Predictors of short-term response and the role of heavy alcohol use in treatment of depression.

BACKGROUND: Depression and alcohol use disorders frequently co-occur. However, research on psychosocial interventions for treating this dual pathology is limited. The Ostrobothnian Depression Study (ODS) aimed to increase the systematic use of evidence-based methods, particularly among patients with comorbid depression and substance use in a naturalistic setting. This is a secondary analysis of the ODS study. The aim of the present study was to explore the predictors of a response to treatment during the first six months of the ODS intervention with a specific focus on the role of comorbid heavy alcohol use. METHODS: The study sample (n = 242) comprised psychiatric specialist care patients with depression (Beck Depression Inventory score ≥ 17) at baseline. Patients with a baseline Alcohol Use Disorders Identification Test (AUDIT) score > 10 (n = 99) were assigned to the AUD (Alcohol Use Disorder) group in this study. The ODS intervention comprised behavioral activation (BA) for all and additional motivational interviewing (MI) for those in AUD group. The predictors of response to treatment (minimum of 50% reduction in depressive symptoms) during the first six months were analyzed with logistic regression models. RESULTS: In the total sample at six months (n = 150), predictors of response to treatment were more severe depression (OR 1.10, CI 1.02-1.18), larger amounts of alcohol consumed (OR = 1.16, CI 1.03-1.31) and antipsychotic medication "not in use" (OR = 0.17, CI 0.07-0.44). In the non-AUD group (n = 100), more severe depression (OR 1.12, CI 1.01-1.25) and antipsychotics "not in use" (OR 0.20, CI 0.06-0.67) also predicted a positive response. Among AUD group patients (n = 50), larger amounts of alcohol consumed (OR 1.54, CI 1.04-2.27) and antipsychotic medication "not in use" (OR 0.12, CI 0.02-0.60) predicted a response to the treatment intervention. CONCLUSIONS: The severity of symptoms and comorbid disorders were found to predict better treatment response, suggesting that the intervention was more effective in patients with severe symptoms. Patients with depression should be treated effectively regardless of having concomitant AUD. The results of this study suggest that BA combined with MI should be one of the treatment options for this dual pathology. TRIAL REGISTRATION: ClinicalTrials.gov Identifier NCT02520271 (11/08/2015).

Clozapine-Related Diarrhea and Colitis: Report of 4 Cases.

BACKGROUND: During clozapine treatment, diarrhea is a rare but clinically relevant adverse effect. Cases of microscopic colitis and eosinophilic colitis have been previously reported. PROCEDURES: We present 4 patients who developed severe diarrhea in early weeks of clozapine therapy. FINDINGS: Two patients had significant peripheral eosinophilia 1 week after diarrhea symptoms. One of these patients also had Charcot-Leyden crystals in stool afterward, confirming the presence of eosinophils in the gut lumen. One of our patients had a confirmed microscopic colitis and later also neutropenia, which required treatment. CONCLUSIONS: Charcot-Leyden crystals in stool may be associated with concurrent diarrhea and eosinophilia during clozapine treatment, which is a previously unreported finding. Occurrence of blood dyscrasias with diarrhea symptoms during clozapine treatment needs further investigation to understand the possible shared mechanisms.

Assessment of alcohol consumption in depression follow-up using self-reports and blood measures including inflammatory biomarkers.

AIMS: Alcohol consumption has been suggested a major role in the pathogenesis and prognosis of depression. However, reliable identification of hazardous drinking continues to be problematic. We compared the accuracy of different biomarkers and self-reports of alcohol consumption in the follow-up study of depression. METHODS: Data from 202 patients with major depressive disorder were obtained through self-reports, AUDIT and AUDIT-C questionnaires and biomarker analyses. The clinical assessments and measurements of biomarkers (GT, CDT, GT-CDT-combination, MCV, ALT, AST, hs-CRP, IL-6) were performed at baseline and after six months of treatment. Based on self-reported alcohol intake at baseline the patients were classified to three subgroups. RESULTS: About 27.2% of patients were categorized to high-risk drinkers, 26.3% low-risk drinkers and 46.5% abstainers. High-risk drinkers showed significantly higher mean values of GT, CDT, GT-CDT-combination and IL-6 than abstainers, diagnostic accuracy being highest with the combined marker of GT-CDT. The accuracy of AUDIT and AUDIT-C to detect high-risk drinking was also significant. During follow-up, the differences observed in the biomarkers at baseline disappeared together with recovery from depression. CONCLUSIONS: Our data suggest the combined use of GT-CDT and AUDIT questionnaires to improve the identification of drinking of patients with depression. This approach could be useful for improving treatment adherence and outcome in depressed patients.

