Analyzing the risk factors for disease progression within 2 years and histological transformation in patients treated with rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone as first-line treatment: A 15-year follow-up of patients with advanced follicular lymphoma in JCOG0203.

Follicular lymphoma (FL) is an indolent lymphoma that becomes aggressive due to histological transformation (HT), leading to reduced survival. Patients with FL have different clinical courses and various treatment options. Some patients exhibit shorter survival and experience disease progression within 24 months of diagnosis/treatment (POD24); the optimal treatment remains an unmet needs. Thus, identifying factors that predict shorter survival is essential to stratify treatment and prolong the survival of patients with FL. To analyze risk factors for POD24 and HT in patients treated with rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) as first-line treatment, we performed this post-hoc analysis of patients with advanced indolent B-cell lymphoma in a randomized clinical trial wherein six cycles of R-CHOP were administered every 2-3 weeks. The primary analysis showed no differences in outcomes, which enabled the analysis of 248 patients with FL, assigned to two arms. All histopathological specimens from the 300 enrolled patients were reviewed by three expert hematopathologists. Multivariable analysis implicated Follicular Lymphoma International Prognostic Index (FLIPI) intermediate (odds ratio [OR] 2.531, 95% confidence interval [CI] 0.676-9.466) and high- (OR 2.236, 95% CI 0.160-31.226) risks, B symptoms (OR 2.091, 95% CI 0.747-5.851), and grade 3A (G3A) (OR 1.833, 95% CI 0.634-5.299) as risk factors for POD24. Furthermore, multivariable analysis through a median follow-up of 15.9 years implicated G3A (OR 2.628, 95% CI 0.806-8.575) and high-risk FLIPI (OR 4.401, 95% CI 0.186-104.377) as risk factors for HT. However, an analysis limited to the first 10 years revealed that the prognostic factors elucidated from the longer-term analysis had a greater impact on HT. G3A and high-risk FLIPI may independently predict POD24 and HT, thereby informing treatment stratification of patients with untreated advanced-stage FL in future trials, particularly to address the unmet needs of patients with POD24.

Concurrent targeting of GSK3 and MEK as a therapeutic strategy to treat pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies worldwide. However, drug discovery for PDAC treatment has proven complicated, leading to stagnant therapeutic outcomes. Here, we identify Glycogen synthase kinase 3 (GSK3) as a therapeutic target through a whole-body genetic screening utilizing a '4-hit' Drosophila model mimicking the PDAC genotype. Reducing the gene dosage of GSK3 in a whole-body manner or knocking down GSK3 specifically in transformed cells suppressed 4-hit fly lethality, similar to Mitogen-activated protein kinase kinase (MEK), the therapeutic target in PDAC we have recently reported. Consistently, a combination of the GSK3 inhibitor CHIR99021 and the MEK inhibitor trametinib suppressed the phosphorylation of Polo-like kinase 1 (PLK1) as well as the growth of orthotopic human PDAC xenografts in mice. Additionally, reducing PLK1 genetically in 4-hit flies rescued their lethality. Our results reveal a therapeutic vulnerability in PDAC that offers a treatment opportunity for patients by inhibiting multiple targets.

Impact of the quality of resected thyroid cancer tissue sample on next-generation sequencing testing.

Activating rearranged during transfection (RET) proto-oncogene alterations can be identified using next-generation sequencing (NGS) of tumor DNA/RNA. We assessed factors associated with NGS (Oncomine Dx Target Test [ODxTT]) success for resected thyroid cancer (TC) specimens, including sample age, processing conditions, and DNA/RNA quality. TC samples were from three Japanese hospitals, with sample age <1-<10 years, fixative 10%/15% neutralized buffered formalin (NBF), and fixation time ≤48 h/>48 h-≤72 h. NGS success rate was defined as the percentage of samples returning validated NGS results (RET fusion-positive/negative [RNA] or RET mutation-positive/negative [DNA], detected using ODxTT). DNA/RNA quality was assessed with indexes based on electrophoresis (DNA/RNA integrity number, DV200 ) and quantitative polymerase chain reaction (DNA/RNA integrity score [ddCq/ΔCq]). NGS success rate (N = 202) was 90%/93% (DNA/RNA) overall, 98%-100% (DNA and RNA) for samples <3 years old, and 91% (DNA and RNA) for samples ≥3-<5 years old fixed in 10% NBF for ≤48 h. Multivariate logistic regression analysis identified ddCq and ΔCq as significant predictors of DNA and RNA NGS success rates, respectively. Quality assessment of nucleic acid extracted from archival tissue samples is important for achieving high NGS success rates in clinical practice, especially for samples ≥3 years old.

