Intratumor transforming growth factor-ß signaling with extracellular matrix-related gene regulation marks chemotherapy-resistant gastric cancer.

Drug-tolerant persister (DTP) cells remain following chemotherapy and can cause cancer relapse. However, it is unclear when acquired resistance to chemotherapy emerges. Here, we compared the gene expression profiles of gastric cancer patient-derived cells (GC PDCs) and their respective xenograft tumors with different sensitivities to 5-fluorouracil (5-FU) by using immunodeficient female BALB/c-nu mice. RNA sequencing analysis of 5-FU-treated PDCs demonstrated that DNA replication/cell cycle-related genes were transiently induced in the earlier phase of DTP cell emergence, while extracellular matrix (ECM)-related genes were sustainably upregulated during long-term cell survival in 5-FU-resistant residual tumors. NicheNet analysis, which uncovers cell-cell signal interactions, indicated the transforming growth factor-ß (TGF-ß) pathway as the upstream regulator in response to 5-FU treatment. This induced ECM-related gene expression in the 5-FU-resistant tumor model. In the 5-FU-resistant residual tumors, there was a marked upregulation of cancer cell-derived TGF-ß1 expression and increased phosphorylation of SMAD3, a downstream regulator of the TGF-ß receptor. By contrast, these responses were not observed in a 5-FU-sensitive tumor model. We further found that TGF-ß-related upregulation of ECM genes was preferentially observed in non-responders to chemotherapy with 5-FU and/or oxaliplatin among 22 patient-derived xenograft tumors. These observations suggest that chemotherapy-induced activation of the TGF-ß1/SMAD3/ECM-related gene axis is a potential biomarker for the emergence of drug resistance in GCs.

Pharmacologic Targeting of Histone H3K27 Acetylation/BRD4-dependent Induction of ALDH1A3 for Early-phase Drug Tolerance of Gastric Cancer.

Anticancer drug-tolerant persister (DTP) cells at an early phase of chemotherapy reshape refractory tumors. Aldehyde dehydrogenase 1 family member A3 (ALDH1A3) is commonly upregulated by various anticancer drugs in gastric cancer patient-derived cells (PDC) and promotes tumor growth. However, the mechanism underlying the generation of ALDH1A3-positive DTP cells remains elusive. Here, we investigated the mechanism of ALDH1A3 expression and a combination therapy targeting gastric cancer DTP cells. We found that gastric cancer tissues treated with neoadjuvant chemotherapy showed high ALDH1A3 expression. Chromatin immunoprecipitation (ChIP)-PCR and ChIP sequencing analyses revealed that histone H3 lysine 27 acetylation was enriched in the ALDH1A3 promoter in 5-fluorouracil (5-FU)-tolerant persister PDCs. By chemical library screening, we found that the bromodomain and extraterminal (BET) inhibitors OTX015/birabresib and I-BET-762/molibresib suppressed DTP-related ALDH1A3 expression and preferentially inhibited DTP cell growth. In DTP cells, BRD4, but not BRD2/3, was recruited to the ALDH1A3 promoter and BRD4 knockdown decreased drug-induced ALDH1A3 upregulation. Combination therapy with 5-FU and OTX015 significantly suppressed in vivo tumor growth. These observations suggest that BET inhibitors are efficient DTP cell-targeting agents for gastric cancer treatment. SIGNIFICANCE: Drug resistance hampers the cure of patients with cancer. To prevent stable drug resistance, DTP cancer cells are rational therapeutic targets that emerge during the early phase of chemotherapy. This study proposes that the epigenetic regulation by BET inhibitors may be a rational therapeutic strategy to eliminate DTP cells.

APC/PIK3CA mutations and ß-catenin status predict tankyrase inhibitor sensitivity of patient-derived colorectal cancer cells.

