Dynamic regulation and requirement for ribosomal RNA transcription during mammalian development.

Ribosomal RNA (rRNA) transcription by RNA polymerase I (Pol I) is a critical rate-limiting step in ribosome biogenesis, which is essential for cell survival. Despite its global function, disruptions in ribosome biogenesis cause tissue-specific birth defects called ribosomopathies, which frequently affect craniofacial development. Here, we describe a cellular and molecular mechanism underlying the susceptibility of craniofacial development to disruptions in Pol I transcription. We show that Pol I subunits are highly expressed in the neuroepithelium and neural crest cells (NCCs), which generate most of the craniofacial skeleton. High expression of Pol I subunits sustains elevated rRNA transcription in NCC progenitors, which supports their high tissue-specific levels of protein translation, but also makes NCCs particularly sensitive to rRNA synthesis defects. Consistent with this model, NCC-specific deletion of Pol I subunits Polr1a, Polr1c, and associated factor Tcof1 in mice cell-autonomously diminishes rRNA synthesis, which leads to p53 protein accumulation, resulting in NCC apoptosis and craniofacial anomalies. Furthermore, compound mutations in Pol I subunits and associated factors specifically exacerbate the craniofacial anomalies characteristic of the ribosomopathies Treacher Collins syndrome and Acrofacial Dysostosis-Cincinnati type. Mechanistically, we demonstrate that diminished rRNA synthesis causes an imbalance between rRNA and ribosomal proteins. This leads to increased binding of ribosomal proteins Rpl5 and Rpl11 to Mdm2 and concomitantly diminished binding between Mdm2 and p53. Altogether, our results demonstrate a dynamic spatiotemporal requirement for rRNA transcription during mammalian cranial NCC development and corresponding tissue-specific threshold sensitivities to disruptions in rRNA transcription in the pathogenesis of congenital craniofacial disorders.

Neuron-specific loss of Ppp6c induces neonatal death and decreases the number of cortical neurons and interneurons.

Protein phosphatase 6 (PP6) is a Ser/Thr protein phosphatase with the catalytic subunit Ppp6c. Recent cell-level studies have revealed that Ppp6c knockdown suppresses neurite outgrowth, suggesting that Ppp6c is involved in the development of the nervous system. We found that the function of PP6 in neurons is essential for mouse survival after birth, as all neural-stem-cell-specific KO (Ppp6cNKO) and neuron-specific KO mice died within 2 days of birth. By contrast, approximately 40 % of oligodendrocyte-specific KO mice died within 2 days of birth, whereas others survived until weaning or later, suggesting that the lethality of PP6 loss differs between neurons and oligodendrocytes. Furthermore, the fetal brain of Ppp6cNKO mice exhibited decreased numbers of neurons in layers V-VI and interneurons in layer I of the neocortex. These results suggest for the first time that Ppp6c is essential for neonatal survival and proper development of neurons and interneurons in the neocortex.

Phase II trial of niraparib for BRCA-mutated biliary tract, pancreatic and other gastrointestinal cancers: NIR-B.

Due to the widespread use of cancer genetic testing in gastrointestinal cancer, the BRCA1/2 genetic mutation has been identified in biliary tract cancer as well as pancreatic cancer. Niraparib is a poly(ADP-ribose) polymerase (PARP) inhibitor, and PARP inhibitors exert their cytotoxicity against cancer cells in the context of homologous recombination deficiency, such as BRCA mutations, via the mechanism of synthetic lethality. The aim of this phase II NIR-B trial is to evaluate the efficacy and safety of niraparib for patients with unresectable advanced or recurrent biliary tract cancer, pancreatic cancer or other gastrointestinal cancers with germline or somatic BRCA1/2 mutations revealed by genetic testing. The primary end point is an investigator-assessed objective response rate in each cohort.Clinical Trial Registration: jRCT2011200023 (ClinicalTrials.gov).

