Effects of Extended Fixation on Advanced Gastric Cancer HER2 Status Assessment Using IHC and FISH.

BACKGROUND/AIM: In gastric cancer, accurate determination of human epidermal growth factor receptor type 2 (HER2) status is crucial for treatment decision-making. However, the optimal formalin fixation time of gastric cancer specimens for HER2 status determination remains unestablished. Here, we investigated real-world data on formalin overfixation and its effect on HER2 status determination in gastric cancer. PATIENTS AND METHODS: We comprehensively analyzed HER2 testing results in 228 gastric cancer specimens, including those subjected to formalin overfixation. Subsequently, we divided 52 resected specimens of advanced gastric cancer into three groups and studied the effects of short-term (6-72 h) and long-term (1 and 2 weeks) fixation on HER2 status determination using immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). RESULTS: A total of 21.5% (49/228) of the specimens were HER2-positive, whereas 78.5% (179/228) were negative. Among the HER2-negative specimens, no biopsies were overfixed, whereas 12.5% (9/72) of the surgical resection specimens were overfixed. The HER2 status of the 6-72-h group was 82.7% and 76.9% identical to that of the 1- and 2-week groups, when determined using IHC, and 73.1% and 36.5%, when determined using FISH, respectively. However, HER2 determination was not feasible in 26.9% and 63.5% of the specimens in the 1- and 2-week groups, respectively. CONCLUSION: Formalin overfixation may hinder the determination of HER2 status and should be avoided in gastric cancer sample preparation.

Steroid-resistant minimal change nephrotic syndrome associated with thymoma treated effectively with rituximab following thymectomy and cyclosporine: a case report.

BACKGROUND: Minimal change nephrotic syndrome (MCNS) can be complicated by thymoma; however, no standard therapy for thymoma-associated MCNS has yet been established. We herein describe a case of steroid-resistant MCNS associated with thymoma, treated effectively with rituximab. CASE PRESENTATION: A 71-year-old Japanese man was referred to our department with severe proteinuria (20 g/gCr). Renal biopsy showed minimal change disease and computed tomography revealed an anterior mediastinal mass. Based on these findings, he was diagnosed with thymoma-associated MCNS. He was treated with oral prednisolone (50 mg/day) and cyclosporine, and underwent thymectomy and plasma exchange. However, no improvement in proteinuria was observed. He therefore received intravenous rituximab 500 mg, resulting in a marked decrease in proteinuria from 5328 to 336 mg/day after 1 week. CONCLUSIONS: This case suggests that rituximab might be an effective therapy in patients with steroid-resistant MCNS associated with thymoma.

High serum proteinase-3 levels predict poor progression-free survival and lower efficacy of bevacizumab in metastatic colorectal cancer.

BACKGROUND: To improve the prognosis of patients with metastatic colorectal cancer (mCRC), investigating predictive biomarkers of their prognosis and chemotherapeutic responsiveness is necessary. This study aimed to analyze the clinical significance of serum proteinase-3 (PRTN3) as a predictor for prognosis and chemosensitivity, especially to bevacizumab therapy, in mCRC. METHODS: This single-center retrospective observational study enrolled 79 patients with mCRC in our hospital and 353 patients with colorectal cancer in the TCGA database. Preoperative serum PRTN3 levels were measured using an enzyme-linked immunosorbent assay. The clinicopathological characteristics and prognosis according to serum PRTN3 levels were then evaluated. PRTN3 expression in tumor and stromal cells was evaluated immunohistochemically. The impact of PRTN3 levels on angiogenesis and bevacizumab sensitivity was evaluated using the tube formation assay. RESULTS: Serum PRTN3 levels were an independent poor prognostic factor for progression-free survival (PFS) (hazard ratio, 2.082; 95% confidence interval, 1.118-3.647; P=0.010) in patients with mCRC. Similarly, prognostic analysis with TCGA data sets showed poorer overall survival in patients with PRTN3 expression than that in patients without PRTN3 expression, especially in patients with stage IV. Immunohistochemical analysis of resected specimens revealed that stromal neutrophils expressed PRTN3, and their expression level was significantly correlated with serum PRTN3 levels. Interestingly, the effectiveness of first-line chemotherapy was significantly poorer in the high serum PRTN3 level group. High serum PRTN3 was significantly associated with poor PFS (hazard ratio, 3.027; 95% confidence interval, 1.175-7.793; P=0.0161) in patients treated with bevacizumab, an anti-angiogenic inhibitor. The tube formation assay revealed that PRTN3 administration notably augmented angiogenesis while simultaneously attenuating the anti-angiogenic influence exerted by bevacizumab therapy. CONCLUSIONS: Serum PRTN3 levels could be a novel predictive biomarker of PFS of first-line chemotherapy, especially for bevacizumab therapy, in patients with mCRC.