BACKGROUND: Aberrant WNT/ß-catenin signaling drives carcinogenesis. Tankyrases poly(ADP-ribosyl)ate and destabilize AXINs, ß-catenin repressors. Tankyrase inhibitors block WNT/ß-catenin signaling and colorectal cancer (CRC) growth. We previously reported that 'short' APC mutations, lacking all seven ß-catenin-binding 20-amino acid repeats (20-AARs), are potential predictive biomarkers for CRC cell sensitivity to tankyrase inhibitors. Meanwhile, 'Long' APC mutations, which possess more than one 20-AAR, do not predict inhibitor-resistant cells. Thus, additional biomarkers are needed to precisely predict the inhibitor sensitivity. METHODS: Using 47 CRC patient-derived cells (PDCs), we examined correlations between the sensitivity to tankyrase inhibitors (G007-LK and RK-582), driver mutations, and the expressions of signaling factors. NOD.CB17-Prkdcscid/J and BALB/c-nu/nu xenograft mice were treated with RK-582. RESULTS: Short APC mutant CRC cells exhibited high/intermediate sensitivities to tankyrase inhibitors in vitro and in vivo. Active ß-catenin levels correlated with inhibitor sensitivity in both short and long APC mutant PDCs. PIK3CA mutations, but not KRAS/BRAF mutations, were more frequent in inhibitor-resistant PDCs. Some wild-type APC PDCs showed inhibitor sensitivity in a ß-catenin-independent manner. CONCLUSIONS: APC/PIK3CA mutations and ß-catenin predict the sensitivity of APC-mutated CRC PDCs to tankyrase inhibitors. These observations may help inform the strategy of patient selection in future clinical trials of tankyrase inhibitors.

[A Case of Pathological Complete Response Induced by Preoperative Gemcitabine plus S-1 with Concurrent Radiation Therapy Followed by Gemcitabine plus S-1 Therapy in Borderline Resectable Pancreatic Cancer].

The patient was a 69-year-old man. He visited our hospital with a complaint of right back pain. An abdominal CT scan confirmed a hypovascular mass 35 mm in diameter in the pancreatic head. He was diagnosed with pancreatic head cancer (cT3, cN0, cM0, cStage ⅡA, borderline resectable-A). Gemcitabine plus S-1(GS)-based chemoradiation therapy(CRT) was performed, followed by 6 courses of GS therapy. Tumor markers were almost normalized, and subtotal stomach-preserving pancreaticoduodenectomy was performed. Histopathological examination of the resected specimen revealed highly atrophic pancreatic tissue with fibrosis and no evidence of residual cancer cells (pathological complete response). The patient remains disease-free 36 months after surgery. There are few reports of pancreatic cancer with pCR after GS-based chemoradiation therapy and subsequent GS therapy. We therefore report this case together with a review of the literature.

Pupil assessment with a new handheld pupillometer in healthy subjects.

OBJECTIVE: To assess the pupil response with a new handheld pupillometer in healthy subjects. METHODS: Sixty-four eyes of 32 healthy subjects (mean age 21.2 years) were tested. After dark adaptation for 10 min, pupil responses to 1 s red and blue light stimuli at 100 cd/m2 were measured in the order from right to left eyes with a 1 min interval. The initial pupil size (D1, mm), minimum pupil size (D2, mm), and constriction rate (CR, %) were obtained. Intra-examiner reproducibility was examined using the coefficient of variation (CV, %) and the Bland-Altman plot. Inter-examiner consistency was examined using the interclass correlation coefficient (ICC) and the agreements with a conventional device, by Pearson's correlation coefficient (r). RESULTS: The CV of all parameters have high reproducibility in the red (11.0-20.7%) and blue (5.5-12.1%) light stimuli. Bland-Altman plot analysis showed no bias with both light stimuli. "Almost perfect" and "substantial" correlations between the examiners were obtained in the red (ICC = 0.78-0.94) and blue (ICC = 0.71-0.89) light stimuli. "Excellent" and "good" correlations between the devices were obtained, except for the CR parameter in the red (D1: r = 0.90; p < 0.001, D2: 0.72; p < 0.001, and CR: 0.08; p = 0.631, respectively) and blue (D1: r = 0.87; p < 0.001, D2: 0.70; p < 0.001, and CR: 0.19; p = 0.274, respectively) light stimuli. CONCLUSION: The novel pupillometer is useful for assessing pupil response. However, because of their different constructions, the CR values cannot be compared directly between the devices.

