstrated that HMGA1 activates AKT function by reducing the activity of the protein phosphatase, phosphatase 2A subunit B (PPP2R2A) via the oncogenic micro (mi) RNA-222. Based on this report we hypothesized that IL-24-mediated AKT inhibition involved the HMGA1/miR-222 axis. To test our hypothesis, in the present study we used a H1299 lung cancer cell line that expressed exogenous human IL-24 when induced with doxycycline (DOX). Induction of IL-24 expression in the tumor cells markedly reduced HMGA1 mRNA and protein levels. Using a mechanistic approach, we found that IL-24 reduced miR-222-3p and -5p levels, as determined by qRT-PCR. Associated with HMGA1 and miR-222 inhibition was a marked increase in PPP2R2A, with a concomitant decrease in phosphorylated AKT T308/S473 expression. SiRNA-mediated knockdown of HMGA1 in combination with IL-24 significantly reduced AKT T308/S473 protein expression and greatly reduced cell migration and invasion compared with individual treatments. Further combination of IL-24 and a miR-222-3p inhibitor significantly increased PPP2R2A expression. Our results demonstrate for the first time that IL-24 inhibits AKT via regulating the HMGA1/miR-222 signaling node in human lung cancer cells and acts as an effective tumor suppressor. Thus, a therapy combining IL-24 with HMGA1 siRNA or miR-222-3p inhibitor should present effective treatment of lung cancer.

Androgen ablation is the standard of care prescribed to patients with advanced or metastatic prostate cancer (PCa) to slow down disease progression. Unfortunately, a majority of PCa patients under androgen ablation progress to castration-resistant prostate cancer (CRPC). Several mechanisms including alternative intra-prostatic androgen production and androgen-independent androgen receptor (AR) activation have been proposed for CRPC progression. Aldo-keto reductase family 1 member C3 (AKR1C3), a multi-functional steroid metabolizing enzyme, is specifically expressed in the cytoplasm of PCa cells; and positive immunoreactivity of the type A γ-aminobutyric acid receptor (GABAAR), an ionotropic receptor and ligand-gated ion channel, is detected on the membrane of PCa cells. We studied a total of 72 radical prostatectomy cases by immunohistochemistry, and identified that 21 cases exhibited positive immunoreactivities for both AKR1C3 and GABAAR. In the dual positive cancer cases, AKR1C3 and GABAAR subunit α1 were either expressed in the same cells or in neighboring cells. Among several possible substrates, AKR1C3 reduces 5α-dihydrotesterone (DHT) to form 5α-androstane-3α, 17β-diol (3α-diol). 3α-diol is a neurosteroid that acts as a positive allosteric modulator of the GABAAR in the central nervous system (CNS). We examined the hypothesis that 3α-diol-regulated pathological effects in the prostate are GABAAR-dependent, but are independent of the AR. In GABAAR-positive, AR-negative human PCa PC-3 cells, 3α-diol significantly stimulated cell growth in culture and the in ovo chorioallantoic membrane (CAM) xenograft model. 3α-diol also up-regulated expression of the epidermal growth factor (EGF) family of growth factors and activation of EGF receptor (EGFR) and Src as measured by quantitative polymerase chain reaction and immunoblotting, respectively. Inclusion of GABAAR antagonists reversed 3α-diol-stimulated tumor cell growth, expression of EGF family members, and activation of EGFR and Src to the level observed in untreated cells. Results from the present study suggest that 3α-diol may act as an alternative intra-prostatic neurosteroid that activates AR-independent PCa progression. The involvement of AKR1C3-mediated steroid metabolisms in modulating GABAAR activation and promoting PCa progression requires continued studies.

BACKGROUND Recent proposals to lengthen the interval in cervical cancer screening highlight the importance of the accurate interpretation of screening tests. Tumor debris present in Papanicolaou (Pap) tests from women with invasive cancer is known to hamper interpretation. The current study evaluated limiting factors in Pap tests from women with invasive cervical cancer. METHODS A total of 3003 women with the spectrum of cervical lesions who had ThinPrep (Hologic Inc, Marlborough, Mass) Pap and human papillomavirus (HPV) genotyping tests performed were grouped by their most severe histologic diagnosis. Cytologic and HPV results were analyzed by cross-sectional analysis. RESULTS The unsatisfactory rate of cytology specimens from patients with cancer (3.1%) was significantly higher than those from patients with cervical intraepithelial neoplasia of type 3 or less (0.8%) (P 30 years can help to identify high-risk women with unsatisfactory Pap tests. Cancer (Cancer Cytopathol) 2014;122:694–701. © 2014 American Cancer Society.

