Conjugation of TLR7 and TLR7/8 agonists onto weak protein antigen via versatile oxime ligation for enhanced vaccine efficacy.

Protein-based subunit vaccines are weakly immunogenic, and developing self-adjuvanting vaccines with adjuvant conjugated to antigen is a promising approach for generating optimal immune responses. Here, we report a novel adjuvant-protein conjugate vaccine based on versatile oxime ligation technique. Firstly, the adjuvant properties of a series of TLR7 and TLR7/8 small molecule agonists in self-adjuvanting vaccines were systematically compared by coupling them to proteins in consistent ratio via p-carboxybenzaldehyde (p-CBA) for the first time. All conjugate vaccines induced cytokine secretion in murine and human macrophages in vitro, and promoted specific antibody production in vivo. Notably, a conjugate containing imidazoquinoline TLR7/8 agonist (TLR7/8a1) showed the greatest enhancement in Th1/2 balanced antibody response. To minimize the interference with the protein antigenic integrity, we further developed a systematic glycoconjugation strategy to conjugate this TLR7/8a1 onto the glycan chains of SARS-CoV-2 S1 glycoprotein via oxime ligation, in which S1 containing different numbers of aldehyde groups were obtained by differential periodate oxidation. The resulting TLR7/8a1-S1 conjugate triggered a potent humoral and cellular immunity in vivo. Together these data demonstrate the promise of these TLR7 and TLR7/8 agonists as effective built-in adjuvants, and the versatile oxime ligation strategy might broaden potential applications in designing different conjugate vaccines.

Rolling Circle Amplification-Based Self-Assembly to Form a "GPS-Nanoconveyor" for In Vitro Targeted Imaging and Enhanced Gene/Chemo (CRISPR/DOX) Synergistic Therapy.

The GPS-Nanoconveyor (MA-NV@DOX-Cas13a) is a targeted nanoplatform designed for the imaging and gene/chemotherapy synergistic treatment of melanoma. It utilizes rolling circle amplification (RCA) products as a scaffold to construct a DNA "Nanoconveyor" (NV), which incorporates a multivalent aptamer (MA) as a "GPS", encapsulates doxorubicin (DOX) in the transporter, and equips it with CRISPR/Cas13a ribonucleoproteins (Cas13a RNP). Carrying MA enhances the ability to recognize the overexpressed receptor nucleolin on B16 cells, enabling targeted imaging and precise delivery of MA-NV@DOX-Cas13a through receptor-mediated endocytosis. The activation of signal transducer and activator of transcription 3 (STAT3) in cancer cells triggers cis-cleavage of CRISPR/Cas13a, initiating its trans-cleavage function. Additionally, deoxyribonuclease I (DNase I) degrades MA-NV, releasing DOX for intracellular imaging and as a chemotherapeutic agent. Experiments demonstrate the superior capabilities of this versatile nanoplatform for cellular imaging and co-treatment while highlighting the advantages of these nanodrug delivery systems in mitigating DOX side effects.

Abemaciclib plus endocrine therapy versus chemotherapy after progression on prior palbociclib in HR+/HER2- metastatic breast cancer: A single center real-world study in China.

BACKGROUND: Cyclin-dependent kinase (CDK) 4/6 inhibitor plus endocrine therapy (ET) become standard-of-care for patients with hormone receptor-positive, human epidermal growth factor receptor-2 negative (HR+/HER2-) metastatic breast cancer (MBC). However, the optimal therapeutic paradigm after progression on CDK4/6 inhibitor remains unclear. This study aimed to evaluate the efficacy and safety of abemaciclib with switching ET versus chemotherapy after progression on prior palbociclib-based ET in Chinese patients with HR+/HER2- MBC. METHODS: From 414 consecutive patients with HR+/HER2- MBC who had been treated with palbociclib plus ET from September 2018 to May 2022 in Peking University Cancer Hospital, we identified 80 patients who received abemaciclib plus switching ET or chemotherapy after progression on palbociclib, matched for age, original stage at diagnosis, disease-free interval, and tumor burden at 1:1 ratio. The primary endpoint was progression-free survival (PFS) compared using the Kaplan-Meier method. A Cox proportional hazard model was performed to identify clinical factors associated with PFS in the abemaciclib group. RESULTS: The median PFS was 6.0 months (95% confidence interval [CI]: 3.94-8.06) in abemaciclib group and 4.0 months (95% CI, 2.52-5.49) in chemotherapy group (p = 0.667). And, there was no difference in median PFS between the sequential and nonsequential arm (6.0 vs. 6.0 months) in the abemaciclib group though fewer lines of prior systemic therapy and longer PFS from prior palbociclib in the sequential arm. However, patients with prior palbociclib as the first-line therapy had a significantly longer median PFS versus prior palbociclib as ≥2nd-line therapy (11.0 vs. 5.0 months, p = 0.043). Based on multivariable analysis, ER+/PR+ was an independent factor associated with longer PFS. There was no significant difference in overall survival between the abemaciclib and chemotherapy groups (p = 0.069). CONCLUSION: Our findings indicate that abemaciclib plus switching ET might be one of feasible treatment options for Chinese patients with HR+/HER2- MBC after progression on prior palbociclib-based therapy in addition to chemotherapy.

