ABSTRACT
Mixed phenotype acute leukaemia (MPAL) is a high-risk subtype of leukaemia with myeloid and lymphoid features, limited genetic characterization, and a lack of consensus regarding appropriate therapy. Here we show that the two principal subtypes of MPAL, T/myeloid (T/M) and B/myeloid (B/M), are genetically distinct. Rearrangement of ZNF384 is common in B/M MPAL, and biallelic WT1 alterations are common in T/M MPAL, which shares genomic features with early T-cell precursor acute lymphoblastic leukaemia. We show that the intratumoral immunophenotypic heterogeneity characteristic of MPAL is independent of somatic genetic variation, that founding lesions arise in primitive haematopoietic progenitors, and that individual phenotypic subpopulations can reconstitute the immunophenotypic diversity in vivo. These findings indicate that the cell of origin and founding lesions, rather than an accumulation of distinct genomic alterations, prime tumour cells for lineage promiscuity. Moreover, these findings position MPAL in the spectrum of immature leukaemias and provide a genetically informed framework for future clinical trials of potential treatments for MPAL.
Subject(s)
Leukemia, Biphenotypic, Acute/genetics , Leukemia, Biphenotypic, Acute/pathology , Cell Lineage/genetics , DNA Mutational Analysis , Female , Genetic Variation/genetics , Genome, Human/genetics , Genomics , Humans , Immunophenotyping , Leukemia, Biphenotypic, Acute/classification , Male , Models, Genetic , Mutation/genetics , Neoplastic Stem Cells/immunology , Neoplastic Stem Cells/metabolism , Neoplastic Stem Cells/pathology , Phenotype , Trans-Activators/geneticsABSTRACT
BACKGROUND: Exophiala dermatitidis is a dematiaceous fungus isolated from various environmental sources. Systemic E. dermatitidis infections can lead to fatal outcomes, and treatment has not yet been standardized. Although E. dermatitidis is also known to cause cutaneous infection, it has not been previously reported to appear as ecthyma gangrenosum (EG), an uncommon cutaneous lesion in neutropenic patients that is mainly caused by Pseudomonas aeruginosa. CASE PRESENTATION: A 2-month-old male infant with mixed-phenotype acute leukemia presented with prolonged fever unresponsive to antibacterial and antifungal agents during myelosuppression due to remission induction therapy. He also presented with skin lesions on the left wrist and left lower quadrant of the abdomen. The abdominal lesion gradually turned black and necrotic, which was consistent with the findings of the EG. E. dermatitidis was isolated from the blood, stool, wrist skin, and endotracheal aspirate. During hematopoietic recovery, consolidation in both lungs was evident. Multiagent antifungal treatment failed to eliminate E. dermatitidis from blood. In order to salvage the central venous catheter, ethanol lock therapy (ELT) was adopted, following which the blood culture became negative. The abdominal lesion that evolved as a necrotic mass connecting the small intestine and subcutaneous tissue adjacent to the skin was surgically resected. After these interventions, the general condition improved. CONCLUSION: Disseminated E. dermatitidis mycosis in the neutropenic infant was successfully managed with a multidisciplinary treatment consisting of multiagent antifungal treatment, ELT, and surgery.
Subject(s)
Ecthyma , Leukemia , Mycoses , Male , Humans , Antifungal Agents/therapeutic use , Mycoses/drug therapy , Ecthyma/drug therapy , Leukemia/drug therapy , Acute Disease , Anti-Bacterial Agents , Ethanol , PhenotypeABSTRACT
Mature teratomas are usually benign tumors that rarely undergo malignant transformation. We report an advanced neuroblastoma arising in a mature teratoma of the ovary. Whole-exome sequencing identified extensive copy-neutral loss of heterozygosity (LOH) in both neuroblastoma and teratoma elements, suggesting that the neuroblastoma evolved from the teratoma. In addition, several truncating germline heterozygous variants in tumor suppressor genes, including RBL2 and FBXW12, became homozygous as a result of LOH. Collectively, we speculate that extensive LOH in teratoma cells may force heterozygous germline variants to become homozygous, which, in turn, may contribute to the development of neuroblastoma with the acquisition of additional chromosomal changes.
