[Clinical Study of Blinatumomab in the Treatment of Adult Relapsed/Refractory Ph-Negative Acute B-Lymphoblastic Leukemia].

OBJECTIVE: To investigate the efficacy and safety of CD19/CD3 bisecific monoclonalantibody (Blinatumomab) in the treatment of adult patients with relapsed / refractory Ph-negative acute B-lymphoblastic leukemia (R/R-B-ALL). METHODS: Ten adult R/R B-ALL patients were all treated with Blinatumomab. Each treatment cycle was administered for 28 days and stopped for 14 days. The dose was 9 µg/day for the first 7 days of cycle 1, and 28 µg/day for days 8-28 if there were no adverse reactions. From the second cycle onwards, the daily dose was 28 µg. The remission, survival time (EFS and OS) and adverse reactions were observed after treatment. RESULTS: Nine patients with curative effect could be evaluated. Four patients achieved CR after one course, and one patient achieved CR after two courses, the overall remission rate was 55.6%(5/9). The median EFS was 4 months (1-12 months), and the median OS was 6 months (2-44 months). Nine of the 10 patients had fever of different degrees. Serum levels of cytokines such as IL-6, IL-10, IL-17 and IFN-γ increased. Two patients resumed medication after 1 week of treatment interruption due to neurotoxicity and CRS, respectively. One patient was discontinued due to grade 3 CRS and died of tropical candidiaemia. CONCLUSION: Blinatumomab has a good response rate in the treatment of relapsed/refractory B-ALL patients, but the duration of remission is shorter. Drug-related adverse reactions are mainly CRS and neurotoxicity. Inflammatory factors IL-6, IL-10, IL-17 and IFN-γ can be used as indicators to monitor CRS. The bisspecificity MAbs provide an opportunity for subsequent allogeneic hematopoietic stem cell transplantation in R/R-B-ALL patients.

[High-Throughput Sequencing Technology for Detection of Gene Mutations in Myeloid Malignancies and Its Clinical Prognostic Significance].

OBJECTIVE: To detect the gene mutations in patients with myeloid malignancies by high-throughput sequencing and explore the correlation between gene mutations and prognosis. METHODS: A retrospective analysis was performed on 56 patients with myeloid malignancies who were hospitalized in the department of hematology, Peking University International Hospital from January 2020 to May 2021. The genetic mutations of the patients were detected by next-generation sequencing technology, and the correlation between the genetic mutations and prognosis of myeloid malignancies was analyzed. RESULTS: In 56 patients, the number of mutated genes detected in a single patient is 0-9, with a median of 3. Sequencing results showed that the most common mutated genes were RUNX1(21.4%), TET2(17.9%), DNMT3A(17.9%), TP53(14.3%) and ASXL1(14.3%), among which the most common mutations occurred in the signaling pathway-related genes (23.3%) and the transcription factor genes (18.3%). 84% of the patients carried multiple mutated genes (≥2), and correlation analysis showed there were obvious co-occurring mutations between WT1 and FLT3, NPM1 and FLT3-ITD, and MYC and FLT3. TP53 mutation was more common in MDS patients.The overall survival time of patients with NRAS mutation was significantly shortened (P =0.049). The prognosis of patients with TP53 mutation was poor compared with those without TP53 mutation, but the difference wasn't statistically significant (P =0.08). CONCLUSION: The application of next-generation sequencing technology is of great significance in myeloid malignancies, which is helpful to better understand the pathogenesis of the disease, to judge the prognosis and to find possible therapeutic targets.

[Pedigree Analysis of ACTN1-Related Thrombocytopenia Attributed to A Novel Mutation].