Effectiveness of gonadotropin-releasing hormone agonists for ovarian function suppression in premenopausal patients with hormone receptor-positive breast cancer: a retrospective single-center real-world study.

PURPOSE: This study evaluated the effectiveness of ovarian function suppression (OFS) of various gonadotropin-releasing hormone agonists (GnRHa) combined with aromatase inhibitors (AI) in premenopausal patients with hormone receptor-positive (HR-positive) breast cancer. Potential risk factors associated with insufficient OFS were analyzed. PATIENTS AND METHODS: Premenopausal HR-positive breast cancer patients who had received AI with GnRHa were studied retrospectively. Patients were divided into different groups according to monthly or trimonthly GnRHa schedules they received, and the effectiveness of OFS was compared between groups. Insufficient OFS was defined as at least one instance of estradiol ≥ 30 pg/ml. Patient data was gathered from medical records for this comparison. RESULTS: Of the 264 patients enrolled in this study, 117 were administered 3.6 mg of goserelin monthly (goserelin 1 M group), 63 received 3.75 mg of leuprorelin monthly (leuprorelin 1 M group) and 84 were given 11.25 mg of leuprorelin every three months (leuprorelin 3 M group). Overall, 7.20% experienced insufficient OFS. The incidence rates in the three GnRHa depot groups were 7.69%, 6.35%, and 7.14%, respectively, without a significant statistical difference (P = 0.900). Notably, younger patients exhibited a higher likelihood of insufficient OFS [OR = 0.900, 95%CI (0.824-0.982), P = 0.018]. CONCLUSION: Insufficient OFS remains a concern during GnRHa and AI treatment. The effectiveness of the three GnRHa depots commonly used in China seems comparable. Younger patients face a heightened risk of insufficient OFS.

Multifunctional Lipidated Protein Carrier with a Built-In Adjuvant as a Universal Vaccine Platform Potently Elevates Immunogenicity of Weak Antigens.

Weak antigens represented by MUC1 are poorly immunogenic, which greatly constrains the development of relevant vaccines. Herein, we developed a multifunctional lipidated protein as a carrier, in which the TLR1/2 agonist Pam3CSK4 was conjugated to the N-terminus of MUC1-loaded carrier protein BSA through pyridoxal 5'-phosphate-mediated transamination reaction. The resulting Pam3CSK4-BSA-MUC1 conjugate was subsequently incorporated into liposomes, which biomimics the membrane structure of tumor cells. The results indicated that this lipidated protein carrier significantly enhanced antigen uptake by APCs and obviously augmented the retention of the vaccine at the injection site. Compared with the BSA-MUC1 and BSA-MUC1 + Pam3CSK4 groups, Pam3CSK4-BSA-MUC1 evoked 22- and 11-fold increases in MUC1-specific IgG titers. Importantly, Pam3CSK4-BSA-MUC1 elicited robust cellular immunity and significantly inhibited tumor growth. This is the first time that lipidated protein was constructed to enhance antigen immunogenicity, and this universal carrier platform exhibits promise for utilization in various vaccines, holding the potential for further clinical application.

Disitamab vedotin (RC48) plus toripalimab for HER2-expressing advanced gastric or gastroesophageal junction and other solid tumours: a multicentre, open label, dose escalation and expansion phase 1 trial.

