Multi-omic cross-sectional cohort study of pre-malignant Barrett's esophagus reveals early structural variation and retrotransposon activity.

Barrett's esophagus is a pre-malignant lesion that can progress to esophageal adenocarcinoma. We perform a multi-omic analysis of pre-cancer samples from 146 patients with a range of outcomes, comprising 642 person years of follow-up. Whole genome sequencing reveals complex structural variants and LINE-1 retrotransposons, as well as known copy number changes, occurring even prior to dysplasia. The structural variant burden captures the most variance across the cohort and genomic profiles do not always match consensus clinical pathology dysplasia grades. Increasing structural variant burden is associated with: high levels of chromothripsis and breakage-fusion-bridge events; increased expression of genes related to cell cycle checkpoint, DNA repair and chromosomal instability; and epigenetic silencing of Wnt signalling and cell cycle genes. Timing analysis reveals molecular events triggering genomic instability with more clonal expansion in dysplastic samples. Overall genomic complexity occurs early in the Barrett's natural history and may inform the potential for cancer beyond the clinically discernible phenotype.

Longitudinal tracking of 97 esophageal adenocarcinomas using liquid biopsy sampling.

BACKGROUND: The incidence of esophageal adenocarcinoma (EAC) is rapidly rising and has a 5-year survival rate of <20%. Beyond TNM (tumor-node-metastasis) staging, no reliable risk stratification tools exist and no large-scale studies have profiled circulating tumor DNA (ctDNA) at relapse in EAC. Here we analyze the prognostic potential of ctDNA dynamics in EAC, taking into account clonal hematopoiesis with indeterminate potential (CHIP). PATIENTS AND METHODS: A total of 245 samples from 97 patients treated with neoadjuvant chemotherapy and surgery were identified from the prospective national UK Oesophageal Cancer Clinical and Molecular Stratification (OCCAMS) consortium data set. A pan-cancer ctDNA panel comprising 77 genes was used. Plasma and peripheral blood cell samples were sequenced to a mean depth of 7082× (range 2196-28 524) and ctDNA results correlated with survival. RESULTS: Characteristics of the 97 patients identified were as follows: 83/97 (86%) male, median age 68 years (SD 9.5 years), 100% cT3/T4, 75% cN+. EAC-specific drivers had higher variant allele fractions than passenger mutations. Using stringent quality criteria 16/79 (20%) were ctDNA positive following resection; recurrence was observed in 12/16 (75%) of these. As much as 78/97 (80%) had CHIP analyses that enabled filtering for CHIP variants, which were found in 18/78 (23%) of cases. When CHIP was excluded, 10/63 (16%) patients were ctDNA positive and 9/10 of these (90%) recurred. With correction for CHIP, median cancer-specific survival for ctDNA-positive patients was 10.0 months versus 29.9 months for ctDNA-negative patients (hazard ratio 5.55, 95% confidence interval 2.42-12.71; P = 0.0003). Similar outcomes were observed for disease-free survival. CONCLUSIONS: We demonstrate in a large, national, prospectively collected data set that ctDNA in plasma following surgery for EAC is prognostic for relapse. Inclusion of peripheral blood cell samples can reduce or eliminate false positives from CHIP. In future, post-operative ctDNA could be used to risk stratify patients into high- and low-risk groups for intensification or de-escalation of adjuvant chemotherapy.

Genomic interaction between ER and HMGB2 identifies DDX18 as a novel driver of endocrine resistance in breast cancer cells.

Breast cancer resistance to endocrine therapies such as tamoxifen and aromatase inhibitors is a significant clinical problem. Steroid receptor coactivator-1 (SRC-1), a coregulatory protein of the oestrogen receptor (ER), has previously been shown to have a significant role in the progression of breast cancer. The chromatin protein high mobility group box 2 (HMGB2) was identified as an SRC-1 interacting protein in the endocrine-resistant setting. We investigated the expression of HMGB2 in a cohort of 1068 breast cancer patients and found an association with increased disease-free survival time in patients treated with endocrine therapy. However, it was also verified that HMGB2 expression could be switched on in endocrine-resistant tumours from breast cancer patients. To explore the function of this poorly characterized protein, we performed HMGB2 ChIPseq and found distinct binding patterns between the two contexts. In the resistant setting, the HMGB2, SRC-1 and ER complex are enriched at promoter regions of target genes, with bioinformatic analysis indicating a switch in binding partners between the sensitive and resistant phenotypes. Integration of binding and gene expression data reveals a concise set of target genes of this complex including the RNA helicase DDX18. Modulation of DDX18 directly affects growth of tamoxifen-resistant cells, suggesting that it may be a critical downstream effector of the HMGB2:ER complex. This study defines HMGB2 interactions with the ER complex at specific target genes in the tamoxifen-resistant setting.

