[Prenatal diagnosis and genetic counselling for a pedigree carrying a large fragment deletion of 13q].

OBJECTIVE: To carry out prenatal diagnosis for a fetus with normal ultrasonographic finding at 20 weeks' gestation but a copy number variant(CNV) of 13q indicated by non-invasive prenatal test (NIPT). METHODS: Karyotyping analysis and chromosomal CNV assay were carried out on the amniotic fluid sample. Parental peripheral blood sample was collected for chromosomal analysis. Detailed fetal ultrasound scan was carried out to rule out structural abnormalities of the fetus. RESULTS: The fetus was detected with a heterozygous 10.14 Mb deletion at 13q21.1q21.32, which has originated from the phenotypically normal mother. No apparent karyotypic abnormality was detected in the fetus and its parents. No ultrasonic abnormality was found in the fetus. CONCLUSION: Both the fetus and its mother have carried a heterozygous 10.14 Mb deletion at 13q21.1q21.32 and presented normal phenotypes.Combined with literature review, the segmental deletion was judged to be a benign variant.

Clinical and Genetic Characteristics of 17 α-Hydroxylase/17, 20-Lyase Deficiency: c.985_987delTACinsAA Mutation of CYP17A1 Prevalent in the Chinese Han Population.

OBJECTIVE: 17 α-hydroxylase/17, 20-lyase deficiency (17-OHD) is a rare recessive hereditary disease that can be attributed to cytochrome P450 17 α-hydroxylase deficiency caused by CYP17A1 gene mutations. METHODS: A large cohort of 10 Chinese Han patients with 17-OHD from 2012 to 2020 were enrolled. The clinical and biochemical features were investigated, and genetic mutations of CYP17A1 were analyzed by polymerase chain reaction-Sanger sequencing. Karyotype identification and the SRY gene test were also carried out. In silico analysis was used to predict the effects of genetic mutations on the protein function. RESULTS: All patients were female. Common complaints were hypertension, hypokalemia, and primary amenorrhea. The karyotype was 46, XY, and the SRY gene was detected in 7 patients; the karyotype was XX in the remaining 3 patients. A total of 7 mutations including Y329N, Y329X, Y329Lfs∗, R96W, A82D, S380N, and A487_P489del have been identified in the CYP17A1 gene. The Y329Lfs∗ mutation was found in 9/10 (90%) of patients with a high allele frequency of 70%. In silico prediction showed that a novel variant of c.1139G>A (S380N) occurs at a conserved residue and can cause disease. CONCLUSION: We presented a detailed description of the clinical and genetic characteristics in Chinese patients with 17-OHD and concluded that Y329Lfs∗ mutation of CYP17A1 is prevalent in the Chinese Han population. Therefore, hotspot screening by polymerase chain reaction-Sanger sequencing for exon 6 of CYP17A1 could contribute to the rapid diagnosis of 17-OHD in China. Genetic counseling based on the genetic diagnosis for at-risk relatives is advised.

[Detection of chromosome aneuploidies in spontaneous abortion villus samples by quantitative fluorescence PCR].

OBJECTIVE: To assess the value of quantitative fluorescence polymerase chain reaction (QF-PCR) for the detection of chromosomal aneuploidies in chorionic villus samples from early abortion. METHODS: One hundred seventy seven specimens were collected. Genomic DNA was extracted, and aneuploidies of 8 chromosomes (13, 15, 16, 18, 21, 22, X and Y) were detected by QF-PCR analysis. RESULTS: The QF-PCR was successful in 176 (99.4%) of the cases. All detection was completed in 48 hours. Sixty three(35.8%) cases have shown abnormal signals, which included 3 cases of trisomy 13, 3 cases of trisomy 15, 14 cases of trisomy 16, 2 cases of trisomy 18, 7 cases of trisomy 22, 3 cases of trisomy 21, 13 cases of 45,X, 1 case of 47,XXX, 2 cases of 47,XXY, 2 cases of haploidy, 11 cases of triploidy, 1 case of trisomy 16 and trisomy 22, 1 case of trisomy 21 and trisomy 22. Trisomy 16 was the most common chromosome aneuploidy (22.22%), which was followed by 45,X (20.63%), triploidy (17.46%) and trisomy 22 (11.11%). CONCLUSION: QF-PCR is a quick and easy method for detecting chromosomal aneuploidies in chorionic villi tissue. The results can provide important information for genetic counseling for spontaneous abortions.