Background: Although the antibody-drug conjugates (ADCs) have significantly improved the survival outcomes of patients with human epidermal receptor 2 (HER2)-expressing gastric or gastroesophageal junction (G/GEJ) cancer, the efficacy of ADC used as a single agent is limited. Therefore, it is necessary to investigate effective and safe combination regimens. Preclinical data indicated a synergetic antitumour effect of RC48 and programmed cell death protein 1 (PD-1) inhibitors. We aimed to evaluate the safety and efficacy of RC48 plus toripalimab in patients with HER2-expressing G/GEJ cancer and other solid tumours. Methods: This was a open-label, multicentre, phase 1 trial performed at three hospitals in China. Eligible patients had advanced G/GEJ cancer or other solid tumours with HER2 IHC≥1 or ISH positivity and were refractory to at least one line of treatment, or standard treatment was intolerable or unavailable for these patients. This study followed a "3 + 3" design with predefined RC48 dosages of 2.0 mg/kg and 2.5 mg/kg plus toripalimab 3 mg/kg, once every 2 weeks (q2w). The primary objectives were to evaluate the safety and determine the recommended phase II dose (RP2D), and the secondary objectives included assessing the pharmacokinetics (PK) and preliminary efficacy. This study was registered with ClinicalTrials.gov, NCT04280341. Findings: Between July 13, 2020 and August 30, 2022, 56 patients, including 30 patients with G/GEJ cancer and 26 patients with other solid tumours, were enrolled and received RC48 plus toripalimab (n = 7 for RC48 2.0 mg/kg, toripalimab 3 mg/kg, q2w; n = 49 for RC48 2.5 mg/kg, toripalimab 3 mg/kg, q2w). No dose-limiting toxic effects occurred. The RP2D was declared as RC48 2.5 mg/kg plus toripalimab 3 mg/kg, q2w. The most common grade 3 adverse events were a decreased neutrophil count (n = 13), and a decreased white blood cell count (n = 7). The efficacy assessment was completed for 52 patients. Among patients with G/GEJ cancer (n = 30), the confirmed objective response rate (ORR) was 43% (12/28, 95% CI 25, 63), median progression-free survival (PFS) was 6.2 months (95% CI 4.0, 6.9), median overall survival (OS) was 16.8 months (95% CI 7.2, NE). The ORR of patients with G/GEJ cancer receiving RP2D (n = 24) reached 50% (11/22, 95% CI 28, 72), with median PFS of 5.1 months (95% CI 1.4, 7.3) and median OS of 14.0 months (95% CI 6.3, NE). Among patients with G/GEJ cancer who received RP2D, a clinical benefit was observed in both HER2-positive and low HER2 expressing populations, with an ORR of 56% (5/9, 95% CI 21, 86) vs. 46% (6/13, 95% CI 19, 75), median PFS of 7.8 months (95% CI 0.9, NE) vs. 5.1 months (95% CI 1.2, 6.9), median OS of NE months (95% CI 4.3, NE) vs. 14.0 months (95% CI 5.1, NE), respectively. Antitumour activity was also observed for other solid tumours, including breast cancer (5/13) and endometrial carcinoma (1/1). Interpretation: Our findings suggested that RC48 plus toripalimab had a manageable safety profile and showed encouraging efficacy in pretreated patients with HER2-positive and low HER2-expressing G/GEJ cancer. The findings of our phase 1 clinical trial support further investigation of HER2-targeted ADC plus immunotherapy in HER2-expressing G/GEJ cancer and pancancer treatment in the future. Funding: Beijing Municipal Medical Research Institutes, Beijing Medical Research Institute (Z200015).

Trastuzumab Biosimilar (HLX02), Pertuzumab Plus Chemotherapy in Patients with HER2-Positive Metastatic Breast Cancer after Progression of Trastuzumab: A Prospective, Phase II Study.

PURPOSE: This study aims to evaluate the efficacy and safety of trastuzumab biosimilar (HLX02) in combination with pertuzumab and chemotherapy in patients with human epidermal growth factor receptor 2 (HER2)-positive metastatic breast cancer (MBC) after progression of trastuzumab. MATERIALS AND METHODS: In this prospective, single-arm, phase II study, patients with HER2-positive MBC after progression of trastuzumab received pertuzuamb, HLX02, and chemotherapy in Beijing Cancer Hospital from March 2020 to December 2022. The primary endpoint was progression-free survival (PFS), and secondary endpoints included objective response rate (ORR), disease control rate (DCR), overall survival (OS), and safety. The study was registered with ClinicalTrials.gov (NCT05188495). RESULTS: A total of 45 patients were included in this study. Twelve patients (26.7%) were treated in second-line and 33 patients (73.3%) were in third-line and later setting. Eighty percent and 15.5% patients had previously received pyrotinib/lapatinib and T-DM1, respectively. With a median follow-up of 24.4 months (range, 1.2 to 43.9 months), the median PFS was 7.6 months (95% confidence interval, 4.3 to 10.9), OS was not reached, the ORR was 31.1%, and DCR was 91.1%. The treatment was well tolerated. CONCLUSION: The combination of trastuzumab biosimilar HLX02, pertuzumab, and chemotherapy exhibited promising efficacy and a favorable safety profile as second- and beyond-line treatment in HER2-positive MBC.

Mucinous Cystadenocarcinoma of the Breast with Bone Metastases: First Case Report and Literature Review.