The Australasian contribution to malaria vaccine development.

Malaria remains a major threat to public health worldwide, despite intense research efforts spanning decades. Much of this work has been directed towards developing an effective malaria vaccine, and scientists from Australasia have made significant contributions to progress in this area. Herein, we review the research undertaken in Australasia and summarize some of the important roles that Australasian researchers have played in malaria vaccine development that has occurred outside the region.

Ets-2 and p160 proteins collaborate to regulate c-Myc in endocrine resistant breast cancer.

Associations between p160 coactivator proteins and endocrine resistance have been described. Though thought to primarily interact with steroid receptors, the p160 proteins can also interact with non-nuclear receptor transcription factors including the MAP kinase effector proteins Ets. Here, we observed that in breast cancer cells resistant and insensitive to endocrine treatment, the growth factor EGF induced Ets-2 but not Ets-1 transcriptional regulation of the oncogene myc. Ets-2 regulation of myc was found to be reliant on the p160 proteins SRC-1 and SRC-3. In support of these molecular observations, strong associations were observed between the transcription factor, Ets-2 and its coactivator SRC-1 (P<0.01) and the target gene myc (P<0.0001) in a cohort of breast cancer patients with locally advanced disease. Expression of Ets-2, SRC-1 and c-Myc individually all associated with reduced disease-free survival (P<0.001, P<0.001 and P=0.002 respectively). There was no association between SRC-3 and disease-free survival (P=0.707). SRC-1 can utilize MAP kinase effector transcription factor Ets-2 to regulate the production of the oncogene myc. These signalling mechanisms may be important in the development of steroid resistant/independent breast cancer.

The determination of the optimal dose of milnacipran in the olfactory bulbectomized rat model of depression.

Olfactory bulbectomy (OB) is associated with a variety of behavioral abnormalities such as hyperactivity in the "open-field" test. Previous studies have shown that chronic administration of antidepressants can reverse this behavioral deficit. The activity of milnacipran (20, 30, and 40 mg/kg, PO bid) administered in two equally divided doses twice daily was assessed in the olfactory bulbectomized rat model of depression. It was found that chronic treatment with milnacipran at the doses of 30 and 40 mg/kg, but not 20 mg/kg, attenuated the lesion-induced hyperactivity of the OB rat in the "open-field" test following 14 days of treatment. In the step-through passive avoidance test, administration of milnacipran at doses of 20, 30, and 40 mg/kg had no effect on the performance deficit associated with olfactory bulbectomy. Olfactory bulbectomy reduced the concentration of noradrenaline (NA) in the frontal cortex. However, chronic milnacipran treatment did not significantly alter this deficit. It is concluded that milnacipran, when administered chronically at doses of 30 and 40 mg/kg, are effective at reversing the "open-field" deficit associated with olfactory bulbectomy, and that a dose of 30 mg/kg is an optimal dose.

Effects of acute and chronic antidepressant administration on phencyclidine (PCP) induced locomotor hyperactivity.

Previously it was found that both acute and chronic antidepressant pre-treatment enhanced the locomotor hyperactivity induced by a challenge injection of the non-competitive NMDA receptor antagonist, dizocilpine (MK-801). In the present study the effects of acute and chronic antidepressant administration on phencyclidine (PCP)-induced locomotor hyperactivity were examined. Phencyclidine (PCP), a non-competitive NMDA receptor antagonist increased locomotor activity in rats. Fluoxetine given acutely increased and prolonged the PCP-induced locomotor hyperactivity, while citalopram, sertraline and paroxetine had no effect on the PCP-induced behavioural effect. Repeated treatment with fluoxetine, citalopram and paroxetine increased the PCP-induced locomotor hyperactivity. In contrast, chronic sertraline administration attenuated the locomotor response to a PCP challenge. These results indicate that these antidepressants which are presumed to have a similar pharmacological profile, differ in their ability to alter PCP-induced hyperactivity. Whether these differences have any bearing on the therapeutic or adverse effects of these drugs remains to be shown.

Acute 3,4-methylenedioxymethamphetamine(MDMA) administration produces a rapid and sustained suppression of immune function in the rat.