A case of renal vein thrombosis associated with autoimmune hemolytic anemia and eosinophilic granulomatous polyangiitis.

Only a few cases of renal vein thrombosis (RVT) occurring in patients with vasculitis have been reported. RVT associated with vasculitis and hemolytic anemia has not been reported yet. We describe here a patient with RVT complicated by pulmonary embolism, autoimmune hemolytic anemia, and eosinophilic granulomatous polyangiitis. A 69-year-old Japanese man who had been treated with corticosteroids was referred to our department for severe proteinuria (4.32 g/gCr). Abdominal ultrasonography showed bilateral RVT, and contrast-enhanced computed tomography showed bilateral pulmonary embolism. Therefore, the patient was diagnosed with RVT complicated by pulmonary embolism. Anticoagulation therapy with heparin followed by apixaban was started. Thereafter, the D-dimer concentration decreased from 8.3 to 1.2 µg/mL, and urinary protein excretion improved to 0.62 g/gCr. Renal function was unchanged with an estimated glomerular filtration rate of 68.8 mL/minute/1.73 m2. The thrombi in both renal veins and pulmonary arteries gradually regressed. Clinicians should be aware of this complication when worsening proteinuria is observed during steroid therapy in patients with autoimmune hemolytic anemia and eosinophilic granulomatous polyangiitis.

IL-6 Levels Correlate with Prognosis and Immunosuppressive Stromal Cells in Patients with Colorectal Cancer.

BACKGROUND: The prognosis for patients with colorectal cancer (CRC) is determined by tumor characteristics as well as the host immune response. This study investigated the relationship between an immunosuppressive state and patient prognosis by evaluating the systemic and tumor microenvironment (TME) interleukin (IL)-6 levels. METHODS: Preoperative serum IL-6 levels were measured using an electrochemiluminescence assay. Expression of IL-6 in tumor and stromal cells was evaluated immunohistochemically in 209 patients with resected CRC. Single-cell analysis of tumor-infiltrating immune cells was performed using mass cytometry in 10 additional cases. RESULTS: Elevated serum IL-6 levels were associated with elevated stromal IL-6 levels and a poor prognosis for patients with CRC. High IL-6 expression in stromal cells was associated with low-density subsets of CD3+ and CD4+ T cells as well as FOXP3+ cells. Mass cytometry analysis showed that IL-6+ cells among tumor-infiltrating immune cells were composed primarily of myeloid cells and rarely of lymphoid cells. In the high-IL-6-expression group, the percentages of myeloid-derived suppressor cells (MDSCs) and CD4+FOXP3highCD45RA- effector regulatory T cells (eTreg) were significantly higher than in the low-IL-6-expression group. Furthermore, the proportion of IL-10+ cells in MDSCs and that of IL-10+ or CTLA-4+ cells in eTregs correlated with IL-6 levels. CONCLUSION: Elevated serum IL-6 levels were associated with stromal IL-6 levels in CRC. High IL-6 expression in tumor-infiltrating immune cells also was associated with accumulation of immunosuppressive cells in the TME.

Serum CCL7 Is a Novel Prognostic Biomarker of Metastatic Colorectal Cancer.

BACKGROUND/AIM: Colorectal cancer is the third most common cancer globally, and the poor prognosis of patients with metastatic colorectal cancer (mCRC) warrants urgent attention. We previously obtained 10 candidate serum biomarkers for mCRC. Our aim with this study was to determine the prognostic performance of the pre-treatment serum C-C motif chemokine ligand 7 (CCL7) concentration in patients with mCRC. PATIENTS AND METHODS: Protein concentrations of CCL7 were examined using ELISA and immunohistochemistry for serum (n=110) and surgical specimens (n=85), respectively, of patients with mCRC. The relationship between protein concentration and prognosis was examined using Cox regression analysis, receiver operator characteristic curve analysis and the Kaplan-Meier method. RESULTS: The overall survival (OS) of patients with high concentrations of serum CCL7 was significantly poorer than that of patients with low concentrations. Patients with a high CCL7 concentration in the stroma had significantly poorer outcomes than those with a low concentration. The concentrations of carcinoembryonic antigen (CEA) and carbohydrate antigen 19-9 were significantly higher in the high-CCL7 group, compared to those in the low-CCL7 group. Univariate and multivariate analysis revealed that serum CCL7 concentration was a significant prognostic factor for mCRC. The combination of serum CCL and CEA concentrations was also useful in this regard (area under the curve=0.71). CONCLUSION: The combined pre-treatment serum levels of CCL7 and CEA are useful prognostic biomarkers for mCRC.

