Sidharth Mahapatra

hPaf1/PD2 interacts with OCT3/4 to promote self-renewal of ovarian cancer stem cells

Cancer stem cells (CSCs), which mediate drug resistance and disease recurrence in several cancers, are therapeutically relevant to ovarian cancer (OC), wherein approximately 80% of patients manifest with tumor recurrence. While there are several markers for ovarian CSCs (OCSCs), the mechanism for their self-renewal maintenance by unique driver/markers is poorly understood. Here, we evaluated the role of hPaf1/PD2, a core component of RNA Polymerase II-Associated Factor (PAF) complex, in self-renewal of OCSCs through marker and functional analyses, including CRISPR/Cas9-silencing of hPaf1/PD2 in OCSCs and provided a possible mechanism for maintenance of OCSCs. Expression of hPaf1/PD2 showed moderate to intense staining in 32.4% of human OC tissues, whereas 67.6% demonstrated basal expression by immunohistochemistry analysis, implying that the minor proportion of cells overexpressing hPaf1/PD2 could be putative OCSCs. Isolated OCSCs showed higher expression of hPaf1/PD2 along with established CSC and self-renewal markers. Knockdown of hPaf1/PD2 in OCSCs resulted in a significant downregulation of CSC and self-renewal markers, and impairment of in vitro tumor sphere (P < 0.05) and colony formation (P = 0.013). Co-immunoprecipitation revealed that OCT3/4 specifically interacts with hPaf1/PD2, and not with other PAF components (Ctr9, Leo1, Parafibromin) in OCSCs, suggesting a complex-independent role for hPaf1/PD2 in OCSC maintenance. Moreover, there was a significant overexpression and co-localization of hPaf1/PD2 with OCT3/4 in OC tissues compared to normal ovary tissues. Our results indicate that hPaf1/PD2 is overexpressed in OCSCs and maintains the self-renewal of OCSCs through its interaction with OCT3/4; thus, hPaf1/PD2 may be a potential therapeutic target to overcome tumor relapse in OC.

MUC4 mucin- a therapeutic target for pancreatic ductal adenocarcinoma

ABSTRACT Introduction: Pancreatic cancer (PC) is characterized by mucin overexpression. MUC4 is the most differentially overexpressed membrane-bound mucin that plays a functional role in disease progression and therapy resistance. Area covered: We describe the clinicopathological significance of MUC4, summarize mechanisms contributing to its deregulated expression, review preclinical studies aimed at inhibiting MUC4, and discuss how MUC4 overexpression provides opportunities for developing targeted therapies. Finally, we discuss the challenges for developing MUC4-based therapeutics, and identify areas where efforts should be directed to effectively exploit MUC4 as a therapeutic target for PC. Expert opinion: Studies demonstrating that abrogation of MUC4 expression reduces proliferation and metastasis of PC cells and enhances sensitivity to therapeutic agents affirm its utility as a therapeutic target. Emerging evidence also supports the suitability of MUC4 as a potential immunotherapy target. However, these studies have been limited to in vitro, ex vivo or in vivo approaches using xenograft tumors in immunodeficient murine models. For translational relevance, MUC4-targeted therapies should be evaluated in murine models with intact immune system and accurate tumor microenvironment. Additionally, future studies evaluating MUC4 as a target for immunotherapy must entail characterization of immune response in PC patients and investigate its association with immunosuppression and survival.

Amelioration of ongoing experimental autoimmune encephalomyelitis with fluoxetine

In patients with multiple sclerosis, the selective serotonin reuptake inhibitor, fluoxetine, resulted in less acute disease activity. We tested the immune modulating effects of fluoxetine in a mouse model of multiple sclerosis, i.e. experimental autoimmune encephalomyelitis (EAE). We show that fluoxetine delayed the onset of disease and reduced clinical paralysis in mice with established disease. Fluoxetine had abrogating effects on proliferation of immune cells and inflammatory cytokine production by both antigen-presenting cells and T cells. Specifically, in CD4 T cells, fluoxetine increased Fas-induced apoptosis. We conclude that fluoxetine possesses immune-modulating effects resulting in the amelioration of symptoms in EAE.

Steroid-induced resolution of refractory pulmonary interstitial emphysema

Abstract Pulmonary interstitial emphysema (PIE) is a familiar complication of mechanical ventilation in premature infants. Its most severe form, marked by life-threatening respiratory and cardiovascular compromise, is a particularly vexing problem in neonatology. Treatment modalities rely on decubitus positioning and reduction of mean airway pressure, but refractory PIE is unresponsive to these maneuvers. Other options such as selective unilateral bronchial intubation, balloon catheter bronchial occlusion, selective lung volume reduction, and mechanical decompression each have clear limitations. In a patient with refractory, life-threatening PIE too unstable for other therapeutic modalities, we describe success with steroid therapy at a familiar dosing regimen.

