Reasons for lack of referral to medical oncology for systemic therapy in stage IV non-small-cell lung cancer: comparison of 2003-2006 with 2010-2011.

INTRODUCTION: Only approximately 25% of stage iv non-small-cell lung cancer (nsclc) patients receive systemic therapy. For such patients, we examined factors affecting referral to a cancer centre (cc) and to medical oncology (mo), and use of systemic therapy. METHODS: Using the Glans-Look Lung Cancer database, we completed a chart review of stage iv nsclc patients diagnosed in Southern Alberta during 2003-2006 and 2010-2011, comparing median overall survival (mos), referral, and treatment in the two cohorts. RESULTS: Of the 922 patients diagnosed in 2003-2006 and the 560 diagnosed in 2010-2011, 94% and 82% respectively were referred to a cc, with 22% and 23% receiving traditional chemotherapy (tctx). Referral to a cc or mo and use of tctx correlated with survival (p < 0.0001): The mos duration was 11.2 months in those receiving tctx and 1.0 months in those not referred to a cc. The overall mos duration was similar in the two cohorts (4.1 months vs. 3.9 months, p = 0.47). Major reasons for lack of referral to mo included poor functional status, rapid decline, and patient wish, which were similar to the reasons for forgoing tctx. In the two cohorts, 87 (9.4%) and 42 (7.5%) patients received epidermal growth factor inhibitors, with a mos duration of 16.2 months. Multivariable analysis showed that male sex [hazard ratio (hr): 1.16; p = 0.008] and pulmonary embolus (hr: 1.2; p = 0.002) correlated with worse survival. In contrast, receipt of chemotherapy (hr: 0.5; p < 0.001) and enrolment in a clinical trial (hr: 0.76; p = 0.049) correlated with better survival. CONCLUSIONS: Our experience confirms that, over time, uptake of systemic therapy, including tctx and targeted therapy, changed little despite their established efficacy. Most of the factors limiting systemic therapy uptake appear to be non-modifiable at the time of referral. Rapid diagnosis and the availability of well-tolerated drugs for all nsclc patients will likely be the most important factors in increasing systemic therapy uptake in this population.

Assessment of treatment patterns associated with injectable disease-modifying therapy among relapsing-remitting multiple sclerosis patients.

BACKGROUND: Availability of oral disease-modifying therapy (DMT) for relapsing-remitting multiple sclerosis (RRMS) may affect injectable DMT (iDMT) treatment patterns. OBJECTIVE: The objective of this paper is to evaluate iDMT persistency, reasons for persistency lapses, and outcomes among newly diagnosed RRMS patients. METHODS: Medical records of 300 RRMS patients initiated on iDMT between 2008 and 2013 were abstracted from 18 US-based neurology clinics. Eligible patients had ≥3 visits: pre-iDMT initiation, iDMT initiation (index), and ≥1 visit within 24 months post-index. MS-related symptoms, relapses, iDMT treatment patterns (i.e. persistency, discontinuation, switching, and restart), and reasons for non-persistency were tracked for 24 months. RESULTS: At 24 months, iDMT persistency was 61.0%; 28.0% of patients switched to another DMT, 8.0% discontinued, and 3.0% stopped and restarted the same iDMT. The most commonly identified reasons for non-persistency were perceived lack of efficacy (22.2%), adverse events (18.8%), and fear of needles/self-injecting (9.4%). At 24 months, 38.0% of patients had experienced a relapse and 11.0% had changes in MRI lesion counts. Patients without MS-related symptoms at index reported increases in the incidence of these symptoms at 24 months. CONCLUSIONS: Non-persistency with iDMT remains an issue in the oral DMT age. Many patients still experienced relapses and disease progression, and should consider switching to more effective therapies.

Adenoid cystic carcinoma of head and neck: clinical predictors of outcome from a Canadian centre.