INTRODUCTION: Mucinous cystadenocarcinoma (MCA) of the breast is an extremely rare type of breast carcinoma. Since its biological characteristics, treatment options, and clinical outcomes are unclear, there is a lack of consensus regarding the optimal management of this disease. Thus, our single case report will aid our understanding of its natural history, prognostic factors, and treatment strategies. CASE PRESENTATION: We presented a 54-year-old woman with a case of advanced MCA of the breast accompanied by a huge breast mass, lymph node involvement, and distant bone metastases. We diagnosed primary breast MCA through clinical examination, imaging, and immunohistochemical assessments. Subsequently, the patient was treated with a regimen of nab-paclitaxel and bevacizumab, resulting in a significant clinical response. Progression-free survival was maintained during the 6-month follow-up period. CONCLUSION: We present the first report worldwide of a rare case of MCA of the breast with a large local mass and bone metastases. Our report adds to the limited literature on this rare breast cancer subtype and highlights the importance of accurate diagnosis and appropriate management of aggressive breast tumors.

Three-Week Versus 4-Week Schedule of nab-Paclitaxel in Patients With Metastatic Breast Cancer: A Randomized Phase II Study.

BACKGROUND: This head-to-head study compared a 3-week versus 4-week schedule of nab-paclitaxel in patients with metastatic breast cancer (mBC). METHODS: Patients with HER2-negative mBC were enrolled and randomly assigned (1:1) to receive nab-paclitaxel for a 3-week schedule (125 mg/m2 on days 1 and 8) or a 4-week schedule (same dose on days 1, 8, and 15) until disease progression or treatment intolerance. Patients with intolerable toxicities were allowed to receive a maintenance regimen after benefiting from nab-paclitaxel. The primary endpoint was progression-free survival (PFS). RESULTS: Ninety-four patients were included in the analysis (n = 47 in each arm). A longer median PFS (mPFS) was observed in the 3-week versus the 4-week schedule in the overall population (not reached vs. 6.8 months; hazard ratio [HR] = 0.44; P = .029). Patients in the 2 arms had a similar overall survival (28.0 vs. 25.8 months), objective response rate (51.1% vs. 48.9%), and disease control rate (93.6% vs. 80.9%). The 3-week schedule was associated with a lower rate of toxicity-related treatment discontinuation (8.5% vs. 29.8%) and dose delays (6.4% vs. 23.4%). CONCLUSION: This study demonstrated the better antitumor activity and safety profile of a 3-week over 4-week nab-paclitaxel schedule in HER2-negative mBC, suggesting that a 3-week schedule may be a better treatment regimen in clinical practice (ClinicalTrials.gov Identifier: NCT04192331).

A phase 1 trial of fuzuloparib in combination with apatinib for advanced ovarian and triple-negative breast cancer: efficacy, safety, pharmacokinetics and germline BRCA mutation analysis.

BACKGROUND: The effect of the combination of an anti-angiogenic agent with a poly (ADP-ribose) polymerase (PARP) inhibitor in cancer treatment is unclear. We assessed the oral combination of fuzuloparib, a PARP inhibitor, and apatinib, a VEGFR2 inhibitor for treating advanced ovarian cancer (OC) or triple-negative breast cancer (TNBC). METHODS: This dose-escalation and pharmacokinetics-expansion phase 1 trial was conducted in China. We used a standard 3 + 3 dose-escalation design, with 7 dose levels tested. Patients received fuzuloparib orally twice daily, and apatinib orally once daily. The study objectives were to determine the safety profile, recommended phase 2 dose (RP2D), pharmacokinetics, preliminary efficacy, and efficacy in relation to germline BRCA mutation (gBRCAmut). RESULTS: Fifty-two pre-treated patients were enrolled (30 OC/22 TNBC). 5 (9.6%) patients had complete response, 14 (26.9%) had partial response, and 15 (28.8%) had stable disease. Objective response rate (ORR) and disease control rate were 36.5% (95% CI 23.6-51.0) and 65.4% (95% CI 50.9-78.0), respectively. At the highest dose level of fuzuloparib 100 mg plus apatinib 500 mg, the ORR was 50.0% (4/8; 95% CI 15.7-84.3); this dose was determined to be the RP2D. Patients with gBRCAmut had higher ORR and longer median progression-free survival (PFS) than those with gBRCAwt, both in OC (ORR, 62.5% [5/8] vs 40.9% [9/22]; PFS, 9.4 vs 6.7 months) and TNBC (ORR, 66.7% [2/3] vs 15.8% [3/19]; PFS, 5.6 vs 2.8 months). Two dose-limiting toxicities occurred: grade 4 febrile neutropenia (fuzuloparib 100 mg plus apatinib 250 mg) and thrombocytopenia (fuzuloparib 100 mg plus apatinib 375 mg). Maximum tolerated dose was not reached. The most common treatment-related grade ≥ 3 toxicities in all patients were hypertension (19.2%), anaemia (13.5%), and decreased platelet count (5.8%). Exposure of apatinib increased proportionally with increasing dose ranging from 250 to 500 mg, when combined with fuzuloparib 100 mg. CONCLUSIONS: Fuzuloparib plus apatinib had acceptable safety in patients with advanced OC or TNBC. Fuzuloparib 100 mg bid plus apatinib 500 mg qd was established as the RP2D. With the promising clinical activity observed, this combination is warranted to be further explored as a potential alternative to chemotherapy. TRIAL REGISTRATION: ClinicalTrials.gov, NCT03075462 (Mar. 9, 2017).