[A Severe Case of Bleeding from Duodenal Invasion Due to Co-Morbid IPMC with Arcuate Ligament Syndrome and IPDA Aneurysm].

An 81-year-old man with a history of left hemiplegia due to a cerebral hemorrhage was admitted to a clinic because of tarry stools. Endoscopic findings revealed an ulcerative lesion with hemorrhage in the descending duodenum. The patient was transferred to our hospital for treatment. Because endoscopic hemostasis was impossible, interventional radiology(IVR) hemostasis was performed using coil embolization for the feeding artery. Simultaneously, angiography showed stenosis of the root of the celiac axis due to arch ligament syndrome and an aneurysm of the inferior pancreaticoduodenal artery (IPDA). Due to the risk of rebleeding, subtotal stomach-preserving pancreatoduodenectomy was performed after the patient's overall condition had stabilized. Despite dissecting the arcuate ligament, the hepatic artery flow did not improve. Hence, a direct arterial anastomosis between the middle colic artery and the gastroduodenal artery was performed. Furthermore, due to the proximity of the IPDA aneurysm to the superior mesenteric artery, IVR embolization for the IPDA aneurysm was performed on postoperative day 8, and he was transferred to a rehabilitation hospital on postoperative day 57. The pathological result was invasive intraductal papillary mucinous carcinoma(IPMC). The patient has been an outpatient with no recurrence 12 months postoperatively.

Wnt signal-dependent antero-posterior specification of early-stage CNS primordia modeled in EpiSC-derived neural stem cells.

The specification of the embryonic central nervous system (CNS) into future brain (forebrain, midbrain, or hindbrain) and spinal cord (SC) regions is a critical step of CNS development. A previous chicken embryo study indicated that anterior epiblast cells marked by Sox2 N2 enhancer activity are specified to the respective brain regions during the transition phase of the epiblast to the neural plate-forming neural primordium. The present study showed that the SC precursors positioned posterior to the hindbrain precursors in the anterior epiblast migrated posteriorly in contrast to the anterior migration of brain precursors. The anteroposterior specification of the CNS precursors occurs at an analogous time (∼E7.5) in mouse embryos, in which an anterior-to-posterior incremental gradient of Wnt signal strength was observed. To examine the possible Wnt signal contribution to the anteroposterior CNS primordium specification, we utilized mouse epiblast stem cell (EpiSC)-derived neurogenesis in culture. EpiSCs maintained in an activin- and FGF2-containing medium start neural development after the removal of activin, following a day in a transitory state. We placed activin-free EpiSCs in EGF- and FGF2-containing medium to arrest neural development and expand the cells into neural stem cells (NSCs). Simultaneously, a Wnt antagonist or agonist was added to the culture, with the anticipation that different levels of Wnt signals would act on the transitory cells to specify CNS regionality; then, the Wnt-treated cells were expanded as NSCs. Gene expression profiles of six NSC lines were analyzed using microarrays and single-cell RNA-seq. The NSC lines demonstrated anteroposterior regional specification in response to increasing Wnt signal input levels: forebrain-midbrain-, hindbrain-, cervical SC-, and thoracic SC-like lines. The regional coverage of these NSC lines had a range; for instance, the XN1 line expressed Otx2 and En2, indicating midbrain characteristics, but additionally expressed the SC-characteristic Hoxa5. The ranges in the anteroposterior specification of neural primordia may be narrowed as neural development proceeds. The thoracic SC is presumably the posterior limit of the contribution by anterior epiblast-derived neural progenitors, as the characteristics of more posterior SC regions were not displayed.

Biological properties of Staphylococcus virus ΦSA012 for phage therapy.