BACKGROUND: To evaluate the efficacy and the limitation of fine-needle aspiration (FNA) biopsy in thyroid bed lesions, a retrospective review was performed of the medical records of thyroid cancer patients who underwent ultrasound-guided FNA biopsy of the thyroid bed at The University of Texas MD Anderson Cancer Center over a 5-year period. METHODS: Data were reviewed on 220 FNA biopsies taken from thyroid bed lesions in 195 patients who had undergone thyroidectomy for thyroid carcinoma. Thyroid bed FNA results were compared with clinical follow-up, including neck dissection results. RESULTS: Recurrent carcinoma was diagnosed by FNA biopsy in 139 of 220 (63%) cases. Neck dissections were performed for 112 sites identified by FNA biopsies, and recurrent carcinoma was confirmed in 110 sites. The concordance between positive and/or suspicious FNA diagnosis and positive neck dissection results was 98% (118 of 120 cases). A false-positive FNA occurred in one patient with follicular thyroid carcinoma. The other discrepancy was attributed to failure to remove the lesion by neck dissection. The diagnostic accuracy of thyroid bed FNA was 100% in papillary and medullary thyroid carcinoma and 93% in follicular thyroid carcinoma. Suspicious and rare false-negative FNA results were attributed to low cellularity and lack of characteristic cytomorphologic features of thyroid carcinoma. CONCLUSIONS: Ultrasound-guided thyroid bed FNA biopsy is accurate and efficient in triaging patients who require post-thyroidectomy follow-up for recurrent thyroid carcinoma. Caution should be taken in the interpretation of FNA specimens that have low cellularity and lack characteristic cytologic features of thyroid carcinoma. Cancer (Cancer Cytopathol) 2013. © 2012 American Cancer Society.

Shear-resistant adhesion and extravasation of disseminated cancer cells at the target organ is a crucial step in hematogenous metastasis. We found that the vascular adhesion molecule E-selectin preferentially promoted the shear-resistant adhesion and transendothelial migration of the estrogen receptor (ER)–/CD44+ hormone-independent breast cancer cells, but not of the ER+/CD44-/low hormone-dependent breast cancer cells. Coincidentally, CD44+ breast cancer cells were abundant in metastatic lung and brain lesions in ER– breast cancer, suggesting that E-selectin supports hematogenous metastasis of ER–/CD44+ breast cancer. In an attempt to prevent hematogenous metastasis through the inhibition of a shear-resistant adhesion of CD44+ cancer cells to E-selectin-expressing blood vessels on the premetastatic niche, an E-selectin targeted aptamer (ESTA) was developed. We demonstrated that a single intravenous injection of ESTA reduced metastases to a baseline level in both syngeneic and xenogeneic forced breast cancer metastasis models without relocating the site of metastasis. The effect of ESTA was absent in E-selectin knockout mice, suggesting that E-selectin is a molecular target of ESTA. Our data highlight the potential application of an E-selectin antagonist for the prevention of hematogenous metastasis of ER–/CD44+ breast cancer.

