Rapid identification of BCR/ABL1-like acute lymphoblastic leukaemia patients using a predictive statistical model based on quantitative real time-polymerase chain reaction: clinical, prognostic and therapeutic implications.

BCR/ABL1-like acute lymphoblastic leukaemia (ALL) is a subgroup of B-lineage acute lymphoblastic leukaemia that occurs within cases without recurrent molecular rearrangements. Gene expression profiling (GEP) can identify these cases but it is expensive and not widely available. Using GEP, we identified 10 genes specifically overexpressed by BCR/ABL1-like ALL cases and used their expression values - assessed by quantitative real time-polymerase chain reaction (Q-RT-PCR) in 26 BCR/ABL1-like and 26 non-BCR/ABL1-like cases to build a statistical "BCR/ABL1-like predictor", for the identification of BCR/ABL1-like cases. By screening 142 B-lineage ALL patients with the "BCR/ABL1-like predictor", we identified 28/142 BCR/ABL1-like patients (19·7%). Overall, BCR/ABL1-like cases were enriched in JAK/STAT mutations (P < 0·001), IKZF1 deletions (P < 0·001) and rearrangements involving cytokine receptors and tyrosine kinases (P = 0·001), thus corroborating the validity of the prediction. Clinically, the BCR/ABL1-like cases identified by the BCR/ABL1-like predictor achieved a lower rate of complete remission (P = 0·014) and a worse event-free survival (P = 0·0009) compared to non-BCR/ABL1-like ALL. Consistently, primary cells from BCR/ABL1-like cases responded in vitro to ponatinib. We propose a simple tool based on Q-RT-PCR and a statistical model that is capable of easily, quickly and reliably identifying BCR/ABL1-like ALL cases at diagnosis

Tailoring CD19xCD3-DART exposure enhances T-cells to eradication of B-cell neoplasms.

Many patients with B-cell malignancies can be successfully treated, although tumor eradication is rarely achieved. T-cell-directed killing of tumor cells using engineered T-cells or bispecific antibodies is a promising approach for the treatment of hematologic malignancies. We investigated the efficacy of CD19xCD3 DART bispecific antibody in a broad panel of human primary B-cell malignancies. The CD19xCD3 DART identified 2 distinct subsets of patients, in which the neoplastic lymphocytes were eliminated with rapid or slow kinetics. Delayed responses were always overcome by a prolonged or repeated DART exposure. Both CD4 and CD8 effector cytotoxic cells were generated, and DART-mediated killing of CD4+ cells into cytotoxic effectors required the presence of CD8+ cells. Serial exposures to DART led to the exponential expansion of CD4 + and CD8 + cells and to the sequential ablation of neoplastic cells in absence of a PD-L1-mediated exhaustion. Lastly, patient-derived neoplastic B-cells (B-Acute Lymphoblast Leukemia and Diffuse Large B Cell Lymphoma) could be proficiently eradicated in a xenograft mouse model by DART-armed cytokine induced killer (CIK) cells. Collectively, patient tailored DART exposures can result in the effective elimination of CD19 positive leukemia and B-cell lymphoma and the association of bispecific antibodies with unmatched CIK cells represents an effective modality for the treatment of CD19 positive leukemia/lymphoma

Extended-spectrum β-lactamase and AmpC β-lactamase Production among Gram-negative Bacilli Isolates Obtained from Urinary Tract Infections and Wound Infections