A clinical study to confirm the efficacy and safety of niraparib for people with advanced biliary tract, pancreatic and other abdominal cancers with the BRCA genetic mutation: the NIR-B trial.BRCA gene is involved in repairing DNA injury and plays an important role in cancer growth. Cells with a mutation in the BRCA gene cannot repair DNA using a method called homologous recombination repair. Niraparib is part of a class of drugs called 'PARP inhibitors' that inhibit enzymes called 'PARP' involved in repairing DNA injury, and has shown efficacy against cancers with BRCA gene mutations. BRCA gene mutations are infrequent but have been found in a variety of cancers. The NIR-B trial is a clinical trial to evaluate the efficacy and safety of niraparib for people with advanced biliary tract, pancreatic and other abdominal cancers with BRCA gene mutations.

Treatment strategy for HER2-negative advanced gastric cancer: salvage-line strategy for advanced gastric cancer.

After immune checkpoint inhibitor (ICI) comes into third-line treatment of advanced gastric cancer, the therapeutic strategy has been dramatically changed. Recent first-line regimen, which consists of ICI and chemotherapeutic agents, prolonged progression-free survival, and subsequent treatment options enabled continuous treatment beyond second-line therapy. Moreover, the advent of vascular endothelial growth factor (VEGF)-targeted agents including angiogenesis inhibitors and TKIs provides an opportunity of considering the interaction between ICI and anti-VEGF agents, and facilitating novel treatment proposal. Although clinical benefit of prolonged VEGF blockade after disease progression has not been confirmed in gastric cancer, combination therapy of cytotoxic agents and anti-VEGF agent, such as irinotecan plus ramucirumab demonstrated favorable objective response rate and progression-free survival in third- or later-line setting. In this review, we discuss recent progress and future directions of later-line treatments of HER2-negative advancer gastric cancer.

Trastuzumab deruxtecan in patients with locally advanced or metastatic HER2-positive gastric cancer: a multicenter, open-label, expanded-access study.

BACKGROUND: Trastuzumab deruxtecan (T-DXd) is an antibody-drug conjugate that consists of an anti-human epidermal growth factor receptor 2 (HER2) antibody bound by a cleavable tetrapeptide-based linker to a cytotoxic topoisomerase I inhibitor. Prior to marketing approval in Japan in September 2020, this expanded-access study was conducted to provide T-DXd to previously treated patients with locally advanced or metastatic HER2-positive gastric or gastroesophageal junction adenocarcinomas. METHODS: This multicenter, open-label, expanded-access study was conducted between March 25 and September 25, 2020 at 17 Japanese sites. Previously treated patients with locally advanced or metastatic HER2-positive gastric or gastroesophageal junction adenocarcinomas received T-DXd 6.4 mg/kg via intravenous infusions at 3-week intervals. Serious adverse events (SAEs), all potential cases of interstitial lung disease (ILD)/pneumonitis, all liver-related events potentially meeting Hy's Law criteria, and all cases of overdose were reported on the case report forms. RESULTS: A total of 64 patients were treated with T-DXd. Among the 17 (26.6%) patients with reported SAEs, 10 (15.6%) had SAEs related to T-DXd treatment. Febrile neutropenia was the most common SAE (n = 6). SAEs led to death in six patients; drug-related SAEs (sepsis and febrile neutropenia) led to death in one patient. Drug-related ILD, as determined by the external Adjudication Committee, occurred in three patients (Grade 1, Grade 2, and Grade 3: all n = 1). CONCLUSION: This expanded-access study provided T-DXd to a broader population of Japanese patients prior to marketing approval in Japan, bridging the gap between clinical trials and drug approval. No new safety concerns were identified.

Phase II study of S-1 plus docetaxel as first-line treatment for older patients with advanced gastric cancer (OGSG 0902).

