Meiosis I arrest abnormalities lead to severe oligozoospermia in meiosis 1 arresting protein (M1ap)-deficient mice.

Meiosis 1 arresting protein (M1ap) is a novel vertebrate gene expressed exclusively in germ cells of the embryonic ovary and the adult testis. In male mice, M1ap expression, which is present from spermatogonia to secondary spermatocytes, is evolutionarily conserved and has a specific spatial and temporal pattern suggestive of a role during germ cell development. To test its function, mice deficient in M1ap were created. Whereas females had histologically normal ovaries, males exhibited reduced testicular size and a myriad of tubular defects, which led to severe oligozoospermia and infertility. Although some germ cells arrested at the zygotene/pachytene stages, most cells advanced to metaphase I before arresting and entering apoptosis. Cells that reached metaphase I were unable to properly align their chromosomes at the metaphase plate due to abnormal chromosome synapses and failure to form crossover foci. Depending on the state of tubular degeneration, all germ cells, with the exemption of spermatogonia, disappeared; with further deterioration, tubules displaying only Sertoli cells reminiscent of Sertoli cell-only syndrome in humans were observed. Our results uncovered an essential role for M1ap as a novel germ cell gene not previously implicated in male germ cell development and suggest that mutations in M1AP could account for some cases of nonobstructive oligozoospermia in men.

Mullerian inhibiting substance preferentially inhibits stem/progenitors in human ovarian cancer cell lines compared with chemotherapeutics.

Cancer stem cells are proposed to be tumor-initiating cells capable of tumorigenesis, recurrence, metastasis, and drug resistance, and, like somatic stem cells, are thought to be capable of unlimited self-renewal and, when stimulated, proliferation and differentiation. Here we select cells by expression of a panel of markers to enrich for a population with stem cell-like characteristics. A panel of eight was initially selected from 95 human cell surface antigens as each was shared among human ovarian primary cancers, ovarian cancer cell lines, and normal fimbria. A total of 150 combinations of markers were reduced to a panel of three--CD44, CD24, and Epcam--which selected, in three ovarian cancer cell lines, those cells which best formed colonies. Cells expressing CD44, CD24, and Epcam exhibited stem cell characteristics of shorter tumor-free intervals in vivo after limiting dilution, and enhanced migration in invasion assays in vitro. Also, doxorubicin, cisplatin, and paclitaxel increased this enriched population which, conversely, was significantly inhibited by Müllerian inhibiting substance (MIS) or the MIS mimetic SP600125. These findings demonstrate that flow cytometry can be used to detect a population which shows differential drug sensitivity, and imply that treatment of patients can be individualized to target both stem/progenitor cell enriched and nonenriched subpopulations. The findings also suggest that this population, amenable to isolation by flow cytometry, can be used to screen for novel treatment paradigms, including biologic agents such as MIS, which will improve outcomes for patients with ovarian cancer.

Mullerian inhibiting substance inhibits invasion and migration of epithelial cancer cell lines.

OBJECTIVE: Given the fact that Mullerian Inhibiting Substance (MIS) causes complex remodeling of the urogenital ridge and regression of the Mullerian ducts during male embryonic development, we examined whether MIS could affect similar cell properties such as migration and invasion that could contribute ultimately to micro-metastasis of cancers arising from Mullerian tissues. MIS receptor expressing cell lines found to be invasive and migratory in vivo are examined in an in vivo assay that is cost-effective. METHODS: We designed in vitro and in vivo experiments to determine if MIS inhibited the movement of cancer lines IGROV-1, HEp3, MDA-MB-231, and HT1080 in cell culture invasion/migration chamber assays and in chick embryo metastasis assays. RESULTS: MIS, at concentrations below those that inhibit cell proliferation, blocked in vitro invasion and in vivo migration of epithelial cancer cells that express the MIS receptor. CONCLUSIONS: While our laboratory has previously established MIS as an inhibitor of cancer cell proliferation using in vitro assays and in vivo xenografts, we now show that MIS can also inhibit in vivo tumor migration.

Normal ovarian surface epithelial label-retaining cells exhibit stem/progenitor cell characteristics.

