Consumption of Solnul™ Resistant Potato Starch Produces a Prebiotic Effect in a Randomized, Placebo-Controlled Clinical Trial.

The effects of resistant starch at high doses have been well-characterized, but the potential prebiotic effects of resistant starch at doses comparable to oligosaccharide prebiotics have not been evaluated. A three-arm randomized, double-blind, placebo-controlled clinical trial was conducted to evaluate the effect of 3.5 g and 7 g daily doses of Solnul™ resistant potato starch (RPS) on beneficial populations of gut bacteria and stool consistency after a 4-week period. The relative abundance of Bifidobacterium and Akkermansia was determined by employing 16Sv4 sequencing of stool samples. To assess the effect of RPS on laxation and bowel movements, stools were recorded and scored using the Bristol Stool Form Scale. Participants consuming 3.5 g/day of RPS experienced significantly greater changes in Bifidobacterium and Akkermansia compared to the placebo after 4 weeks. The number of diarrhea- and constipation-associated bowel movements were both significantly lower in the 3.5 g RPS arm compared to the placebo group. Participants consuming 7 g of RPS responded similarly to those in the 3.5 g arm. Our analyses demonstrate that Solnul™ RPS has a prebiotic effect when consumed for 4 weeks at the 3.5 g per day dose, stimulating increases in beneficial health-associated bacteria and reducing diarrhea- and constipation-associated bowel movements when compared to the placebo group.

Playing a social dilemma game as an exploratory learning activity before instruction improves conceptual understanding.

Society's most pressing problems involve social dilemmas, yet few individuals recognize and understand their core components. We examined how a serious social dilemma game used in an educational setting impacted understanding of a classic social dilemma, the tragedy of the commons. Participants (N = 186) were randomly assigned to one of two gameplay conditions or a Lesson-Only condition without the game (traditional lesson with a reading). In the Explore-First condition, participants played the game as an exploratory learning activity before the lesson. In the Lesson-First condition, participants played the game after the lesson. Both gameplay conditions were rated as more interesting than the Lesson-Only condition. However, participants in the Explore-First condition exhibited higher conceptual understanding and spontaneous transfer to real-world dilemmas than the other conditions, which did not differ. These benefits were selective to social concepts (e.g., self-interest, interdependency) explored via gameplay. These benefits did not occur for ecological concepts (e.g., scarcity, tragedy), which were taught to everyone during the beginning instructions. Policy preferences were equal across conditions. Serious social dilemma games offer a promising educational tool for conceptual development when students can explore the complexities of social dilemmas for themselves. (PsycInfo Database Record (c) 2023 APA, all rights reserved).

The Hidden Cost of the Opioid Epidemic in the United States: Drug Screening in Insurance Claims.

BACKGROUND: The opioid crisis has had a substantial financial impact on the health care system in the United States. This study evaluates how health plans have been affected financially and shows how a laboratory benefit management (LBM) program can be used to address related drug testing in an outpatient setting. METHODS: Monthly claims data from private health plans were collected from June 1, 2016 to February 29, 2020. The total number of claims (units) for definitive and presumptive drug testing were calculated and the number of paid claims recorded. Claims distribution by laboratory type and medical code billed, the paid rate and compound annual growth rate, and the test distribution and paid rate of rendering providers who had submitted a minimum of 1000 claims were determined. RESULTS: In total, 2,004,230 drug testing claims were submitted. After the LBM program was implemented, the percentage of paid claims for definitive drug testing (Healthcare Common Procedure Coding System code G0483) decreased and the paid rate for the low-cost tests (HCPCS code G0480) in physician office and independent laboratory settings increased. The compound annual growth rate for G0483 claims submitted indicated a 70.5% and 31.9% decrease in payments to physician offices and independent laboratories, respectively, for the period ending February 2020. CONCLUSIONS: An LBM program can positively address policy enforcement while reducing unnecessarily complex tests and limiting potential fraud, waste, and abuse by directing testing toward laboratories amenable to cost-efficient contractual savings. Moreover, for definitive drug testing, the enforcement of the use of Healthcare Common Procedure Coding System codes and a move toward more cost-efficient tests (G0480), when clinically applicable, supported by clinical practice guidelines, or evidence-based medicine, is an approach to providing medical benefits while maintaining health costs.

Increasing levels of Parasutterella in the gut microbiome correlate with improving low-density lipoprotein levels in healthy adults consuming resistant potato starch during a randomised trial.