AbstractObjective: To investigate the clinical phenotype and genotype of an ACTN1-associated thrombocytopenic family and explore its molecular pathogenesis. METHODS: All the family members' peripheral blood was collected for routine blood tests, blood smear, coagulation function, and platelet aggregation test. Flow cytometry was used to detect the expression of platelet CD41 and CD61. The proband and her father were tested bone marrow cytomorphology. Whole-exome sequencing techniques were performed to detect and uncover mutant loci of suspected pathogenic genes. Bioinformatics was used to assess the conserved nature of the mutated loci and to analyze the effect of the mutated genes leading to the function of the corresponding amino acid sequences. RESULTS: The platelet count of the proband was 88×109/L, and the blood smear showed dumbbell-shaped platelets, snake-shaped platelets and platelets of various sizes. Her bone marrow cytomorphology revealed normal megakaryocyte morphology with a count of 270. The platelet count of the proband's father was 74×109/L, with large platelets and platelets of various sizes observed in the blood smear, and the morphology of megakaryocytes was normal in bone marrow with a megakaryocyte count of 239. Her grandfather had a platelet count of 83×109/L, with snake-shaped platelets and platelets of various sizes on blood smears. Other family members were normal in all tests. The missense mutation c.2396G > A in exon 20 of the ACTN1 gene in the proband resulted in the mutation of 799 amino acids of the encoded protein, i.e., Arg, to His. The sequencing results of her father and grandfather at this locus were found to be consistent with her. Furthermore, bioinformatics analysis indicated that the locus was highly conserved across species and that variation in this locus might lead to functional impairment of the protein. The protein model analysis demonstrated that α-actin-1 at position 799 Arg and Glu at position 811 could form a critical salt bridge which stabilizes the conformation of the Ca2+ binding loop within the calmodulin-like motif. the mutation of R799H lost this critical salt bridge and destabilized this structural domain. CONCLUSION: In the present study, the newly uncovered missense mutation c.2396G＞A in exon 20 of the ACTN1 gene is potentially the molecular mechanism for the thrombocytopenia.

Candidates of Genomic Tests in HR+/HER2- Breast Cancer Patients With 1-2 Positive Sentinel Lymph Node Without Axillary Lymph Node Dissection: Analysis From Multicentric Cohorts.

BACKGROUND: The genomic tests such as the MammaPrint and Oncotype DX test are being gradually applied for hormone receptor positive/HER-2 negative (HR+/HER2-) breast cancer patients with up to three positive axillary lymph nodes (ALNs). The first results from RxPONDER trial suggested that Oncotype DX could be applied to patients with 1-2 positive sentinel lymph nodes (SLNs) without axillary lymph node dissection (ALND), which constituted 37.4% of the intent-to-treat population. However, there was no distinctive research on how to apply genomic tests precisely to HR+/HER2- patients with 1-2 positive SLNs without ALND. The purpose was to construct a nomogram using the multi-center retrospective data to predict precisely which HR+/HER2- candidates with 1-2 positive SLNs could be subjected to genomic tests (≤ 3 positive lymph nodes). METHODS: We conducted a retrospective analysis of 18,600 patients with stage I-III breast cancer patients treated with sentinel lymph node biopsy (SLNB) in Shandong Cancer Hospital, Fudan University Shanghai Cancer Center, and West China Hospital. The univariate and multivariate logistic regression analysis was conducted to identify the independent predictive factors of having ≤ 3 positive nodes among patients with 1-2 positive SLNs. A nomogram was developed based on variables in the final model with p<0.05. Calibration of the nomogram was carried out by internal validation using the bootstrap resampling approach and was displayed using a calibration curve. The discrimination of the model was evaluated using the ROC curve. RESULTS: Based on the database of the three institutions, a total of 18,600 breast cancer patients were identified undergoing SLNB between May 2010 and 2020. Among the 1817 HR+/HER2- patients with 1-2 positive SLNs undergoing ALND, 84.2% harbored ≤ 3 totals metastatic ALNs. The multivariate logistic regression analysis identified imaging abnormal nodes (OR=0.197, 95%CI: 0.082-0.472), the number of positive SLNs (OR=0.351, 95%CI: 0.266-0.464), the number of negative SLNs (OR=1.639, 95%CI: 1.465-1.833), pathological tumor stage (OR=0.730, 95%CI: 0.552-0.964), and lympho-vascular invasion (OR=0.287, 95%CI: 0.222-0.398) as independent predictors for the proportion of patients with ≤ 3 total metastatic ALNs (all p<0.05). These five predictors were used to create a predictive nomogram. The AUC value was 0.804 (95%CI: 0.681-0.812, p<0.001). The calibration curve showed a satisfactory fit between the predictive and actual observation based on internal validation with a bootstrap resampling frequency of 1000. CONCLUSION: The nomogram based on the multi-centric database showed a good accuracy and could assist the oncologist in determining precisely which HR+/HER2- candidates with 1-2 positive SLNs without ALND could perform genomic tests. In the era of SLNB and precision medicine, the combined application of genomic tests and SLNB could provide patients with a better strategy of dual de-escalation management, including the de-escalation of both surgery and systemic treatment.