Staphylococcus virus ΦSA012 has a wide host range and efficient lytic activity. Here, we assessed the biological stability of ΦSA012 against temperature, freeze-thawing, and pH to clinically apply the phage. In addition, inoculation of ΦSA012 through i.p. and i.v. injections into mice revealed that phages were reached the limit of detection in serum and accumulated notably spleens without inflammation at 48 h post-inoculation. Furthermore, inoculation of ΦSA012 through s.c. injections in mice significantly induced IgG, which possesses neutralizing activity against ΦSA012 and other Staphylococcus viruses, ΦSA039 and ΦMR003, but not Pseudomonas viruses ΦS12-3 and ΦR18 or Escherichia viruses T1, T4, and T7 in vitro. Immunoelectron microscopic analysis showed that purified anti-phage IgG recognizes the long-tail fiber of staphylococcus viruses. Although S. aureus inoculation resulted in a 25% survival rate in a mouse i.p. model, ΦSA012 inoculation (i.p.) improved the survival rate to 75%; however, the survival rate of ΦSA012-immunized mice decreased to less than non-immunized mice with phage i.v. injection at a MOI of 100. These results indicated that ΦSA012 possesses promise for use against staphylococcal infections but we should carefully address the appropriate dose and periods of phage administration. Our findings facilitate understandings of staphylococcus viruses for phage therapy.

Risk factors for non-gastric-cancer-related death after gastrectomy in elderly patients.

Aim: To identify preoperative factors, especially other diseases that cause death, that are associated with the prognosis of gastrectomy in elderly patients with gastric cancer. Methods: This retrospective study included a total of 211 consecutive patients aged ≥75 years who underwent radical gastrectomy due to gastric cancer. Time-dependent receiver operating characteristic curve analysis was performed to determine the optimal cutoff values for various perioperative factors. Risk factors for the overall survival and death from other diseases were analyzed using the Cox proportional hazards model. Results: Among the all perioperative factors, sex, neutrophil-to-lymphocyte ratio, skeletal muscle mass index, and lymph node dissection in accordance with guidelines or not extracted as independent risk factors for death from other diseases. In an analysis restricted to the preoperative factors, sex, neutrophil-to-lymphocyte ratio, and skeletal muscle mass index of the patients were extracted as independent risk factors for death from other diseases and overall survival. We divided the patients into four groups according to the number of preoperative risk factors for death from other diseases and found that the 5-year non-gastric-cancer-related survival was different among the four groups (risk factor 0, 91.7%; risk factor 1, 83.3%; risk factor 2, 56.3%; risk factor 3, 27.2%; P < 0.001). Conclusion: Male sex, low skeletal muscle mass index, and high neutrophil-to-lymphocyte ratio are risk factors for non-gastric-cancer-related death and the overall survival of elderly patients undergoing gastrectomy. Cautious treatment strategies are needed for elderly gastric cancer patients with many risk factors.

CD4 and FOXP3 as predictive markers for the recurrence of T3/T4a stage II colorectal cancer: applying a novel discrete Bayes decision rule.

BACKGROUND: We recently reported the relapse-free survival (RFS) significance of the combination of CD4+ and forkhead box P3+ (FOXP3) T-cell densities identified by immunohistochemistry in patients with stage I, II, and III colorectal cancer (CRC) who underwent curative resections. This study was designed to determine the optimal combination of markers that predict recurrence in patients with T factors of T3/T4a stage II CRC by applying a novel Bayes decision rule. METHODS: Using 137 cancer tissue specimens from T3/T4a stage II patients, 12 clinicopathologic and immune factors were analysed as predictive candidates for recurrence. RESULTS: Our study showed that the combination of low CD4+ and low FOXP3+ T-cell densities resulted in extremely poor RFS. CONCLUSIONS: Adjuvant chemotherapy may be considered for patients with a combination of low CD4+ and low FOXP3+ T-cell densities. The discovery of this new prognostic indicator is important for the appropriate management of patients undergoing curative resection for T3/T4a stage II CRC.

[Tumor Immunogenicity and Immune Checkpoint Inhibitors].

Cancer immunotherapy including immune checkpoint inhibitors(ICIs)have established itself as the fourth cancer therapy. However, the response rate of ICIs is still only about 20%, and tumors resistant to ICIs are often so-called"cold-tumor"with low tumor immunogenicity. Therefore, research and development is being conducted worldwide on how to convert cold- tumors into hot-tumors with high immunogenicity. In this paper, we review the relationship between tumor immunogenicity and ICI, as well as therapeutic methods to enhance tumor immunogenicity, and introduce our research about novel cancer peptide vaccination therapy.

Elevated expression of RAB3B plays important roles in chemoresistance and metastatic potential of hepatoma cells.