OBJECTIVES: Adenoid cystic carcinoma (acc) is often treated with surgery, with or without adjuvant radiation therapy (rt). We evaluated disease characteristics, treatments, and potentially prognostic variables in patients with acc. METHODS: Our retrospective analysis considered consecutive cases of acc presenting at a tertiary care hospital between 2000 and 2014. Factors predictive of overall survival (os) and disease-free survival (dfs) were identified by univariate analysis. RESULTS: The 60 patients analyzed had a mean age of 58 years (range: 22-88 years), with a 2:1 female:male ratio. Tumour locations included the major salivary glands (40% parotid, 17% submandibular and sublingual), the oro-nasopharyngeal cavity (27%), and other locations (16%). Of the 60 patients, 35 (58%) received surgery with adjuvant rt; 12 (20%), rt only; 13 (22%), surgery only. Of 18 patients (30%) who experienced a recurrence within 5 years, 3 (5%) developed local recurrence only, and the remaining 15 (25%), distant metastasis. The 5-year os and dfs were 64.5% [95% confidence interval (ci): 45.9% to 78.1%] and 46.2% (95% ci: 29.7% to 61.2%) respectively. In patients without recurrence, 5-year os was 77% (95% ci: 52.8% to 89.9%), and in patients with recurrence, it was 42.7% (95% ci: 15.8% to 67.6%). Patients treated with rt only had a 5-year os of 9.2%. Predictors of 5-year dfs were TNM stage, T stage, nodal status, treatment received, and margin status; age, nodal status, treatment received, and margin status predicted 5-year os. CONCLUSIONS: Despite surgery and rt, one third of patients with acc experience distant recurrence. Patients whose tumours are not amenable to surgery have a poor prognosis, indicating a need for alternative approaches to improve outcomes.

First-, second-, third-line therapy for mRCC: benchmarks for trial design from the IMDC.

BACKGROUND: Limited data exist on outcomes for metastatic renal cell carcinoma (mRCC) patients treated with multiple lines of therapy. Benchmarks for survival are required for patient counselling and clinical trial design. METHODS: Outcomes of mRCC patients from the International mRCC Database Consortium database treated with 1, 2, or 3+ lines of targeted therapy (TT) were compared by proportional hazards regression. Overall survival (OS) and progression-free survival (PFS) were calculated using different population inclusion criteria. RESULTS: In total, 2705 patients were treated with TT of which 57% received only first-line TT, 27% received two lines of TT, and 16% received 3+ lines of TT. Overall survival of patients who received 1, 2, or 3+ lines of TT were 14.9, 21.0, and 39.2 months, respectively, from first-line TT (P<0.0001). On multivariable analysis, 2 lines and 3+ lines of therapy were each associated with better OS (HR=0.738 and 0.626, P<0.0001). Survival outcomes for the subgroups were as follows: for all patients, OS 20.9 months and PFS 7.2 months; for those similar to eligible patients in the first-line ADAPT trial, OS 14.7 months and PFS 5.6 months; for those similar to patients in first-line TIVO-1 trial, OS 24.8 months and PFS 8.2 months; for those similar to patients in second-line INTORSECT trial, OS 13.0 months and PFS 3.9 months; and for those similar to patients in the third-line GOLD trial, OS 18.0 months and PFS 4.4 months. CONCLUSIONS: Patients who are able to receive more lines of TT live longer. Survival benchmarks provide context and perspective when interpreting and designing clinical trials.

Differential apoptotic activities of wild-type FOXL2 and the adult-type granulosa cell tumor-associated mutant FOXL2 (C134W).

Some mutations in FOXL2 result in premature ovarian failure accompanied by blepharophimosis, ptosis, epicanthus inversus syndrome type I disease, and FOXL2-null mice exhibit developmental defects in granulosa cells. Recently, FOXL2 c.402C>G, a new somatic mutation that leads to a p.C134W change, was found in the majority of adult-type ovarian granulosa cell tumors (GCTs). In this study, we investigated the possible mechanisms by which the C134W mutation contributes to the development of GCTs. Wild-type (WT) and mutant FOXL2 displayed differential apoptotic activities. Specifically, WT FOXL2 induced significant granulosa cell death, but the mutant exhibited minimal cell death. The FOXL2-induced apoptotic response was greatly dependent on caspase 8, BID and BAK because the depletion of any of these three proteins inhibited FOXL2 from eliciting the full apoptotic response. Activation of caspase 8 and subsequent increased production of truncated BID, and oligomerization of BAK, and release of cytochrome c were all associated with the apoptosis induced by WT FOXL2 expression. In contrast, the mutant FOXL2 was unable to elicit the full array of apoptotic signaling responses. In addition, we found differential TNF-R1 (tumor necrosis factor-receptor 1) and Fas (CD95/APO-1) upregulation between the WT and the mutant, and the silencing of TNF-R1 or Fas and the blockage of the death signaling mediated by TNF-R1 or Fas using TNF-Fc or Fas-Fc, respectively, resulted in significant attenuations of FOXL2-induced apoptosis. Moreover, granulosa cells that expressed either WT FOXL2 or mutant exhibited distinct cell death sensitivities on activation of death receptors and deprivation of serum. Thus, the differential activities of FOXL2 and its mutant may partially account for the pathophysiology of GCT development.