Doublecortin-like kinase 1 (DCLK1) is a putative pancreatic stem cell marker and is upregulated in pancreatic cancer, colorectal cancer, and many other solid tumors. It marks tumor stem cells in mouse models of intestinal neoplasia. Here we sought to determine whether DCLK1 protein can be detected in the bloodstream and if its levels in archived serum samples could be quantitatively assessed in pancreatic cancer patients. DCLK1 specific ELISA, western blotting, and immunohistochemical analyses were used to determine expression levels in the serum and staining intensity in archived tumor tissues of pancreatic ductal adenocarcinoma (PDAC) patients and in pancreatic cancer mouse models. DCLK1 levels in the serum were elevated in early stages of PDAC (stages I and II) compared to healthy volunteers (normal controls). No differences were observed between stages III/IV and normal controls. In resected surgical tissues, DCLK1 expression intensity in the stromal cells was significantly higher than that observed in tumor epithelial cells. Circulating tumor cells were isolated from KPCY mice and approximately 52% of these cells were positive for Dclk1 staining. Dclk1 levels in the serum of KPC mice were also elevated. We have previously demonstrated that DCLK1 plays a potential role in regulating epithelial mesenchymal transition (EMT). Given the increasingly recognized role of EMT derived stem cells in cancer progression and metastasis, we hypothesize that DCLK1 may contribute to the metastatic process. Taken together, our results suggest that DCLK1 serum levels and DCLK1 positive circulating tumor cells should be further assessed for their potential diagnostic and prognostic significance.

// Nathaniel Weygant 1 , Dongfeng Qu 1,2,3 , Randal May 1,2 , Ryan M. Tierney 1 , William L. Berry 4 , Lichao Zhao 5 , Shweta Agarwal 5 , Parthasarathy Chandrakesan 1 , Harisha R. Chinthalapally 1 , Nicholas T. Murphy 1 , James D. Li 1 , Sripathi M. Sureban 1,2,3 , Michael J. Schlosser 6 , James J. Tomasek 4 and Courtney W. Houchen 1,2,3,6 1 Department of Medicine, University of Oklahoma Health Sciences Center, Oklahoma, OK 2 Department of Veterans Affairs Medical Center, Oklahoma, OK 3 Peggy and Charles Stephenson Oklahoma Cancer Center, Oklahoma, OK 4 Department of Cell Biology, University of Oklahoma Health Sciences Center, Oklahoma, OK 5 Department of Pathology, University of Oklahoma Health Sciences Center, Oklahoma, OK 6 COARE Biotechnology, Oklahoma, OK Correspondence: Courtney W. Houchen, email: // Keywords : DCLK1, Renal Carcinoma, Tumor Stem Cell, Focal Adhesion, EMT Received : October 31, 2014 Accepted : December 10, 2014 Published : December 11, 2014 Abstract Renal clear cell carcinoma (RCC) is the most common type of kidney cancer and the 8 th most common cancer overall in the US. RCC survival rates drop precipitously with regional and distant spread and recent studies have demonstrated that RCC presents an epithelial-mesenchymal transition (EMT) phenotype linked to increased recurrence and decreased survival. EMT is a key characteristic of tumor stem cells (TSCs) along with chemo-resistance and radio-resistance, which are also phenotypic of RCC. Targeting these factors is key to increasing the survival of RCC patients. Doublecortin-like kinase 1 (DCLK1) marks TSCs in pancreatic and colorectal cancer and regulates EMT and stemness. Analysis of the Cancer Genome Atlas’ RCC dataset revealed that DCLK1 is overexpressed and dysregulated on the mRNA and epigenetic level in more than 93% of RCC tumors relative to adjacent normal tissue. Immunohistochemistry using α-DCLK1 antibody confirmed overexpression and demonstrated a major increase in immunoreactivity in stage II-III tumors compared to normal kidney and stage I tumors. Small-interfering RNA (siRNA) mediated knockdown of DCLK1 resulted in decreased expression of EMT and pluripotency factors and significantly reduced invasion, migration, focal adhesion, drug-resistance, and clonogenic capacity. These findings suggest that DCLK1 is a novel, overexpressed factor in RCC progression that may be targeted to suppress EMT, metastasis, and stemness in early-stage and advanced RCC to increase patient survival. Moreover, the possibility that DCLK1 may mark a population of tumor stem-like cells in RCC should be further investigated in light of these findings.