Ovulation induces cyclic rupture and regenerative repair of the ovarian coelomic epithelium. This process of repeated disruption and repair accompanied by complex remodeling typifies a somatic stem/progenitor cell-mediated process. Using BrdU incorporation and doxycycline inducible histone2B-green fluorescent protein pulse-chase techniques, we identify a label-retaining cell population in the coelomic epithelium of the adult mouse ovary as candidate somatic stem/progenitor cells. The identified population exhibits quiescence with asymmetric label retention, functional response to estrous cycling in vivo by proliferation, enhanced growth characteristics by in vitro colony formation, and cytoprotective mechanisms by enrichment for the side population. Together, these characteristics identify the label-retaining cell population as a candidate for the putative somatic stem/progenitor cells of the coelomic epithelium of the mouse ovary.

Endothelial cells promote migration and proliferation of enteric neural crest cells via beta1 integrin signaling.

Enteric neural crest-derived cells (ENCCs) migrate along the intestine to form a highly organized network of ganglia that comprises the enteric nervous system (ENS). The signals driving the migration and patterning of these cells are largely unknown. Examining the spatiotemporal development of the intestinal neurovasculature in avian embryos, we find endothelial cells (ECs) present in the gut prior to the arrival of migrating ENCCs. These ECs are patterned in concentric rings that are predictive of the positioning of later arriving crest-derived cells, leading us to hypothesize that blood vessels may serve as a substrate to guide ENCC migration. Immunohistochemistry at multiple stages during ENS development reveals that ENCCs are positioned adjacent to vessels as they colonize the gut. A similar close anatomic relationship between vessels and enteric neurons was observed in zebrafish larvae. When EC development is inhibited in cultured avian intestine, ENCC migration is arrested and distal aganglionosis results, suggesting that ENCCs require the presence of vessels to colonize the gut. Neural tube and avian midgut were explanted onto a variety of substrates, including components of the extracellular matrix and various cell types, such as fibroblasts, smooth muscle cells, and endothelial cells. We find that crest-derived cells from both the neural tube and the midgut migrate avidly onto cultured endothelial cells. This EC-induced migration is inhibited by the presence of CSAT antibody, which blocks binding to beta1 integrins expressed on the surface of crest-derived cells. These results demonstrate that ECs provide a substrate for the migration of ENCCs via an interaction between beta1 integrins on the ENCC surface and extracellular matrix proteins expressed by the intestinal vasculature. These interactions may play an important role in guiding migration and patterning in the developing ENS.

Characterization of the chromosome 1q41q42.12 region, and the candidate gene DISP1, in patients with CDH.

Cytogenetic and molecular cytogenetic studies demonstrate association between congenital diaphragmatic hernia (CDH) and chromosome 1q41q42 deletions. In this study, we screened a large CDH cohort (N=179) for microdeletions in this interval by the multiplex ligation-dependent probe amplification (MLPA) technique, and also sequenced two candidate genes located therein, dispatched 1 (DISP1) and homo sapiens H2.0-like homeobox (HLX). MLPA analysis verified deletions of this region in two cases, an unreported patient with a 46,XY,del(1)(q41q42.13) karyotype and a previously reported patient with a Fryns syndrome phenotype [Kantarci et al., 2006]. HLX sequencing showed a novel but maternally inherited single nucleotide variant (c.27C>G) in a patient with isolated CDH, while DISP1 sequencing revealed a mosaic de novo heterozygous substitution (c.4412C>G; p.Ala1471Gly) in a male with a left-sided Bochdalek hernia plus multiple other anomalies. Pyrosequencing demonstrated the mutant allele was present in 43%, 12%, and 4.5% of the patient's lymphoblastoid, peripheral blood lymphocytes, and saliva cells, respectively. We examined Disp1 expression at day E11.5 of mouse diaphragm formation and confirmed its presence in the pleuroperitoneal fold, as well as the nearby lung which also expresses Sonic hedgehog (Shh). Our report describes the first de novo DISP1 point mutation in a patient with complex CDH. Combining this finding with Disp1 embryonic mouse diaphragm and lung tissue expression, as well as previously reported human chromosome 1q41q42 aberrations in patients with CDH, suggests that DISP1 may warrant further consideration as a CDH candidate gene.

Development of an efficiently cleaved, bioactive, highly pure FLAG-tagged recombinant human Mullerian Inhibiting Substance.