Efficacy of apatinib 250 mg combined with chemotherapy in patients with pretreated advanced breast cancer in a real-world setting.

Objectives: This study evaluated the efficacy and safety of apatinib (an oral small-molecule tyrosine kinase inhibitor targeting VEGFR-2) 250 mg combined with chemotherapy in patients with pretreated metastatic breast cancer in a real-world setting. Patients and methods: A database of patients with advanced breast cancer who received apatinib between December 2016 and December 2019 in our institution was reviewed, and patients who received apatinib combined with chemotherapy were included. Progression-free survival (PFS), overall survival (OS), the objective response rate (ORR), the disease control rate (DCR), and treatment-related toxicity were analyzed. Results: In total, 52 evaluated patients with metastatic breast cancer previously exposed to anthracyclines or taxanes who received apatinib 250 mg combined with chemotherapy were enrolled in this study. Median PFS and OS were 4.8 (95% confidence interval [CI] = 3.2-6.4) and 15.4 months (95% CI = 9.2-21.6), respectively. The ORR and DCR were 25% and 86.5%, respectively. Median PFS for the previous line of treatment was 2.1 months (95% CI = 0.65-3.6), which was significantly shorter than that for the apatinib-chemotherapy combination (p < 0.001). No significant difference was identified in the ORR and PFS among the subgroups(subtypes, target lesion, combined regimens and treatment lines). The common toxicities related to apatinib were hypertension, hand-foot syndrome, proteinuria, and fatigue events. Conclusion: Apatinib 250 mg combined with chemotherapy provided favorable efficacy in patients with pretreated metastatic breast cancer regardless of molecular types and treatment lines. The toxicities of the regimen were well tolerated and manageable. This regimen could be a potential treatment option in patients with refractory pretreated metastatic breast cancers.

Pegylated liposomal doxorubicin (Duomeisu®) monotherapy in patients with HER2-negative metastatic breast cancer heavily pretreated with anthracycline and taxanes: a single-arm, phase II study.

PURPOSE: To evaluate the efficacy and safety of pegylated liposomal doxorubicin (PLD) in patients with human epidermal growth factor receptor 2 (HER2)-negative metastatic breast cancer (MBC) heavily pretreated with anthracycline and taxanes. METHODS: In this single-arm, phase II study, patients with HER2-negative MBC previously treated with anthracycline and taxanes as second- to fifth chemotherapy received PLD (Duomeisu®, generic doxorubicin hydrochloride liposome) 40 mg/m2 every 4 weeks until disease progression, unacceptable toxicity, or completion of six cycles. Primary endpoint was progression-free survival (PFS). Secondary endpoints included overall survival (OS), objective response rate (ORR), disease control rate (DCR), clinical benefit rate (CBR), and safety. RESULTS: Of 44 enrolled patients (median age, 53.5 years; range, 34-69), 41 and 36 were evaluable for safety and efficacy, respectively. In total, 59.1% (26/44) of patients had ≥ 3 metastatic sites, 86.4% (38/44) had visceral disease, and 63.6% (28/44) had liver metastases. Median PFS was 3.7 months (95% confidence interval [CI] 3.3-4.1) and median OS was 15.0 months (95% CI 12.1-17.9). ORR, DCR, and CBR were 16.7%, 63.9%, and 36.1%, respectively. The most common adverse events (AEs) were leukopenia (53.7%), fatigue (46.3%), and neutropenia (41.5%), with no grade 4/5 AEs. The most common grade 3 AEs were neutropenia (7.3%) and fatigue (4.9%). Patients experienced palmar-plantar-erythrodysesthesia (24.4%, 2.4% grade 3), stomatitis (19.5%, 7.3% grade 2), and alopecia (7.3%). One patient displayed a left ventricular ejection fraction decline of 11.4% from baseline after five cycles of PLD therapy. CONCLUSION: PLD (Duomeisu®) 40 mg/m2 every 4 weeks was effective and well-tolerated in patients with HER2-negative MBC heavily pretreated with anthracycline and taxanes, revealing a potentially viable treatment option for this population. Trial registration Chinese Clinical Trial Registry: ChiCTR1900022568.