Polymorphisms of the PD-L1 gene 3'-untranslated region are associated with the expression of PD-L1 in non-small cell lung cancer.

Recent results show that polymorphisms of programmed death ligand 1 (PD-L1, also known as CD274 or B7-H1) might be used as a possible marker for effectiveness of chemotherapy and cancer risk. However, the effect of PD-L1 gene variations on PD-L1 expression remain unclear. Given the post-transcriptional machinery in tumor PD-L1 expression, we investigated single nucleotide polymorphisms (SNPs) in the 3'-untranslated region (3'-UTR) of the PD-L1 gene, rs4143815 and rs4742098, using formalin-fixed paraffin-embedded sections of 154 patients with non-small cell lung cancers (NSCLCs). In rs4143815, the GG genotype showed significant association with PD-L1 expression (P = 0.032). In rs4742098, the AA genotype was significantly associated with histology and PD-L1 expression (P = 0.022 and P = 0.008, respectively). In multivariate logistic regression analysis, the AA genotype in rs4742098 was correlated with PD-L1 expression (odds ratio 0.408, P = 0.048). Interestingly, approximately 10% of the NSCLC cases showed somatic mutation when we compared genotypes of these SNPs between NSCLC tissues and non-tumor tissues from the same patients. In addition, cases with somatic mutation showed higher levels of PD-L1 expression than cases with germline mutation in rs4143815 GG. In conclusion, we demonstrated that the rs4143815 and rs4742098 SNPs in the 3'-UTR of PD-L1 were associated with tumor PD-L1 expression in NSCLCs.

Type 1 Cryoglobulinemic Vasculitis Due to Monoclonal Gammopathy of Undetermined Significance Successfully Treated by Bortezomib Plus Dexamethasone.

Cryoglobulins are immunoglobulins that precipitate in cold conditions. Type I cryoglobulinemic vasculitis is associated with hematological malignancies. We herein report a case of steroid-resistant type 1 cryoglobulinemic vasculitis associated with monoclonal gammopathy of undetermined significance (MGUS) in a 47-year-old woman. By immunofixation of cryoglobulin, we found that the main component of cryoglobulin was the M protein due to MGUS, so treatment of MGUS was needed. Bortezomib+dexamethasone therapy resulted in a rapid decrease in cryoglobulin and improvement in the symptoms of cryoglobulinemic vasculitis. In refractory type I cryoglobulinemic vasculitis, treatment of the underlying gammaglobulinopathy should be considered.

Expression Analysis of Early Metastatic Seeding of Colorectal Cancer.

BACKGROUND: Distant metastasis is the leading cause of death in patients with colorectal cancer (CRC). Tumor dissemination for metastasis formation occurs in advanced cancers and also during early stages of tumorigenesis. Here, we investigated the genes involved in early metastatic seeding of CRC using gene expression analysis. PATIENTS AND METHODS: We performed a cDNA microarray using specimens resected from stages I-II CRC with and without metachronous metastatic recurrence. For the candidate genes, we immunohistochemically validated protein expression using a tissue microarray of stages I-III CRC. RESULTS: The expression of TROP2, VWCE, and BMP7 was upregulated in the recurrence group rather than in the non-recurrence group. Protein expression analysis revealed significant association of these genes with distant metastatic recurrence. The specimens with high expression of BMP7 showed worse recurrence-free survival (RFS; p = 0.02). Those with high expression of TROP2 and VWCE showed worse overall survival (OS) and RFS (TROP2: p = 0.01 and p = 0.03; VWCE: p < 0.05 and p < 0.001, respectively). In the multivariate analysis, high expression of VWCE and BMP7 was an independent predictor of recurrence [VWCE: hazard ratio (HR) 3.41, p < 0.001; BMP7: HR 2.93, p = 0.005]. In contrast, TROP2 was an independent prognostic factor for OS (HR 4.58, p = 0.03). CONCLUSIONS: Gene expression analysis revealed that TROP2, VWCE, and BMP7 were involved in early metastatic seeding. The high expression of these genes may warrant careful surveillance or adjuvant therapy, even in stages I-II CRC cases.