Subject(s)
Germ-Line Mutation , Loss of Heterozygosity , Neoplasms, Multiple Primary/genetics , Neuroblastoma/genetics , Ovarian Neoplasms/genetics , Teratoma/genetics , Adolescent , F-Box Proteins/genetics , Female , Homozygote , Humans , Neoplasms, Multiple Primary/drug therapy , Neoplasms, Multiple Primary/pathology , Neuroblastoma/drug therapy , Neuroblastoma/pathology , Ovarian Neoplasms/drug therapy , Ovarian Neoplasms/pathology , Retinoblastoma-Like Protein p130/genetics , Teratoma/drug therapy , Teratoma/pathology , Exome SequencingABSTRACT
Hospital-acquired Clostridium difficile infection (CDI) may cause life-threatening colitis for children with cancer, making identification of risk factors important. We described characteristics of pediatric cancer patients with primary and recurring CDI, and evaluated potential risk factors. Among 189 cancer patients, 51 cases (27%) of CDI and 94 matched controls of cancer patients without CDI were analyzed. Multivariable logistic regression was used to evaluate the association between CDI and several potential risk factors. Median age of CDI cases was lower (3.3 y; 0.60 to 16.2) than controls (7.7 y; 0.4 to 20.5). Median duration of neutropenia before CDI was longer for CDI cases (10.0 d; 0.0 to 30.0) compared with duration calculated from reference date in controls (6.0 d; 0.0 to 29.0). Multivariable analysis showed that older age was associated with reduced risk (≥7 vs. 0 to 3 y, odds ratio=0.11; 95% confidence interval, 0.02-0.54), and prolonged neutropenia was associated with increased risk (odds ratio=1.11; 95% confidence interval, 1.01-1.22). CDI recurred in 26% of cases. Younger age and prolonged neutropenia were risk factors for CDI in children with cancer. Increasing awareness to these risk factors will help to identify opportunities for CDI prevention in cancer patients.
Subject(s)
Clostridium Infections/etiology , Neoplasms/complications , Adolescent , Age Factors , Case-Control Studies , Child , Child, Preschool , Clostridioides difficile , Clostridium Infections/virology , Humans , Iatrogenic Disease , Infant , Neutropenia/complications , Risk Factors , Young AdultSubject(s)
Precursor Cell Lymphoblastic Leukemia-Lymphoma , Acute Disease , Antineoplastic Combined Chemotherapy Protocols , Humans , Injections, Spinal , Methotrexate/therapeutic use , Precursor Cell Lymphoblastic Leukemia-Lymphoma/complications , Precursor Cell Lymphoblastic Leukemia-Lymphoma/drug therapyABSTRACT
Infantile fibrosarcoma is a non-rhabdomyosarcoma soft-tissue sarcoma that occurs in infancy and which has a relatively good prognosis. A vincristine and dactinomycin (VA) regimen has been shown to be effective, although the duration of chemotherapy has not been well defined. We describe the case of a 4-month-old boy with a mass at the left dorsum of the foot who was diagnosed with infantile fibrosarcoma after resection of the tumor, the margin of which was macroscopically positive. VA treatment was carried out with careful monitoring of response and adverse effects. Pancytopenia was seen during the second cycle, and therapy was reduced thereafter. The treatment was continued for 38 weeks (12 cycles). There was no functional impairment, and no evidence of recurrence at 18 months after therapy.
Subject(s)
Antineoplastic Agents/therapeutic use , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Dactinomycin/therapeutic use , Fibrosarcoma/drug therapy , Foot/surgery , Soft Tissue Neoplasms/drug therapy , Vincristine/therapeutic use , Chemotherapy, Adjuvant , Fibrosarcoma/surgery , Humans , Infant , Male , Soft Tissue Neoplasms/surgeryABSTRACT
We describe the case of a 13-year-old girl with multifocal disseminated Ewing sarcoma family of tumor (ESFT) who received a 5/8 human leukocyte antigen-matched haploidentical hematopoietic cell transplantation to generate a graft-versus-tumor effect. The patient had grade 2 acute graft-versus-host disease (GVHD) of the skin and chronic GVHD nausea and abdominal pain that required prednisolone for 17 months, but has been free from ESFT for 3 years 10 months after therapy. The present case suggests a beneficial effect of haploidentical hematopoietic cell transplantation in disseminated ESFT.