A nucleolin-activated polyvalent aptamer nanoprobe for the detection of cancer cells.

Sensitive detection of cancer cells plays a critical role in early cancer diagnosis. Nucleolin, overexpressed on the surface of cancer cells, is regarded as a candidate biomarker for cancer diagnosis. Thus, cancer cells can be detected through the detection of membrane nucleolin. Herein, we designed a nucleolin-activated polyvalent aptamer nanoprobe (PAN) to detect cancer cells. In brief, a long single-stranded DNA with many repeated sequences was synthesized through rolling circle amplification (RCA). Then the RCA product acted as a scaffold chain to combine with multiple AS1411 sequences, which was doubly modified with fluorophore and quenching group, respectively. The fluorescence of PAN was initially quenched. Upon binding to target protein, the conformation of PAN changed, leading to the recovery of fluorescence. The fluorescence signal of cancer cells treated with PAN was much brighter compared with that of monovalent aptamer nanoprobes (MAN) at the same concentration. Furthermore, the binding affinity of PAN to B16 cells was proved to be 30 times higher than that of MAN by calculating the dissociation constants. The results indicated that PAN could specifically detect target cells, and this design concept has potential to become promising in cancer diagnosis.

The prognostic value of neutrophil-to-lymphocyte ratio in de novo stage IV breast cancer: a retrospective cohort study.

Background: The presence of a high neutrophil-to-lymphocyte ratio (NLR) has been associated with increased mortality in several malignancies. And the majority of studies on breast cancer (BC) analyzed patients with early-stage. Fewer studies focused on metastatic BC (MBC). De novo stage IV BC with no prior treatment is more suitable for analyzing prognostic factors. Herein, we examined the prognostic value of baseline NLR in de novo stage IV BC patients. Methods: We retrospectively screened the medical records of female patients who were diagnosed with de novo stage IV BC at Peking University Cancer Hospital between January 2011 and December 2020. All patients were followed up by telephone every 6 months. Receiver operating characteristic (ROC) curve analysis was used to determine the optimal cutoff value of NLR for progression-free survival (PFS). Peripheral blood lymphocyte subsets and tumor infiltrating lymphocytes (TILs) were analyzed by flow cytometry and immunohistochemistry, respectively. Correlations of PFS and overall survival (OS) with NLR and other clinicopathological factors were evaluated using Kaplan-Meier method and Cox regression analyses. Results: A total of 128 patients between January 2011 and December 2020 were enrolled. 70 (54.7%) cases were hormone receptor (HR)-positive/human epidermal growth factor receptor 2 (HER2)-negative, 79 (61.7%) patients had visceral metastasis and 67 (52.3%) patients had more than 2 metastatic sites. The cutoff values of NLR were 2.9, optimized by ROC curve analysis. Totals of 77 and 51 patients were assigned to the NLR-low (≤2.9) and NLR-high (>2.9) groups, respectively. Compared with NLR-high patients, the NLR-low patients had significantly longer median PFS (14.8 vs. 7.2 months; hazard ratio =1.791; P=0.003). The OS showed no significant difference (64.1 vs. 56.0 months, P=0.980). The patients with NLR-low had a higher level of peripheral CD3+ T cells (P=0.028) and a lower level of peripheral CD4+CD25+ regulatory T (Treg) cells (P=0.041). Patient samples with NLR-low also demonstrated higher levels of TILs than those with NLR-high (P=0.025). Conclusions: The baseline NLR-high is associated with adverse PFS in patients with de novo stage IV BC. The NLR-high status may indicate immune suppression status, which can help identify patients with unfavorable prognosis and assist with physicians' treatment decision.

Rolling Circle Amplification-Based DNA Nano-Assembly for Targeted Drug Delivery and Gene Therapy.

Combining the killing ability of chemotherapy drugs on tumor cells with the inhibiting ability of genetic drugs on tumor cell growth, a dual drug delivery system loaded with therapy drugs and siRNA has gradually received more and more research and extensive attention. In this paper, we designed a DNA nano-assembly based on rolling circle amplification that can co-deliver doxorubicin (Dox) and siRNA simultaneously. In order to fully exploit the potential of the dual loading system in cancer treatment, we selected STAT3 gene as a target and used siRNA to target STAT3 of mRNA and reduce the STAT3 expression in mouse melanoma cell line (B16); meanwhile, Dox as a chemotherapy drug was combined with multivalent aptamers specifically targeting B16 to achieve efficient delivery of siRNA and Dox. The results showed that the synergistic delivery system could achieve high efficiency in targeting and inhibiting proliferation in mouse melanoma cells. In addition, the synergistic effect of the dual delivery system on apoptosis of cancer cells was significantly better than that of single drug delivery systems.