BACKGROUND: Prebiotics, defined as a substrate that is selectively utilized by host microorganisms conferring a health benefit, present a potential option to optimize gut microbiome health. Elucidating the relationship between specific intestinal bacteria, prebiotic intake, and the health of the host remains a primary microbiome research goal. OBJECTIVE: To assess the correlations between gut microbiota, serum health parameters, and prebiotic consumption in healthy adults. METHODS: We performed ad hoc exploratory analysis of changes in abundance of genera in the gut microbiome of 75 participants from a randomized, placebo-controlled clinical trial that evaluated the effects of resistant potato starch (RPS; MSPrebiotic®, N = 38) intervention versus a fully digestible placebo (N = 37) for which primary and secondary outcomes have previously been published. Pearson correlation analysis was used to identify relationships between health parameters (ie. blood glucose and lipids) and populations of gut bacteria. RESULTS: Abundance of Parasutterella (phylum Proteobacteria) tended to increase in the gut microbiome of individuals consuming RPS and those increases in Parasutterella were correlated with reductions in low-density lipoprotein (LDL) levels in participants consuming RPS but not placebo. Segregating RPS-consuming individuals whose LDL levels decreased (ie "Responders") from those who did not (ie. "Non-Responders") revealed that LDL Responders had significantly higher levels of Parasutterella both at baseline and after 12 weeks of consuming RPS. CONCLUSION: Our analyses suggest that RPS may help improve LDL levels depending upon the levels of Parasutterella in an individual's gut microbiome. TRIAL REGISTRATION: This study protocol was reviewed and approved by Health Canada (Submission #188517; "Notice of Authorization" dated 06/05/13) and registered as NCT01977183 (10/11/13) listed on NIH website: ClinicalTrials.gov. Data generated in this study have been submitted to NCBI ( http://www.ncbi.nlm.nih.gov/bioproject/381931 ). FUNDING: MSP Starch Products Inc.

Agricultural Exposures and Breast Cancer Among Latina in the San Joaquin Valley of California.

OBJECTIVE: The aim of this study was to assess the role of agricultural work, pesticide exposure, and age at first farm labor exposure in breast cancer (BC) risk among Hispanic women in Central California. METHODS: A BC case control study was conducted. Latina BC cases were identified through the California Cancer Registry and controls were recruited. Both cases and controls completed a detailed questionnaire. Pesticide exposure data were obtained by linking the crops, work locations, and dates worked in specific farm jobs with the California Department of Pesticide Regulation (DPR) Pesticide Use Reports (PUR). RESULTS: Chemicals associated with BC risk included organophosphates, organochlorines, and a phthalimide, Captan. Age at first work in farm labor was younger in cases than controls (P = 0.03). CONCLUSIONS: Agricultural work may be associated with the increased BC risk in female Hispanic farm workers.

Formulation and evaluation of mixed polymeric micelles of quercetin for treatment of breast, ovarian, and multidrug resistant cancers.

BACKGROUND: Quercetin (QCT), a naturally occurring flavonoid has a wide array of pharmacological properties such as anticancer, antioxidant and anti-inflammatory activities. QCT has low solubility in water and poor bioavailability, which limited its use as a therapeutic molecule. Polymeric micelles (PMs) is a novel drug delivery system having characteristics like smaller particle size, higher drug loading, sustained drug release, high stability, increased cellular uptake and improved therapeutic potential. In the present study, we have formulated and characterized mixed PMs (MPMs) containing QCT for increasing its anticancer potential. METHODS: The MPMs were prepared by thin film hydration method, and their physicochemical properties were characterized. The in vitro anticancer activity of the MPMs were tested in breast (MCF-7 and MDA-MB-231, epithelial and metastatic cancer cell lines, respectively), and ovarian (SKOV-3 and NCI/ADR, epithelial and multi-drug resistant cell lines, respectively) cancer. RESULTS: The optimal MPM formulations were obtained from Pluronic polymers, P123 and P407 with molar ratio of 7:3 (A16); and P123, P407 and TPGS in the molar ratio of 7:2:1 (A22). The size of the particles before lyophilization (24.83±0.44 nm) and after lyophilisation (37.10±4.23 nm), drug loading (8.75±0.41%), and encapsulation efficiency (87.48±4.15%) for formulation A16 were determined. For formulation A22, the particle size before lyophilization, after lyophilization, drug loading and encapsulation efficiency were 26.37±2.19 nm, 45.88±13.80 nm, 9.01±0.11% and 90.07±1.09%, respectively. The MPMs exhibited sustained release of QCT compared to free QCT as demonstrated from in vitro release experiments. The solubility of QCT was markedly improved compared to pure QCT. The MPMs were highly stable in aqueous media as demonstrated by their low critical micelle concentration. The concentration which inhibited 50% growth (IC50) values of both micellar preparations in all the cancer cell lines were significantly less compared to free QCT. CONCLUSION: Both the MPMs containing QCT could be used for effective delivery to different type of cancer and may be considered for further development.

