Mouse model for endometriosis is characterized by proliferation and inflammation but not epithelial-to-mesenchymal transition and fibrosis.

Endometriosis is a common disorder of unknown etiology, and non-surgical therapies are still a challenge. To understand the pathogenesis and preclinical testing of drugs for endometriosis, animal models are highly desirous. Herein, we carried out longitudinal characterization of a mouse model for endometriosis where uterine tissue was transplanted onto the intestinal mesentery. During the course of lesion development from day 15 to 60 post-induction, the ectopic endometrium became pale, fluid-filled and the animals developed peritoneal adhesions. Most lesions resembled a well-differentiated type of endometriosis and ~13% of animals had mixed type of lesions. There was extensive stromal compaction in the ectopic tissue. During the progression of endometriosis, there was increased proliferation of epithelial and stromal cells as evident by PCNA staining. Cyp19a1 (aromatase) mRNA was detected in the ectopic lesions on day 15 and 30 post-induction of endometriosis, by day 60 the expression was reduced. As compared to the control endometrium, the mRNA levels of Esr1 progressively reduced while the levels of inflammation associated genes (Esr2, Ifng, Tnf and Il1b) increased in the ectopic lesions. Infiltration of macrophages and polymorphonuclear leucocytes was also observed in the ectopic lesions indicative of inflammation. As compared to control, there was no change in levels of Cytokeratin and E-cadherin in the epithelial cells of ectopic endometrium. We did not observe excessive collagen deposition or α -SMA positive myofibroblasts in the stroma of the ectopic endometrium. Thus, epithelial-to-mesenchymal transition and fibrosis are not detected in the mouse model of endometriosis. Our results show that the mouse model of endometriosis mimics some but not all the features of human endometriosis.

Humanized mouse models of immunological diseases and precision medicine.

With the increase in knowledge resulting from the sequencing of the human genome, the genetic basis for the underlying differences in individuals, their diseases, and how they respond to therapies is starting to be understood. This has formed the foundation for the era of precision medicine in many human diseases that is beginning to be implemented in the clinic, particularly in cancer. However, preclinical testing of therapeutic approaches based on individual biology will need to be validated in animal models prior to translation into patients. Although animal models, particularly murine models, have provided significant information on the basic biology underlying immune responses in various diseases and the response to therapy, murine and human immune systems differ markedly. These fundamental differences may be the underlying reason why many of the positive therapeutic responses observed in mice have not translated directly into the clinic. There is a critical need for preclinical animal models in which human immune responses can be investigated. For this, many investigators are using humanized mice, i.e., immunodeficient mice engrafted with functional human cells, tissues, and immune systems. We will briefly review the history of humanized mice, the remaining limitations, approaches to overcome them and how humanized mouse models are being used as a preclinical bridge in precision medicine for evaluation of human therapies prior to their implementation in the clinic.

Dual Targeting of EGFR and IGF1R in the TNFAIP8 Knockdown Non-Small Cell Lung Cancer Cells.

Aberrant regulation of EGFR is common in non-small cell lung carcinomas (NSCLC), and tumor resistance to targeted therapies has been attributed to emergence of other co-occurring oncogenic events, parallel bypass receptor tyrosine kinase pathways including IGF1R, and TNFα-driven adaptive response via NF-κB. TNFAIP8, TNFα-inducible protein 8, is an NF-κB-activated prosurvival and oncogenic molecule. TNFAIP8 expression protects NF-κB-null cells from TNFα-induced cell death by inhibiting caspase-8 activity. Here, we demonstrate that knockdown of TNFAIP8 inhibited EGF and IGF-1-stimulated migration in NSCLC cells. TNFAIP8 knockdown cells showed decreased level of EGFR and increased expression of sorting nexin 1 (SNX1), a key regulator of the EGFR trafficking through the endosomal compartments, and treatment with SNX1 siRNA partially restored EGFR expression in these cells. TNFAIP8 knockdown cells also exhibited downregulation of IGF-1-induced pIGF1R and pAKT, and increased expression of IGF-1-binding protein 3 (IGFBP3), a negative regulator of the IGF-1/IGF1R signaling. Consistently, treatment of TNFAIP8 knockdown cells with IGFBP3 siRNA restored pIGF1R and pAKT levels. TNFAIP8 knockdown cells had enhanced sensitivities to inhibitors of EGFR, PI3K, and AKT. Furthermore, IHC expression of TNFAIP8 was associated with poor prognosis in NSCLC. These findings demonstrate TNFAIP8-mediated regulation of EGFR and IGF1R via SNX1 and IGFBP3, respectively. We posit that TNFAIP8 is a viable, multipronged target downstream of the TNFα/NF-κB axis, and silencing TNFAIP8 may overcome adaptive response in NSCLC. IMPLICATIONS: TNFAIP8 and its effectors SNX1 and IGFBP3 may be exploited to improve the efficacy of molecular-targeted therapies in NSCLC and other cancers.Visual Overview: http://mcr.aacrjournals.org/content/molcanres/17/5/1207/F1.large.jpg.