Mullerian Inhibiting Substance (MIS), a member of the TGF-beta family, causes regression of the Mullerian duct in male embryos, after binding to Mullerian Inhibiting Substance Receptor II (MISRII). It has also been extensively demonstrated that it can inhibit proliferation of various cancer cell lines such as ovarian, prostate, and breast cancer in vitro and in vivo. Hence, the availability of a recombinant, epitope tagged, bioactive MIS is important for the selection of patients for treatment and for probing novel molecular targets for MIS in various tissues. To this end, we have expressed a recombinant, internally FLAG-tagged form of hMIS with the tag (DYKDDDDK) immediately after the cleavage site (427-428) of MIS at the C-terminus with a modified dibasic cleavage motif sequence. We show that this construct results in a highly pure, endogenously processed (cleaved) FLAG MIS, that causes complete regression of the Mullerian Duct in an organ culture assay. In addition, purified FLAG MIS was able to bind and affinity purify both transfected and endogenous MIS type II receptor. The availability of this fully functional, epitope tagged form of MIS should facilitate scale-up for preclinical and clinical use and should also be used for the study of MIS binding proteins and for tracking in pharmacokinetic studies.

Late complications of newborn circumcision: a common and avoidable problem.

PURPOSE: The purpose of this paper is to study the types of operative and post-operative late complications resulting from newborn circumcisions and to make recommendations to prevent them. METHODS: After obtaining IRB approval, a retrospective review of the late complications resulting from newborn circumcisions treated at the MassGeneral Hospital for Children from January 2003 to December 2007 was undertaken. The source used was the consultation notes and operative reports of affected patients. Additionally, cases seen in the outpatient Pediatric Urology Clinic from April 2007 to April 2008 were reviewed. RESULTS: A total of 8,967 children were operated during the study period, of which 424 (4.7%) were for complications resulting from previous neonatal circumcision. Penile adhesions, skin bridges, meatal stenosis, redundant foreskin (incomplete circumcision with uncircumcised appearance), recurrent phimosis, buried penis and penile rotation were the most frequent complications. At the outpatient clinic of the Section of Pediatric Urology, 127 boys with concerns following newborn circumcision were evaluated, representing 7.4% of the total volume of cases seen in this clinic. CONCLUSIONS: Our results indicate the need to undertake a collaborative study to define the incidence of complications following newborn circumcisions, which should be performed by practitioners with adequate training in the technique of their choice and its post-operative care.

Megacalycosis: a rare condition.

Megacalycosis is an extremely rare condition. We report our experience with two cases and discuss its pathogenesis, diagnosis and management in children. Our two patients had presented a prior diagnosis of congenital hydronephrosis. An increased number of calyces with a significant disproportion between the degree of calyceal dilatation and a mildly dilated renal pelvis were found in each case. Megacalycosis must be considered in the differential diagnosis of congenital hydronephrosis, polycalycosis, and infundibular stenosis. The diagnosis is suggested by ultrasound and confirmed by diuretic renography, intravenous pyelography or magnetic resonance urography. Voiding cystourethrography should be performed to rule out vesicoureteral reflux. A high index of suspicion is needed for the diagnosis of this condition.

Clinical clearance of the cervical spine in blunt trauma patients younger than 3 years: a multi-center study of the american association for the surgery of trauma.

BACKGROUND: Cervical spine clearance in the very young child is challenging. Radiographic imaging to diagnose cervical spine injuries (CSI) even in the absence of clinical findings is common, raising concerns about radiation exposure and imaging-related complications. We examined whether simple clinical criteria can be used to safely rule out CSI in patients younger than 3 years. METHODS: The trauma registries from 22 level I or II trauma centers were reviewed for the 10-year period (January 1995 to January 2005). Blunt trauma patients younger than 3 years were identified. The measured outcome was CSI. Independent predictors of CSI were identified by univariate and multivariate analysis. A weighted score was calculated by assigning 1, 2, or 3 points to each independent predictor according to its magnitude of effect. The score was established on two thirds of the population and validated using the remaining one third. RESULTS: Of 12,537 patients younger than 3 years, CSI was identified in 83 patients (0.66%), eight had spinal cord injury. Four independent predictors of CSI were identified: Glasgow Coma Score <14, GCSEYE = 1, motor vehicle crash, and age 2 years or older. A score of <2 had a negative predictive value of 99.93% in ruling out CSI. A total of 8,707 patients (69.5% of all patients) had a score of <2 and were eligible for cervical spine clearance without imaging. There were no missed CSI in this study. CONCLUSIONS: CSI in patients younger than 3 years is uncommon. Four simple clinical predictors can be used in conjunction to the physical examination to substantially reduce the use of radiographic imaging in this patient population.