Rac1 Dosage Is Crucial for Normal Endochondral Bone Growth.

Rac1, a member of the small Rho GTPase family, plays multiple cellular roles. Studies of mice conditionally lacking Rac1 have revealed essential roles for Rac1 in various tissues, including cartilage and limb mesenchyme, where Rac1 loss produces dwarfism and long bone shortening. To gain further insight into the role of Rac1 in skeletal development, we have used transgenic mouse lines to express a constitutively active (ca) Rac1 mutant protein in a Cre recombinase-dependent manner. Overexpression of caRac1 in limb bud mesenchyme or chondrocytes leads to reduced body weight and shorter bones compared with control mice. Histological analysis of growth plates showed that caRac1;Col2-Cre mice displayed ectopic hypertrophic chondrocytes in the proliferative zone and enlarged hypertrophic zones. These mice also displayed a reduced proportion of proliferating cell nuclear antigen-positive cells in the proliferative zone and nuclear ß-catenin localization in the ectopic hypertrophic chondrocytes. Importantly, overexpression of caRac1 partially rescued the phenotypes of Rac1fl/fl;Col2-Cre and Rac1fl/fl;Prx1-Cre conditional knockout mice, including body weight, bone length, and growth plate disorganization. These results suggest that tight regulation of Rac1 activity is necessary for normal cartilage development.

Loss of the mammalian DREAM complex deregulates chondrocyte proliferation.

Mammalian DREAM is a conserved protein complex that functions in cellular quiescence. DREAM contains an E2F, a retinoblastoma (RB)-family protein, and the MuvB core (LIN9, LIN37, LIN52, LIN54, and RBBP4). In mammals, MuvB can alternatively bind to BMYB to form a complex that promotes mitotic gene expression. Because BMYB-MuvB is essential for proliferation, loss-of-function approaches to study MuvB have generated limited insight into DREAM function. Here, we report a gene-targeted mouse model that is uniquely deficient for DREAM complex assembly. We have targeted p107 (Rbl1) to prevent MuvB binding and combined it with deficiency for p130 (Rbl2). Our data demonstrate that cells from these mice preferentially assemble BMYB-MuvB complexes and fail to repress transcription. DREAM-deficient mice show defects in endochondral bone formation and die shortly after birth. Micro-computed tomography and histology demonstrate that in the absence of DREAM, chondrocytes fail to arrest proliferation. Since DREAM requires DYRK1A (dual-specificity tyrosine phosphorylation-regulated protein kinase 1A) phosphorylation of LIN52 for assembly, we utilized an embryonic bone culture system and pharmacologic inhibition of (DYRK) kinase to demonstrate a similar defect in endochondral bone growth. This reveals that assembly of mammalian DREAM is required to induce cell cycle exit in chondrocytes.

Small molecule-induced mitochondrial disruption directs prostate cancer inhibition via UPR signaling.

We previously identified SMIP004 (N-(4-butyl-2-methyl-phenyl) acetamide) as a novel inducer of cancer-cell selective apoptosis of human prostate cancer cells. SMIP004 decreased the levels of positive cell cycle regulators, upregulated cyclin-dependent kinase inhibitors, and resulted in G1 arrest, inhibition of colony formation in soft agar, and cell death. However, the mechanism of SMIP004-induced cancer cell selective apoptosis remained unknown. Here, we used chemical genomic and proteomic profiling to unravel a SMIP004-induced pro-apoptotic pathway, which initiates with disruption of mitochondrial respiration leading to oxidative stress. This, in turn, activates two pathways, one eliciting cell cycle arrest by rapidly targeting cyclin D1 for proteasomal degradation and driving the transcriptional downregulation of the androgen receptor, and a second pathway that activates pro-apoptotic signaling through MAPK activation downstream of the unfolded protein response (UPR). SMIP004 potently inhibits the growth of prostate and breast cancer xenografts in mice. Our data suggest that SMIP004, by inducing mitochondrial ROS formation, targets specific sensitivities of prostate cancer cells to redox and bioenergetic imbalances that can be exploited in cancer therapy.