(+)-3,4-Methylenedioxymethamphetamine (MDMA;'Ecstasy') is a ring substituted phenylisopropylamine that is structurally related to both amphetamines and hallucinogens. The unique behavioural activating properties of MDMA have led to its widespread abuse. MDMA induces many neurochemical, behavioural and endocrine alterations which closely resemble those elicited by exposure to acute stress, suggesting that MDMA could be regarded as a 'chemical stressor'. In addition to the neurochemical, behavioural and endocrine effects of stressor exposure, it has been reported that stress produces alterations in immune function. However, to date the effects of MDMA on immune function have been restricted to in vitro investigations. In this study we report, for the first time, that acute in vivo administration of MDMA (20 mg/kg, i.p.) produced a rapid (within 30 min) suppression of Con A-induced lymphocyte proliferation and a profound reduction in the total leucocyte count in rats that persisted for at least 6 h following injection. These alterations in immune function were accompanied by a significant increase in plasma corticosterone concentrations 30 min post MDMA administration which had returned to baseline values within 6 h of drug administration. In addition, there was a significant depletion in cortical 5-HT concentrations both 30 min and 6 h after MDMA administration. The results of this study provide evidence that in addition to the well established toxic effects of MDMA on the central serotonergic system, a single administration of this widely abused drug induces a rapid and sustained suppression of immune function.

Behavioural and neurochemical effects of dizocilpine in the olfactory bulbectomized rat model of depression.

The activity of dizocilpine (MK-801; 0.1 and 0.3 mg/kg) administered once daily intraperitoneally (I.P.) was assessed in the olfactory bulbectomized rat model of depression. Olfactory bulbectomy (OB) is associated with a variety of behavioural abnormalities, such as hyperactivity in the "open field" test. Previous studies have shown that chronic administration of antidepressants can reverse this behavioural deficit. In the present study, chronic treatment with 0.1 and 0.3 mg/kg of dizocilpine (I.P.) antagonized the lesion-induced hyperactivity in the "open field" test. Acute treatment with dizocilpine was associated with an increase in locomotor activity in both sham-operated and OB rats, with a greater response in the sham-operated group. Following chronic treatment, this hyperactivity was found to be greater in the OB-treated animals compared with the sham-treated animals. Olfactory bulbectomy reduced serotonin (5-HT), noradrenaline (NA), and dopamine (DA) concentrations in the frontal cortex. Chronic dizocilpine administration did not alter the 5-HT or NA response. In contrast, chronic administration of dizocilpine to OB animals did attenuate the OB-related deficit in DA. In the OB-operated control animals, there was an increase in DOPAC levels. In conclusion, chronic dizocilpine administration displays antidepressant-like activity in the OB rat model of depression. However, unlike conventional antidepressants, dizocilpine does not correct the 5-HT and NA neurotransmitter deficits that occur in this model.

The effects of the 5-HT1A agonist flesinoxan, in three paradigms for assessing antidepressant potential in the rat.

5-HT1A receptor agonists have been shown to be effective clinically in the treatment of depression and anxiety. Flesinoxan is an example which is highly selective for the 5-HT1A receptor subtype. The objective of this study was to appraise the antidepressant potential of flesinoxan (1 and 3 mg/kg s.c.) in three tests which are indicative of antidepressant activity. These are (1) the forced swim test, following sub-acute administration, (2) 'open field' activity in the olfactory bulbectomised (OB) rat, following chronic administration, and (3) 8-OH-DPAT-induced hypothemia following chronic treatment. Both doses of flesinoxan significantly reduced the immobility time in the sham and OB groups when compared to their respective controls. In the 'open field', there was a significant increase in the ambulation of the OB control group. The higher dose of flesinoxan significantly reduced this deficit. In addition both doses of flesinoxan significantly attenuated the 8-OH-DPAT-induced hypothermic response. These effects of flesinoxan are quantitatively similar to those seen following the chronic administration of antidepressants. These studies illustrate the potential antidepressant properties of flesinoxan, and hence further emphasise the role of the 5-HT1A receptor in the pathogenesis of depression.

Prediction of the resting metabolic rate in obese patients.

Resting metabolic rate (RMR) was measured in 154 women and 48 men before the beginning of a weight reduction program. In both sexes there were significant univariate correlations between RMR and fat-free mass, body fat, weight, fat cell weight, and fat cell number (from total body water). Women also showed significant correlations between RMR and fat cell number (from total body potassium), free triiodothyronine index, and fasting and postglucose insulin levels. Multiple regression analysis showed that both fat-free mass and fat cell weight and number were significant predictors of RMR. The contribution of fat-free mass was three to five times greater per kg than that of body fat. There was no significant contribution of thyroid hormones or insulin to the prediction of RMR. Fat cell number and fat cell weight were significant predictors of RMR, whether determined from body water, body potassium, or a formula using both water and potassium. There was no significant difference in regression coefficients between men and women. Thus the difference in RMR between the sexes is probably caused by the higher proportion of fat-free mass in men. The effect of age was small and not statistically significant.