Subject(s)
Bone Neoplasms/therapy , Hematopoietic Stem Cell Transplantation/methods , Pelvic Bones , Sarcoma, Ewing/therapy , Adolescent , Female , Graft vs Host Disease , HumansABSTRACT
Ewing sarcoma/primitive neuroectodermal tumor (ES/PNET) of the kidney is extremely rare, and is usually diagnosed after nephrectomy without neoadjuvant chemotherapy. Although ifosfamide and etoposide improve survival to a great extent in ES/PNET, the use of nephrotoxic agent, particularly ifosfamide, is a concern after nephrectomy. We describe the case of a 14-year-old female patient with abdominal mass who was diagnosed with ES/PNET of the right kidney after nephrectomy. Adjuvant chemotherapy including ifosfamide and etoposide were given. The estimated glomerular filtration rate decreased to 75% after the end of therapy. There was no evidence of recurrence 70 months after initial diagnosis.
Subject(s)
Ifosfamide/therapeutic use , Kidney Neoplasms/drug therapy , Sarcoma, Ewing/drug therapy , Adolescent , Antineoplastic Agents, Alkylating/therapeutic use , Female , Humans , Kidney Neoplasms/diagnosis , Magnetic Resonance Imaging , Neuroectodermal Tumors, Primitive , Sarcoma, Ewing/diagnosis , Tomography, X-Ray ComputedABSTRACT
Thiopurine S-methyltransferase (TPMT) and nucleoside diphosphate-linked moiety X-type motif 15 (NUDT15) variants are considered to be genes responsible for severe myelotoxicity induced by 6-mercaptopurine (6MP). We report a 4-year-old girl with acute lymphoblastic leukemia, who developed the complication of severe 6MP-induced myelotoxicity due to homozygous NUDT15 variant alleles. In early consolidation therapy containing 6MP, her course was complicated by severe neutropenia (Grade 4) and chemotherapy had to be discontinued for 33 days. The 6MP dose was subsequently adjusted based on the white blood cell count. The ratios of the prescribed 6MP dose over the protocol dose in early consolidation, central nervous system (CNS) prophylaxis, re-induction, interim maintenance and maintenance therapy were 63%, 27%, 4%, 26% and 7%, respectively. Suspension of therapy was required during early consolidation, CNS prophylaxis and interim maintenance therapy. We investigated candidate genes for 6MP-associated myelotoxicity and found homozygous NUDT15 variant alleles and a heterozygous inosine triphosphate pyrophosphatase (ITPA) variant allele. In patients with homozygous NUDT15 variants, drastic reduction (less than 10%) of the 6MP dose from the protocol dose might be required not only during maintenance therapy, but also during other treatment courses containing 6MP. Screening of candidate genes at diagnosis is recommended in order to avoid serious adverse events.
Subject(s)
Mercaptopurine/adverse effects , Mutation, Missense , Neutropenia/chemically induced , Precursor Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Pyrophosphatases/genetics , Child, Preschool , Female , Homozygote , Humans , Precursor Cell Lymphoblastic Leukemia-Lymphoma/geneticsABSTRACT
Formalin-fixed and paraffin-embedded (FFPE) sections mounted on microscope slides are one of the largest available resources for retrospective research on various diseases, but quantitative phosphoproteome analysis of FFPE sections has never been achieved because of the extreme difficulty of procuring sufficient phosphopeptides from the limited amounts of proteins on the slides. Here, we present the first protocol for quantitative phosphoproteome analysis of FFPE sections by utilizing phase-transfer surfactant-aided extraction/tryptic digestion of FFPE proteins followed by high-recovery phosphopeptide enrichment via lactic acid-modified titania chromatography. We established that FFPE sections retain a similar phosphoproteome to fresh tissue specimens during storage for at least 9 months, confirming the utility of our method for evaluating phosphorylation profiles in various diseases. We also verified that chemical labeling based on reductive dimethylation of amino groups was feasible for quantitative phosphoproteome analysis of FFPE samples on slides. Furthermore, we improved the LC-MS sensitivity by miniaturizing nanoLC columns to 25 µm inner diameter. With this system, we could identify 1090 phosphopeptides from a single FFPE section obtained from a microscope slide, containing 25.2 ± 5.4 µg of proteins. This protocol should be useful for large-scale phosphoproteome analysis of archival FFPE slides, especially scarce samples from patients with rare diseases.