Loss of the Prader-Willi obesity syndrome protein necdin promotes adipogenesis.

We investigated the role of necdin during adipogenic differentiation. Necdin is one of several genes inactivated in children with Prader-Willi syndrome, who are predisposed to increased adiposity at the expense of lean mass. Necdin promotes neuronal and muscle differentiation and survival through interactions with a variety of proteins, including cell surface receptors, modifiers of protein stability, and transcription factors. In pre-adipocytes, necdin over-expression inhibits adipogenesis, while reducing necdin levels enhances adipogenic differentiation in tissue culture cells. We now directly demonstrate a role for necdin in inhibiting adipogenesis using cells derived from necdin deficient mice.

Comparative proteomic analysis of a cytosolic fraction from ß3 integrin-deficient cells.

Integrins are heterodimeric transmembrane receptors involved in sensing and transmitting informational cues from the extracellular environment to the cell. This study explored sub-proteome changes in response to elimination of the ß3 integrin using a knockout murine model. Cleavable isotope-coded affinity tagging (cICAT) in combination with sub-cellular fractionation, multiple dimensions of separation and tandem mass spectrometry (MS/MS) were used to characterize differentially expressed proteins among ß3 integrin(-/-) (ß3(-/-)) mouse embryonic fibroblasts and isogenic wild-type (WT) controls. From a cytosolic protein fraction, 48 proteins were identified, in which expression differed by > 1.5-fold. Predominant ontological groups included actin-binding/cytoskeletal proteins and protease/protease inhibitors. Interestingly, ß3 integrin expression was inversely correlated with expression of cathepsin B, a lysosomal cysteine protease, as its expression was greater by over 3.5-fold in the ß3(-/-) cells. This inverse correlation was also observed in stable heterologous cells transfected with ß3 integrin, where the intracellular expression and activity of cathepsin B was lower compared to control cells. Our data suggests that the composition of the cellular proteome is influenced by integrin expression patterns and reveals a strong functional relationship between ß3 integrin and cathepsin B.

Higher population-based incidence rates of triple-negative breast cancer among young African-American women : Implications for breast cancer screening recommendations.

BACKGROUND: Differences in the breast cancer burden of African-American women compared with white American women are well documented. Recent controversies have emerged regarding age-appropriate mammographic screening guidelines, and these surveillance recommendations may influence future breast cancer disparities. The objective of the current study was to evaluate age-specific breast cancer stage distributions and incidence rates of triple-negative breast cancer (TNBC) in a population-based tumor registry. METHODS: The authors analyzed breast cancers from the California Cancer Registry (CCR) that were diagnosed between 1988 and 2006. The results were stratified by age and race/ethnicity, with white Americans identified as non-Hispanic whites (NHWs) and African Americans identified as non-Hispanic blacks (NHBs). Breast cancer stage distributions and TNBC incidence rates also were analyzed. RESULTS: In total, 375,761 invasive breast cancers were evaluated (including 276,938 in NHWs and 21,681 in NHBs). NHBs and Hispanics tended to be younger than NHWs (median ages 57 years, 54 years, and 64 years, respectively). Lifetime incidence rates were higher for NHWs compared with NHBs and Hispanics; however, for women aged <44 years, incidence was highest among NHBs. NHBs also had higher incidence rates of stage III and IV disease and a higher incidence of TNBC in all age categories. CONCLUSIONS: Population-based data demonstrated that African-American women had a more advanced stage distribution for breast cancer compared with white American women and higher incidence rates for TNBC. These patterns were observed for women ages 40 to 49 years and for older women, and they suggest that mammographic screening for the early detection of breast cancer will be particularly relevant for younger African-American women.

Loss of Necdin impairs myosin activation and delays cell polarization.