Hereditary thrombocythemia due to splicing donor site mutation of THPO in a Japanese family.

Thrombopoietin (THPO) is an essential factor for platelet production. Hereditary thrombocythemia (HT) is caused by a germline mutation of THPO, MPL, or JAK2 and is inherited in an autosomal-dominant manner. We identified a Japanese family with HT due to a point mutation of the splicing donor site of the THPO gene (THPO c.13 + 1G > A). Bone marrow biopsy showed increased megakaryocytes mimicking essential thrombocythemia. One affected family member developed chronic myeloid leukemia. We cloned the mutation and developed mutated and wild type THPO expression vectors. Molecular analysis showed that the mutation causes an exon 3 skipping transcript of THPO that abrogates a suppressive untranslated upstream open reading frame. Although the transcript levels of THPO mRNA were comparable, mutated transcripts were more efficiently translated and THPO protein expression was significantly higher than that of the wild type.

R-CHOP treatment for patients with advanced follicular lymphoma: Over 15-year follow-up of JCOG0203.

Anti-CD20 antibody in combination with chemotherapy extends overall survival (OS) in untreated advanced-stage follicular lymphoma (FL), yet the optimal associated therapy is unclear. Data on the cumulative incidence of secondary malignancies postrelapse after conventional immunochemotherapy are scarce. A long-term analysis of rituximab combined with cyclophosphamide, doxorubicin, vincristine and prednisone (R-CHOP) as first-line treatment was conducted in a randomised clinical trial. A six-cycle R-CHOP regimen was administered every 2 or 3 weeks without rituximab maintenance. A prespecified evaluation was conducted 15 years after the completion of enrolment, following initial analysis results that showed no significant differences in outcomes at the 3-year mark. In-depth analyses were performed on the cohort of 248 patients with FL who were allocated to the two treatment arms. With a median follow-up period of 15.9 years, the 15-year OS was 76.2%. There were no protocol treatment-related deaths, nor were there any fatal infections attributable to subsequent lymphoma treatment. At 15 years, the cumulative incidence of non-haematological and haematological malignancies was 12.8% and 3.7% respectively. Histological transformation appeared after a median of 8 years. R-CHOP maintains safety and efficacy in patients with advanced FL over extended follow-up, making it a viable first-line option for patients with advanced-stage FL.

A case of Hodgkin lymphoma-type Richter syndrome presenting as small-intestinal perforation.

Hodgkin lymphoma type of Richter syndrome (HL-type RS) is a rare disease that arises in patients with chronic lymphocytic leukemia (CLL). HL-type RS lesions can manifest in various sites and are often accompanied by related symptoms. This is the first case report to describe diagnosis of HL-type RS after emergency surgery for gastrointestinal perforation caused by the development of a HL-type RS lesion. A 47-year-old man diagnosed with CLL three years prior began treatment with ibrutinib due to worsening anemia and splenomegaly two months prior to the emergency department presentation. Although splenomegaly improved, lymphocytopenia, anemia, and a newly arising mesenteric lymphadenopathy continued to worsen. He presented to the emergency department with abdominal pain, and subsequent surgery revealed small intestinal perforation and mesenteric lymphadenopathy with HL-type RS confirmed by histopathological examination of the resected small intestine. He subsequently received brentuximab vedotin, doxorubicin, vinblastine, and dacarbazine (A + AVD), which effectively managed the HL-type RS. If CLL clinical presentation deviates from the typical course, an early tissue biopsy should be considered to evaluate for HL-type RS. Given the adoption of the A + AVD regimen as the standard treatment for Hodgkin lymphoma, further research is needed to evaluate its efficacy in HL-type RS.