Negative Cooperativity in Guest Binding of a Ditopic Self-Folding Biscavitand.

A brand-new self-folding biscavitand was synthesized from a feet-to-feet-connected bisresorcinarene. The X-ray crystal structure of the biscaivtand showed that the two cavities are tightly connected with four butylene linkages. The conformationally coupled two cavities accommodated two cationic guests, showing a homotropic negative cooperativity in nonpolar solvents (toluene and chloroform). A polar tetrahydrofuran solvent weakened the cyclic hydrogen bonding interactions of the biscavitand, which resulted in noncooperative guest binding.

Neutralization of the induced VEGF-A potentiates the therapeutic effect of an anti-VEGFR2 antibody on gastric cancer in vivo.

The vascular endothelial growth factor (VEGF)/VEGF receptor (VEGFR) axis is an essential regulator of angiogenesis and important therapeutic target in cancer. Ramucirumab is an anti-VEGFR2 monoclonal antibody used for the treatment of several cancers. Increased circulating VEGF-A levels after ramucirumab administration are associated with a worse prognosis, suggesting that excess VEGF-A induced by ramucirumab negatively affects treatment efficacy and that neutralizing VEGF-A may improve treatment outcomes. Here, we evaluated the effect of combination treatment with an anti-VEGFR2 antibody and anti-VEGF-A antibody on gastric tumor progression and normal tissues using a preclinical BALB/c-nu/nu mouse xenograft model. After anti-VEGFR2 antibody treatment in mice, a significant increase in plasma VEGF-A levels was observed, mirroring the clinical response. The elevated VEGF-A was host-derived. Anti-VEGF-A antibody co-administration enhanced the anti-tumor effect of the anti-VEGFR2-antibody without exacerbating the toxicity. Mechanistically, the combination treatment induced intra-tumor molecular changes closely related to angiogenesis inhibition and abolished the gene expression changes specifically induced by anti-VEGFR2 antibody treatment alone. We particularly identified the dual treatment-selective downregulation of ZEB1 expression, which was critical for gastric cancer cell proliferation. These data indicate that the dual blockade of VEGF-A and VEGFR2 is a rational strategy to ensure the anti-tumor effect of angiogenesis-targeting therapy.

Serum IL-8 level as a candidate prognostic marker of response to anti-angiogenic therapy for metastatic colorectal cancer.

PURPOSE: Liver metastasis (LM) is associated with poor prognosis in patients with metastatic colorectal cancer (mCRC). Here, we investigated the prognostic utility of several serum factors in mCRC patients with or without LM who were treated with anti-angiogenic agents in first-line (FL) or salvage-line (SL) settings. METHODS: A combined cohort of 125 patients was analyzed in this single institute pooled analysis: FL cohort receiving bevacizumab (n = 71) and SL cohort receiving regorafenib (n = 54). Blood samples were obtained at baseline (BL) and during treatment, and serum factors were measured by ELISA. Overall survival (OS) was analyzed using Kaplan-Meier curves, the log-rank test, and Cox proportional hazard regression methods. RESULTS: In univariate analysis of the combined cohort, right-sided CRC, primary unresected tumor, wild-type KRAS, LM, ≥ 2 metastatic sites, and SL were associated with shorter OS; in multivariable analysis, LM and SL remained significant. Serum angiopoietin-2 (Ang-2) levels ≥ 2190.3 pg/ml and interleukin (IL)-8 levels ≥ 15.1 pg/ml at BL were significantly associated with LM. Using these cut-off values, patients with higher Ang-2 or IL-8 levels at BL had shorter OS than those with lower BL levels (Ang-2: hazard ratio [HR] 2.57, 95% confidence interval [CI] 1.47-4.51, P = 0.001; IL-8: HR 4.31, 95%CI 2.11-8.79, P < 0.001). High serum IL-8 level remained a significant predictor of shorter OS in multivariable analysis (HR 3.24, 95%CI 1.47-7.16, P = 0.004). CONCLUSION: Circulating IL-8 and Ang-2 levels are associated with LM in mCRC patients. IL-8 may be a prognostic marker of response to anti-angiogenic therapy, regardless of the treatment timing.

