Volume 105 · Issue 06 · 2025

Fetal nucleated red blood cells (fNRBCs) in the peripheral blood of pregnant women which contain the complete genetic information of the fetus, emerge as the ideal cells for fetal genetic diseases screening and prediction. These cells have been proven useful for fetal sex identification and the detection of chromosomal aneuploidy. Given the strong heterogeneity and diverse mutation sites and types of monogenic diseases, several bottleneck issues, such as rare cell enrichment and sorting, cell origin identification, sequencing of minute amounts of DNA, and bioinformatics analysis, merits further investigation prior to utilizing fNRBCs for testing. Addressing these challenges can contribute to obtaining highly purified fetal cells, high-coverage, high-fidelity, and unbiased whole-genome amplification products, as well as accurate genetic mutation detection and analysis results. In light of the aforementioned issues, this article primarily focuses on the advancements and challenges in the non-invasive prenatal testing of monogenic diseases using fNRBCs.

High-throughput sequencing technology is playing an increasingly important role in the prevention and control of hereditary birth defects. Among these technologies based on next-generation sequencing (NGS), non-invasive prenatal testing (NIPT), low-coverage whole genome sequencing (copy number variation sequencing, CNV-seq), and exome sequencing (ES) have been widely applied in clinical practice. In recent years, the research and clinical application of third generation sequencing (TGS) have yielded numerous results. With its long-read sequencing advantages, TGS has solved many challenges in the three-level prevention of complex and difficult genetic diseases, further improving the technical system for the prevention and control of hereditary birth defects. This article introduces the application of high-throughput sequencing technology in various stages of hereditary birth defect prevention and explores the key technologies and application trends in each field.

Chemotherapy-induced anemia (CIA) is one of the most common complications in cancer patients. If CIA treatment is not effective, it will affect the normal implementation of chemotherapy regimens, and reduce the quality of life of patients and shorten clinical survival. At present, CIA has serious shortcomings in treatment rate and unsatisfactory therapeutic effect. It is often forced to reduce the dose of chemotherapy or delay chemotherapy due to anemia that cannot be rapidly and effectively treated in clinical practice. These problems are mainly related to factors such as complex etiology of the disease, late timing of clinical intervention, and certain limitations in existing treatment options. Therefore, disease management needs to further strengthen the concept of early diagnosis and early treatment to avoid the increased clinical burden due to the severity of anemia. Meanwhile, with the in-depth understanding of the mechanism of anemia, new drug research and development based on the comprehensive regulatory mechanism of hypoxia-inducible factor (HIF) has begun in clinical practice. A phase Ⅲ clinical study of Roxostat in the treatment of chemotherapy-induced anemia in patients with non-medullary malignancies was presented at the European Society of Medical Oncology (ESMO) conference in 2023. This hypoxia-inducible factor prolyl hydroxylase inhibitor (HIF-PHI) drug is an oral preparation, which has shown good efficacy and safety in the study, and has superior compliance compared with recombinant human erythropoietin-α (rHuEPO-α). Starting from the re-understanding of the mechanism of CIA, this review analyzes the current diagnosis and treatment dilemmas of CIA, puts forward suggestions in combination with the "Guildlines of Chinese Society of Clinical Oncology (CSCO) clinical practice in tumor-related anemia (2024)", and introduces and looks forward to HIF-PHI, a new therapeutic drug in the future.

Genetic factors are the primary causes of congenital hearing loss, and prevention of hereditary hearing loss is the main breakthrough point in the control and prevention of deafness . The progress in molecular diagnostic technology has effectively improved the diagnostic rate of hereditary hearing loss. Moreover, the findings of population-based molecular epidemiological studies on hearing loss provide theoretical support for the implementation of carrier screening, which aims to identify high-risk families that may give birth to deaf children, and thus lay the foundation for the large-scale prevention of hereditary hearing loss. In view of the lack of unified normative documents on the prevention of hereditary hearing loss in clinical application, a multidisciplinary team of experts consulted the latest evidence-based medicine at home and abroad, and has reached a consensus on the applicable population, strategy, technology, prevention period selection, and clinical process for the pre-pregnancy/prenatal prevention of hereditary hearing loss. The goal is to provide a reference for the clinical standardization implementation of hereditary hearing loss prevention and insight for the research and development of prevention technologies.