Exacerbation of Intimal Fibrosis and Endarteritis in a Kidney Transplant Recipient with Chronic Active Antibody-Mediated Rejection and COVID-19: A Case Report.

Kidney transplant recipients are immunocompromised hosts at risk for comorbidity and mortality due to infection. Currently, there are no established guidelines for the management of immunosuppressed transplant recipients with coronavirus disease 2019 (COVID-19). The impact of COVID-19 and its therapeutic management on chronic active antibody-mediated rejection (CAAMR) are still unclear. Here, we report a case of CAAMR exacerbation with endarteritis and intimal fibrosis after COVID-19. A 41-year-old female kidney transplant recipient with CAAMR was admitted to a local hospital with moderately severe COVID-19. Her doses of tacrolimus and mycophenolate mofetil were reduced, and she was administered methylprednisolone pulse and antiviral drugs. This resulted in a good clinical course and she was discharged in 15 days. During and after hospitalization, the immunosuppressants were gradually returned to the baseline levels. However, about 1.5 months after discharge, the serum creatinine level became elevated. An indication kidney biopsy showed CAAMR with intimal fibrosis and endarteritis in all interlobular arteries. An increase of immunosuppressant led to a decrease of the serum creatinine level. Factors contributing to CAAMR with intimal fibrosis and endarteritis may include (1) insufficient immunosuppression due to changes in the levels of immunosuppressive; (2) overlap with endothelial cell injury caused by COVID-19, and (3) an immune-activated state associated with COVID-19. COVID-19 is a life-threatening disease that can result in unexpected changes in immunological status. Possible allograft rejection should be carefully managed in such patients.

Drosophila Screening Identifies Dual Inhibition of MEK and AURKB as an Effective Therapy for Pancreatic Ductal Adenocarcinoma.

Significant progress has been made in understanding the pathogenesis of pancreatic ductal adenocarcinoma (PDAC) by generating and using murine models. To accelerate drug discovery by identifying novel therapeutic targets on a systemic level, here we generated a Drosophila model mimicking the genetic signature in PDAC (KRAS, TP53, CDKN2A, and SMAD4 alterations), which is associated with the worst prognosis in patients. The '4-hit' flies displayed epithelial transformation and decreased survival. Comprehensive genetic screening of their entire kinome revealed kinases including MEK and AURKB as therapeutic targets. Consistently, a combination of the MEK inhibitor trametinib and the AURKB inhibitor BI-831266 suppressed the growth of human PDAC xenografts in mice. In patients with PDAC, the activity of AURKB was associated with poor prognosis. This fly-based platform provides an efficient whole-body approach that complements current methods for identifying therapeutic targets in PDAC. SIGNIFICANCE: Development of a Drosophila model mimicking genetic alterations in human pancreatic ductal adenocarcinoma provides a tool for genetic screening that identifies MEK and AURKB inhibition as a potential treatment strategy.

Long-term follow-up after R-High CHOP/CHASER/LEED with Auto-PBSCT in untreated mantle cell lymphoma-Final analysis of JCOG0406.

Progression-free survival after R-High CHOP/CHASER/LEED with auto-PBSCT in untreated mantle cell lymphoma in JCOG0406 study. A continuous pattern of relapse was observed.

Assessment for the timing of comprehensive genomic profiling tests in patients with advanced solid cancers.