Background: High mobility group A1 (HMGA1), a member of the non-histone chromosomal proteins and commonly referred to as architectural transcription factor, regulates transcription of various genes involved in cell growth and survival. Overexpression of HMGA1 has been shown to be associated with tumor progression and metastasis in several cancers, including human lung cancer. A recent study demonstrated that HMGA1 activates AKT function by reducing the activity of the protein phosphatase, phosphatase 2A subunitB (PPPR2A) via the oncogenic micro (mi) RNA-222. We demonstrated that interleukin (IL)-24, a novel tumor suppressor/cytokine, inhibited AKT in lung cancer cells. However, the molecular mechanism of AKT inhibition by IL-24 remains elusive. Aim: To determine the molecular mechanism of IL-24-mediated AKT inhibition involved the HMGA1/miR-222 axis. Methods: Human H1299 lung tumor cell line was stably transfected with a tetracycline-inducible plasmid vector carrying the IL-24. After the induced expression of IL-24 protein, expression levels of HMGA1 and its downstream molecular mechanisms were analyzed at the RNA and protein levels in lung cancer cell lines. The inhibitory effect of IL-24 on HMG A1/miR222 /AKT axis in the lung cancer cells is determined by RT-qPCR, western blot, reporter assay, and immunocytochemistry. Mechanistic approaches on overexpression and knockdown of HMGA1 and or miR-222 were utilized and the consequences of its inhibition/overexpression were analyzed on HMGA1/miR222 /AKT signaling axis and in vitro migration and invasion. Results: Upon induction of IL-24 expression in the H1299 lung tumor cells, we observed a marked reduction in HMGA1protein and mRNA levels. Using a mechanistic approach, we found that IL-24 reduced miR-222-3p and -5p levels, as determined by qRT-PCR. Associated with HMGA1 and miR-222 inhibition was a marked increase in PPP2R2A, with a concomitant decrease in phosphorylated AKTT308/S473 expression. SiRNA-mediated knockdown of HMGA1in combination with IL-24 significantly reduced AKT T308/S473 protein expression and greatly reduced cell migration and invasion compared with individual treatments. Further combination of IL-24 and a miR-222-3p inhibitor significantly increased PPP2R2A expression. Conclusion: Our results demonstrate for the first time that IL-24 inhibits AKT via regulating the HMGA1/miR-222 signaling node in human lung cancer cells and acts as an effective tumor suppressor. HMGA1 should present a novel target for the effective treatment of lung cancer. Citation Format: Janani Panneerselvam, Akhil Srivastava, Ranganayaki Muralidharan, Qi Wang, Wei Zheng, Lichao Zhao, Allshine Chen, Yan Zhao, Anupama Munshi, Rajagopal Ramesh. IL-24 modulates the high mobility group (HMG) A1/miR222 /AKT signaling in lung cancer cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3738.

HuR, an mRNA binding protein regulates the stability of many oncoproteins associated with cell survival, proliferation, migration and angiogenesis. HuR overexpression is a marker for poor prognosis in patients diagnosed with cancer of lung, ovary, breast and colon. We hypothesized that the silencing of HuR using small interfering RNA (siRNA) could be a promising approach for lung cancer therapy. To test our hypothesis, we developed a tumor-targeted nanoparticle (NP) system that is targeted to transferrin receptor (TfR) for delivering HuRsiRNA (HuR-TfNP) in human lung cancer cells. Human lung cancer cells (A549, HCC827) and normal lung fibroblast (MRC-9) cell lines expressing varying levels of TfR were used in the present study. TfR expression was highest in A549, moderate in HCC827, and low to undetectable in MRC9 cells. In vitro studies demonstrated enhanced uptake of Tf-NP (51%) in TfR overexpressing A549 cells, compared to the non-targeted NP. Specificity studies using desferrioxamine (DFO; 100 µM), a stimulator of TfR, showed a two-fold increased uptake of Tf-NP whereas blocking TfR with exogenous transferrin (1 µg/well) reduced the uptake by 3 fold in A549 cells. Further, HuR-TfNP treatment reduced HuR expression and significantly suppressed cell proliferation at 24h and 48h compared to control siRNA containing NP (C-Tf-NP) in tumor cells but not in normal cells. Greatest inhibition was observed in A549 cells (23% and 30% inhibition at 24 and 48 h respectively) compared to 15% and 25% in HCC827. In MRC-9 cells, only 4% inhibition was observed. HuR-TfNP induced G1 cell-cycle arrest in tumor cells that correlated with marked reduction in Cyclin D1, and Cyclin E protein expression. Further, tumor cell migration and invasion was significantly inhibited in HuR-TfNP treated tumor cells compared to C-TfNP treatment (p