To explore the clinical value of expanded carrier screening (ECS) for monogenic diseases in Chinese pregnant individuals.

The clinical data of pregnant individuals with ECS for 334 single-gene recessive (AR or X-linked) diseases (434 pregnant woman and 315 spouses) at West China Second University Hospital of Sichuan University from May 2021 to February 2024 were retrospectively collected, including the general information, gestational age, carrier rate, detection of pathogenic/likely pathogenic (P/LP) variation with Human Gene Mutation Database (HGMD), and detection rate of at-risk couple (ARC). The diagnostic gestational weeks for ARC with concurrent screening and sequential screening mode were compared.

This study included 749 individuals (434 females and 315 males) with a median (Q₁, Q₃) age of 31 (28, 34) years. The carrier rate for carrying at least one P/LP variant was 65.6% (491/749). When only the P/LP variation included in HGMD was reported, detection rate of ARC was 8.3% (26/315). The actual detection rate of ARC (including P/LP variation in HGMD and not in HGMD) increased to 9.5 % (30/315). A total of 86 genes overlapped with the T3 screening genes recommended by American College of Medical Genetics and Genomics (ACMG-T3), and the detection rate of ARC was 7.0% (22/315). Compared with ACMG-T3, 334 gene screening packages covered an additional 248 genes, and the detection rate of ARC was 2.5% (8/315). The gestational age of ARC at prenatal diagnosis with concurrent screening was significantly earlier than that of sequential screening [(19.7±3.3) weeks vs (22.2±2.2) weeks, P=0.049].

Concurrent screening for monogenic disease carriers of multiple diseases during pregnancy will evidently advance the timing of diagnosis.

To explore the genesis of the mitochondrial A1555G mutation and its correlation with the severity of hearing impairment.

A cross-sectional study was conducted. Individuals with the mitochondrial A1555G mutation who were screened for neonatal deafness genes at the Molecular Diagnostic Center for Deafness, PLA General Hospital, between January 2020 and December 2023 were selected to analyze the proportions of heterogeneous and homogeneous variants. Meanwhile, individuals with the mitochondrial A1555G heterogeneous mutation identified through next-generation sequencing in the outpatient database of the Department of Otolaryngology, Head and Neck Surgery, PLA General Hospital from January 2016 to December 2023 were selected for analysis of their genetic origins and phenotypic characteristics. The correlation between the severity of hearing loss and the A1555G mutation load was further explored by examining individuals with known mutation load and their corresponding hearing phenotypes.

Among the 65 942 newborns, 157 individuals (63 males and 94 females) were found to carry the A1555G mutation (2.4 per thousand individuals) (157/65 942). There were 48 individuals carrying heterogeneous mutation (7 per ten thousand individuals) (48/65 942), and 30.6% (48/157) of the individuals carrying A1555G mutation had heterogeneous mutation. Among the A1555G heteroplasmy individuals, there were five cases of neonatal mutation, 14 cases of clear mutation load, and the mutation load ranged from 1.0% to 99.4%. Two cases showed bilateral extremely severe hearing loss, and one case exhibited unilateral extremely severe hearing loss. Analysis of 118 cases involving the A1555G mutation load and hearing phenotype, including heteroplasmy individuals, revealed a positive correlation between the degree of hearing loss and the mutation load (r_s=0.489, 95%CI: 0.350-0.606, P<0.001). Additionally, the mutation load in individuals with moderate to severe hearing loss was greater than 50% in all cases (52/52).

The current study indicates that some of the genetic sources of A1555G heterogenous mutation individuals are de novo mutations. There is a wide variation range in the mutation load among individuals carrying the A1555G variant, and a positive correlation exists between the mutation load and the severity of hearing loss, particularly in cases of moderate to severe hearing impairment.

To analyze the prenatal diagnostic results in families with de novo monogenic diseases and the mutation origins in affected children from families with reproductive history of children with recurrent de novo mutations (DNMs).