BACKGROUND: Cancer stem cells (CSCs) are thought to play important roles in carcinogenesis, recurrence, metastasis, and therapy-resistance. We have successfully induced cancer stem-like sphere cells (CSLCs) which possess enhanced chemoresistance and metastatic potential. To enable the development of targeted therapy against CSLCs, we identified a gene responsible for this phenotype in CSLC. METHODS: Human hepatoma cell line SK-HEP-1 was used for CSLC induction with a unique sphere inducing medium, and HuH-7 cells were used as non-sphere forming cells in the same condition. RNA-sequencing was performed followed by validation with quantitative RT-PCR and western blotting. Knockdown experiments were done by using CRISPR-Cas9 genome-editing, and the rescue experiments were performed using the expressing plasmid vector. Chemoresistance and liver metastasis of the cells, was studied following the splenic injection of cells to severely immune deficient mice and evaluated using the MTS assay. Quantification of exosomes in the medium was done using ELISA. RESULTS: RAB3B was identified as an up-regulated gene in both CSLCs and prognostically poor hepatocellular carcinoma (HCC) by RNA-sequencing. RAB3B-KD cells showed altered CSLC phenotypes such as sphere formation, chemoresistance, and metastatic potentials, and those were rescued by RAB3B complementation. Increased exosome secretion was observed in CSLCs, and it was not observed in the RAB3B-KD cells. In addition, the RAB3B expression correlated with the expression of ABCG2, APOE, LEPR, LXN, and TSPAN13. CONCLUSION: The up regulation of RAB3B may play an important role in the chemoresistance and metastatic potential of CSLCs.

Live imaging of avian epiblast and anterior mesendoderm grafting reveals the complexity of cell dynamics during early brain development.

Despite previous intensive investigations on epiblast cell migration in avian embryos during primitive streak development before stage (st.) 4, this migration at later stages of brain development has remained uninvestigated. By live imaging of epiblast cells sparsely labeled with green fluorescence protein, we investigated anterior epiblast cell migration to form individual brain portions. Anterior epiblast cells from a broad area migrated collectively towards the head axis during st. 5-7 at a rate of 70-110 µm/h, changing directions from diagonal to parallel and forming the brain portions and abutting head ectoderm. This analysis revised the previously published head portion precursor map in anterior epiblasts at st. 4/5. Grafting outside the brain precursor region of mCherry-expressing nodes producing anterior mesendoderm (AME) or isolated AME tissues elicited new cell migration towards ectopic AME tissues. These locally convergent cells developed into secondary brains with portions that depended on the ectopic AME position in the anterior epiblast. Thus, anterior epiblast cells are bipotent for brain/head ectoderm development with given brain portion specificities. A brain portion potential map is proposed, also accounting for previous observations.

[A Case in Which FOLFOXIRI Was Useful as Preoperative Chemotherapy for Locally Advanced Rectal Cancer with Difficulty in Securing CRM].

The patient is a 65-year-old woman. Colonoscopy performed for close examination of constipation and lower abdominal pain revealed a circumferential type 3 lesion in the rectum Ra. A CT scan showed invasion of the primary lesion into the extramural and sacral front and multiple metastases in the mesorectal lymph nodes but no distant metastasis. Staging laparoscopy was performed. As the mesorectum around the primary lesion was tightly adherent, it was difficult to R0 resection; hence, only construction of colostomy was performed. We have introduced chemotherapy(FOLFOXIRI plus bevacizumab therapy), and 4 courses were administered. Post-treatment CT scan showed that the peri-invasiveness of the primary tumor had disappeared and the enlarged lymph nodes had shrunk. Furthermore, SUVmax of PET-CT for main lesion was decreased, dramatically. On day 109 after the initial surgery, laparoscopic low anterior resection was performed. Although the left hypogastric nerve was resected, other areas could be dissected and R0 resection could be performed. FOLFOXIRI therapy has shown good early-tumor shrinkage and depth of response and may be useful for patients with locally advanced rectal cancer who have difficulty securing circumferential resection margin(CRM).

[A Case of Anastomotic Recurrence of Rectal Cancer Treated by Laparoscopic Total Pelvic Exenteration after Neoadjuvant Chemoradiation].