Erythrocyte ghost-mediated gene delivery for prolonged and blood-targeted expression.

This study reports the use of erythrocyte ghosts (EG) as a biocompatible nonviral delivery system for extended circulation and prolonged expression of plasmid DNA in the blood. Murine interleukin-2-expressing plasmid DNA was efficiently loaded to EG by electroporation in hypotonic condition. The presence of plasmid DNA in EG was confirmed by fluorescence-labeled plasmid DNA. At 21 min after intravenous administration into mice, the level of plasmid DNA in the blood was 92 000-fold higher following EG-mediated delivery as compared to the injection of naked form. EG-mediated gene delivery revealed higher and more prolonged mRNA expression levels of plasmid DNA in the blood until 9 days after the single intravenous injection. Moreover, plasmid DNA-loaded EG showed gene expression targeted to the blood cells. At 3 days post-dose, substantial expression levels of plasmid DNA delivered in EG were observed only in the blood and not in the other organs. Of the blood cells, the subpopulation containing granulocytes showed higher expression of plasmid DNA than mononuclear cells. These results indicate the potential of EG as a safe, prolonged and blood-targeted delivery system of therapeutic genes.

Enhanced immunogenicity of human papillomavirus 16 L1 genetic vaccines fused to an ER-targeting secretory signal peptide and RANTES.

To increase the potency of human papillomavirus (HPV) DNA vaccines, we constructed a series of HPV16 L1 vaccines genetically fused with a secretion signal and/or immune cell-recruiting RANTES. The DNA vaccines encoding secretory HPV L1 were constructed by inserting HPV L1 gene into a vector with an ER-targeting secretory signal sequence. The expression plasmid encoding secretory HPV L1 (pER/L1) was fused with cDNA of RANTES, generating pER/L1/R. For comparison, HPV L1 genes were cloned into pVAX1 vector with no signal sequence (pL1), and further linked to the N-terminus (pL1/R) or C-terminus of RANTES (pR/L1). The secretion of L1 proteins was observed in the pER/L1, pER/L1/R, and pR/L1-transfected cells, except the pL1/R-transfected group. Cytoplasmic localization of L1 protein was observed in the cells transfected with pL1/R, but not with pER/L1/R at 48 h after transfection. In mice, RANTES-fused vaccines more effectively elicited the levels of HPV16 L1-specific IgG and IgG2a antibodies than pL1. Of RANTES-fused vaccines, pER/L1/R encoding the secreted fusion protein induced the highest humoral and CD8(+) T-cell-stimulating responses. These results suggest that the immunogenicity of HPV L1 DNA vaccines could be enhanced by genetic fusion to a chemokine and secretory signal peptide sequences.

Polyethylenimine-mediated cellular uptake, nucleus trafficking and expression of cytokine plasmid DNA.

Although polyethylenimine (PEI) has been widely used as a nonviral vector, there is little mechanistic understanding on PEI-mediated delivery. Here, we studied whether the expression of murine interleukin-2 (mIL-2) plasmids could be improved by complexation with PEI at various N/P ratios, and whether the cellular uptake, nuclear translocation, and retention of plasmids could be affected by the N/P ratios. Compared with the naked mIL-2, PEI/mIL-2 complexes showed at least two orders of magnitude higher expression at Raw264 cells in the N/P ratio-dependent manner. PEI-mediated cellular uptake and nuclear trafficking of plasmids, quantitated by competitive polymerase chain reaction, also depended on the N/P ratios showing the highest cell and nuclear levels of plasmids at 10/1. The higher cellular levels of plasmid DNA after PEI-mediated delivery were also observed in other cell lines. Unlike naked plasmids, PEI/mIL-2 complexes (N/P ratios >/=4/1) showed prolonged cellular and nuclear retention of mIL-2 plasmids. The nuclear translocation and higher cellular level of plasmids given in PEI complexes were similarly observed by fluorescence microscopy. Moreover, PEI/mIL-2 complexes revealed high stability against DNase I, partly explaining the prolonged subcellular retention. These results indicate that the expression of plasmid mIL-2 might be highly enhanced by complexation with PEI and that such increased expression could be attributed by the higher cellular uptake, nuclear translocation and prolonged retention.