Extended-spectrum β-lactamases (ESBLs) and AmpC β-lactamases continue to be a major problem in healthcare settings. Due to the scarcity of information regarding the antibiotic susceptibility patterns particularly from urinary tract infection (UTI) and wound infections, the current study was carried out to assist the clinicians to prescribe appropriate antibiotics against Gram-negative clinical isolates. In the current study, urine (n = 620) and pus (n = 228) samples were collected from different sites (at various clinical departments) and subjected to direct microscopic examination, culture and antibiotic susceptibility testing (AST). In the AST testings, the isolates that exhibited reduced zone of inhibition to one or more of the antibiotics such as cefotaxime (≤27 mm), ceftriaxone (≤25 mm), ceftazidime (≤22 mm), cefpodoxime (≤17 mm) and aztreonam (≤27 mm) were considered as potential ESBL producers and the ESBL production was confirmed using phenotypic screening test (double-disk synergy test) and phenotypic confirmatory test (combined-disk test). However, isolates showing resistance or decreased sensitivity to cefoxitin, cefotaxime, ceftriaxone, ceftazidime, cefpodoxime or aztreonam and sensitive to cefepime were considered as a screen positive AmpC producer and subjected to AmpC disk tests. The current study concluded that 72.41% and 21.76% of ESBL and AmpC producers were detected, respectively in our hospital. It was also observed that the double-disk synergy and combined-disk tests were equally effective for ESBL detection. Further, AmpC disk test is simple, easy to perform and interpret, requiring less expertise for the rapid detection of AmpC isolates

The novel lncRNA BlackMamba controls the neoplastic phenotype of ALK- anaplastic large cell lymphoma by regulating the DNA helicase HELLS.

The molecular mechanisms leading to the transformation of anaplastic lymphoma kinase negative (ALK-) anaplastic large cell lymphoma (ALCL) have been only in part elucidated. To identify new culprits which promote and drive ALCL, we performed a total transcriptome sequencing and discovered 1208 previously unknown intergenic long noncoding RNAs (lncRNAs), including 18 lncRNAs preferentially expressed in ALCL. We selected an unknown lncRNA, BlackMamba, with an ALK- ALCL preferential expression, for molecular and functional studies. BlackMamba is a chromatin-associated lncRNA regulated by STAT3 via a canonical transcriptional signaling pathway. Knockdown experiments demonstrated that BlackMamba contributes to the pathogenesis of ALCL regulating cell growth and cell morphology. Mechanistically, BlackMamba interacts with the DNA helicase HELLS controlling its recruitment to the promoter regions of cell-architecture-related genes, fostering their expression. Collectively, these findings provide evidence of a previously unknown tumorigenic role of STAT3 via a lncRNA-DNA helicase axis and reveal an undiscovered role for lncRNA in the maintenance of the neoplastic phenotype of ALK-ALCL

Whole-genome characterization of myoepithelial carcinomas of the soft tissue

Myoepithelial carcinomas (MECs) of soft tissue are rare and aggressive tumors affecting young adults and children, but their molecular landscape has not been comprehensively explored through genome sequencing. Here, we present the whole-exome sequencing (WES), whole-genome sequencing (WGS), and RNA sequencing findings of two MECs. Patients 1 and 2 (P1, P2), both male, were diagnosed at 27 and 37 yr of age, respectively, with shoulder (P1) and inguinal (P2) soft tissue tumors. Both patients developed metastatic disease, and P2 died of disease. P1 tumor showed a rhabdoid cytomorphology and a complete loss of INI1 (SMARCB1) expression, associated with a homozygous SMARCB1 deletion. The tumor from P2 showed a clear cell/small cell morphology, retained INI1 expression and strong S100 positivity. By WES and WGS, tumors from both patients displayed low tumor mutation burdens, and no targetable alterations in cancer genes were detected. P2's tumor harbored an EWSR1::KLF15 rearrangement, whereas the tumor from P1 showed a novel ASCC2::GGNBP2 fusion. WGS evidenced a complex genomic event involving mainly Chromosomes 17 and 22 in the tumor from P1, which was consistent with chromoplexy. These findings are consistent with previous reports of EWSR1 rearrangements (50% of cases) in MECs and provide a genetic basis for the loss of SMARCB1 protein expression observed through immunohistochemistry in 10% of 40% of MEC cases. The lack of additional driver mutations in these tumors supports the hypothesis that these alterations are the key molecular events in MEC evolution. Furthermore, the presence of complex structural variant patterns, invisible to WES, highlights the novel biological insights that can be gained through the application of WGS to rare cancers

Repeat pneumococcal polysaccharide vaccination does not impair functional immune responses among Indigenous Australians.