Hepatocellular Carcinoma in Transplantable Child-Pugh A Cirrhotics: Should Cost Affect Resection vs Transplantation?

BACKGROUND: There is no consensus regarding the optimal surgical treatment for transplantable hepatocellular carcinoma (HCC) patients with well-compensated cirrhosis. Our aim was to compare outcomes between Child-Pugh A (CPA) cirrhotics who underwent liver resection or transplantation for HCC. METHODS: Clinicopathologic data were retrospectively collected for all surgically treated HCC patients between 7/1992 and 12/2015. Disease-free survival (DFS) and overall survival (OS) were calculated from the time of operation or diagnosis (intention-to-treat analysis including patients removed from the transplant list). The average overall cost including pre-operative and post-operative procedures was calculated for each group. RESULTS: Of the 513 surgically treated HCC patients, 184 had CPA cirrhosis and fulfilled the Milan criteria (MC). Of those, 95 (52%) were resected and 89 (48%) were transplanted. Twenty-two patients were removed from the transplant list. Transplanted patients were younger (p < 0.001), had a higher MELD score (p < 0.001) and a higher frequency of hepatitis C (p < 0.001). Length of stay and postoperative complication rates were similar between groups. DFS was longer for transplanted patients (3-, 5-, and 10-year DFS rates 48, 44, 31% vs 96, 94, 94%, respectively, p < 0.001). OS was similar between groups (3-, 5-, and 10-year OS rates 76, 62, 41% vs 82, 77, 53%, respectively, p = 0.07). Only size of greatest lesion and T stage were independent predictors of OS. The cost was much higher for the transplant group, even when accounting for the treatment of recurrences ($37,391 vs $137,996). CONCLUSIONS: Since OS is similar between CPA cirrhotics within the MC undergoing resection or transplantation for HCC, but cost is significantly higher for transplantation. Resection should be considered for first-line treatment.

c-Jun N-terminal kinase inhibitor II (SP600125) activates Mullerian inhibiting substance type II receptor-mediated signal transduction.

Müllerian inhibiting substance (MIS), the hormone required for Müllerian duct regression in fetal males, is also expressed in both adult males and females, but its physiological role in these settings is not clear. The expression of the MIS type II receptor (MISRII) in ovarian cancer cells and the ability of MIS to inhibit proliferation of these cells suggest that MIS might be a promising therapeutic for recurrent ovarian cancer. Using an MISRII-dependent activity assay in a small-molecule screen for MIS-mimetic compounds, we have identified the c-Jun N-terminal kinase inhibitor SP600125 as an activator of the MIS signal transduction pathway. SP600125 increased the activity of a bone morphogenetic protein-responsive reporter gene in a dose-dependent manner and exerted a synergistic effect when used in combination with MIS. This effect was specific for the MISRII and was not seen with other receptors of the TGFbeta family. Moreover, treatment of mouse ovarian cancer cells with a combination of SP600125 and paclitaxel, an established chemotherapeutic agent used in the treatment of ovarian cancer, or with MIS enabled inhibition of cell proliferation at a lower dose than with each treatment alone. These results offer a strong rationale for testing the therapeutic potential of SP600125, alone or in combination with already established drugs, in the treatment of recurrent ovarian cancer with a much-needed decrease in the toxic side effects of currently employed therapeutic agents.

Hospital readmission after distal pancreatectomy is predicted by specific intra- and post-operative factors.

BACKGROUND: Distal pancreatectomy (DP) continues to carry a significant risk of morbidity resulting in hospital readmissions and increased costs. Prognostic factors predicting 30-day readmission after DP were evaluated. METHODS: Data were collected from 946 patients undergoing DP at the University of Verona Hospital Trust and the Massachusetts General Hospital between 2004 and 2014. Patients were divided into a derivation and a validation cohort. RESULTS: The 30-day readmission rate was 13.9%. Predictors of readmission were age over 60 years (OR 1.8), intraoperative transfusions (OR 2.02), CR-POPF (OR 2.4), abdominal abscesses (OR 3.9), and urinary tract infections (OR 5.9). The score generated by the derivation cohort was validated identifying three different categories with a progressively increased risk for readmission. CONCLUSION: One out of seven patients undergoing DP will be readmitted within 30 days of discharge. Comorbidities seems not to affect the risk. A 10-point score predicts the risk of 30-days readmission.