NDN is one of several genes inactivated in Prader-Willi syndrome (PWS), a developmental disorder characterized by obesity, hypotonia, and developmental delay. We demonstrate that loss of Necdin in murine and human fibroblasts impairs polarity initiation through a Cdc42-myosin-dependent pathway, thereby reducing cell migration. We identified defective polarization in both primary neuron cultures and in the developing limb in Ndn-null mice. Ndn-null neurons fail to activate myosin light chain and display defective polarization with respect to a brain-derived neurotrophic factor gradient. Pax3+ muscle progenitors in Ndn-null developing forelimbs display defective polarization, do not adequately migrate into the dorsal limb bud, and extensor muscles are consequently smaller. These results provide strong evidence that Necdin is a key protein regulating polarization of the cytoskeleton during development. Furthermore, this is the first demonstration of a cellular defect in PWS and suggests a novel molecular mechanism to explain neurological and muscular pathophysiologies in PWS.

Assessment of erlotinib in chemoresponse assay.

BACKGROUND: There is a need to identify the subset of patients sensitive to epidermal growth factor receptor (EGFR) inhibition prior to using such treatments. MATERIALS AND METHODS: Three non-small cell lung cancer (NSCLC) cell lines (H292, H358, and Calu3) and 34 primary human lung tumor specimens were tested for chemoresponse to erlotinib. RESULTS: The assay distinguished responsiveness to erlotinib among NSCLC cell lines and human lung tumor explants. The H292 cells were responsive, the Calu3 cells were intermediate responsive and the H358 cells were non-responsive. These results were consistent with published tumor growth inhibition by erlotinib in xenografts derived from these cell lines. Out of the 34 patient specimens, 3 (8.8%) were responsive to erlotinib, 7 (20.6%) were intermediate responsive and 24 (70.6%) were non-responsive. CONCLUSION: The in vitro chemoresponse assay profile was similar to that noted for human tumors in clinical trials. Chemoresponse testing may help predict patient response to erlotinib and assist chemotherapy decision-making.

Feasibility assessment of a chemoresponse assay to predict pathologic response in neoadjuvant chemotherapy for breast cancer patients.

BACKGROUND: For chemosensitivity and resistance assays to be clinically useful in predicting patient outcome, they should require small amounts of tissue and be highly reproducible and reliable. PATIENTS AND METHODS: Expanded tumor cells from transcutaneous biopsies of breast lesions (n=62) were tested for chemoresponse using the cell-based ChemoFx assay. Pathologic complete response (pCR) was determined on a subset of patients (n=34). Assay score and pCR were determined independently in a blinded manner. Logistic regression models were used to select predictors for response. RESULTS: Tumor cells were successfully isolated from 83.9% of patients. Chemoresponse profiles were robust and reproducible with coefficient of variance of <3%. In a limited initial patient outcome correlation, assay score of docetaxel/capecitabine significantly predicted pCR; the cross-validated model was 75% accurate. CONCLUSION: It is feasible to assess the chemoresponsiveness of small breast lesions using the ChemoFx assay to assist in choosing neoadjuvant chemotherapy for breast cancer patients.

Comparison of angiogenesis-related factor expression in primary tumor cultures under normal and hypoxic growth conditions.

BACKGROUND: A localized hypoxic environment occurs during tumor growth necessitating an angiogenic response or tumor necrosis results. Novel cancer treatment strategies take advantage of tumor-induced vascularisation by combining standard chemotherapeutic agents with angiogenesis-inhibiting agents. This has extended the progression-free interval and prolonged survival in patients with various types of cancer. We postulated that the expression levels of angiogenesis-related proteins from various primary tumor cultures would be greater under hypoxic conditions than under normoxia. METHODS: Fifty cell sources, including both immortalized cell lines and primary carcinoma cells, were incubated under normoxic conditions for 48 hours. Then, cells were either transferred to a hypoxic environment (1% O2) or maintained at normoxic conditions for an additional 48 hours. Cell culture media from both conditions was collected and analyzed via an ELISA-based assay to determine expression levels of 11 angiogenesis-related factors: VEGF, PDGF-AA, PDGF-AA/BB, IL-8, bFGF/FGF-2, EGF, IP-10/CXCL10, Flt-3 ligand, TGF-beta1, TGF-beta2, and TGF-beta3. RESULTS: A linear correlation between normoxic and hypoxic growth conditions exists for expression levels of eight of eleven angiogenesis-related proteins tested including: VEGF, IL-8, PDGF-AA, PDGF-AA/BB, TGF-beta1, TGF-beta2, EGF, and IP-10. For VEGF, the target of current therapies, this correlation between hypoxia and higher cytokine levels was greater in primary breast and lung carcinoma cells than in ovarian carcinoma cells or tumor cell lines. Of interest, patient cell isolates differed in the precise pattern of elevated cytokines. CONCLUSION: As linear correlations exist between expression levels of angiogenic factors under normoxic and hypoxic conditions in vitro, we propose that explanted primary cells may be used to probe the in vivo hypoxic environment. Furthermore, differential expression levels for each sample across all proteins examined suggests it may be possible to build a predictor for angiogenesis-related anticancer agents, as each sample has a unique expression profile. Further studies should be performed to correlate in vitro protein expression levels of angiogenesis-related factors with in vivo patient response.