High-Pressure Synthesis and Crystal Structure of the Sulfur-Richest Chromium Sulfide CrS3 Composed of Cr(III) and Disulfide Ions.

The new binary chromium sulfide CrS3 has been synthesized by reaction of Cr3S4 and sulfur mixtures at 800 °C and 13 GPa. CrS3 crystallizes in an orthorhombic unit cell with a = 4.6742(7) Å, b = 5.7315(8) Å, c = 10.603(2) Å, and V = 284.873(4) Å3. It has a novel structure composed of Cr2S10 edge-shared octahedral dimers, which share all of their corners to form a three-dimensional structure. All of the sulfur atoms form S22- disulfide ions with a S-S distance of 2.063(5) or 2.068(8) Å. The structure of CrS3 is a derivative of the crystal structure of marcasite FeS2, in which one in three metal sites of the marcasite structure is vacant in the CrS3 structure.

[A Case of a Huge, Mixed-Type IPMC with Suspected Rupture of the Omental Bursa That Was Effectively Treated with Gemcitabine plus Nab-Paclitaxel].

A 53-year-old woman was referred to our hospital because of upper abdominal pain and expansion of the pancreatic main duct. Enhanced computed tomography revealed expansion of the main pancreatic duct from the head to the tail; in addition, a 30 mm cystic tumor was observed in the pancreatic head and a 56 mm tumor was observed in the ventral side of the pancreatic body. Endoscopy revealed fistula formation in the duodenum of the Vater papilla on the oral side. The patient was diagnosed with an intraductal papillary mucinous carcinoma(IPMC). In addition, PET-CT revealed accumulation of FDG in the ventral side of the pancreatic body, and a disseminated nodule in the omental bursa was suspected. We administered 6 courses of gemcitabine plus nab-paclitaxel therapy, after which, the tumor in the ventral side of the pancreatic body disappeared. We then performed sub-stomach-preserving pancreatoduodenectomy. The results of abdominal cavity washing cytology were negative, and there were no disseminated nodules in the omental bursa. Therefore, we could perform R0 excision.

Comparison of unpalatable meal contents between patients who underwent total and distal gastrectomies.

BACKGROUND: Unpalatable meal contents have several impacts on the dietary life of patients who undergo gastrectomy. However, few studies have focused on the unpalatable meal contents according to surgical procedure. This study aims to clarify the differences in the unpalatable meal contents between the patients who underwent total and distal gastrectomies (TG and DG, respectively). METHODS: This study involved patients (n = 341) who underwent TG or DG within 5 years, and a questionnaire of unpalatable meal contents was used. The data on the demographics, operation types, Dysfunction After Upper Gastrointestinal Surgery 20 (DAUGS20) scoring system, and nutrition conditions were confirmed. Furthermore, these were analyzed using descriptive statistics and compared between TG (n = 180) and DG (n = 161) groups. RESULTS: The unpalatable meal contents that were significantly different between two groups were big in size (p = 0.013), solid (p = 0.040), rough (p = 0.041), and dry (p = 0.045), which were more difficulty in the TG group. Furthermore, the strong sour taste was more difficulty in the DG group (p = 0.031). DISCUSSION: The meals which the TG group had difficulty eating were characteristic of sticking or stagnating in the digestive tract, making the passage of food more difficult in the TG patients. This was because they had a smaller storage for foods and a narrower space at the anastomotic region than those who underwent DG. CONCLUSION: The meal contents were assumed to have been influenced by the surgical procedure. We conclude that the patients in the TG group felt more difficulty in eating the meal contents that could stagnate at the anastomotic region than those in the DG group.