This study was a cross-sectional study. A total of 41 cases with adverse pregnancy history of de novo monogenic diseases who underwent genetic counseling and prenatal diagnosis from January 2021 to December 2023 at the Obstetrics and Gynecology Hospital of Fudan University were included. Prenatal diagnosis and other clinical data were reviewed, and peripheral blood of the parents, peripheral blood or tissue of the probands, and amniotic fluid or chorionic villus were collected. For families with reproductive history of children with recurrent DNMs, additional saliva and semen were collected from all the parents. Targeted high-throughput sequencing was performed to assess parental somatic mosaicism and male germline mosaicism. As for the cases in which the mutation was undetected in the semen, Sanger sequencing was utilized to search for single nucleotide polymorphism (SNP) sites upstream and downstream of the mutation site and clarify the mutation origins in combination with TA cloning.

A total of 41 families were included, with male age of (34.1±3.9) years (41 cases) and female age of (33.0±3.9) years (41 cases). Moreover, 32 causative genes were involved, with neurodevelopmental disorders, hereditary myopathies, hereditary bone diseases, hereditary ophthalmopathies, hereditary cardiovascular diseases and other multisystem diseases accounting for 53.7% (22/41), 12.2% (5/41), 7.3% (3/41), 4.9% (2/41), 2.4% (1/41), and 19.5% (8/41), respectively. One DNMs was detected in 37 families who underwent prenatal diagnosis during the second trimester. Four families with reproductive history of children with recurrent DNMs were analyzed for the mutation origins, of which two families had a low proportion of mosaicism detected in paternal semen, with variant allele fraction (VAF) of 3.7% and 12.8%, respectively, and the origins were from the parents detected by Sanger sequencing in combination with TA cloning in another two families.

DNMs are at risk of recurrence. The"targeted high-throughput sequencing+Sanger sequencing+TA cloning"process is conducive to identifying the parental origin of the mutation.

To explore the value of noninvasive prenatal diagnosis for monogenic disorders (NIPD-M) based on relative haplotype dosage (RHDO) and Bayes factor (BF) for pregnant women with high-risk recessive genetic disease in the first trimester.

A total of 206 pregnant women with high-risk recessive genetic disease and pedigree samples at the First Affiliated Hospital of Zhengzhou University between September 2022 and November 2023 were collected. The cell-free DNA (cfDNA) was extracted from the pregnant woman′s plasma and the genomic DNA (gDNA) was extracted from the pedigree blood samples. The designed capture panel covered 10 genes (DMD, SMN1, PAH, MMACHC, MMUT, F8, F9, SLC26A4, GJB2, CYP21A2). Sequencing of target regions was performed through high-throughput sequencing platforms, informative single nucleotide polymorphism (SNP) was screened, and pathogenic haplotypes were constructed. The fetal genotype was determined based on the dose change of the informative SNPs in cfDNA combined with the BF algorithm. The circular binary segmentation (CBS) algorithm was used to exclude the interference of recombination events. All NIPD-M results were validated by invasive prenatal diagnosis or newborn genetic testing.

Among the recruited families, the median (Q₁, Q₃) age of the 206 pregnant women was 32 (27, 38) years, and the earliest blood collection was at 7 weeks of pregnancy, with the median (Q₁, Q₃) blood collection time of 8⁺² (8, 9⁺²) weeks and fetal fraction (FF) of 5.21% (3.56%, 7.64%). A total of 190 cases were successfully tested, while 16 failed the test, with a success rate of 92.2% (190/206). The NIPD-M results were consistent with the invasive prenatal diagnosis and newborn genetic testing, with the accuracy rate of 100% (190/190).

NIPD-M provides an earlier and safer means of prenatal diagnosis for high-risk pregnant women with recessive genetic diseases, with high accuracy and rapid response.

A total of 112 carriers (51 males and 61 females) with Robertsonian translocations who underwent 148 cycles of preimplantation genetic testing for structural rearrangements (PGT-SR) between February 2017 and December 2023 at the First Center of General Hospital of Chinese PLA were recruited. The heterologous type rob (13; 14) was the most common form of translocation, which accounted for 65.2% (73/112). The total euploidy rate of blastocysts was 42.1% (303/720). Male carriers had higher rate of euploid embryo than female carriers [50.0% (149/298) vs 36.5% (154/422), P<0.001]. Likewise, the clinical pregnancy rate in male carriers was higher than female carriers, but the difference was not statistically significant [62.9% (39/62) vs 49.4% (38/77), P=0.154]. There were 139 transferrable embryos, with a clinical pregnancy rate of 55.4% (77/139) and abortion rate of 14.3% (11/77). The current study indicates that male carriers with Robertsonian translocations had higher rate of euploid embryo than female carriers in PGT-SR cycles.