Indigenous Australians experience one of the highest rates of pneumococcal disease globally. In the Northern Territory of Australia, a unique government-funded vaccination schedule for Indigenous Australian adults comprising multiple lifetime doses of the pneumococcal polysaccharide vaccine is currently implemented. Despite this programme, rates of pneumococcal disease do not appear to be declining, with concerns raised over the potential for immune hyporesponse associated with the use of this vaccine. We undertook a study to examine the immunogenicity and immune function of a single and repeat pneumococcal polysaccharide vaccination among Indigenous adults compared to non-Indigenous adults. Our results found that immune function, as measured by opsonophagocytic and memory B-cell responses, were similar between the Indigenous groups but lower for some serotypes in comparison with the non-Indigenous group. This is the first study to document the immunogenicity following repeat 23-valent pneumococcal polysaccharide vaccine administration among Indigenous Australian adults, and reinforces the continued need for optimal pneumococcal vaccination programmes among high-risk populations

ZFX Mediates Non-canonical Oncogenic Functions of the Androgen Receptor Splice Variant 7 in Castrate-Resistant Prostate Cancer

Androgen receptor splice variant 7 (AR-V7) is crucial for prostate cancer progression and therapeutic resistance. We show that, independent of ligand, AR-V7 binds both androgen-responsive elements (AREs) and non-canonical sites distinct from full-length AR (AR-FL) targets. Consequently, AR-V7 not only recapitulates AR-FL's partial functions but also regulates an additional gene expression program uniquely via binding to gene promoters rather than ARE enhancers. AR-V7 binding and AR-V7-mediated activation at these unique targets do not require FOXA1 but rely on ZFX and BRD4. Knockdown of ZFX or select unique targets of AR-V7/ZFX, or BRD4 inhibition, suppresses growth of castration-resistant prostate cancer cells. We also define an AR-V7 direct target gene signature that correlates with AR-V7 expression in primary tumors, differentiates metastatic prostate cancer from normal, and predicts poor prognosis. Thus, AR-V7 has both ARE/FOXA1 canonical and ZFX-directed non-canonical regulatory functions in the evolution of anti-androgen therapeutic resistance, providing information to guide effective therapeutic strategies. By cistrome profiling of endogenous androgen receptor (AR) versus an AR splice variant, AR-V7, Cai et al. uncovered non-canonical pathways uniquely targeted by AR-V7 and ZFX, a previously unknown AR-V7 partner. Targeting cofactors (ZFX or BRD4) or non-canonical downstream pathways of AR-V7 provides potential therapeutic ways for treating prostate cancer

Small Cell Carcinoma of the Ovary, Hypercalcemic Type (SCCOHT) beyond SMARCA4 Mutations: A Comprehensive Genomic Analysis.

Small cell carcinoma of the ovary, hypercalcemic type (SCCOHT) is an aggressive malignancy that occurs in young women, is characterized by recurrent loss-of-function mutations in the SMARCA4 gene, and for which effective treatments options are lacking. The aim of this study was to broaden the knowledge on this rare malignancy by reporting a comprehensive molecular analysis of an independent cohort of SCCOHT cases. We conducted Whole Exome Sequencing in six SCCOHT, and RNA-sequencing and array comparative genomic hybridization in eight SCCOHT. Additional immunohistochemical, Sanger sequencing and functional data are also provided. SCCOHTs showed remarkable genomic stability, with diploid profiles and low mutation load (mean, 5.43 mutations/Mb), including in the three chemotherapy-exposed tumors. All but one SCCOHT cases exhibited 19p13.2-3 copy-neutral LOH. SMARCA4 deleterious mutations were recurrent and accompanied by loss of expression of the SMARCA2 paralog. Variants in a few other genes located in 19p13.2-3 (e.g., PLK5) were detected. Putative therapeutic targets, including MAGEA4, AURKB and CLDN6, were found to be overexpressed in SCCOHT by RNA-seq as compared to benign ovarian tissue. Lastly, we provide additional evidence for sensitivity of SCCOHT to HDAC, DNMT and EZH2 inhibitors. Despite their aggressive clinical course, SCCOHT show remarkable inter-tumor homogeneity and display genomic stability, low mutation burden and few somatic copy number alterations. These findings and preliminary functional data support further exploration of epigenetic therapies in this lethal disease