BACKGROUND: Although there is insufficient evidence for the treatment of older patients with advanced gastric cancer, fluorouracil combined with platinum chemotherapy has been recognized as a standard first-line treatment for such populations in Japan despite the lack of efficacy and toxicity data. METHODS: Patients aged 75 years or older with advanced gastric cancer were enrolled. S-1 plus docetaxel (docetaxel: 40 mg/m2, day 1; S-1: 80 mg/m2, days 1-14; q21 days) was repeated every 3 weeks. The primary endpoint was overall response rate. Secondary endpoints were safety, progression-free survival, time to treatment failure, and overall survival. The sample size was calculated as 30 under the hypothesis of an expected response rate of 40% and a threshold response rate of 20%, at a power of 90% and a two-sided alpha value of 5%. RESULTS: From February 2010 to January 2015, 31 patients were enrolled and assessed for efficacy and toxicity. The response rate was 45.2% (95% CI 27.3%-64.0%; p = 0.001) and it exceeded the expected response rate set at 40%. Median progression-free survival was 5.8 months, the 1-year survival rate was 58.1%, and the median survival time was 16.1 months. The major grade 3/4 adverse events were neutropenia (58%), febrile neutropenia (13%), anemia (10%), anorexia (10%), and fatigue (6%). CONCLUSIONS: These findings indicate that S-1 plus docetaxel as first-line treatment for older patients is feasible and that it has promising efficacy against advanced gastric cancer.

ISSLS PRIZE in Basic Science 2024: superiority of nucleus pulposus cell- versus mesenchymal stromal cell-derived extracellular vesicles in attenuating disc degeneration and alleviating pain.

PURPOSE: To investigate the therapeutic potential of extracellular vesicles (EVs) derived from human nucleus pulposus cells (NPCs), with a specific emphasis on Tie2-enhanced NPCs, compared to EVs derived from human bone marrow-derived mesenchymal stromal cells (BM-MSCs) in a coccygeal intervertebral disc degeneration (IDD) rat model. METHODS: EVs were isolated from healthy human NPCs cultured under standard (NPCSTD-EVs) and Tie2-enhancing (NPCTie2+-EVs) conditions. EVs were characterized, and their potential was assessed in vitro on degenerative NPCs in terms of cell proliferation and senescence, with or without 10 ng/mL interleukin (IL)-1ß. Thereafter, 16 Sprague-Dawley rats underwent annular puncture of three contiguous coccygeal discs to develop IDD. Phosphate-buffered saline, NPCSTD-EVs, NPCTie2+-EVs, or BM-MSC-derived EVs were injected into injured discs, and animals were followed for 12 weeks until sacrifice. Behavioral tests, radiographic disc height index (DHI) measurements, evaluation of pain biomarkers, and histological analyses were performed to assess the outcomes of injected EVs. RESULTS: NPC-derived EVs exhibited the typical exosomal morphology and were efficiently internalized by degenerative NPCs, enhancing cell proliferation, and reducing senescence. In vivo, a single injection of NPC-derived EVs preserved DHI, attenuated degenerative changes, and notably reduced mechanical hypersensitivity. MSC-derived EVs showed marginal improvements over sham controls across all measured outcomes. CONCLUSION: Our results underscore the regenerative potential of young NPC-derived EVs, particularly NPCTie2+-EVs, surpassing MSC-derived counterparts. These findings raise questions about the validity of MSCs as both EV sources and cellular therapeutics against IDD. The study emphasizes the critical influence of cell type, source, and culture conditions in EV-based therapeutics.

Assessment of Tie2-Rejuvenated Nucleus Pulposus Cell Transplants from Young and Old Patient Sources Demonstrates That Age Still Matters.