Gene amplification and expression of the DNA repair enzyme, N-methylpurine-DNA glycosylase (MPG) in HPV-infected cervical neoplasias.

BACKGROUND: Lethal and mutagenic damages of DNA is caused by a variety of agents including viruses. It is known that HPV is one of the major causes of cervical carcinogenesis and that cells eliminate DNA lesions with DNA repair enzymes. However, the role of N-methylpurine-DNA glycosylase (MPG) is not known in the development of cervical cancer. MATERIALS AND METHODS: Multiplex polymerase chain reaction (PCR) was used for the detection and typing of HPV in the biopsy. Gene amplification of MPG was measured by a PCR-based assay. The mRNA levels of MPG were determined by reverse transcription-PCR using hypoxanthine-guanine phosphoribosyl transferase as the reference gene. An immunohistochemical technique was used to examine the distribution of MPG in the tissues. RESULTS: Of 68 Korean cervical neoplasia patients, 86.8% showed HPV infection. High-risk HPV 16/18 were the most prevalent but positive only in 47.3% of the invasive cancer patients. Gene amplification of MPG was significantly increased in high-risk HPV-infected tissues as compared to low-risk HPV-infected and normal tissues (p < 0.05). The mRNA levels of MPG were higher in HPV-infected invasive carcinoma than normal cervical tissues. Immunohistochemical staining revealed that the intracellular expression and distribution (localization) of MPG altered in the cervical neoplasia. Interestingly, MPG expression in CIN III and invasive carcinoma (IC) was much higher than normal and CIN I. Granular positivity of MPG was notable in the perinuclear regions of the cytoplasm in HPV-infected invasive cancer. CONCLUSION: This is the first report on MPG expression in cervical neoplasia. Our results indicate that the gene amplification and expression of MPG were increased in high-risk HPV-infected cervical neoplasias and the intracellular distribution of MPG protein was altered, suggesting a role of MPG in carcinogenesis.

Soluble adhesion molecules and cytokines in tumor-associated tissue eosinophilia of nasopharyngeal carcinoma.

The phenomenon of tumor-associated tissue eosinophilia (TATE) is seen in some cases of nasopharyngeal carcinoma (NPC) and is characterized by the eosinophils breaking through the vascular wall and pervading the tumor stroma. The margination and trans-endothelial migration of eosinophils in a typical inflammatory reaction depend on the activating effects of certain cytokines and the expression of adhesion molecules on the eosinophils and endothelial cells. In order to investigate whether the adhesion molecules and activating cytokines play a role in eosinophil tumor infiltration, we measured the serum levels of 3 adhesion molecules, intercellular adhesion molecule-1, E-selectin and vascular cell adhesion molecule-1, and 2 cytokines, IL-3 and IL-5, in 60 NPC patients and 40 normal healthy subjects. We found that the NPC patients had higher serum levels of all three soluble adhesion molecules than the normal subjects but the levels of adhesion molecules failed to correlate with the TATE phenomenon. The levels of IL-3 and IL-5 appeared not to differ between the NPC and control groups. We postulate that the three soluble adhesion molecules do not play a major role in TATE and that their elevation in serum may be due to local and/or systemic immune responses.

Nasal absorption and biodistribution of plasmid DNA: an alternative route of DNA vaccine delivery.

Nasal administration is emerging as a new route of DNA vaccine delivery. We aimed to study the extent of absorption and biodistribution of intranasally administered plasmid DNA. After intranasal administration, the level of plasmid DNA in the serum peaked at 1.5 h. The ratio of the area under the concentration (AUC) after intranasal administration of DNA over the AUC after intravenous administration was 0.14. At 15 min post inoculation, the highest organ distribution was observed in the liver and the cervical lymph nodes showed the highest level among the lymph nodes. At 24 h a higher localization of plasmids to the brain than to the lung and spleen was notable. A significant level of mRNA expression was observed in the lymph nodes. These results suggest that plasmid DNA can be substantially absorbed and distributed to the lymph nodes after intranasal administration, partly explaining the systemic immunogenicity of intranasally administered plasmid DNA vaccines.

Increased expression and localization of a serine protease inhibitor, protease nexin-1 (PN-1), in the ovary and uterus during implantation in rat.