Recombinant human Mullerian inhibiting substance inhibits long-term growth of MIS type II receptor-directed transgenic mouse ovarian cancers in vivo.

PURPOSE: Mullerian inhibiting substance (MIS) is a glycoprotein hormone that causes Mullerian duct regression in male embryos. In short-term experiments, recombinant human MIS (rhMIS) inhibits xenotransplanted human ovarian cancer cell lines that are thought to be of Mullerian origin. Because this highly lethal cancer has a high recurrence rate after conventional chemotherapy, new treatments are warranted. We examined whether rhMIS as a novel, nontoxic, naturally occurring growth inhibitor can be an effective anticancer drug in long-term studies in vivo against allograft tumors that recapitulate human ovarian carcinoma. EXPERIMENTAL DESIGN: Mouse ovarian carcinoma (MOVCAR) cell lines expressing the early region of the SV40 virus, including the large and small T-antigen genes under transcriptional control of a portion of the murine MIS receptor type II (MISRII) gene promoter, were derived from TgMISIIR-TAg transgenic mice. rhMIS was tested against MOVCAR cells in growth inhibition assays in vitro, and in vivo in 6-week-old female nude mice. Tumor growth in animals was measured at weekly intervals for up to 20 weeks. RESULTS: MOVCAR cells and tumors express MISRII by Western blot, immunohistochemical, and Northern blot analyses. rhMIS significantly inhibited MOVCAR cell growth in vitro and in vivo in three separate long-term allotransplantation experiments. CONCLUSIONS: Because rhMIS is an effective anticancer agent in in vitro and in long-term in vivo preclinical experiments against MISRII-positive tumors, we predict that rhMIS can be used safely and effectively to treat human ovarian malignancies.

Pancreatic ductal adenocarcinoma: long-term survival does not equal cure.

BACKGROUND: Pancreatic ductal adenocarcinoma represents 90% of pancreatic cancers and is an important cause of cancer death in the United States. Operative resection remains as the only treatment providing prolonged survival, but even after a curative resection, 5-year survival rates are low. Our aim was to identify the prognostic factors for long-term survival after resection of pancreatic ductal adenocarcinoma related to patients, treatments, and tumor biology. METHODS: Retrospective review identified 959 patients who underwent resection of their pancreatic adenocarcinoma between February 1985 and December 2010, of whom 499 were resected before November 2006 and represent the cohort we describe in this study. Patient, tumor, and treatment-related variables were assessed for their associations with 5- and 10-year overall survival. RESULTS: Of the 499 patients, 49% were female and median age was 65 years. The majority of patients had stage IIb disease (60%). Actual 5-year survival after resection of pancreatic adenocarcinoma was 19% (95/499), and actual 10-year survival was 10% (33/329). Significant clinicopathologic factors predicting 5- and 10-year survival were negative margins and negative nodal status. Interestingly, 41% (39/95) of long-term survivors had positive nodes and 24% (23/95) had positive margins. CONCLUSION: Pancreatic ductal adenocarcinoma demonstrates a very heterogeneous biology, but patients with negative resection margins and node negative cancers are more likely to survive 5 years after resection. However, our series demonstrates that the biology of the cancer rather than simple pathologic factors determine a patient's prognosis.

Renal abscess in previously healthy girl.

We describe the case of a previously healthy 11-year-old girl with a Staphylococcal aureus renal abscess. The diagnosis was confirmed by renal ultrasonography and abdominal computed tomography. We emphasize the need to obtain an abdominal ultrasound scan and computed tomography scan in patients with abdominal or flank pain, and laboratory evidences of infection to avoid diagnostic and therapeutic delays.

Circumcised hypospadias.

PURPOSE: Patients with circumcised hypospadias have been of significant concern over many decades due to the belief that prior circumcision might negatively affect the results of hypospadias' repair. We evaluated outcomes in consecutive males with anterior, distal penile, and the megameatus with intact prepuce variant of hypospadias (MIP). METHODS: After IRB approval a retrospective of 48 consecutive males with circumcised hypospadias was reviewed. In all cases the urethroplasty was accomplished with either urethral plate tubularization or a MAGPI procedure. No skin flaps were used. RESULTS: A total of 48 patients with circumcised hypospadias (anterior, MIP variant, and distal penile) underwent operative reconstruction by one of us (RVP). All patients were followed for at least 8 months. CONCLUSIONS: Prior circumcision did not negatively affect the results of subsequent urethroplasty in patients with anterior, distal penile, and the MIP variant of hypospadias. The use of the tubularized incised plate urethroplasty (TIP) has virtually eliminated the need for skin flaps in anterior hypospadias repair.