Cell transplantation is being actively explored as a regenerative therapy for discogenic back pain. This study explored the regenerative potential of Tie2+ nucleus pulposus progenitor cells (NPPCs) from intervertebral disc (IVD) tissues derived from young (<25 years of age) and old (>60 years of age) patient donors. We employed an optimized culture method to maintain Tie2 expression in NP cells from both donor categories. Our study revealed similar Tie2 positivity rates regardless of donor types following cell culture. Nevertheless, clear differences were also found, such as the emergence of significantly higher (3.6-fold) GD2 positivity and reduced (2.7-fold) proliferation potential for older donors compared to young sources. Our results suggest that, despite obtaining a high fraction of Tie2+ NP cells, cells from older donors were already committed to a more mature phenotype. These disparities translated into functional differences, influencing colony formation, extracellular matrix production, and in vivo regenerative potential. This study underscores the importance of considering age-related factors in NPPC-based therapies for disc degeneration. Further investigation into the genetic and epigenetic alterations of Tie2+ NP cells from older donors is crucial for refining regenerative strategies. These findings shed light on Tie2+ NPPCs as a promising cell source for IVD regeneration while emphasizing the need for comprehensive understanding and scalability considerations in culture methods for broader clinical applicability.

Human resources for administrative work to carry out a comprehensive genomic profiling test in Japan.

Comprehensive genomic profiling (CGP) tests have been nationally reimbursed in Japan since June 2019 under strict restrictions, and over 46,000 patients have taken the test. Core Hospitals and Designated Hospitals host molecular tumor boards, which is more time-consuming than simply participating in them. We sent a questionnaire to government-designated Cancer Genomic Medicine Hospitals, including all 12 Core Hospitals, all 33 Designated Hospitals, and 117 of 188 Cooperative Hospitals. The questionnaire asked how much time physicians and nonphysicians spent on administrative work for cancer genomic medicine. For every CGP test, 7.6 h of administrative work was needed. Physicians spent 2.7 h/patient, while nonphysicians spent 4.9 h/patient. Time spent preparing for molecular tumor boards, called Expert Panels, was the longest, followed by time spent participating in Expert Panels. Assuming an hourly wage of ¥24,000/h for physicians and ¥2800/h for nonphysicians, mean labor cost was ¥78,071/patient. On a monthly basis, more time was spent on administrative work at Core Hospitals compared with Designated Hospitals and Cooperative Hospitals (385 vs. 166 vs. 51 h/month, respectively, p < 0.001). Consequently, labor cost per month was higher at Core Hospitals than at Designated Hospitals and Cooperative Hospitals (¥3,951,854 vs. ¥1,687,167 vs. ¥487,279/month, respectively, p < 0.001). Completing a CGP test for a cancer patient in Japan is associated with significant labor at each hospital, especially at Core Hospitals. Streamlining the exchange of information and simplifying Expert Panels will likely alleviate this burden.

The Phase II Study of Panitumumab in Chemotherapy-Naïve Frail or Elderly Patients with RAS Wild-type Colorectal Cancer: OGSG 1602 Final Results.

BACKGROUND: We previously reported the response rate of a phase II OGSG1602 study on panitumumab in chemotherapy-naive frail or elderly patients with RAS wild-type unresectable colorectal cancer (CRC) [Terazawa T, Kato T, Goto M, et al. Oncologist. 2021;26(1):17]. Herein, we report a survival analysis. METHODS: Patients aged ≥65 years and considered unsuitable for intensive chemotherapy or aged ≥76 years were enrolled. Primary tumors located from the cecum to the transverse colon were considered right-sided tumors (RSTs); those located from the splenic flexure to the rectum were considered left-sided tumors (LSTs). RESULTS: Among the 36 enrolled patients, 34 were included in the efficacy analysis, with 26 and 8 having LSTs and RSTs, respectively. The median progression-free survival (PFS) and overall survival (OS) were 6.0 [95% CI, 5.4-10.0] and 17.5 months (95% CI, 13.8-24.3), respectively. Although no significant differences existed in PFS between patients with LST and RST {6.6 (95% CI, 5.4-11.5) vs. 4.9 months [95% CI, 1.9-not available (NA), P = .120]}, there were significant differences in OS [19.3 (95% CI, 14.2-NA) vs.12.3 months (95% CI, 9.9-NA), P = .043]. CONCLUSION: Panitumumab showed favorable OS in frail or elderly patients with RAS wild-type CRC and no prior exposure to chemotherapy. Panitumumab may be optimal for patients with LSTs (UMIN Clinical Trials Registry Number UMIN000024528).