Subject(s)
Formaldehyde/chemistry , Paraffin Embedding , Phosphoproteins/metabolism , Proteome , Tissue Fixation , Animals , Chromatography, Liquid , Mice , Mice, Inbred C57BL , Tandem Mass SpectrometryABSTRACT
The bone marrow microenvironment (BME) drives drug resistance in acute lymphoblastic leukemia (ALL) through leukemic cell interactions with bone marrow (BM) niches, but the underlying mechanisms remain unclear. Here, we show that the interaction between ALL and mesenchymal stem cells (MSCs) through integrin ß1 induces an epithelial-mesenchymal transition (EMT)-like program in MSC-adherent ALL cells, resulting in drug resistance and enhanced survival. Moreover, single-cell RNA sequencing analysis of ALL-MSC co-culture identifies a hybrid cluster of MSC-adherent ALL cells expressing both B-ALL and MSC signature genes, orchestrated by a WNT/ß-catenin-mediated EMT-like program. Blockade of interaction between ß-catenin and CREB binding protein impairs the survival and drug resistance of MSC-adherent ALL cells in vitro and results in a reduction in leukemic burden in vivo. Targeting of this WNT/ß-catenin-mediated EMT-like program is a potential therapeutic approach to overcome cell extrinsically acquired drug resistance in ALL.
Subject(s)
Epithelial-Mesenchymal Transition , Precursor Cell Lymphoblastic Leukemia-Lymphoma , Humans , beta Catenin , Precursor Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Coculture Techniques , Drug Resistance , Cell Proliferation , Tumor MicroenvironmentABSTRACT
During postnatal life, the bone marrow (BM) supports both self-renewal and differentiation of hematopoietic stem cells (HSCs) in specialized microenvironments termed stem cell niches. Cell-cell and cell-extracellular matrix interactions between HSCs and their niches are critical for the maintenance of HSC properties. Here, we analyzed the function of N-cadherin in the regulation of the proliferation and long-term repopulation activity of hematopoietic stem/progenitor cells (HSPCs) by the transduction of N-cadherin shRNA. Inhibition of N-cadherin expression accelerated cell division in vitro and reduced the lodgment of donor HSPCs to the endosteal surface, resulting in a significant reduction in long-term engraftment. Cotransduction of N-cadherin shRNA and a mutant N-cadherin that introduced the silent mutations to shRNA target sequences rescued the accelerated cell division and reconstitution phenotypes. In addition, the requirement of N-cadherin for HSPC engraftment appears to be niche specific, as shN-cad-transduced lineage(-)Sca-1(+)c-Kit(+) cells successfully engrafted in spleen, which lacks an osteoblastic niche. These findings suggest that N-cad-mediated cell adhesion is functionally required for the establishment of hematopoiesis in the BM niche after BM transplantation.
Subject(s)
Cadherins/genetics , Graft Survival/genetics , Hematopoietic Stem Cell Transplantation , Animals , Cadherins/physiology , Cell Adhesion/genetics , Cell Adhesion/physiology , Cell Movement/genetics , Cell Movement/physiology , Cell Survival/genetics , Cells, Cultured , Gene Knockdown Techniques , Hematopoietic Stem Cells/physiology , Mice , Mice, Congenic , Mice, Inbred C57BL , NIH 3T3 Cells , Stem Cell Niche/physiology , Time FactorsABSTRACT
The endosteal niche is critical for the maintenance of hematopoietic stem cells (HSCs). However, it consists of a heterogeneous population in terms of differentiation stage and function. In this study, we characterized endosteal cell populations and examined their ability to maintain HSCs. Bone marrow endosteal cells were subdivided into immature mesenchymal cell-enriched ALCAM(-)Sca-1(+) cells, osteoblast-enriched ALCAM(+)Sca-1(-), and ALCAM(-)Sca-1(-) cells. We found that all 3 fractions maintained long-term reconstitution (LTR) activity of HSCs in an in vitro culture. In particular, ALCAM(+)Sca-1(-) cells significantly enhanced the LTR activity of HSCs by the up-regulation of homing- and cell adhesion-related genes in HSCs. Microarray analysis showed that ALCAM(-)Sca-1(+) fraction highly expressed cytokine-related genes, whereas the ALCAM(+)Sca-1(-) fraction expressed multiple cell adhesion molecules, such as cadherins, at a greater level than the other fractions, indicating that the interaction between HSCs and osteoblasts via cell adhesion molecules enhanced the LTR activity of HSCs. Furthermore, we found an osteoblastic marker(low/-) subpopulation in ALCAM(+)Sca-1(-) fraction that expressed cytokines, such as Angpt1 and Thpo, and stem cell marker genes. Altogether, these data suggest that multiple subsets of osteoblasts and mesenchymal progenitor cells constitute the endosteal niche and regulate HSCs in adult bone marrow.