Comprehensive genomic profiling (CGP) tests have been covered by public insurance in Japan for patients with advanced solid tumors who have completed or are completing standard treatments or do not have them. Therefore, genotype-matched drug candidates are often unapproved or off-label, and improving clinical trial access is critical, involving the appropriate timing of CGP tests. To address this issue, we analyzed the previous treatment data for 441 patients from an observational study on CGP tests discussed by the expert panel at Hokkaido University Hospital between August 2019 and May 2021. The median number of previous treatment lines was two; three or more lines accounted for 49%. Information on genotype-matched therapies was provided to 277 (63%). Genotype-matched clinical trials were ineligible because of an excess number of previous treatment lines or use of specific agents were found in 66 (15%) patients, with the highest proportion in breast and prostate cancers. Many patients met the exclusion criteria of one to two or more treatment lines across cancer types. In addition, previous use of specific agents was a frequent exclusion criterion for breast, prostate, colorectal, and ovarian cancers. The patients with tumor types with a low median number (two or fewer) of previous treatment lines, including most rare cancers, primary unknown cancers, and pancreatic cancers, had significantly fewer ineligible clinical trials. The earlier timing of CGP tests may improve access to genotype-matched clinical trials, with their proportion varying by cancer type. Each relevant society needs to advocate the desirable timing of CGP testing nationwide.

Zoobiquity experiments show the importance of the local MMP9-plasminogen axis in inflammatory bowel diseases in both dogs and patients.

Using a zoobiquity concept, we directly connect animal phenotypes to a human disease mechanism: the reduction of local plasminogen levels caused by matrix metalloproteinase-9 (MMP9) activity is associated with the development of inflammation in the intestines of dogs and patients with inflammatory bowel disease. We first investigated inflammatory colorectal polyps (ICRPs), which are a canine gastrointestinal disease characterized by the presence of idiopathic chronic inflammation, in Miniature Dachshund (MD) and found 31 missense disease-associated SNPs by whole-exome sequencing. We sequenced them in 10 other dog breeds and found five, PLG, TCOF1, TG, COL9A2 and COL4A4, only in MD. We then investigated two rare and breed-specific missense SNPs (T/T SNPs), PLG: c.477G > T and c.478A>T, and found that ICRPs with the T/T SNP risk alleles showed less intact plasminogen and plasmin activity in the lesions compared to ICRPs without the risk alleles but no differences in serum. Moreover, we show that MMP9, which is an NF-κB target, caused the plasminogen reduction and that intestinal epithelial cells expressing plasminogen molecules were co-localized with epithelial cells expressing MMP9 in normal colons with the risk alleles. Importantly, MMP9 expression in patients with ulcerous colitis or Crohn's disease also co-localized with epithelial cells showing enhanced NF-κB activation and less plasminogen expression. Overall, our zoobiquity experiments showed that MMP9 induces the plasminogen reduction in the intestine, contributing to the development of local inflammation and suggesting the local MMP9-plasminogen axis is a therapeutic target in both dogs and patients. Therefore, zoobiquity-type experiments could bring new perspectives for biomarkers and therapeutic targets.

Subcutaneous Panniculitis-like T-cell Lymphoma Lacking Subcutaneous Tumor Mimicking Adult-onset Still's Disease.

We herein report a case of subcutaneous panniculitis-like T-cell lymphoma (SPTCL) resembling adult-onset Still's disease (AOSD). A 40-year-old woman presented with a fever, erythema, and painful subcutaneous nodules on the trunk. Laboratory data and a bone marrow analysis showed hemophagocytic syndrome. Although AOSD was suspected, based on a histopathological evaluation of the erythema, she was diagnosed with SPTCL. She was refractory to combination chemotherapy but achieved durable remission with cyclosporine monotherapy. Genetic testing revealed a homozygous HAVCR2 c.245A>G variant (rs184868814) that had caused NLRP3 inflammasome activation. SPTCL and AOSD share a pathogenesis in terms of NLRP3 inflammasome activation, so the clinical phenotype of SPTCL reasonably mimics AOSD.

Skin biopsies for diagnosing neuronal intranuclear inclusion disease: A retrospective study of 12 cases.