Trastuzumab Deruxtecan in Previously Treated HER2-Positive Gastric Cancer.

BACKGROUND: Trastuzumab deruxtecan (DS-8201) is an antibody-drug conjugate consisting of an anti-HER2 (human epidermal growth factor receptor 2) antibody, a cleavable tetrapeptide-based linker, and a cytotoxic topoisomerase I inhibitor. The drug may have efficacy in patients with HER2-positive advanced gastric cancer. METHODS: In an open-label, randomized, phase 2 trial, we evaluated trastuzumab deruxtecan as compared with chemotherapy in patients with HER2-positive advanced gastric cancer. Patients with centrally confirmed HER2-positive gastric or gastroesophageal junction adenocarcinoma that had progressed while they were receiving at least two previous therapies, including trastuzumab, were randomly assigned in a 2:1 ratio to receive trastuzumab deruxtecan (6.4 mg per kilogram of body weight every 3 weeks) or physician's choice of chemotherapy. The primary end point was the objective response, according to independent central review. Secondary end points included overall survival, response duration, progression-free survival, confirmed response (response persisting ≥4 weeks), and safety. RESULTS: Of 187 treated patients, 125 received trastuzumab deruxtecan and 62 chemotherapy (55 received irinotecan and 7 paclitaxel). An objective response was reported in 51% of the patients in the trastuzumab deruxtecan group, as compared with 14% of those in the physician's choice group (P<0.001). Overall survival was longer with trastuzumab deruxtecan than with chemotherapy (median, 12.5 vs. 8.4 months; hazard ratio for death, 0.59; 95% confidence interval, 0.39 to 0.88; P = 0.01, which crossed the prespecified O'Brien-Fleming boundary [0.0202 on the basis of number of deaths]). The most common adverse events of grade 3 or higher were a decreased neutrophil count (in 51% of the trastuzumab deruxtecan group and 24% of the physician's choice group), anemia (38% and 23%, respectively), and decreased white-cell count (21% and 11%). A total of 12 patients had trastuzumab deruxtecan-related interstitial lung disease or pneumonitis (grade 1 or 2 in 9 patients and grade 3 or 4 in 3), as adjudicated by an independent committee. One drug-related death (due to pneumonia) was noted in the trastuzumab deruxtecan group; no drug-related deaths occurred in the physician's choice group. CONCLUSIONS: Therapy with trastuzumab deruxtecan led to significant improvements in response and overall survival, as compared with standard therapies, among patients with HER2-positive gastric cancer. Myelosuppression and interstitial lung disease were the notable toxic effects. (Funded by Daiichi Sankyo; DESTINY-Gastric01 ClinicalTrials.gov number, NCT03329690.).

Impact of tumor shrinkage pattern with biweekly triplet gemcitabine+cisplatin+S-1 regimen for biliary tract cancers: Implications for neoadjuvant therapy from the data of KHBO1401 (KHBO1401-1A study).

BACKGROUND: The multicenter randomized phase III KHBO1401 study (gemcitabine+cisplatin+S-1 [GCS] versus GC in biliary tract cancers [BTC]) demonstrated that GCS not only prolonged patient survival but also achieved a high response rate and that it should be good for neoadjuvant therapy. Therefore, to explore the possibilities of neoadjuvant therapy, we investigated the tumor shrinkage pattern. METHODS: Among the total of 246 patients enrolled in the KHBO1401, the tumor shrinkage pattern and survival were investigated in patients with measurable BTC (n=183, 74%; GCS, n=91; GC, n=92). RESULTS: The tumor shrinkage pattern could be divided to 4 categories based on the response at 100 days after enrollment: category A (<-30% in size), B (-30% to 0%), C (0% to +20%), and D (>+20%). The GCS arm included more category A and B cases (61 [67%] vs. 33 [36%], P<0.0001). Each category predicted best response and overall survival (P<0.0001). Category A showed sustained tumor response compared with category B; in GCS, the time to maximum tumor response was 165 ± 76 days in category A and 139 ± 78 in category B. Categories C and D did not achieve tumor shrinkage. The maximum tumor shrinkage size in category A was -53% in the GCS arm and -65% in the GC arm (P=0.0892). Twenty percent of patients in the GCS showed tumor regrowth 154 ± 143 days later. CONCLUSION: GCS provided faster and greater tumor shrinkage with better survival in comparison to GC, although 20% of patients showed re-growth after 6 cycles.