Protease nexin-1 (PN-1) is a serine protease inhibitor (serpin) that inactivates several proteases, including thrombin, urokinase, plasminogen activators (PA), and plasmin. It also plays a role in regulating proteolytic activity generated by PA system. PN-1 is known to be involved in tissue remodeling, cellular invasiveness, matrix degradation, and tumor growth. However, the role of PN-1 in female reproductive tracts, such as the uterus, ovary, and oviduct, during pregnancy is not known. The present study was designed to investigate the changes of PN-1 mRNA level and localization in the tracts during implantation and early pregnancy by using reverse transcription (RT)-polymerase chain reaction (PCR) and in situ hybridization. We found that PN-1 mRNA levels were coordinately regulated during early pregnancy in a stage- and tissue-specific manner, such that an increased expression of PN-1 gene appeared at the time of the implantation period in the uterus and ovary. Both the uterus and ovary synthesized PN-1 mRNA and their maximal PN-1 expression occurred on Day 6.5 postcoitum (p.c.). On 13.5 days of pregnancy, PN-1 level was low in the uterus and ovary. On the other hand, PN-1 mRNA in the oviduct did not show after 6.5 days of pregnancy. It appears that PN-1 mRNA in the uterus and ovary was highly regulated during early pregnancy, which might have an important role in implantation of rat blastocysts. PN-1 was localized in endometrial stromal cells of the uterus and in granulosa cells of the unstimulated primary follicles in the ovary during periimplantation period. Also, PN-1 mRNA expression was higher at implantation period than that at nonimplantation period of pregnancy. In conclusion, PN-1 is expressed in female reproductive tracts and highly regulated during implantation and early pregnancy.

Embryotropic role of hemoglobin and ethylenediaminetetraacetic acid in preimplantation development of ICR mouse 1-cell embryos.

OBJECTIVE: To evaluate embryotropic action of hemoglobin (Hb) and ethylenediaminetetraacetic acid (EDTA) on preimplantation embryo development. DESIGN: In vitro model study using mouse embryos. SETTING: University affiliated hospital, Pochon CHA University. ANIMALS: Four-week-old block strain ICR mice naturally mated after superovulation. INTERVENTION(S): One-cell embryos were cultured in serum-free, modified preimplantation-1 medium, to which 1 microg/ml Hb and/or 0.1 mM EDTA were added. MAIN OUTCOME MEASURE(S): Preimplantation development and blastomere number. RESULT(S): More (P<.05) 1-cell embryos developed to the 4-cell (52% vs. 67%-84%), 8-cell (48% vs. 65%-81%), and blastocyst (40% vs. 61%-79%) stages after the addition of hemoglobin (Hb) and/or EDTA than after no addition. Highest proportion of embryos developed to each stage after the combined addition of Hb+EDTA. EDTA specifically stimulated the development before the 8-cell stage, which was as similar as Hb+EDTA. On the contrary, higher ratio of morula to blastocyst transformation was obtained after the addition of Hb or Hb+EDTA than after no addition (0.76 vs. 0.96-0.98). Significant increases in the cell number of blastocysts (46.5-47.2 vs. 53.2 cells), inner cell mass (ICM) cells (16.7-17.5 vs. 21 cells), and the ratio of ICM cells to trophoblasts (0.3-0.37 to 0.39) were found after the combined addition of Hb+EDTA, compared with no addition or with the addition of EDTA or Hb alone. CONCLUSIONS: Hb and EDTA have stage-specific effects on supporting preimplantation embryo development; Hb promotes both the development before the 8-cell stage and the morula to blastocyst transformation, whereas EDTA mainly promotes the development to the 8-cell stage. The combined exposure of embryos to Hb and EDTA improves not only preimplantation development but also the growth and quality of blastocysts.

Spatial expression of a DNA repair gene, N-methylpurine-DNA glycosylase (MPG) during development in mice.

BACKGROUND: DNA repair is a crucial phenomenon that maintains the chromosome integrity of genome which are continuously damaged by endogenous and exogenous alkylating agents. If the damaged DNA is not repaired, it may lead to mutation, chromosomal aberration, aging and cancer. N-methylpurine-DNA glycosylase (MPG), a ubiquitous DNA repair enzyme, removes N-methylpurine and other damaged purines in DNA. MATERIALS AND METHODS: MPG mRNA expression was revealed at various stages of mouse development from day 7.5 p.c. (post coitum) embryo to day 400 mature adult by Northern blot hybridization or RT-PCR. RESULTS: MPG transcripts were abundant in the mouse embryo during pregnancy and in adult testis and ovary. The MPG mRNA level in the testis was low in 1-week-old mice, but the level showed its maximum among the organs tested in 4-week-old young adults. In placenta, the level of MPG mRNA continuously decreased from day 7.5 p.c. to day 17.5 p.c. CONCLUSIONS: The spatial expression of MPG gene is highly regulated. Transcription of MPG is maximum in rapidly dividing and growing tissues during development. These data suggest that an elevated rate of MPG transcription is required for DNA replication.