Laboratory indicators predict axillary nodal pathologic complete response after neoadjuvant therapy in breast cancer.

The purpose was to integrate clinicopathological and laboratory indicators to predict axillary nodal pathologic complete response (apCR) after neoadjuvant therapy (NAT). The pretreatment clinicopathological and laboratory indicators of 416 clinical nodal-positive breast cancer patients who underwent surgery after NAT were analyzed from April 2015 to 2020. Predictive factors of apCR were examined by logistic analysis. A nomogram was built according to logistic analysis. Among the 416 patients, 37.3% achieved apCR. Multivariate analysis showed that age, pathological grading, molecular subtype and neutrophil-to-lymphocyte ratio were independent predictors of apCR. A nomogram was established based on these four factors. The area under the curve (AUC) was 0.758 in the training set. The validation set showed good discrimination, with AUC of 0.732. In subtype analysis, apCR was 23.8, 47.1 and 50.8% in hormone receptor-positive/HER2-, HER2+ and triple-negative subgroups, respectively. According to the results of the multivariate analysis, pathological grade and fibrinogen level were independent predictors of apCR after NAT in HER2+ patients. Except for traditional clinicopathological factors, laboratory indicators could also be identified as predictive factors of apCR after NAT. The nomogram integrating pretreatment indicators demonstrated its distinguishing capability, with a high AUC, and could help to guide individualized treatment options.

Lay abstract The purpose of this study was to integrate more pretreatment indicators, including clinicopathological factors and simple laboratory indicators, to predict axillary nodal pathologic complete response (apCR) after neoadjuvant therapy for breast cancer. The authors collected the pretreatment clinicopathological factors and laboratory indexes of 416 nodal-positive patients with breast cancer. The authors then built a nomogram to predict the therapeutic effect in axillary lymph nodes. Among 416 patients, 37.3% (155 of 416) achieved apCR. The results showed that age, pathological grading, molecular subtype and neutrophil-to-lymphocyte ratio were independent predictors of apCR. Based on these four factors, a nomogram was then built. This nomogram helped to predict apCR. In addition to traditional clinicopathological factors, laboratory indicators were also identified as predictive factors of apCR after neoadjuvant therapy. Integrating pretreatment indicators might help to predict apCR and guide individualized treatment options.

Neo-adjuvant chemotherapy and axillary de-escalation management for patients with clinically node-negative breast cancer.

This study aimed to explore the optimal time of sentinel lymph node biopsy (SLNB) and neo-adjuvant chemotherapy (NAC) and to assess the feasibility of selective elimination of axillary surgery after NAC in clinically node-negative (cN0) patients. From April 2010 to August 2018, 845 patients undergoing surgery after NAC were included in this retrospective study to analyze the correlation between different clinicopathological characteristics of cN0 patients and negative axillary lymph node after NAC (ypN0). Among the 148 cN0 patients, 83.1% (123/148) were ypN0. The rates of ypN0 in patients with hormone receptor positive (HR+)/HER2-, HR+/HER2+, HR-/HER2+, and triple-negative (TN) breast cancer were 75.4% (46/61), 82.6% (19/23), 85.2% (23/27), and 94.6% (35/37), respectively (P < 0.001). The rates of ypN0 in TN and HER2+ patients were 94.6% and 95.5%, which were significantly higher than that in HR+/HER2- patients (P < 0.05). Molecular subtypes, clinical stage, radiologic complete response, and pathologic complete response (bpCR) of the breast tumor correlated with ypN0 after full-course NAC (P < 0.05). Molecular subtypes (OR = 2.374, P = 0.033), clinical stage (OR = 0.320, P = 0.029), and bpCR (OR = 0.454, P = 0.012) were independent predictors for ypN0. The optimal time of SLNB and NAC in cN0 patients might be different among different molecular subtypes: it would be preferable to perform SLNB prior to NAC for HR+/HER2- patients, and SLNB after NAC for TN and HER2+ patients to reduce the risk of axillary lymph node dissection. In view of the high ypN0 rate in cN0 patients, axillary surgical staging might be selectively eliminated, especially for HER2+ and TN patients.