Behavioral activation versus treatment as usual in naturalistic sample of psychiatric patients with depressive symptoms: a benchmark controlled trial.

BACKGROUND: More systematic use of evidence-based brief therapies is needed in the treatment of depression within psychiatric care. The aim of this study was to explore the impact of behavioral activation therapy (BA) for patients with depressive symptoms in a routine clinical setting of secondary psychiatric care. METHODS: The BA-treated intervention group (n = 242) comprised patients with depressive symptoms (Beck Depression Inventory (BDI) score ≥ 17 at baseline). The control group (n = 205) patients received treatment as usual in the same catchment area. The groups were matched at baseline using BDI and Alcohol Use Disorders Identification Test scores and inpatient/outpatient status. The groups were compared at 6-, 12- and 24-month follow-up points on functional outcome (Global Assessment of Functioning scale), service use, dropout and deaths. The Montgomery-Åsberg Depression Rating Scale was used to assess depressive symptoms in the intervention group. RESULTS: The estimated difference in GAF score between intervention and control group patients was significant at 12- and 24-months follow-up points in favor of intervention group (GAF score difference 4.85 points, p = 0.006 and 5.71 points, p = 0.005, respectively; estimate for patient group 2.26, p = 0.036). The rates of dropout, mortality and service use were similar between the groups. Among the intervention group patients, the estimated improvement in MADRS score compared to baseline was statistically significant throughout the follow-up (p < 0.001 at all follow-up points). CONCLUSIONS: The systematic use of BA among secondary psychiatric care depressive patients provides encouraging results despite the patients had various comorbid non-psychotic disorders. TRIAL REGISTRATION: ClinicalTrials.gov , Identifier NCT02520271, Release Date: 06/27/2015, retrospectively registered.

Temperament and character profiles are associated with depression outcome in psychiatric secondary care patients with harmful drinking.

BACKGROUND: Temperament and character profiles have been associated with depression outcome and alcohol abuse comorbidity in depressed patients. How harmful alcohol use modifies the effects of temperament and character on depression outcome is not well known. Knowledge of these associations could provide a method for enhancing more individualized treatment strategies for these patients. METHODS: We screened 242 depressed patients with at least moderate level of depressive symptoms. The Alcohol Use Disorders Identification Test (AUDIT) was used for identifying patients with marked alcohol use problems (AUP, AUDIT≥11). After 6 weeks of antidepressive treatment 173 patients were assessed using the Montgomery-Åsberg Depression Rating Scale (MADRS), and the Temperament and Character Inventory (TCI-R). Outcome of depression (MADRS scores across three follow-up points at 6 weeks, 6 months and 24 months) was predicted with AUP, gender, and AUP x Gender and AUP x Time interactions together with temperament and character dimension scores in a linear mixed effects model. RESULTS: Poorer outcome of depression (MADRS scores at 6 weeks, 6 months and 24 months) was predicted by AUP × Time interaction (p = 0.0002) together with low Reward Dependence (p = 0.003). Gender and all other temperament and character traits were non-significant predictors of the depression outcome in the mixed effects model. CONCLUSIONS: Possibly due to the modifying effect of alcohol use problems, high Reward Dependence was associated with better depression treatment outcome at 6 months. Harm Avoidance and Self-Directedness did not predict depression outcome when alcohol use problems were controlled.

Outcome of patients with dual diagnosis in secondary psychiatric care.