Effect of the number of cycles of docetaxel + S-1 therapy on long-term survival in adjuvant chemotherapy for stage III gastric cancer. A pooled analysis of the OGSG0604 and OGSG1002 trials.

BACKGROUND: S-1 plus docetaxel (DS) therapy followed by S-1 is the standard of care in Japan in postoperative adjuvant chemotherapy for stage III gastric cancer, but long-term survival and the number of DS cycles required are unclear. The purpose of this study was to investigate the impact of the number of cycles of DS therapy on the 5-year survival in stage III gastric cancer in a pooled analysis of two phase II trials (OGSG0604 and OGSG1002). PATIENTS AND METHODS: Patients with histologically confirmed stage III gastric cancer who underwent gastrectomy with D2 lymphadenectomy were enrolled in this pooled analysis. They received DS therapy for four or eight cycles, followed by S-1 until 1 year postgastrectomy. The 5-year overall survival (OS) and the 5-year disease free survival (DFS) by the landmark analysis was evaluated. RESULTS: In total, 113 patients from the OGSG0604 and OGSG1002 trials were enrolled in this study. The landmark analysis showed a 5-year OS that was better with four to eight cycles of DS therapy than with one to three cycles of DS therapy, with the best 5-year OS of 77.4% (95% confidence interval, 66.5-90.1%) for eight cycles. The 5-year DFS was approximately 66% when four or eight cycles of DS therapy were given. CONCLUSION: Although eight cycles of DS therapy may prolong prognosis, the present study did not provide a clear conclusion as to how many DS therapy cycles are needed to improve prognosis after D2 gastrectomy for stage III gastric cancer. TRIAL REGISTRATION: Registration number: UMIN00000714 and UMIN000004440.

Phase I dose-escalation study on irinotecan, cisplatin, and S-1 combination in chemotherapy-naïve patients with HER2-negative advanced gastric cancer (HERBIS-4B, OGSG 1106).

BACKGROUND: The development of triplet regimens for advanced gastric cancer is challenging. The aim of this phase I dose-escalation study was to determine the maximum tolerated dose and recommended dose of the combination of irinotecan, cisplatin, and S-1 in chemotherapy-naïve patients with HER2-negative advanced gastric cancer. METHODS: The 3 + 3 design was adopted. Every 4 weeks, patients received an escalating dose of intravenous irinotecan (100-150 mg/m2) on day 1 and fixed doses of intravenous cisplatin (60 mg/m2) on day 1 and oral S-1 (80 mg/m2) on days 1 to 14. RESULTS: Twelve patients were enrolled in two dose level cohorts. In the level 1 cohort (irinotecan 100 mg/m2, cisplatin 60 mg/m2, and S-1 80 mg/m2), dose-limiting toxicity including grade 4 neutropenia and febrile neutropenia occurred in one of six patients, whereas in the level 2 cohort (irinotecan 125 mg/m2, cisplatin 60 mg/m2, and S-1 80 mg/m2), dose-limiting toxicities including grade 4 neutropenia developed in two of six patients. Thus, the level 1 and 2 doses were determined to be the recommended and maximum tolerated doses, respectively. Common grade 3 or higher adverse events were neutropenia (75%; n = 9), anemia (25%; n = 3), anorexia (8%; n = 1), and febrile neutropenia (17%; n = 2). Irinotecan, cisplatin, and S-1 combination therapy achieved an overall response rate of 67% with a median progression-free survival and overall survival of 19.3 and 22.4 months, respectively. CONCLUSIONS: The potential treatment efficacy of this triplet regimen in HER2-negative advanced gastric cancer warrants further evaluation, especially in patients requiring intensive chemotherapy.