Columnar Organization of Carbo[5]helicene Directed by Peripheral Steric Perturbation.

Three carbo[5]helicenes ([5]CHs) containing benzylmaleimide groups displayed columnar stacked organizations, including dislocated, parallel, and alternating (P)- and (M)-helix molecular arrangements in the solid state. The peripheral benzyl groups resulted in very small steric interactions that are responsible for the formation of columnar stacked arrangements in the crystalline state.

ALDH1A3-mTOR axis as a therapeutic target for anticancer drug-tolerant persister cells in gastric cancer.

Tumors consist of heterogeneous cell populations that contain cancer cell subpopulations with anticancer drug-resistant properties called "persister" cells. While this early-phase drug tolerance is known to be related to the stem cell-like characteristic of persister cells, how the stem cell-related pathways contribute to drug resistance has remained elusive. Here, we conducted a single-cell analysis based on the stem cell lineage-related and gastric cell lineage-related gene expression in patient-derived gastric cancer cell models. The analyses revealed that 5-fluorouracil (5-FU) induces a dynamic change in the cell heterogeneity. In particular, cells highly expressing stem cell-related genes were enriched in the residual cancer cells after 5-FU treatment. Subsequent functional screening identified aldehyde dehydrogenase 1A3 (ALDH1A3) as a specific marker and potential therapeutic target of persister cells. ALDH1A3 was selectively overexpressed among the ALDH isozymes after treatment with 5-FU or SN38, a DNA topoisomerase I inhibitor. Attenuation of ALDH1A3 expression by RNA interference significantly suppressed cell proliferation, reduced the number of persister cells after anticancer drug treatment and interfered with tumor growth in a mouse xenograft model. Mechanistically, ALDH1A3 depletion affected gene expression of the mammalian target of rapamycin (mTOR) cell survival pathway, which coincided with a decrease in the activating phosphorylation of S6 kinase. Temsirolimus, an mTOR inhibitor, reduced the number of 5FU-tolerant persister cells. High ALDH1A3 expression correlated with worse prognosis of gastric cancer patients. These observations indicate that the ALDH1A3-mTOR axis could be a novel therapeutic target to eradicate drug-tolerant gastric cancer cells.

In silico chemical screening identifies epidermal growth factor receptor as a therapeutic target of drug-tolerant CD44v9-positive gastric cancer cells.

BACKGROUND: Tumours consist of heterogeneous cancer cells and are likely to contain drug-tolerant cell subpopulations, causing early relapse. However, treatment strategies to eliminate these cells have not been established. METHODS: We established gastric cancer patient-derived cells (PDCs) to examine the contribution of CD44 splicing variant 9 (CD44v9)-positive cells in gastric cancer drug tolerance. We performed gene expression signature-based in silico screening using JFCR_LinCAGE, our anticancer compound gene expression database and subsequent validation in BALB/c-nu/nu mouse xenograft to identify agents targeting the drug-tolerant cancer cells. RESULTS: CD44v9-positive cancer cells were enriched among residual cancer cells after treatment with SN-38, an active metabolic of irinotecan. CD44v9 protein was responsible for this drug resistance. We identified epidermal growth factor receptor (EGFR) inhibitors as agents that can target CD44v9-positive cell populations in gastric cancer PDCs. CD44v9 promoted cell proliferation, and EGFR inhibition attenuated CD44v9 protein expression through downregulation of the AKT and the ERK signalling pathways, leading to preferential suppression of CD44v9-positive cells. Importantly, EGFR inhibitors significantly reduced the number of residual cancer cells after cytotoxic anticancer drug treatment and enhanced the antitumor effect of irinotecan in vivo. CONCLUSIONS: EGFR inhibitors could be potential agents to eradicate cytotoxic anticancer drug-tolerant gastric cancer cell populations.

Serum VEGF-A and CCL5 levels as candidate biomarkers for efficacy and toxicity of regorafenib in patients with metastatic colorectal cancer.