Despite the success of CD20 antibody rituximab in immunotherapy, acquired resistance is one of the prime obstacles for the successful treatment of B-cell malignancies. There is an urgent need to intensify efforts against resistance in cancer treatment. Growing evidence indicated that lysosomes may form an “Achilles heel” for cancer cells by sensitizing them to death pathways. Here, we uncover an important role of CD20 in initiation of ceramide/lysosomal membrane permeabilization (LMP)-mediated cell death, showing that colocalization of CD20-TNFR1 after type II CD20 antibody ligation can stimulate de novo ceramide synthesis by ceramide synthase and consequently induce remarkable lysosomal permeabilization (LMP) and lysosome-mediated cell death. Further studies show that the potent lysosome-mediated cell death induced by CD20 antibodies exhibits a profound killing effect against both rituximab-sensitive and -resistant (RR) lymphoma. Furthermore, engineering of rituximab by introducing a point mutation endows it with the ability to induce potent ceramide/LMP-mediated cell death in both RR lymphoma and primary B-cell malignancies from patients with rituximab-refractory, suggesting the potential clinical application to combat rituximab resistance.

A 66-year-old man presented with diffuse slate-gray skin discoloration and multiorgan failure. Diagnostic workup showed disseminated bone and visceral lesions and positive Sporothrix serology. He was treated with antifungals for disseminated sporotrichosis but he died shortly after. Autopsy revealed metastatic melanoma with diffuse melanosis and no Sporothrix infection.

Superior Vena Cava-Aorta Ganglionated Plexus. Introduction: The mechanism underlying spontaneous rapid superior vena cava (SVC) firing that initiates atrial fibrillation (AF) remains poorly understood. We investigated the role of the SVC-aorta-ganglionated plexus (SVC-Ao-GP) in AF initiated by rapid firing from the SVC. Methods and Results: In 42 dogs, a circular catheter was positioned above the SVC-atrial junction. Multielectrode catheters were sutured on atria, atrial appendages and pulmonary veins. The effective refractory period (ERP) and window of vulnerability (WOV) for AF were measured at all sites in the baseline state, during cervical vagosympathetic trunk stimulation and during SVC-Ao-GP stimulation, before and after SVC-Ao-GP ablation. AF inducibility was also assessed by delivering high-frequency stimulation (HFS) within myocardial refractory period to the SVC before and after SVC-Ao-GP ablation. HFS applied to the SVC-Ao-GP slowed the sinus rate and/or atrioventricular conduction. HFS of the SVC-Ao-GP induced more significant shortening of ERP and a greater increase in WOV at the SVC than other sites. Ablation of the SVC-Ao-GP significantly increased the baseline ERP and decreased the baseline WOV only at the SVC. AF induced at the SVC by HFS during refractoriness was eliminated by ablation of the SVC-Ao-GP but was not altered by ablation of the 4 major atrial GP. Direct injection of acetylcholine into the SVC-Ao-GP initiated rapid firing from the SVC in every case. Conclusions: The SVC-Ao-GP preferentially modulates the electrophysiological function of the SVC sleeves and may contribute to rapid firing from the SVC. (J Cardiovasc Electrophysiol, Vol. 21, pp. 1392-1399, December 2010)

Background— The mechanism(s) for acute changes in electrophysiological properties of the atria during rapid pacing induced atrial fibrillation (AF) is not completely understood. We sought to evaluate the contribution of the intrinsic cardiac autonomic nervous system in acute atrial electrical remodeling and AF induced by 6-hour rapid atrial pacing. Methods and Results— Continuous rapid pacing (1200 bpm, 2� threshold [TH]) was performed at the left atrial appendage. Group 1 (n=7) underwent 6-hour pacing immediately followed by ganglionated plexi (GP) ablation; group 2 (n=7) underwent GP ablation immediately followed by 6-hour pacing; and group 3 (n=4) underwent administration of autonomic blockers, atropine (1 mg/kg), and propranolol (0.6 mg/kg) immediately followed by 6-hour pacing. The effective refractory period (ERP) and window of vulnerability (WOV, in milliseconds), ie, the difference between the longest and the shortest coupling interval of the premature stimulus that induced AF, were measured at 2�TH and 10�TH at the left atrium, right atrium, and pulmonary veins every hour before and after GP ablation or autonomic blockade. In group 1, ERP was markedly shortened in the first 2 hours and then stabilized both at 2�TH and 10�TH; however, WOV was progressively widened throughout the 6-hour period. After GP ablation, ERP was significantly longer than before ablation and AF could not be induced (WOV=0) at either 2�TH or 10�TH. In groups 2 and 3, rapid atrial pacing failed to shorten the ERP. AF could not be induced in 6 of 7 dogs in group 2 and all 4 dogs in group 3 during the 6-hour pacing period. Conclusion— The intrinsic cardiac autonomic nervous system plays a crucial role in the acute stages of atrial electrical remodeling induced by rapid atrial pacing.