An amyloidogenic hexapeptide derived from amylin attenuates inflammation and acute lung injury in murine sepsis

Although the accumulation of amyloidogenic proteins in neuroinflammatory conditions is generally considered pathologic, in a murine model of multiple sclerosis, amyloid-forming fibrils, comprised of hexapeptides, are anti-inflammatory. Whether these molecules modulate systemic inflammatory conditions remains unknown. We hypothesized that an amylin hexapeptide that forms fibrils can attenuate the systemic inflammatory response in a murine model of sepsis. To test this hypothesis, mice were pre-treated with either vehicle or amylin hexapeptide (20 μg) at 12 hours and 6 hours prior to intraperitoneal (i.p.) lipopolysaccharide (LPS, 20 mg/kg) administration. Illness severity and survival were monitored every 6 hours for 3 days. Levels of pro- (IL-6, TNF-α, IFN-γ) and anti-inflammatory (IL-10) cytokines were measured via ELISA at 1, 3, 6, 12, and 24 hours after LPS (i.p.). As a metric of lung injury, pulmonary artery endothelial cell (PAEC) barrier function was tested 24 hours after LPS administration by comparing lung wet-to-dry ratios, Evan’s blue dye (EBD) extravasation, lung histology and caspase-3 activity. Compared to controls, pretreatment with amylin hexapeptide significantly reduced mortality (p<0.05 at 72 h), illness severity (p<0.05), and pro-inflammatory cytokine levels, while IL-10 levels were elevated (p<0.05). Amylin pretreatment attenuated LPS-induced lung injury, as demonstrated by decreased lung water and caspase-3 activity (p<0.05, versus PBS). Hence, in a murine model of systemic inflammation, pretreatment with amylin hexapeptide reduced mortality, disease severity, and preserved lung barrier function. Amylin hexapeptide may represent a novel therapeutic tool to mitigate sepsis severity and lung injury.

Triheptanoin: A Rescue Therapy for Cardiogenic Shock in Carnitine-acylcarnitine Translocase Deficiency

Carnitine-acylcarnitine translocase (CACT) deficiency is a rare long-chain fatty acid oxidation disorder (LC-FAOD) with high mortality due to cardiomyopathy or lethal arrhythmia. Triheptanoin (UX007), an investigational drug composed of synthetic medium odd-chain triglycerides, is a novel therapy in development for LC-FAOD patients. However, cases of its safe and efficacious use to reverse severe heart failure in CACT deficiency are limited. Here, we present a detailed report of an infant with CACT deficiency admitted in metabolic crisis that progressed into severe cardiogenic shock who was successfully treated by triheptanoin. The child was managed, thereafter, on triheptanoin until her death at 3 years of age from a cardiopulmonary arrest in the setting of acute respiratory illness superimposed on chronic hypercarbic respiratory failure.

Diagnostic Dilemma in the Treatment of a Fatal Case of Bloody Diarrhea

Although diarrhea is the most commonly reported pediatric illness in the United States, mortality is usually a rare and unexpected event. We report the case of a healthy 13-month-old male that succumbed to a diarrheal illness of unclear etiology. Presenting signs included frequent nonbloody stools that progressed to frankly bloody stools over 72 hours. Associated symptoms included fever, tenesmus, relief with stool passage, and significant fatigue. On examination, the patient appeared tired and lay with legs curled toward his chest. The abdominal exam was remarkable for hypoactive bowel sounds, diffuse tenderness to palpation without guarding or rebound pain, and intermittent prolapse of rectal tissue. Abdominal plain films demonstrated a paucity of bowel gas, especially in the rectum; and ultrasound revealed thickening of bowel loops in the left lower quadrant. Abdominal computed tomography scan showed decreased enhancement of the mucosa of the rectosigmoid colon. The patient deteriorated rapidly with cardiorespiratory arrest occurring 48 hours after admission. Despite a protracted effort at cardiopulmonary resuscitation, perfusing heart rate or rhythm could not be reestablished. Autopsy revealed infarction and necrosis of the rectosigmoid colon with invasive gram-negative bacilli. Here we present his perplexing case, diagnostic evaluations, and suggest a unifying diagnosis.