Prognostic Value of Pretreatment Neutrophil-to-Lymphocyte Ratio in HER2-Positive Metastatic Breast Cancer.

This study aimed to examine the prognostic value of the neutrophil-to-lymphocyte ratio (NLR) and other clinicopathological features in HER2+ MBC patients who received first-line anti-HER2 therapy. A total of 129 patients were assigned to NLR-low and NLR-high groups based on a cutoff value of 3.0 at baseline. Peripheral blood lymphocyte subsets and gene mutations in circulating tumor DNA were analyzed by flow cytometry and Next-generation sequencing, respectively. Survival was evaluated by the Kaplan−Meier method and Cox regression analysis. Of the 129 patients, 77 and 52 were assigned to the NLR-low (≤3) and NLR-high (>3) groups, respectively. Compared with NLR-high patients, the NLR-low patients had significantly longer median progression-free survival (PFS) (11.7 vs. 7.7 months) (p = 0.001, HR = 2.703 95% CI 1.543−4.736 and overall survival (OS) (37.4 vs. 28.7 months) (p = 0.044, HR = 2.254 95% CI 1.024−4.924). Furthermore, this association was independent of metastatic sites or estrogen receptor status. Peripheral blood CD3+ (p = 0.034) and CD4+ (p = 0.010) T cell numbers were significantly higher in the NLR-low group than the NLR-high group. The mutational profile of MBC was generally similar between the two groups. Baseline NLR was a prognostic factor of PFS and OS for patients with HER2+ MBC in the first-line setting. These results may facilitate the selection of patients who will benefit most from anti-HER2 treatment.

"Willow Branch" DNA Self-Assembly for Cancer Dual-Target and Proliferation Inhibition.

DNA nanotechnology is beginning to yield unique advantages in the area of drug delivery. For the dual-targeting and proliferation suppression of cancer cells, a "willow branch" DNA assembly based on rolling circle amplification (RCA) was built. Three single-stranded DNAs, including antibody modified cDNAs, aptamer cDNAs, and simple cDNAs, were employed in the DNA self-assembly, along with the RCA scaffolds (every 63 bases is a repeat unit). "Willow branch" DNA (WB DNA) assembly successfully linked multiple antibodies and aptamers together to achieve dual targeting of cancer cells. Binding of CD44 antibodies and S2.2 aptamers to receptors on the cell membrane inhibits both pathways, ß-catenin signaling and nuclear factor-kappa B-specific transcription activity, through feedback regulation. Results demonstrated that WB DNA assembly could effectively exert multivalency clustering cell-surface receptors, modulating signal pathways and inhibiting proliferation. This study proposes a new approach for cancer dual-target and proliferation inhibition by clustering multivalent receptors.

Risk factors for cardiac rupture after acute ST-segment elevation myocardial infarction during the percutaneous coronary intervention era: a retrospective case-control study.

Background: In percutaneous coronary intervention (PCI) era, more clinically valuable risk factors are still needed to determine the occurrence of cardiac rupture (CR). Therefore, we aimed to provide evidence for the early identification of CR in ST-segment elevation myocardial infarction (STEMI). Methods: A total of 22,016 consecutive patients with STEMI admitted to Cangzhou Central Hospital and Tianjin Chest Hospital from January 2013 to July 2021 were retrospectively included, among which 195 patients with CR were included as CR group. From the rest 21,820 STEMI patients without CR, 390 patients at a ratio of 1:2 were included as the control group. A total of 66 patients accepted PCI in the CR group, and 132 patients who accepted PCI in the control group at a ratio of 1:2 were included. The status of first medical contact, laboratory examinations, and PCI characteristics were recorded. Multivariate logistic regression analysis was used to investigate the risk factors related to CR. Results: There was a higher proportion of patients with myocardial infarction (MI) in the high lateral wall in the CR group (23.6% vs. 8.2%, P<0.001). The proportion of single lesions was lower in the CR group (24.2% vs. 45.5%, P=0.004). Female (OR =2.318, 95% CI: 1.431-3.754, P=0.001), age (OR =1.066, 95% CI: 1.041-1.093, P<0.001), smoking (OR =1.750, 95% CI: 1.086-2.820, P=0.022), total chest pain time (OR =1.017, 95% CI: 1.000-1.035, P=0.049), recurrent acute chest pain (OR =2.750, 95% CI: 1.535-4.927, P=0.001), acute myocardial infarction (AMI) in the high lateral wall indicated by ECG (OR =5.527, 95% CI: 2.798-10.918, P<0.001), acute heart failure (OR =3.585, 95% CI: 2.074-6.195, P<0.001), and NT-proBNP level (OR =1.000, 95% CI: 1.000-1.000, P=0.023) were risk factors for CR in all patients. In patients who accepted PCI, single lesion (OR =0.421, 95% CI: 0.204-0.867, P=0.019), preoperative thrombolysis in myocardial infarction (TIMI) grade (OR =0.358, 95% CI: 0.169-0.760, P=0.007), and postoperative TIMI grade (OR =0.222, 95% CI: 0.090-0.546, P=0.001) were risk factors for CR. Conclusions: Non-single lesions and preoperative and postoperative TIMI grades were risk factors for CR in patients who accepted PCI. In addition to previously reported indicators, we found that AMI in the high lateral wall maybe helpful in early and accurate identification and prevention of possible CR.