Pregnancies and deliveries after in vitro maturation culture followed by in vitro fertilization and embryo transfer without stimulation in women with polycystic ovary syndrome.

OBJECTIVE: To improve the efficacy of an IVF-ET program for unstimulated patients with polycystic ovary syndrome (PCOS) with the use of culture for oocyte maturation. DESIGN: Prospective studies with the comparison of different ET procedures from March 1995 through February 1998. SETTING: University-affiliated hospital. PATIENT(S): Ninety-four cycles in 64 consenting patients with PCOS. INTERVENTION(S): Immature oocytes were retrieved from unstimulated patients with PCOS and subsequently cultured and fertilized in vitro. Zygote intrafallopian transfer (ZIFT), uterine ET, or a combined approach of ZIFT + uterine ET was subsequently performed. MAIN OUTCOME MEASURE(S): Laboratory and clinical data. RESULT(S): Among 1, 280 immature oocytes (13.6 +/- 7.5 oocytes per patient) retrieved, 89% (1,139) were morphologically normal, and 62.2% (708/1,139) of the normal oocytes matured in vitro after culture for 48 hours. When intracytoplasmic sperm injection was performed, 68% (481/708) developed to the normal pronuclear stage, and 88.1% of the embryos cocultured with Vero cells (266/302) cleaved. Eighty-five ET cycles were conducted and pregnancy was established in 23 cycles (27.1%), which consisted of 8 after uterine ET and 15 after a combined approach. Seventeen patients delivered 20 normal infants. CONCLUSION(S): The IVF-ET method using no ovarian stimulation followed by in vitro maturation culture can be a feasible assisted reproductive technology for treatment of PCOS with various complications.

In vitro blastocyst formation of human oocytes obtained from unstimulated and stimulated cycles after vitrification at various maturational stages.

OBJECTIVE: To evaluate the developmental competence and chromosomal normality of oocytes vitrified at various times after maturation culture. DESIGN: In vitro model study. SETTING: A university-affiliated hospital. PATIENT(S): Unstimulated women who underwent cesarean section or oophorectomy and infertile women who underwent a long protocol of GnRH stimulation. INTERVENTION(S): Retrieved oocytes were vitrified at 0 or 48 hours after culture in unstimulated cycles and at 0, 8-15, or 24-28 hours after culture in stimulated cycles. MAIN OUTCOME MEASURE(S): Postthaw morphologic normality, maturation, fertilization, cleavage, blastocyst formation, and chromosome number. RESULT(S): In the 53 oocytes that were obtained from unstimulated cycles, no statistically significant differences were found in rates of morphologic normality (range, 56%-63%) or fertilization (range, 31%-37%) according to the time of vitrification. In the 50 oocytes that were obtained from stimulated cycles, more of those that were vitrified at 24-28 hours were morphologically normal than those that were vitrified at 0 or 8-15 hours. Regardless of these differences, high cleavage rates (83%-100%) were obtained that did not differ significantly among the treatment groups. In both cycles, 20%-43% of cleaved oocytes developed to the blastocyst stage by 6 days after IVF. All the karyotyped blastocysts, three from unstimulated cycles and four from stimulated cycles, had a normal number of chromosomes. CONCLUSION(S): Vitrified and thawed oocytes from unstimulated or stimulated cycles developed to the blastocyst stage, regardless of when vitrification occurred; the number of chromosomes in the blastocysts was normal.

Freezing immature oocytes.

The establishment of a long-term preservation system for mammalian oocytes is important for the development of both biological and medical sciences. A number of efforts have been made to develop this system. In human reproductive medicine, the development of an oocyte cryopreservation system can improve the efficacy of the current assisted reproductive technology (ART) for infertile patients with severe reproductive disorders. In this article, the technical development of cryopreservation programs for human oocytes and its biological background were reviewed. Clinical outcome after the use of this technology was further introduced.