Subject(s)
Bone Marrow Cells/metabolism , Hematopoietic Stem Cells/cytology , Mesenchymal Stem Cells/metabolism , Osteoblasts/cytology , Osteoblasts/metabolism , Activated-Leukocyte Cell Adhesion Molecule/metabolism , Animals , Antigens, Differentiation/metabolism , Biomarkers/metabolism , Blotting, Western , Bone and Bones/cytology , Bone and Bones/metabolism , Cell Differentiation , Colony-Forming Units Assay , Gene Expression Profiling , Immunoenzyme Techniques , Mice , Mice, Inbred C57BL , Oligonucleotide Array Sequence Analysis , RNA, Messenger/genetics , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
ZNF384-rearranged fusion oncoproteins (FO) define a subset of lineage ambiguous leukemias, but their mechanistic role in leukemogenesis and lineage ambiguity is poorly understood. Using viral expression in mouse and human hematopoietic stem and progenitor cells (HSPC) and a Ep300::Znf384 knockin mouse model, we show that ZNF384 FO promote hematopoietic expansion, myeloid lineage skewing, and self-renewal. In mouse HSPCs, concomitant lesions, such as NRASG12D, were required for fully penetrant leukemia, whereas in human HSPCs, expression of ZNF384 FO drove B/myeloid leukemia, with sensitivity of a ZNF384-rearranged xenograft to FLT3 inhibition in vivo. Mechanistically, ZNF384 FO occupy a subset of predominantly intragenic/enhancer regions with increased histone 3 lysine acetylation and deregulate expression of hematopoietic stem cell transcription factors. These data define a paradigm for FO-driven lineage ambiguous leukemia, in which expression in HSPCs results in deregulation of lineage-specific genes and hematopoietic skewing, progressing to full leukemia in the context of proliferative stress. SIGNIFICANCE: Expression of ZNF384 FO early in hematopoiesis results in binding and deregulation of key hematopoietic regulators, skewing of hematopoiesis, and priming for leukemic transformation. These results reveal the interplay between cell of origin and expression of ZNF384 FO to mediate lineage ambiguity and leukemia development. This article is highlighted in the In This Issue feature, p. 171.
Subject(s)
Leukemia, Myeloid, Acute , Oncogene Proteins, Fusion , Animals , Hematopoiesis/genetics , Hematopoietic Stem Cells/metabolism , Humans , Leukemia, Myeloid, Acute/genetics , Mice , Oncogene Proteins, Fusion/genetics , Trans-Activators/genetics , Transcription Factors/geneticsABSTRACT
Hemophagocytic lymphohistiocytosis (HLH) is a severe complication after allogeneic hematopoietic cell transplantation (HCT) and can cause graft failure or multi-organ failure. Here, we report two children with refractory HCT-associated HLH treated with ruxolitinib. In the first patient, ruxolitinib resolved fever, cytopenia and hyperferritinemia. In another patient, although severe hepatic failure, which developed and worsened before the administration of ruxolitinib, was irreversible, rapid improvement in fever, leukopenia and hyperferritinemia was observed. Of note, multiplex cytokine profiling showed amelioration of cytokine storm in both patients. Ruxolitinib may be an encouraging option for HCT-associated HLH.