Free Vascularized Fibula Salvage of Failed CPH in Pediatric Sarcoma Patients.

Background: Due to extended life expectancy and recent improvements in surgical techniques, limb salvage has replaced amputation as the gold standard and is now performed in 90-95% of upper extremity malignancies. However, many of these salvage procedures are associated with significant postsurgical complications. In particular, the clavicula pro humero (CPH) procedure is associated with high rates of nonunion. We present our experience with upper extremity salvage using the free vascularized fibular flap (VFF) after failure or nonunion of the original CPH procedure in the pediatric population. Methods: Five patients under the age of 18 diagnosed with upper extremity sarcoma who underwent tumor resection with immediate CPH reconstruction complicated with nonunion, and subsequent revision with free VFF were included. Data on patient demographics, oncologic characteristics, surgical procedures, intraoperative details, postoperative complications, and time to graft union were recorded. Results: Five patients (average age = 8.4 years; range = 5-10 years at surgery date) underwent secondary limb salvage procedure with free VFF reconstruction following failed CPH reconstruction for proximal humeral osteosarcoma (n = 4) or Ewing sarcoma (n = 1). The mean follow-up was 3.7 years. Complications occurred in five patients (100%), with three patients requiring reoperation (60%). Four patients achieved graft union (average union time = 3.7 months) and successful limb reconstruction. Four patients were alive with no local recurrence of the disease. One patient did not achieve union and was lost to follow-up. Conclusion: Primary bone tumors in the pediatric population require wide surgical resection, and reconstruction often has high complication rates that can warrant further procedures. A free VFF is a viable option for upper extremity salvage after previously failed reconstructions because it provides vascularized tissue to a scarred tissue bed and allows for the replacement or augmentation of large bony defects.

Germline Mutational Landscape in Chinese Patients With Advanced Breast Cancer.

Background: Genetic testing for breast cancer (BC) patients may shift the paradigm towards more personalized management and treatment strategies. While gene alterations may be ethnic-specific in breast cancer, our understanding of genetic epidemiology of BC remains mainly driven by data from Caucasian populations and further limited to selected handful of genes. Methods: We collected whole blood samples from 356 BC patients at metastatic first line BC and primary stage IV disease at Beijing Cancer Hospital between Jan. 2013 to Dec. 2019. A comprehensive 600-gene cancer panel was used to detect germline variants in the covered genes with a median 300x sequencing depth. Variants were classified into pathogenic, likely pathogenic, variant of uncertain significance, likely benign and benign groups according to the ACMG/AMP Standards and Guidelines. Pathogenic and likely pathogenic variants were considered as deleterious mutations. Results: The median age of 356 BC patients was 49 years (range, 21-87 years) at the first diagnosis of BC. Deleterious germline mutations across 48 cancer-related genes were identified in 21.6% (77/356) of the patients. The most prevalent mutations were BRCA1/2 mutations (7.0%), followed by ATM and RAD50 mutations (1.4% each). In addition, patients with family history were more likely to carry BRCA1 mutations (P=0.04). Moreover, patients with triple-negative breast cancer (TNBC) were more likely to harbor BRCA1 mutations than those with HR+ or HER2+ breast cancer (P=0.006). While there was no significant survival difference observed in BRCA1/2 carriers relative to non-carriers, patients with DNA damage repair (DDR) gene mutations (mostly frequently BRCA, ATM, RAD50) had worse disease-free survival (P=0.02). Conclusions: The most prevalent germline mutations in a large cohort of Chinese patients with advanced BC were BRCA1/2 mutations, followed by ATM and RAD50 mutations. In total, approximately 16.0% (57/356) of patients carry deleterious mutations in DDR pathway. Patients with breast or ovarian cancer family history were more likely to carry BRCA1/2 mutations, and ones with DDR mutations had worse survival. These findings suggest that DDR mutations are prevalent in Chinese BC patients who may potentially benefit from treatment with Poly (ADP-ribose) polymerase inhibitors.