GM2 ganglioside storage disease is a rare genetic disease, and Sandhoff disease is a subtype of this disease. This paper reports a case of adult Sandhoff disease, and after literature review, it is concluded that the first symptom of this disease is mostly limb weakness, others include muscle atrophy, ataxia, spasticity, speech disorder, etc. Electromyography is mostly chronic neurogenic damage myoelectric changes, and the prognosis of patients depends on enzyme activity. At present, this disease can only be treated symptomatically, and the treatment of causes is also being explored, such as gene supplementation therapy, substrate reduction therapy, and neuroprotective therapy.

Preimplantation genetic testing (PGT) has been widely used in the field of assisted reproduction. On the basis of the continuous development of molecular genetics detection technology, PGT has more and more broad application prospect. Non-invasive non-invasive PGT technology can avoid embryo damage caused by biopsy. DNA methylation screening based on preimplantation blastocyst trophoblastic ectoderm cells and RNA-based PGT technology provide new methods and theoretical support for embryo developmental potential prediction and genetic screening. This review summarizes the application status and research progress of new PGT technology, and provides evidence for clinical application.

Immunosuppressive tumor microenvironment (TME) is a key factor that limits the efficacy of immunotherapy in solid tumors and can be used as a target for tumor therapy. In recent years, a number of studies have proposed a new concept of immunomodulatory vaccine. By selecting tumor microenvironment antigen (TMA) co-expressed on the surface of suppressive immune cells and stromal cells in tumor cells and TME as the target, immunogenic tumor vaccine can be prepared to induce strong endogenous antigen-specific T cell response. Immunomodulatory vaccines, which combine the dual effects of killing tumor cells and targeted clearance of inhibitory immune components, can effectively improve immunosuppressive TME and are a promising novel immunotherapy model. This paper reviews the classification of existing TMA and the immunomodulatory vaccines developed based on them, summarizes the latest progress of preclinical and clinical research related to immunomodulatory vaccines, and analyzes the rationality of their combination with other immunotherapy strategies, finally providing new ideas for lifting the immune restriction of TME in solid tumors and improving the immunotherapy effect.

The rapid development of immunotherapy has made a great innovation in tumor treatment, and it has been successfully applied in clinical tumor treatment. Three-dimensional tumor models can simulate the tumor microenvironment (TME) and microvascular system in vitro, and more realistically reflect the immunotherapy process. Three-dimensional tumor model provides a new way for the research of tumor immunotherapy, and has a broad application prospect in the fields of reconstructing tumor immune microenvironment in vitro, screening immune drugs, guiding clinical individualized medication, etc. This paper will briefly introduce the three-dimensional culture models and their application in immunotherapy.

The patient, a 58-year-old female, went to the Department of Ophthalmology of Beijing Tiantan Hospital in January 2023 due to "repeated friction and bleeding of both eyelid masses for more than 20 days". The patient was recently treated with camrelizumab (SHR-1210, Cam) for stage IV lung adenocarcinoma surgery. Four days after injection, two mulberry-shaped vascular lesions were found growing on the skin of the upper eyelid of the left eye and the lower eyelid of the right eye (shown by arrows in Figure 1, and the photo was published with the consent of the patient), and within 20 days, multiple dome-shaped bright red lesions of 0.2 to 6.0 mm appeared on the breast, neck, and shoulder and back. Due to the frequent friction and bleeding of eyelid lesions, the patient required surgery, and the preoperative examination such as improving coagulation function showed that the eyelid lesions were normal, and the eyelid lesions were removed in Beijing Tiantan Hospital. Tissue HE staining revealed lobulated proliferation of dermal capillaries and vascular endothelial cells (Figure 2, ×100). Immunohistochemical results showed positive staining for CD34 and erythroblast transformation-specific-1 related gene (ERG) in vascular endothelial cells. Therefore, CAM-induced reactive cutaneous capillary hyperplasia (RCCEP) was diagnosed. After 1 month of follow-up, the eyelid lesions did not recur, and RCCEP in other parts of the body gradually resolved after discontinuation.