BACKGROUND: Dual diagnosis (DD) is a common co-morbidity of mental illness and substance use disorder (SUD) and patients with DD are prone to complications. Better knowledge on the outcome, mortality and management of patients with DD in usual secondary psychiatric care would help to inform improved treatment strategies in the future. AIMS: To explore the functional outcome and mortality of patients with DD receiving psychiatric treatment. To assess the recognition of substance use disorders (SUDs) in terms of diagnosis, and the associations of clinically diagnosed SUDs with treatment-related variables. METHODS: The sample of 330 patients was collected by screening all currently treated patients with the Alcohol Use Disorders Identification Test (AUDIT) and a question about other substances used. The inclusion criteria were AUDIT >7 and/or reported use of other substances during the preceding 12 months. The Global Assessment of Functioning scale was used to assess functional outcomes during a 2-year follow-up. Information concerning treatment and patient characteristics was collected retrospectively. RESULTS: Level of functioning remained stable among all study patients during follow-up. The mortality rate was not increased. Effective medication use was associated with improved functional outcomes. SUDs were underdiagnosed. A clinically diagnosed SUD seemed to have an impact on the regularity of appointments and the doses of prescribed medications. CONCLUSIONS: Given our results suggesting a stable level of functioning, patients with DD appear to be well managed within secondary psychiatric care. Attention should be paid to more precise diagnostics of SUDs and to effective use of medication.

[Assessment of an addiction psychiatric patient within primary health care]. / Psyyke plus ja päihteitä mukana: päihdepsykiatrisen potilaan arviointi perusterveydenhuollossa.

Assessment of addiction psychiatric patients during the hectic work at a community health center is a challenging task. A more precise survey of the substance abuse and mental problem can be started by using various screening questionnaires. In devising the treatment plan, focus will initially be on the main problem, and upon prolonged cooperation a more precise survey of the drug and mental problem will be carried out. The patient's overall picture should be considered, avoiding a split treatment of the drug problem and the mental problem. Assessment of psychotic addiction patients belongs to a psychiatrist and the treatment in most cases to specialized care.

Detection of Experimental Colorectal Peritoneal Metastases by a Novel PDGFRß-Targeting Nanobody.

Peritoneal metastases in colorectal cancer (CRC) belong to Consensus Molecular Subtype 4 (CMS4) and are associated with poor prognosis. Conventional imaging modalities, such as Computed Tomography (CT) and Fluorodeoxyglucose-Positron Emission Tomography (FDG-PET), perform very poorly in the detection of peritoneal metastases. However, the stroma-rich nature of these lesions provides a basis for developing molecular imaging strategies. In this study, conducted from 2019 to 2021, we aimed to generate a Platelet-Derived Growth Factor Receptor beta (PDGFRB)-binding molecular imaging tracer for the detection of CMS4 CRC, including peritoneal metastases. The expression of PDGFRB mRNA discriminated CMS4 from CMS1-3 (AUROC = 0.86 (95% CI 0.85-0.88)) and was associated with poor relapse-free survival. PDGFRB mRNA and protein levels were very high in all human peritoneal metastases examined (n = 66). Therefore, we generated a PDGFRB-targeting llama nanobody (VHH1E12). Biotin-labelled VHH1E12 bound to immobilized human and mouse PDGFRB with high affinity (EC50 human PDGFRB = 7 nM; EC50 murine PDGFRB = 0.8 nM), and to PDGFRB-expressing HEK293 cells grown in vitro. A pharmacokinetic analysis of IRDye-800CW-conjugated VHH1E12 in mice showed that the plasma half-life was 6 min. IRDye-800CW-conjugated VHH1E12 specifically accumulated in experimentally induced colorectal cancer peritoneal metastases in mice. A tissue analysis subsequently demonstrated co-localization of the nanobody with PDGFRB expression in the tumour stroma. Our results demonstrate the potential value of PDGFRB-targeted molecular imaging as a novel strategy for the non-invasive detection of CMS4 CRC, in particular, peritoneal metastases.

Impact of Comorbid Alcohol Use Disorder on Health-Related Quality of Life Among Patients With Depressive Symptoms.