Neuronal intranuclear inclusion disease (NIID) is a progressive neurodegenerative condition. Skin biopsies taken from the lower leg were reported to be a standard diagnostic procedure for NIID; however, no studies have addressed the optimal skin biopsy locations. We retrospectively analyzed 12 cases in which skin biopsies were performed for diagnosing NIID. We collected clinical information including age, sex, skin biopsy site, the presence of nuclear inclusion bodies, the results of p62 immunostaining, the final diagnosis from the department of neurology, and the presence of abnormal GGC repeats in the NOTCH2NLC gene. Four of the 12 cases had a final diagnosis of NIID. One of the four cases was biopsied from the lower leg, whereas the other three cases were biopsied from the abdomen or thigh. Biopsy specimens of the four definite NIID cases revealed the average rates of nuclear inclusion body-positive cells in adipocytes, sweat gland cells, and fibroblasts to be 13.2%, 10.3%, and 6.3%, respectively. GGC repeat abnormalities in the NOTCH2NLC gene were observed in two of the four cases. The present study indicates that sites with ample subcutaneous fat tissue could be promising for diagnostic skin biopsies for NIID.

Evaluation of 95-Gene Classifier of Formalin-fixed Paraffin-embedded Tissues in ER-positive, HER2-negative, and Node-negative Breast Cancer.

BACKGROUND/AIM: A subset of patients with estrogen receptor (ER)-positive, HER2-negative, and node-negative breast cancer experience recurrences. Predicting patients who will have recurrences within 5 years of surgery is essential so that patients can be selected to receive adjuvant chemotherapy. The 95-gene classifier (95-GC) has been validated as a method to differentiate patients into high and low-risk groups for early recurrence. PATIENTS AND METHODS: In this study, we performed 95-GC analysis on 56 formalin-fixed paraffin-embedded (FFPE) tissue samples from patients who underwent surgery for ER-positive, HER2-negative, and node-negative breast cancer and did not receive adjuvant chemotherapy. We associated the obtained high- and low-risk groups with clinicopathological characteristics and recurrence-free survival (RFS). RESULTS: We classified 12 out of 56 patients into the high-risk recurrence group. We found significantly higher KI67 scores in patients in the high-risk group. Other clinicopathological characteristics were not associated with the 95-GC risk groups. Patients in the 95-GC low-risk group had a significantly better prognosis than those in the high-risk group (p=0.0387). The 5-year RFS rate was 97.6% in the low-risk group and 74.1% in the high-risk group, while the 10-year RFS rates were 90.1% and 74.1%, respectively. CONCLUSION: The 95-GC score can accurately predict RFS within 5 years of surgery for ER-positive, HER2-negative, and node-negative breast cancer using FFPE tissue samples. These prediction models could help assign patients to the most effective treatment regimen.

A Case of Intraocular Metastasis of Lung Cancer Diagnosed Using Cell Block Preparation of the Vitreous Humor.

BACKGROUND/AIM: Masquerade syndrome is characterized by uveitis-like manifestations caused by tumor cell infiltration into the ocular tissues. The aim of the study was to report a lung cancer patient with persistent unilateral vitreous opacity, who was eventually diagnosed with masquerade syndrome using cell block preparation. CASE REPORT: An 82-year-old female complained of blurred vision in her left eye (OS). Because of pulmonary adenocarcinoma, she had previously received anticancer drug treatment at another Hospital and achieved partial remission. Ophthalmic examinations revealed anterior chamber inflammation and vitreous opacity OS. Corticosteroid eye drops were administered, but the inflammation did not improve, and was referred to the Hokkaido University Hospital. The left best-corrected visual acuity was 0.1 with normal intraocular pressure. Anterior chamber inflammation was 2+ cells, and vitreous haze was 4+ OS. B-mode ultrasonography showed diffuse heterogeneous high echoic changes in the vitreous cavity. She underwent vitrectomy, and cell block preparation of the vitreous infusion fluids was performed. Cytopathology revealed adenocarcinoma cells with a high nuclear/cytoplasmic ratio and glandular formation. The immunocytochemical study showed that tumor cells were positive for thyroid transcription factor-1 (TTF-1), napsin A, and CK7, therefore diagnosis of masquerade syndrome due to intraocular metastasis of pulmonary adenocarcinoma was reached. Chemoradiotherapy was administered, and the eye got phthisis bulbi after irradiation 2 years after diagnosis. CONCLUSION: Cell block preparation using vitreous humor may be useful in the diagnosis and management of intraocular metastasis of pulmonary adenocarcinoma in patients with prolonged vitreous opacity.