Renal clear cell carcinoma (RCC) is the most common type of kidney cancer and the 8th most common cancer overall in the US. RCC survival rates drop precipitously with regional and distant spread and recent studies have demonstrated that RCC presents an epithelial-mesenchymal transition (EMT) phenotype linked to increased recurrence and decreased survival. EMT is a key characteristic of tumor stem cells (TSCs) along with chemo-resistance and radio-resistance, which are also phenotypic of RCC. Targeting these factors is key to increasing the survival of RCC patients. Doublecortin-like kinase 1 (DCLK1) marks TSCs in pancreatic and colorectal cancer and regulates EMT and stemness. Analysis of the Cancer Genome Atlas’ RCC dataset revealed that DCLK1 is overexpressed and dysregulated on the mRNA and epigenetic level in more than 93% of RCC tumors relative to adjacent normal tissue. Immunohistochemistry using α-DCLK1 antibody confirmed overexpression and demonstrated a major increase in immunoreactivity in stage II-III tumors compared to normal kidney and stage I tumors. Small-interfering RNA (siRNA) mediated knockdown of DCLK1 resulted in decreased expression of EMT and pluripotency factors and significantly reduced invasion, migration, focal adhesion, drug-resistance, and clonogenic capacity. These findings suggest that DCLK1 is a novel, overexpressed factor in RCC progression that may be targeted to suppress EMT, metastasis, and stemness in early-stage and advanced RCC to increase patient survival. Moreover, the possibility that DCLK1 may mark a population of tumor stem-like cells in RCC should be further investigated in light of these findings. Citation Format: Nathaniel Weygant, Dongfeng Qu, Randal May, Ryan M. Tierney, William L. Berry, Lichao Zhao, Shweta Agarwal, Parthasarathy Chandrakesan, Sripathi M. Sureban, Michael J. Schlosser, James J. Tomasek, Courtney W. Houchen. DCLK1 is a broadly dysregulated target against epithelial-mesenchymal transition, focal adhesion, and stemness in clear cell renal carcinoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4374. doi:10.1158/1538-7445.AM2015-4374

Although trastuzumab has succeeded in breast cancer treatment, acquired resistance is one of the prime obstacles for breast cancer therapies. There is an urgent need to develop novel HER2 antibodies against trastuzumab resistance. Here, we first rational designed avidity-imporved trastuzumab and pertuzumab variants, and explored the correlation between the binding avidity improvement and their antitumor activities. After characterization of a pertuzumab variant L56TY with potent antitumor activities, a bispecific immunoglobulin G-like CrossMab (Tras-Permut CrossMab) was generated from trastuzumab and binding avidity-improved pertuzumab variant L56TY. Although, the antitumor efficacy of trastuzumab was not enhanced by improving its binding avidity, binding avidity improvement could significantly increase the anti-proliferative and antibody-dependent cellular cytotoxicity (ADCC) activities of pertuzumab. Further studies showed that Tras-Permut CrossMab exhibited exceptional high efficiency to inhibit the progression of trastuzumab-resistant breast cancer. Notably, we found that calreticulin (CRT) exposure induced by Tras-Permut CrossMab was essential for induction of tumor-specific T cell immunity against tumor recurrence. These data indicated that simultaneous blockade of HER2 protein by Tras-Permut CrossMab could trigger CRT exposure and subsequently induce potent tumor-specific T cell immunity, suggesting it could be a promising therapeutic strategy against trastuzumab resistance.