Background and Aim: In psychiatric clinical practice, comorbidity of depression and alcohol use disorder (AUD) is common. Both disorders have a negative impact on health-related quality of life (HRQoL) in general population. However, research on the impact of comorbid AUD on HRQoL among clinically depressed patients is limited. The purpose of this study was to explore the impact of a psychosocial treatment intervention on HRQoL for depressive patients in specialized psychiatric care with a special focus on the impact of AUD on HRQoL. Material and Methods: Subjects were 242 patients of the Ostrobothnia Depression Study (ClinicalTrials.gov Identifier NCT02520271). Patients referred to specialized psychiatric care who scored at least 17 points on the Beck Depression Inventory at baseline and who had no psychotic disorders were included in the ODS. The treatment intervention in ODS comprised behavioral activation for all but began with motivational interviewing for those with AUD. HRQoL was assessed regularly during 24-month follow-up by the 15D instrument. In the present study, HRQoL of ODS patients with or without AUD was compared and the factors explaining 15D score analyzed with a linear mixed model. In order to specify the impact of clinical depression on HRQoL during the early phase of treatment intervention, a general population sample of the Finnish Health 2011 Survey was used as an additional reference group. Results: HRQoL improved among all ODS study sample patients regardless of comorbid AUD during the first year of follow-up. During 12-24 months of follow-up the difference between groups was seen as HRQoL continued to improve only among the non-AUD patients. A combination of male gender, anxiety disorder, and AUD was associated with the poorest HRQoL in this sample. In combined sample analyses with the reference group, clinical depression had an impact on HRQoL in short-term follow-up regardless of the treatment intervention. Conclusions: This study suggests that, in terms of improvement in HRQoL, the heterogenous group of depressive patients in specialized psychiatric care can be successfully treated with behavioral activation in combination with motivational interviewing for those with AUD. Clinical Trial Registration: ClinicalTrials.gov Identifier: NCT02520271. Ostrobothnia Depression Study (ODS). A Naturalistic Follow-up Study on Depression and Related Substance Use Disorders. (2015). Available online at: https://clinicaltrials.gov/ct2/show/NCT02520271.

Status of inflammation and alcohol use in a 6-month follow-up study of patients with major depressive disorder.

INTRODUCTION: The studies on the relationship between alcohol consumption, the status of inflammation, and depression have produced conflicting results. In this study, we followed patients with major depressive disorders by monitoring biomarkers of inflammation together with biomarkers of heavy alcohol use. METHOD: The levels of IL-6 (interleukin-6), IL-8 (interleukin-8), hs-CRP (high sensitivity C-reactive protein), YKL-40 (also known as Chitinase-3-like protein 1 or CHI3L1), and biomarkers of alcohol consumption and liver status (GT, CDT, ALT, alkaline phosphatase) were measured at baseline and after 6 months of psychiatric treatment from 242 patients suffering from current major depressive disorder (MDD) with (n = 99) or without (n = 143) alcohol use disorder (AUD). RESULTS: At baseline, the patients with MDD + AUD showed higher levels of inflammatory biomarkers IL-6 (p < 0.001), hs-CRP (p < 0.01), YKL-40 (p < 0.05), and biomarkers of alcohol consumption, than the corresponding group of non-AUD patients. These differences disappeared during follow-up and recovery from depression. The level of IL-8 decreased significantly in both AUD (p < 0.05) and non-AUD (p < 0.05) patients. During follow-up, the biomarkers of alcohol consumption, GT and CDT, in AUD patients were found to decrease in parallel with serum YKL-40 levels. CONCLUSIONS: Alcohol consumption appears to modulate the status of inflammation in depressive patients. A more systematic use of biomarkers of inflammation together with biomarkers of alcohol consumption and liver status may prove to be of value in a more comprehensive assessment and treatment of patients with depression.

Temperament clusters associate with anxiety disorder comorbidity in depression.