Enhanced circulation half-life of site-specific PEGylated rhG-CSF: optimization of PEG molecular weight.

Recombinant human granulocyte colony stimulating factor (rhG-CSF) and its PEGylated product "mono-PEG20-GCSF" have already been widely used for treatment of all kinds of neutropenia. However, the high required dosage of mono-PEG20-GCSF made it relatively expensive in clinical use. We postulated that an N-terminal site-specific PEGylated rhG-CSF with higher PEG Mw (PEG30 kDa) might be able to achieve longer circulation half-life while retaining its bioactivity, allowing the reduction of dosage for clinical use. rhG-CSF was PEGylated at the N-terminus by 5 kDa, 10 kDa, 20 kDa and 30 kDa methoxy-poly(ethylene glycol)-propionaldehyde (mPEG-ALD), and the four PEGylates were compared with respect to reaction, separation, characterization and also in vivo/in vitro activity, results showed that the mPEG-ALD of higher Mw demonstrated better N-terminal site-specific selectivity, separation purity and yield. The production cost and in vitro activity of mono-PEG30-GCSF and mono-PEG20-GCSF were almost the same, while mono-PEG30-GCSF showed longer in vivo circulation half-life and 60% higher drug bioavailability than mono-PEG20-GCSF. Consequently, mono-PEG30-GCSF shall be administered at a lower dosage than mono-PEG20-GCSF while retaining the same therapeutic efficacy.

Poly (ethylene glycol) prodrug for anthracyclines via N-Mannich base linker: design, synthesis and biological evaluation.

Poly (ethylene glycol)s (PEGs) are potential drug carriers for improving the therapeutic index of anticancer agents. In this work, a novel methodology for constructing PEG prodrug of anthracycline anticancer drugs was developed based on N-Mannich base of salicylamide and its 2-acyloxymethylated derivative. The resultant conjugates first subjected to in vitro hydrolysis testing, which revealed the release behavior of newly synthesized PEG prodrugs could be adjusted by the status of 2-hydroxy group of salicylamide. These PEG prodrugs also demonstrated superior cytotoxicity in antiproliferative assay. O-blocked doxorubicin prodrug with PEG20k as carrier was selected for further in vivo assessments and presented longer circulating life in pharmacokinetic experiment. This high molecular prodrug was also found to be more efficacious against S-180 xenografted tumor than equivalent amount of doxorubicin.

Thermosensitive polymer-conjugated albumin nanospheres as thermal targeting anti-cancer drug carrier.

Thermosensitive Poly(N-isopropylacrylamide-co-acrylamide-co-allylamine) (PNIPAM-AAm-AA)-conjugated albumin nanospheres (PAN) was developed as a new thermal targeting anti-cancer drug carrier by conjugating PNIPAM-AAm-AA on the surface of albumin nanospheres (AN). AN with diameter below 200nm and narrow size distribution was successfully prepared in the first step with desolvation technique. PNIPAM-AAm-AA with different molecular weight (M(w)) was synthesized in the second step by radical polymerization and conjugated onto the surface of AN. Anti-cancer drug adriamycin (ADR) was then entrapped into the AN and PAN during the particle preparation. Compared with AN, the release rate of ADR from PAN in trypsin solution was slower, and decreased with increasing the conjugation amounts (hairy density) or M(w) of PNIPAM-AAm-AA (hairy length). Moreover, the release of ADR from PAN above the cloud-point temperature (T(cp)) of PNIPAM-AAm-AA became faster due to shrinkage of hairy thermosensitive polymer. To testify the thermal targetability in vivo, PAN was incubated with HepG2 cells. As expected, PAN can target cancer cells above the T(cp) of PNIPAM-AAm-AA, whereas it cannot below the T(cp). These results might reflect that PAN may selectively accumulate onto solid tumors that are maintained above physiological temperature due to local hyperthermia.