Axillary and internal mammary sentinel lymph node biopsy in breast cancer after neoadjuvant chemotherapy.

With the improvement of neoadjuvant chemotherapy (NAC), the proportion of pathological complete response (pCR) in the breast and axillary lymph node (ALN) is increasing. The evaluation of pCR does not include the status of internal mammary lymph node (IMLN). This study is to evaluate the roles of both axillary sentinel lymph node biopsy (ASLNB) and internal mammary sentinel lymph node biopsy (IM-SLNB) in breast cancer patients after NAC. There were 74 patients enrolled into this study. IM-SLNB was performed on patients with radioactive internal mammary sentinel lymph node (IM-SLN). Patients (n = 8) with cN0 and ycN0 received ASLNB, and axillary lymph node dissection (ALND) in cases of positive axillary sentinel lymph node (ASLN). Patients (n = 48) with cN+ but ycN0 received ASLNB and ALND. Patients (n = 18) with ycN+ received ALND without ASLNB. The visualization rate of IM-SLN was 56.8% (42/74). The success rate of IM-SLNB was 97.6% (41/42) and the metastasis rate of IM-SLN was 7.3% (3/41). The success rate of ASLNB was 100% (56/56). The false negative rate (FNR) of ASLNB was 17.2% (5/29). The FNR in patients with 1, 2 and ≥ 3ASLNs examined was 27.3% (3/11), 20.0% (2/10) and 0% (0/8) respectively. ASLNB could be performed on ycN0 after NAC, and ALND should be performed on initially ALN-positive patients. IM-SLNB should be considered after NAC, especially for patients with clinically positive axillary nodes before NAC, which might help make clear of the pathological nodal staging of both ALN and IMLN, improve the definition of nodal pCR, and guide the individual adjuvant regional and systemic therapy.

Validation study for the hypothesis of internal mammary sentinel lymph node lymphatic drainage in breast cancer.

According to axilla sentinel lymph node lymphatic drainage pattern, we hypothesized that internal mammary sentinel lymph node (IM-SLN) receives lymphatic drainage from not only the primary tumor area, but also the entire breast parenchyma. Based on the hypothesis a modified radiotracer injection technique was established and could increase the visualization rate of the IM-SLN significantly. To verify the hypothesis, two kinds of tracers were injected at different sites of breast. The radiotracer was injected with the modified technique, and the fluorescence tracer was injected in the peritumoral intra-parenchyma. The location of IM-SLN was identified by preoperative lymphoscintigraphy and intraoperative gamma probe. Then, internal mammary sentinel lymph node biopsy (IM-SLNB) was performed. The fluorescence status of IM-SLN was identified by the fluorescence imaging system. A total of 216 patients were enrolled from September 2013 to July 2015. The overall visualization rate of IM-SLN was 71.8% (155/216). The success rate of IM-SLNB was 97.3% (145/149). The radiotracer and the fluorescence tracer were identified in the same IM-SLN in 127 cases, the correlation and the agreement is significant (Case-base, rs=0.836, P<0.001; Kappa=0.823, P<0.001). Different tracers injected into the different sites of the intra-parenchyma reached the same IM-SLN, which demonstrates the hypothesis that IM-SLN receives the lymphatic drainage from not only the primary tumor area but also the entire breast parenchyma.