BACKGROUND: Individual temperament is associated with psychiatric morbidity and could explain differences in psychiatric comorbidities. We investigated the association of temperament profile clusters with anxiety disorder comorbidity in patients with depression. METHODS: We assessed the temperament of 204 specialized care-treated depressed patients with the Temperament and Character Inventory (TCI-R) and their diagnoses with the Mini International Neuropsychiatric Interview. Two-step cluster analysis was used for defining patients' temperament profiles and logistic regression analysis was used for predicting different anxiety disorders for various temperament profiles. RESULTS: Four temperament clusters were found: 1) Novelty seekers with highest Novelty Seeking scores (n = 56),2) Persistent with highest Persistence scores (n = 36), 3) Reserved with lowest Novelty Seeking scores (n = 66) and 4) Wearied with highest Harm avoidance, lowest Reward Dependence and lowest Persistence scores (n = 58). After adjusting for clinical variables, panic disorder and/or agoraphobia were predicted by Novelty seekers' temperament profile with odds ratio [OR] = 3.5 (95% confidence interval [CI] = 1.8 - 6.9, p < 0.001), social anxiety disorder was predicted by Wearied temperament profile with OR = 3.4 (95% CI = 1.6 - 7.5, p = 0.002), and generalized anxiety disorder was predicted by Reserved temperament profile with OR = 2.6 (95% CI = 1.2 - 5.3, p = 0.01). LIMITATIONS: The patients' temperament profiles were assessed while displaying depressive symptoms, which may have affected results. CONCLUSIONS: Temperament clusters with unique dimensional profiles were specifically associated with different anxiety disorders in this study. These results suggest that TCI-R could offer a valuable dimensional method for predicting the risk of anxiety disorders in diverse depressed patients.

Temperament and character profiles associated with depression and treatment response in patients with or without comorbid substance abuse.

There is limited knowledge on the relationship between temperament and character profiles and substance abuse comorbidity in depressed patients. We recruited 127 depressed patients without alcohol use problems (non-AUP) and 89 depressed patients with alcohol use problems (AUP). We assessed all patients using the Temperament and Character Inventory (TCI-R) at baseline and after 6 weeks of treatment. Using univariate general linear models (GLMs), we analyzed differences in TCI-R between AUP and non-AUP. GLMs were also used in analyzing the associations between TCI-R changes and antidepressive treatment responses measured with changes in Montgomery Åsberg Depression Rating Scale score (ΔMADRS). Alcohol use explained independently significant proportions of the variation in Novelty Seeking, Self-Directedness, and Persistence. Reward Dependence score change explained 14.1% of the ΔMADRS in AUP, but was non-significant in non-AUP. Character score changes in Self-Directedness and Self-Transcendence explained together 14.1% of ΔMADRS in non-AUP, whereas they were all non-significant in AUP. AUP compared with non-AUP patients had lower Self-Directedness and Persistence and higher Novelty Seeking scores. Detected changes in Reward Dependence and lower Self-Directedness in AUP patients could be reflective of different biological mechanisms associated with depressive symptomatology in alcohol abuse. Changes in character are associated with acute treatment response in non-AUP.

Phenotypic rescue of a Drosophila model of mitochondrial ANT1 disease.

A point mutation in the Drosophila gene that codes for the major adult isoform of adenine nuclear translocase (ANT) represents a model for human diseases that are associated with ANT insufficiency [stress-sensitive B(1) (sesB(1))]. We characterized the organismal, bioenergetic and molecular phenotype of sesB(1) flies then tested strategies to compensate the mutant phenotype. In addition to developmental delay and mechanical-stress-induced seizures, sesB(1) flies have an impaired response to sound, defective male courtship, female sterility and curtailed lifespan. These phenotypes, excluding the latter two, are shared with the mitoribosomal protein S12 mutant, tko(25t). Mitochondria from sesB(1) adults showed a decreased respiratory control ratio and downregulation of cytochrome oxidase. sesB(1) adults exhibited ATP depletion, lactate accumulation and changes in gene expression that were consistent with a metabolic shift towards glycolysis, characterized by activation of lactate dehydrogenase and anaplerotic pathways. Females also showed downregulation of many genes that are required for oogenesis, and their eggs, although fertilized, failed to develop to the larval stages. The sesB(1) phenotypes of developmental delay and mechanical-stress-induced seizures were alleviated by an altered mitochondrial DNA background. Female sterility was substantially rescued by somatic expression of alternative oxidase (AOX) from the sea squirt Ciona intestinalis, whereas AOX did not alleviate developmental delay. Our findings illustrate the potential of different therapeutic strategies for ANT-linked diseases, based on alleviating metabolic stress.