Worsening ambulation in elderly patients with cervical odontoid fractures: A nationwide multicenter study in Japan.

BACKGROUND: Despite the increasing prevalence of cervical odontoid fractures in older adults, the treatment strategy is controversial. The objectives of the current study are to investigate the prognosis and complications of cervical odontoid fractures in elderly patients and to identify factors associated with worsening of ambulation after 6 months. METHODS: This multicenter, retrospective study included 167 patients aged 65 years or older with odontoid fractures. Patient demographic and treatment data were investigated and compared according to the treatment strategy. To determine associations with worsening ambulation after 6 months, we focused on the treatment strategies (nonsurgical treatment [collar immobilization or halo vest], conversion to surgery, or initial surgery) and patients' background. RESULTS: Patients who received nonsurgical treatment were significantly older, and patients who underwent surgery had more Anderson-D'Alonzo type 2 fractures. Of the patients initially treated nonsurgically, 26% later underwent surgery. Numbers of complications, including death, and degrees of ambulation after 6 months did not differ significantly among treatment strategies. Patients who had worsened ambulation after 6 months were significantly more likely to be older than 80 years, to have needed assistance with walking before injury, and to have cerebrovascular disease. Multivariable analysis showed that a score of ≥2 on the 5-item modified frailty index (mFI-5) was significantly associated with worsening ambulation. CONCLUSIONS: Preinjury mFI-5 scores of ≥2 were significantly associated with worsening ambulation 6 months after treatment of cervical odontoid fractures in older adults.

Concepts of Regeneration for Spinal Diseases in 2023.

It is our pleasure to announce the publication of the Special Issue "Regeneration for Spinal Diseases 3.0" in the International Journal of Molecular Sciences (ISSN 1422-0067) [...].

Investigation of the Mitigation of DMSO-Induced Cytotoxicity by Hyaluronic Acid following Cryopreservation of Human Nucleus Pulposus Cells.

To develop an off-the-shelf therapeutic product for intervertebral disc (IVD) repair using nucleus pulposus cells (NPCs), it is beneficial to mitigate dimethyl sulfoxide (DMSO)-induced cytotoxicity caused by intracellular reactive oxygen species (ROS). Hyaluronic acid (HA) has been shown to protect chondrocytes against ROS. Therefore, we examined the potential of HA on mitigating DMSO-induced cytotoxicity for the enhancement of NPC therapy. Human NPC cryopreserved in DMSO solutions were thawed, mixed with equal amounts of EDTA-PBS (Group E) or HA (Group H), and incubated for 3-5 h. After incubation, DMSO was removed, and the cells were cultured for 5 days. Thereafter, we examined cell viability, cell proliferation rates, Tie2 positivity (a marker of NP progenitor cells), and the estimated numbers of Tie2 positive cells. Fluorescence intensity of DHE and MitoSOX staining, as indicators for oxidative stress, were evaluated by flow cytometry. Group H showed higher rates of cell proliferation and Tie2 expressing cells with a trend toward suppression of oxidative stress compared to Group E. Thus, HA treatment appears to suppress ROS induced by DMSO. These results highlight the ability of HA to maintain NPC functionalities, suggesting that mixing HA at the time of transplantation may be useful in the development of off-the-shelf NPC products.

Recombinant Laminin-511 Fragment (iMatrix-511) Coating Supports Maintenance of Human Nucleus Pulposus Progenitor Cells In Vitro.