The Prader-Willi syndrome protein necdin interacts with the E1A-like inhibitor of differentiation EID-1 and promotes myoblast differentiation.

Proliferation and differentiation of muscle precursors are controlled by the activation of muscle-specific genes and inactivation of inhibitors of differentiation. Necdin is a multi-functional protein that is up-regulated during neural and myogenic differentiation. Necdin facilitates cell cycle exit and differentiation during development, but the role of necdin in embryonic myogenesis has not been described. In a cytoplasmic two-hybrid screen, we identified a novel interaction between necdin and the E1A-like inhibitor of differentiation (EID-1). EID-1 inhibits transcriptional activation of genes required for myogenic differentiation, and is degraded in myoblasts upon cell cycle exit. In a transactivation assay, necdin had no direct effect on myoD-responsive promoters in the presence of MyoD, but necdin did relieve the EID-1-dependent inhibition of these same promoters. In vivo, a normal number of MyoD-expressing myoblasts was present in primary embryonic limb bud cultures from mouse embryos with congenital necdin deficiency. In contrast, the number of myosin heavy chain-expressing myotubes in differentiating limb bud cultures cultured for 5 days was reduced compared with cultures from wild-type littermate controls. In the presence of necdin, steady-state levels of EID-1 were increased and the half-life of EID-1 was extended, and EID-1 was re-localized from the nucleus to the cytoplasm when necdin was co-expressed in transfected cells. Collectively, these data are consistent with a model whereby necdin promotes myoblast differentiation at least in part by relieving the inhibitory effect of EID-1.

The ChemoFx assay: an ex vivo chemosensitivity and resistance assay for predicting patient response to cancer chemotherapy.

The ChemoFx Assay is an ex vivo assay designed to predict the sensitivity and resistance of a given patient's solid tumor to a variety of chemotherapy agents. A portion of a patient's solid tumor, as small as a core biopsy, is mechanically disaggregated and established in primary culture where malignant epithelial cells migrate out of tumor explants to form a monolayer. Cultures are verified as epithelial and exposed to increasing doses of selected chemotherapeutic agents. The number of live cells remaining post-treatment is enumerated microscopically using automated cell-counting software. The resultant cell counts in treated wells are compared with those in untreated control wells to generate a dose-response curve for each chemotherapeutic agent tested on a given patient specimen. Features of each dose-response curve are used to score a tumor's response to each ex vivo treatment as "responsive," "intermediate response," or "non-responsive." Collectively, these scores are used to assist an oncologist in making treatment decisions.

Ex vivo enrichment of malignant carcinoma cells in primary culture.

AIMS: Establishing and maintaining human tumours in primary culture can be challenging. In this application, a short-term primary culture process is desired to ensure cells maintained in culture are representative of the in vivo tumour for the purpose of chemoresponse testing. To ensure the appropriate cells are being grown, the cultures must be evaluated for malignancy. The clinical gold standard determination of malignancy is cytological evaluation by a cytopathologist. METHODS: Fifty human tumour specimens (breast, colon, lung, ovary) were established and maintained in primary culture. Cytospins were prepared upon initiation of culture and again at completion of the culture process. Cytospins were stained (Diff-Quik, Papanicolaou) and evaluated by a cytopathologist for the percentage of malignant cells at both times. RESULTS: An increase in the percentage of malignant cells was noted in 86% (43/50) of the cultures evaluated; 8% (4/50) of the cultures maintained the same percentage of malignant cells throughout the culture period, and 6% (3/50) displayed a decrease in malignant cells. On average, the percentage of malignant cells increased by 37% and was not associated with the length of culture (range 5-28 days). CONCLUSIONS: The described primary culture process enriches for malignant cells, which is desirable for further evaluation such as chemoresponse testing.