Patient is 69-year-old man, who underwent a high anterior resection with laparoscopic support for rectal cancer. The patient was diagnosed with anastomotic recurrent rectal cancer after 14 months after surgery. The pelvic MRI scan showed invasion of the prostate and seminal vesicles, so NACRT was performed. Tumors were found to have decreased in size, although there was still some residual invasion of the prostate and seminal vesicle. Laparoscopic total pelvic exenteration (Lap-TPE), and combined excision of the anal elevator muscle and bladder were performed. Preoperative diagnosis was ycT4b, N0, M0, ycStage Ⅱ, and pathological diagnosis was pT4b (prostate and seminal vesicles), INF b, Ly2, v2, Pn1b, pPM0, pDM0, pRM0, and pN0. Laparoscopic surgery allowed to operate safely, with minimal blood loss and a good field of vision. After postoperative adjuvant chemotherapy, lung and liver metastasis appeared after 6 months after surgery, but there was no local recurrence. The patient is treated with chemotherapy, and the metastases are under control. The patient is survive 17 months after Lap-TPE.

[A Case of Gastrointestinal Stromal Tumor of the Lower Rectum That Enabled Minimally Invasive Surgery with Imatinib Mesylate as Neoadjuvant Chemotherapy].

A 67-year-old woman was admitted with melena. A colonoscopy detected a 50 mm submucosal tumor close to the dentate line. We diagnosed the rectal gastrointestinal stromal tumor by EUS-FNA. With the expectation of tumor shrinkage and strong hope of the patient, we started imatinib mesylate as neoadjuvant chemotherapy. A CT scan after 3 months after administration of imatinib mesylate showed the reduction of the size to 35 mm. We operated transanal endoscopic surgery considering the localization of the tumor. From histopathological findings, the tumor was low risk in the modified-Fletcher classification, and low risk in the Miettinen classification. Eight months after the operation, no recurrence was observed without further adjuvant chemotherapy. In this case, we were able to resect the tumor without injuring the film of tumor by operating transanal endoscopic surgery, because of tumor shrinkage with imatinib mesylate as neoadjuvant chemotherapy. I considered that using imatinib mesylate preoperatively was contributed to minimally invasive surgery.

[Postoperative Management of Refractory Pleural Effusion in a Patient with Esophageal Cancer Accompanied by Cirrhosis].

This study reports a case of a 61-year-old man with a chief complaint of anemia. The patient was diagnosed with esophageal cancer(Stage Ⅰ). Preoperative examination revealed alcoholic liver cirrhosis(Child-Pugh A, liver damage B). After a period of abstinence to improve liver function, minimally invasive esophagectomy, retrosternal reconstruction with a gastric tube, and two-field lymph node dissection were performed. The thoracic duct was preserved during the operation. Post- surgery, the bill pleural effusion was increased. Drainage was initiated using thoracentesis with frosemide, spironolactone, and tolvaptan. On post-operating day(POD)35, the patient was discharged; however, right pleural effusion continued to increase. Therefore, cell-free and concentrated reinfusion therapy for right pleural effusion was performed on POD 56. After the treatment, the pleural effusion was well-controlled with 20 mg of frosemide. This case suggested that cell-free and concentrated pleural effusion reinfusion therapy contributed to the management of refractory pleural effusion in patients with esophageal cancer accompanied by cirrhosis.

[A Resected Case of Advanced Lower Rectal Cancer with Neoadjuvant Chemotherapy by FOLFOXIRI plus Cetuximab].

Although the current standard of care for patients with lower rectal cancer in Japan includes total mesorectal resection with lateral lymph node dissection, postoperative local and distant recurrence rates are high. Multidisciplinary treatment is important to improve the prognosis. A man in his 30s was diagnosed with lower rectal cancer due to bloody stool and referred to our department. He was diagnosed as cT3N3M0, cStage Ⅲc with right obturator lymph node metastasis. Four courses of neoadjuvant chemotherapy(NAC)with FOLFOXIRI plus cetuximab were performed. Because Grade 3 neutropenia was observed in the first cycle(CTCAE v5.0), pegfilgrastim was administered in the second and subsequent cycles, and NAC was completed without dose reduction. The patient underwent laparoscopy-assisted intersphincteric rectal resection and D3+rtLD2 dissection. Histopathological resection margins were negative, and the resection was R0. Lymph node metastasis was found only in No. 263d-rt, and the pathological diagnosis was ypT3N3M0, pStage Ⅲc. Histological evaluation of response to treatment was Grade 2. The postoperative course was good and the patient was discharged on postoperative day 15. The patient received 8 courses of adjuvant chemotherapy with mFOLFOX6 from the 7th postoperative week and is alive and recurrence-free 6 months after surgery.