The angiopoietin-1 receptor (Tie2) marks specific nucleus pulposus (NP) progenitor cells, shows a rapid decline during aging and intervertebral disc degeneration, and has thus sparked interest in its utilization as a regenerative agent against disc degeneration. However, the challenge of maintaining and expanding these progenitor cells in vitro has been a significant hurdle. In this study, we investigated the potential of laminin-511 to sustain Tie2+ NP progenitor cells in vitro. We isolated cells from human NP tissue (n = 5) and cultured them for 6 days on either standard (Non-coat) or iMatrix-511 (laminin-511 product)-coated (Lami-coat) dishes. We assessed these cells for their proliferative capacity, activation of Erk1/2 and Akt pathways, as well as the expression of cell surface markers such as Tie2, GD2, and CD24. To gauge their regenerative potential, we examined their extracellular matrix (ECM) production capacity (intracellular type II collagen (Col2) and proteoglycans (PG)) and their ability to form spherical colonies within methylcellulose hydrogels. Lami-coat significantly enhanced cell proliferation rates and increased Tie2 expression, resulting in a 7.9-fold increase in Tie2-expressing cell yields. Moreover, the overall proportion of cells positive for Tie2 also increased 2.7-fold. Notably, the Col2 positivity rate was significantly higher on laminin-coated plates (Non-coat: 10.24% (±1.7%) versus Lami-coat: 26.2% (±7.5%), p = 0.010), and the ability to form spherical colonies also showed a significant improvement (Non-coat: 40.7 (±8.8)/1000 cells versus Lami-coat: 70.53 (±18.0)/1000 cells, p = 0.016). These findings demonstrate that Lami-coat enhances the potential of NP cells, as indicated by improved colony formation and proliferative characteristics. This highlights the potential of laminin-coating in maintaining the NP progenitor cell phenotype in culture, thereby supporting their translation into prospective clinical cell-transplantation products.

[A Case of Squamous Cell Carcinoma of the Lower Rectum].

A 61-year-old man presented with dyschezia, and further examination revealed squamous cell carcinoma of the lower rectum invading the bladder and seminal vesicles. The clinical diagnosis was squamous cell carcinoma of the lower rectum, cT4b(bladder and seminal vesicle)N0M0, cStage Ⅱc. Neoadjuvant chemoradiotherapy was administered with external irradiation of the entire pelvis(50.4 Gy/28 Fr)and chemotherapy with 5-fluorouracil, Leucovorin, and oxaliplatin(FOLFOX). Once tumor shrinkage was observed 3 months after chemoradiotherapy, laparoscopic total pelvic exenteration with TaTME approach was performed. The patient was discharged on the 26th postoperative day without any postoperative complications. Histopathological examination showed only squamous cell carcinoma component with Grade 1a histological treatment effect. The pathological diagnosis was ypT4b(bladder, seminal vesicle)ypN0cM0, ypStage Ⅱc. The patient was alive without any recurrence 6 months after surgery.

Chronological improvement in precision oncology implementation in Japan.

In Japan, comprehensive genomic profiling (CGP) tests for refractory cancer patients have been approved since June 2019, under the requirement that all cases undergoing CGP tests are annotated by the molecular tumor board (MTB) at each government-designated hospital. To investigate improvement in precision oncology, we evaluated and compared the proportion of cases receiving matched treatments according to CGP results and those recommended to receive genetic counseling at all core hospitals between the first period (11 hospitals, June 2019 to January 2020) and second period (12 hospitals, February 2020 to January 2021). A total of 754 and 2294 cases underwent CGP tests at core hospitals in the first and second periods, respectively; 28 (3.7%) and 176 (7.7%) patients received matched treatments (p < 0.001). Additionally, 25 (3.3%) and 237 (10.3%) cases were recommended to receive genetic counseling in the first and second periods, respectively (p < 0.001). The proportion was associated with the type of CGP test: tumor-only (N = 2391) vs. tumor-normal paired (N = 657) analysis (10.0% vs. 3.5%). These results suggest that recommendations regarding available clinical trials in networked MTBs might contribute to increasing the numbers of matched treatments, and that tumor-normal paired rather than tumor-only tests can increase the efficiency of patient referrals for genetic counseling.