Sexual dimorphism of G-protein subunit Gng13 expression in the cortical region of the developing mouse ovary.

In our search for genes required for the development and function of mouse gonads, we identified Gng13 (guanine nucleotide binding protein 13, gamma), a gene with an embryonic expression pattern highly restricted to the ovary. Based on reverse transcriptase-polymerase chain reaction (RT-PCR) and whole-mount in situ hybridization, Gng13 is expressed in both XX and XY gonads at embryonic day (E) 11.5, but becomes up-regulated in the XX gonad by E12.5. Expression is retained after treatment with busulfan, a chemical known to eliminate germ cells, pointing to the soma as a site of Gng13 transcription. In situ hybridization of embryonic ovarian tissue sections further localized the expression to the cortex of the developing XX gonad. Gng13 expression in the adult is also highly restricted. Northern blot analyses and Genomic Institute of the Novartis Research Foundation expression profiling of adult tissues detected very high expression in the cerebrum and cerebellum, in addition to, a weaker signal in the ovary. Gng13 belongs to a well-known family of signal transduction molecules with functions in many aspects of development and organ physiology. Here, we report that, in the developing mouse embryo, expression of Gng13 mRNA is highly restricted to the cortex of the XX gonad during sexual differentiation, suggesting a role for this gene during ovarian development.

Expression analysis and evolutionary conservation of the mouse germ cell-specific D6Mm5e gene.

During our search for genes required for gonadal development and function in the mouse, we identified D6Mm5e (DNA segment, Chr 6. Miriam Meisler 5, expressed), a gene with an expression pattern highly restricted to the embryonic ovary and the postnatal testis. Based on RT-PCR, Northern blot, and in situ hybridization analyses, we show that D6Mm5e is expressed in the germ cells of the female embryo upon their initial entry into meiosis, and in male germ cells during the last stages of spermatogenesis. Two transcripts are detected in the gonads resulting from the alternative splicing of exon 8. This splicing event does not introduce a frame shift, and creates an mRNA product that uses the same stop codon as the longer transcript. Although D6Mm5e does not belong to any known protein family and does not contain any known protein signature motifs, the high level of evolutionary conservation and the cellular and temporal expression suggest that D6Mm5e may have a role in male and female gametogenesis. Here we report the gonad-restricted mRNA expression profile of D6Mm5e in the mouse, and the evolutionary conservation of its amino acid sequence.

Ovarian cancer side population defines cells with stem cell-like characteristics and Mullerian Inhibiting Substance responsiveness.

The recent identification of "side population" (SP) cells in a number of unrelated human cancers and their normal tissue sources has renewed interest in the hypothesis that cancers may arise from somatic stem/progenitor cells. The high incidence of recurrence attributable to multidrug resistance and the multiple histologic phenotypes indicative of multipotency suggests a stem cell-like etiology of ovarian cancer. Here we identify and characterize SP cells from two distinct genetically engineered mouse ovarian cancer cell lines. Differential efflux of the DNA-binding dye Hoechst 33342 from these cell lines defined a human breast cancer-resistance protein 1-expressing, verapamil-sensitive SP of candidate cancer stem cells. In vivo, mouse SP cells formed measurable tumors sooner than non-SP (NSP) cells when equal numbers were injected into the dorsal fat pad of nude mice. The presence of Mullerian Inhibiting Substance (MIS) signaling pathway transduction molecules in both SP and NSP mouse cells led us to investigate the efficacy of MIS against these populations in comparison with traditional chemotherapies. MIS inhibited the proliferation of both SP and NSP cells, whereas the lipophilic chemotherapeutic agent doxorubicin more significantly inhibited the NSP cells. Finally, we identified breast cancer-resistance protein 1-expressing verapamil-sensitive SPs in three of four human ovarian cancer cell lines and four of six patient primary ascites cells. In the future, individualized therapy must incorporate analysis of the stem cell-like subpopulation of ovarian cancer cells when designing therapeutic strategies for ovarian cancer patients.