Regorafenib is an oral multi-kinase inhibitor used as salvage therapy for metastatic colorectal cancer (mCRC). We tested whether serum cytokine levels are associated with clinical outcome in the mCRC patients receiving regorafenib. Serum samples were collected before treatment start, day 21, and progressive disease, and eleven angiogenic and inflammatory cytokine serum levels were examined. Fifty-four patients of a total of 62 enrolled patients were eligible for the analyses. The chemokine ligand 5 (CCL5) levels ≤ cut-off value (59959 pg/ml) at baseline was associated with relative tumor shrinkage (P = 0.021), better progression-free survival (PFS) (P = 0.036) and overall survival (OS) (P = 0.019). Vascular endothelial growth factor A (VEGF-A) levels showing a decrease on day 21 were significantly associated with a better PFS (P = 0.021). CCL5 levels ≤ cut-off was associated with any grade hand-foot skin reaction (HFSR) (P = 0.025) and thrombocytopenia (P = 0.013). Low chemokine ligand 2 levels at baseline were associated with grade 2 ≤ HFSR. High angiopoietin-2 and basic fibroblast growth factor (bFGF) levels at baseline were associated with grade 3 ≤ total bilirubin increase and transaminases increase, respectively. Low bFGF levels at baseline were associated with grade 3 ≤ hypertension. No correlation with severe events was observed. Baseline serum CCL5 levels and decrease of the serum VEGF-A levels may serve as potential predictive markers for survival or treatment-specific toxicities in mCRC patients receiving regorafenib.

[A case of triple negative recurrent breast cancer successfully treated with capecitabine+docetaxel combination chemotherapy].

A 73-year-old woman underwent pectoralis-preserving mastectomy for left breast cancer (papillotubular carcinoma, f, T2, ly0, v0, N1 (21/21), T2N1M0 (Stage IIB), ER (-), PgR (-), HER2 (-)) in August 2004. It was called a triple negative breast cancer. She received systemic chemotherapy using AC followed by paclitaxel. In February 2006 (disease- free interval of one year and five months), skin and chest wall recurrences in the left breast were revealed. Systemic chemotherapy using capecitabine (1,800 mg/body/day) monotherapy resulted in PD after 4 courses. Subsequently, treatment with capecitabine+cyclophosphamide combination therapy resulted in PD after 6 courses. Since November 2006, treatment with capecitabine+docetaxel combination chemotherapy was initiated. Each course consisted of capecitabine at a dosage of 1,800 mg/body/day for 2 weeks and docetaxel at a dosage of 60 mg/body (day 8 only) followed by withdrawal for 1 week. After 3 courses, a marked response was seen, and a total of 6 courses were performed. No serious side effect was revealed, and a marked response has been maintained. It is suspected that capecitabine+docetaxel combination therapy is useful for a triple negative recurrent breast cancer which is refractory to systemic chemotherapy.

[Results of treatment of far advanced and recurrent stomach cancer with TS-1].

We used TS-1 as first-line therapy to treat 44 patients with far advanced or recurrent gastric cancer, and assessed the results and safety. One treatment cycle consisted of TS-1, 80 mg/m2/day, for 28 days followed by a 14-day rest period. The efficacy rate in the cases capable of being evaluated was 30.1% (11/36), and 25.0%, (7/28) when TS-1 was used as monotherapy. The efficacy rate was lower than in a phase II study, however, the median survival time (MST) of 10.7 months for the patients as a whole, the 1-year survival rate of 43.2%, and the 2-year survival rate of 20.5% were favorable. There were many NC cases in which long-term therapy was possible, and they contributed to the long-term survival. The incidence of adverse events was 84.1%, but the incidence of grade 3 or more events was low at 13.6%. Since TS-1 is highly efficacious and safe, as well as convenient because of being an oral preparation, it appears that it can be ranked as the drug of first choice for chemotherapy of far advanced or recurrent gastric cancer.