Tumor hypoxia as a driving force in genetic instability.

Sub-regions of hypoxia exist within all tumors and the presence of intratumoral hypoxia has an adverse impact on patient prognosis. Tumor hypoxia can increase metastatic capacity and lead to resistance to chemotherapy and radiotherapy. Hypoxia also leads to altered transcription and translation of a number of DNA damage response and repair genes. This can lead to inhibition of recombination-mediated repair of DNA double-strand breaks. Hypoxia can also increase the rate of mutation. Therefore, tumor cell adaptation to the hypoxic microenvironment can drive genetic instability and malignant progression. In this review, we focus on hypoxia-mediated genetic instability in the context of aberrant DNA damage signaling and DNA repair. Additionally, we discuss potential therapeutic approaches to specifically target repair-deficient hypoxic tumor cells.

PTEN deletion in prostate cancer cells does not associate with loss of RAD51 function: implications for radiotherapy and chemotherapy.

PURPOSE: PTEN deletions in prostate cancer are associated with tumor aggression and poor outcome. Recent studies have implicated PTEN as a determinant of homologous recombination (HR) through defective RAD51 function. Similar to BRCA1/2-defective tumor cells, PTEN-null prostate and other cancer cells have been reported to be sensitive to PARP inhibitors (PARPi). To date, no direct comparison between PTEN and RAD51 expression in primary prostate tumors has been reported. EXPERIMENTAL DESIGN: Prostate cancer cell lines and xenografts with known PTEN status (22RV1-PTEN(+/+), DU145-PTEN(+/-), PC3-PTEN(-/-)) and H1299 and HCT116 cancer cells were used to evaluate how PTEN loss affects RAD51 expression and PARPi sensitivity. Primary prostate cancers with known PTEN status were analyzed for RAD51 expression. RESULTS: PTEN status is not associated with reduced RAD51 mRNA or protein expression in primary prostate cancers. Decreased PTEN expression did not reduce RAD51 expression or clonogenic survival following PARPi among prostate cancer cells that vary in TP53 and PTEN. PARPi sensitivity instead associated with a defect in MRE11 expression. PTEN-deficient cells had only mild PARPi sensitivity and no loss of HR or RAD51 recruitment. Clonogenic cell survival following a series of DNA damaging agents was variable: PTEN-deficient cells were sensitive to ionizing radiation, mitomycin-C, UV, H(2)O(2), and methyl methanesulfonate but not to cisplatin, camptothecin, or paclitaxel. CONCLUSIONS: These data suggest that the relationship between PTEN status and survival following DNA damage is indirect and complex. It is unlikely that PTEN status will be a direct biomarker for HR status or PARPi response in prostate cancer clinical trials.

The receptor tyrosine kinase inhibitor amuvatinib (MP470) sensitizes tumor cells to radio- and chemo-therapies in part by inhibiting homologous recombination.

BACKGROUND AND PURPOSE: RAD51 is a key protein involved in homologous recombination (HR) and a potential target for radiation- and chemotherapies. Amuvatinib (formerly known as MP470) is a novel receptor tyrosine kinase inhibitor that targets c-KIT and PDGFRα and can sensitize tumor cells to ionizing radiation (IR). Here, we studied amuvatinib mechanism on RAD51 and functional HR. MATERIALS AND METHODS: Protein and RNA analyses, direct repeat green fluorescent protein (DR-GFP) assay and polysomal fractioning were used to measure HR efficiency and global translation in amuvatinib-treated H1299 lung carcinoma cells. Synergy of amuvatinib with IR or mitomycin c (MMC) was assessed by clonogenic survival assay. RESULTS: Amuvaninib inhibited RAD51 protein expression and HR. This was associated with reduced ribosomal protein S6 phosphorylation and inhibition of global translation. Amuvatinib sensitized cells to IR and MMC, agents that are selectively toxic to HR-deficient cells. CONCLUSIONS: Amuvatinib is a promising agent that may be used to decrease tumor cell resistance. Our work suggests that this is associated with decreased RAD51 expression and function and supports the further study of amuvatinib in combination with chemotherapy and radiotherapy.