Genetic variations of hOCT1 gene and CYP3A4/A5 genes and their association with imatinib response in Chronic Myeloid Leukemia.

There is an increasing body of evidence demonstrating that mechanisms independent of BCR/ABL gene also contribute to imatinib resistance in Chronic Myeloid Leukemia (CML). It has been extensively reported that polymorphisms of the genes associated with imatinib metabolization and imatinib influx/efflux play an important role in the disease resistance. We investigated the impact of 12 genetic variants of the two genes, CYP3A4/A5 and the human cation transporter 1 gene (hOCT1) on the clinical outcome, in a cohort of 106 newly diagnosed CML patients. In the patient cohort investigated, only 6 variant alleles could be detected. The others were not present and could not be investigated. Two polymorphisms, CYP3A5*3 (rs776746)and hOCT1 M408V (rs628031), were significantly associated with the Complete Cytogenetic Response (CCyR) at 6 months and Major Molecular Response (MMR) at 12 months. The presence of favourable alleles at M408V and M420del in combination was associated with a MMR at 12 months. Functional polymorphisms of the genes associated with imatinib influx and metabolization may play a role in predicting primary response to imatinib and treatment outcome.

Evolution of BCR/ABL gene mutation in CML is time dependent and dependent on the pressure exerted by tyrosine kinase inhibitor.

BACKGROUND: Mutations in the ABL kinase domain and SH3-SH2 domain of the BCR/ABL gene and amplification of the Philadelphia chromosome are the two important BCR/ABL dependent mechanisms of imatinib resistance. Here, we intended to study the role played by TKI, imatinib, in selection of gene mutations and development of chromosomal abnormalities in Indian CML patients. METHODS: Direct sequencing methodology was employed to detect mutations and conventional cytogenetics was done to identify Philadelphia duplication. RESULTS: Among the different mechanisms of imatinib resistance, kinase domain mutations (39%) of the BCR/ABL gene were seen to be more prevalent, followed by mutations in the SH3-SH2 domain (4%) and then BCR/ABL amplification with the least frequency (1%). The median duration of occurrence of mutation was significantly shorter for patients with front line imatinib than those pre-treated with hydroxyurea. Patients with high Sokal score (p = 0.003) showed significantly higher incidence of mutations, as compared to patients with low/intermediate score. Impact of mutations on the clinical outcome in AP and BC was observed to be insignificant. Of the 94 imatinib resistant patients, only 1 patient exhibited duplication of Philadelphia chromosome, suggesting a less frequent occurrence of this abnormality in Indian CML patients. CONCLUSION: Close monitoring at regular intervals and proper analysis of the disease resistance would facilitate early detection of resistance and thus aid in the selection of the most appropriate therapy.

A novel 5-way translocation t(9;11;13;19;22) in a case of chronic-phase chronic myeloid leukemia.

Chronic myeloid leukemia (CML) is a hematopoietic stem cell disorder that occurs because of t(9;22)(q34;q11) translocations. Complex translocations have been reported in CML. We report a novel 5-way translocation 46,XY,t(9;11;13;19;22)(9q34.12;11p11.12;13q21.31;19q13.12;22q11.21) using GTG banding, fluorescence in situ hybridization, and spectral karyotyping in a case of chronic-phase CML. Molecular analysis revealed the presence of 2 types of transcripts (b3a2, b2a2). The patient was responding to the imatinib treatment. However, the patient needs to be carefully monitored at various intervals.

Deletion of ABL/BCR on der(9) associated with severe basophilia.

Chronic basophilic leukemia is a rare form in chronic myeloid leukemia patients. Only limited number of reports are available. Herein, we describe a patient who presented with fatigue, weight loss, leucocytosis, prominent basophilia, and mild eosinophilia. On biopsy, bone marrow was hypercellular with marked basophils. The immunophenotype showed abnormal expression of CD7, which is suggestive of basophilic maturation. Chromosomal analysis from GTG-banded metaphases revealed Ph positivity, and fluorescence in situ hybridization (FISH) with BCR/ABL dual color, dual fusion probe showed single fusion on the der(22) chromosome and ABL/BCR fusion was deleted on the der(9) chromosome. The deletion (ABL/BCR) on der(9) may be associated with basophilia which may be also indicative of the transformation of CML to acute myeloid leukemia.

Recent developments in drug resistance mechanism in chronic myeloid leukemia: a review.

A revolution in medical science was marked with the advent of imatinib, a site-specific drug for the management of patients with chronic myeloid leukemia (CML). Imatinib mesylate (also known as Glivec, Gleevec, STI-571, CGP57148), an orally administered 2-phenylaminopyrimidine derivative approved by FDA in 2001 for the treatment for CML, is highly effective in treating the early stages of CML, but remission induced in advanced phase was observed to be relatively short-lived. The primary cause of resistance in patients with CML is the mutations in the BCR-ABL kinase domain. This review discusses the different mechanisms leading to imatinib resistance and various treatment options to over-ride imatinib resistance.