Inhibition of FGF receptor blocks adaptive resistance to RET inhibition in CCDC6-RET-rearranged thyroid cancer.

Genetic alterations in RET lead to activation of ERK and AKT signaling and are associated with hereditary and sporadic thyroid cancer and lung cancer. Highly selective RET inhibitors have recently entered clinical use after demonstrating efficacy in treating patients with diverse tumor types harboring RET gene rearrangements or activating mutations. In order to understand resistance mechanisms arising after treatment with RET inhibitors, we performed a comprehensive molecular and genomic analysis of a patient with RET-rearranged thyroid cancer. Using a combination of drug screening and proteomic and biochemical profiling, we identified an adaptive resistance to RET inhibitors that reactivates ERK signaling within hours of drug exposure. We found that activation of FGFR signaling is a mechanism of adaptive resistance to RET inhibitors that activates ERK signaling. Combined inhibition of FGFR and RET prevented the development of adaptive resistance to RET inhibitors, reduced cell viability, and decreased tumor growth in cellular and animal models of CCDC6-RET-rearranged thyroid cancer

Single-cell profiling reveals an endothelium-mediated immunomodulatory pathway in the eye choroid

The activity and survival of retinal photoreceptors depend on support functions performed by the retinal pigment epithelium (RPE) and on oxygen and nutrients delivered by blood vessels in the underlying choroid. By combining single-cell and bulk RNA sequencing, we categorized mouse RPE/choroid cell types and characterized the tissue-specific transcriptomic features of choroidal endothelial cells. We found that choroidal endothelium adjacent to the RPE expresses high levels of Indian Hedgehog and identified its downstream target as stromal GLI1+ mesenchymal stem cell-like cells. In vivo genetic impairment of Hedgehog signaling induced significant loss of choroidal mast cells, as well as an altered inflammatory response and exacerbated visual function defects after retinal damage. Our studies reveal the cellular and molecular landscape of adult RPE/choroid and uncover a Hedgehog-regulated choroidal immunomodulatory signaling circuit. These results open new avenues for the study and treatment of retinal vascular diseases and choroid-related inflammatory blinding disorders.Funding for this study was provided by National Institutes of Health grants EY08538 and GM34107 (E. Rodriguez-Boulan); EY027038 (R.F. Mullins); 1R21CA224391-01A1 (J.H. Zippin); and 1R01CA194547, 1U24CA210989, and P50CA211024 (O. Elemento); National Cancer Institute grant R01CA192176 and cancer center support grant P30 CA008748-48 (A.L. Joyner); Comunidad Autónoma de Madrid grant 2017-T1/BMD-5247 (I. Benedicto); Agencia Nacional Argentina de Promoción Cient´ıfica y Tecnológica grant PICT 2014-3687 and Fundación Sales (G.A. Rabinovich); a Pew Latin American Fellowship (G.L. Lehmann); Calder Research Scholar Award Vitiligo/Pigment Cell Disorders (J.H. Zippin); Starr Foundation Tri-Institutional Stem Cell Initiative award 2013-028; NYSTEM contract C32596GG; and Research to Prevent Blindness and Dyson Foundation departmental grants. The CNIC is supported by the Instituto de Salud Carlos III, the Ministerio de Ciencia e Innovación, and the Pro CNIC Foundation and is a Severo Ochoa Center of Excellence (SEV-2015-0505).S