Subject(s)
Hematopoietic Stem Cell Transplantation/adverse effects , Lymphohistiocytosis, Hemophagocytic/drug therapy , Lymphohistiocytosis, Hemophagocytic/etiology , Pyrazoles/therapeutic use , Antineoplastic Combined Chemotherapy Protocols/adverse effects , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Child, Preschool , Disease Management , Disease Susceptibility , Female , Hematopoietic Stem Cell Transplantation/methods , Humans , Infant , Lymphohistiocytosis, Hemophagocytic/diagnosis , Male , Nitriles , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/complications , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/diagnosis , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Pyrazoles/administration & dosage , Pyrazoles/adverse effects , Pyrimidines , Transplantation, Homologous , Treatment OutcomeABSTRACT
Hematopoietic stem cells (HSCs) reside in hypoxic areas of the bone marrow. However, the role of hypoxia in the maintenance of HSCs has not been fully characterized. We performed xenotransplantation of human cord blood cells cultured in hypoxic or normoxic conditions into adult NOD/SCID/IL-2Rgamma(null) (NOG) mice. Hypoxic culture (1% O(2)) for 6 days efficiently supported the maintenance of HSCs, although cell proliferation was suppressed compared to the normoxic culture. In contrast, hypoxia did not affect in vitro colony-forming ability. Upregulation of a cell cycle inhibitor, p21, was observed in hypoxic culture. Immunohistochemical analysis of recipient bone marrow revealed that engrafted CD34(+)CD38(-) cord blood HSCs were hypoxic. Taken together, these results demonstrate the significance of hypoxia in the maintenance of quiescent human cord blood HSCs.
Subject(s)
Cell Differentiation , Fetal Blood/cytology , Hematopoietic Stem Cells/physiology , Anaerobiosis , Animals , Antigens, CD34/metabolism , Cell Culture Techniques , Cells, Cultured , Cyclin-Dependent Kinase Inhibitor p21/metabolism , Fetal Blood/physiology , Hematopoietic Stem Cells/cytology , Humans , Mice , Mice, Mutant Strains , Transplantation, Heterologous , Up-RegulationABSTRACT
Somatic genetic alterations of IKZF1, which encodes the lymphoid transcription factor IKAROS, are common in high-risk B-progenitor acute lymphoblastic leukemia (ALL) and are associated with poor prognosis. Such alterations result in the acquisition of stem cell-like features, overexpression of adhesion molecules causing aberrant cell-cell and cell-stroma interaction, and decreased sensitivity to tyrosine kinase inhibitors. Here we report coding germline IKZF1 variation in familial childhood ALL and 0.9% of presumed sporadic B-ALL, identifying 28 unique variants in 45 children. The majority of variants adversely affected IKZF1 function and drug responsiveness of leukemic cells. These results identify IKZF1 as a leukemia predisposition gene, and emphasize the importance of germline genetic variation in the development of both familial and sporadic ALL.
Subject(s)
Germ-Line Mutation , Ikaros Transcription Factor/genetics , Ikaros Transcription Factor/metabolism , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/genetics , Animals , Child , Female , Frameshift Mutation , Genetic Predisposition to Disease , Humans , Male , Mice , Neoplasm Transplantation , Pedigree , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/metabolism , Sequence Analysis, DNAABSTRACT
The carcinoembryonic antigen glypican-3 (GPC3) is a good target of anticancer immunotherapy against pediatric solid tumors expressing GPC3. In this non-randomized, open-label, phase I clinical trial, we analyzed the safety and efficacy of GPC3-peptide vaccination in patients with pediatric solid tumors. Eighteen patients with pediatric solid tumors expressing GPC3 underwent GPC3-peptide vaccination (intradermal injections every 2 weeks), with the primary endpoint being the safety of GPC3-peptide vaccination and the secondary endpoints being immune response, as measured by interferon (IFN)-γ enzyme-linked immunospot assay and Dextramer staining, and the clinical outcomes of tumor response, progression free survival (PFS), and overall survival (OS). Our findings indicated that GPC3 vaccination was well tolerated. We observed disease-control rates [complete response (CR)+partial response+stable disease] of 66.7% after 2 months, and although patients in the progression group unable to induce GPC3-peptide-specific cytotoxic T lymphocytes (CTLs) received poor prognoses, patients in the partial-remission and remission groups or those with hepatoblastoma received good prognoses. The GPC3-peptide vaccine induced a GPC3-specific CTL response in seven patients, with PFS and OS significantly longer in patients with high GPC3-specific CTL frequencies than in those with low frequencies. Furthermore, we established GPC3-peptide-specific CTL clones from a resected-recurrent tumor from one patient, with these cells exhibiting GPC3-peptide-specific cytokine secretion. The results of this trial demonstrated that the GPC3-peptide-specific CTLs induced by the GPC3-peptide vaccine infiltrated tumor tissue, and use of the GPC3-peptide vaccine might prevent the recurrence of pediatric solid tumors, especially hepatoblastomas, after a second CR.