Sentinel lymph node biopsy in patients with breast ductal carcinoma in situ: Chinese experiences.

The axillary treatment of patients with ductal carcinoma in situ (DCIS) remains controversial. The aim of the present study was to evaluate the roles of sentinel lymph node biopsy (SLNB) in patients with breast DCIS. A database containing the data from 262 patients diagnosed with breast DCIS and 100 patients diagnosed with DCIS with microinvasion (DCISM) who received SLNB between January 2002 and July 2014 was retrospectively analyzed. Of the 262 patients with DCIS, 9 presented with SLN metastases (3 macrometastases and 6 micrometastases). Patients with large tumors diagnosed by ultrasound or with tumors of high histological grade had a higher positive rate of SLNs than those without (P=0.037 and P<0.0001, respectively). Of the 100 patients with DCISM, 11 presented with metastases. Younger patients had a higher positive rate of SLNs (P=0.028). According to the results of this study and the systematic review of recent studies, the indications of SLNB for patients with DCIS are as follows: SLNB should be performed in all DCISM patients and in those DCIS patients who received mastectomy, and could be avoided in those who received breast-conserving surgery. However, SLNB should be recommended to patients who have high risks of harboring invasive components. The risk factors include a large, palpable tumor, a mammographic mass or a high histological grade.

Validation study of the modified injection technique for internal mammary sentinel lymph node biopsy in breast cancer.

According to the hypothesis of internal mammary sentinel lymph node (IM-SLN) lymphatic drainage pattern, a modified radiotracer injection technique (periareolar intraparenchyma, high volume, and ultrasonographic guidance) was established. To verify the accuracy of the hypothesis and validate the modified radiotracer injection technique and to observe whether the lymphatic drainage of the whole breast parenchyma could reach to the same IM-SLN, different tracers were injected into different locations of the breast. The validation study results showed that the correlation and the agreement of the radiotracer and the fluorescence tracer are significant (case-base, r s =0.808, P<0.001; Kappa =0.79, P<0.001). It proved that the lymphatic drainage from different location of the breast (the primary tumor, the subareolar plexus) reached the same IM-SLNs and the hypothesis of IM-SLN lymphatic drainage pattern (ie, IM-SLN receives lymphatic drainage from not only the primary tumor area, but also the entire breast parenchyma). In other words, it validated the accuracy of our modified radiotracer injection technique.

Risk factors for sentinel lymph node metastasis and validation study of the MSKCC nomogram in breast cancer patients.

OBJECTIVE: To evaluate the risk factors for sentinel lymph node metastasis and validate the value of the Memorial Sloan-Kettering Cancer Center nomogram for the prediction of sentinel lymph node metastasis in breast cancer patients. METHODS: A sentinel lymph node biopsy database containing 1227 consecutive breast cancer patients (416 patients with at least one positive sentinel lymph node) was retrospectively analyzed. The predictive value of the Memorial Sloan-Kettering Cancer Center nomogram was calculated by the trend line and the area under the receiver-operator characteristic curve. Meanwhile, predictors for sentinel lymph node metastasis were also evaluated. RESULTS: Tumor size, histological grade, lymphovascular invasion, mulifocality, estrogen receptor and progesterone receptor status were significant independent predictors for sentinel lymph node metastasis (all P<0.01). The Memorial Sloan-Kettering Cancer Center nomogram presented an area under the receiver-operator characteristic curve value of 0.730. Patients with predictive value<16% had a frequency of sentinel lymph node metastasis of 0.9%. Those with values larger than 70% had a frequency of 96.2%. CONCLUSIONS: The risk factors for sentinel lymph node metastasis in our study were consistent with those in the Memorial Sloan-Kettering Cancer Center nomogram. The Memorial Sloan-Kettering Cancer Center nomogram is a useful tool that could accurately predict the probability of sentinel lymph node metastasis in our breast cancer patients. Axillary surgical staging might be avoided in patients with a predictive value of <16% and axillary lymph node dissection might be done directly in those with a predictive value >70%, while other patients should still accept sentinel lymph node biopsy.