Tumor cell kill by c-MYC depletion: role of MYC-regulated genes that control DNA double-strand break repair.

MYC regulates a myriad of genes controlling cell proliferation, metabolism, differentiation, and apoptosis. MYC also controls the expression of DNA double-strand break (DSB) repair genes and therefore may be a potential target for anticancer therapy to sensitize cancer cells to DNA damage or prevent genetic instability. In this report, we studied whether MYC binds to DSB repair gene promoters and modulates cell survival in response to DNA-damaging agents. Chromatin immunoprecipitation studies showed that MYC associates with several DSB repair gene promoters including Rad51, Rad51B, Rad51C, XRCC2, Rad50, BRCA1, BRCA2, DNA-PKcs, XRCC4, Ku70, and DNA ligase IV. Endogenous MYC protein expression was associated with increased RAD51 and KU70 protein expression of a panel of cancer cell lines of varying histopathology. Induction of MYC in G(0)-G(1) and S-G(2)-M cells resulted in upregulation of Rad51 gene expression. MYC knockdown using small interfering RNA (siRNA) led to decreased RAD51 expression but minimal effects on homologous recombination based on a flow cytometry direct repeat green fluorescent protein assay. siRNA to MYC resulted in tumor cell kill in DU145 and H1299 cell lines in a manner independent of apoptosis. However, MYC-dependent changes in DSB repair protein expression were not sufficient to sensitize cells to mitomycin C or ionizing radiation, two agents selectively toxic to DSB repair-deficient cells. Our results suggest that anti-MYC agents may target cells to prevent genetic instability but would not lead to differential radiosensitization or chemosensitization.

Contextual synthetic lethality of cancer cell kill based on the tumor microenvironment.

Acute and chronic hypoxia exists within the three-dimensional microenvironment of solid tumors and drives therapy resistance, genetic instability, and metastasis. Replicating cells exposed to either severe acute hypoxia (16 hours with 0.02% O(2)) followed by reoxygenation or moderate chronic hypoxia (72 hours with 0.2% O(2)) treatments have decreased homologous recombination (HR) protein expression and function. As HR defects are synthetically lethal with poly(ADP-ribose) polymerase 1 (PARP1) inhibition, we evaluated the sensitivity of repair-defective hypoxic cells to PARP inhibition. Although PARP inhibition itself did not affect HR expression or function, we observed increased clonogenic killing in HR-deficient hypoxic cells following chemical inhibition of PARP1. This effect was partially reversible by RAD51 overexpression. PARP1(-/-) murine embryonic fibroblasts (MEF) showed a proliferative disadvantage under hypoxic gassing when compared with PARP1(+/+) MEFs. PARP-inhibited hypoxic cells accumulated γH2AX and 53BP1 foci as a consequence of altered DNA replication firing during S phase-specific cell killing. In support of this proposed mode of action, PARP inhibitor-treated xenografts displayed increased γH2AX and cleaved caspase-3 expression in RAD51-deficient hypoxic subregions in vivo, which was associated with decreased ex vivo clonogenic survival following experimental radiotherapy. This is the first report of selective cell killing of HR-defective hypoxic cells in vivo as a consequence of microenvironment-mediated "contextual synthetic lethality." As all solid tumors contain aggressive hypoxic cells, this may broaden the clinical utility of PARP and DNA repair inhibition, either alone or in combination with radiotherapy and chemotherapy, even in tumor cells lacking synthetically lethal, genetic mutations.