Axillary response and outcome in breast cancer patients after neoadjuvant treatment: The role of radiotherapy in reducing recurrence in ypN0 patients with initially cN+ stage.

Objective: We aim to explore the clinicopathological features associated with axillary node response and recurrence in breast cancer patients undergoing neoadjuvant treatment (NAT). Methods: We retrospectively reviewed the medical records of 486 stage I to III breast cancer patients who received NAT and surgery between 2016 and 2021. Results: A total of 486 cases were reviewed and 154 (31.7%) patients achieved breast pathological complete response (pCR) (ypT0/Tis). Of the 366 cases with initially cN+, 177 (48.4%) cases reach ypN0. Breast pCR is in high accordance to axillary pCR (81.5%). Hormone receptor (HR)-/HER2+ breast cancer patients have the highest axillary pCR rate (78.3%). Patients achieve axillary pCR have a significantly better disease-free survival (DFS) (P=0.0004). Further analysis reveals that the DFS of ypN0 and ypN1 cases are similar (P=0.9049). Moreover, DFS in patients with ypN0 (P<0.0001) and ypN1 (P<0.0001) is significantly better than that in patients with ypN2-3. For post-mastectomy ypN0 cases, radiation could only improve DFS in patients with initially cN+ stage (P=0.0499). Multivariate Cox regression analysis shows that radiation is an independent factor to improve DFS (Hazard ratio (HR): 0.288(0.098-0.841), P=0.0230). Radiation does not improve DFS in pre-cN0/ypN0 patients (P=0.1696). Conclusion: Axillary pCR rate is higher than breast pCR rate. HR-/HER2+ patients have the highest axillary pCR rate. Axillary pCR is associated with better DFS. Radiation could further improve DFS in ypN0 patients with initially positive nodal disease.

Tacrolimus Causes Hypertension by Increasing Vascular Contractility via RhoA (Ras Homolog Family Member A)/ROCK (Rho-Associated Protein Kinase) Pathway in Mice.

BACKGROUND: To provide tacrolimus is first-line treatment after liver and kidney transplantation. However, hypertension and nephrotoxicity are common tacrolimus side effects that limit its use. Although tacrolimus-related hypertension is well known, the underlying mechanisms are not. Here, we test whether tacrolimus-induced hypertension involves the RhoA (Ras homolog family member A)/ROCK (Rho-associated protein kinase) pathway in male C57Bl/6 mice. METHODS: Intra-arterial blood pressure was measured under anesthesia. The reactivity of renal afferent arterioles and mesenteric arteries were assessed in vitro using microperfusion and wire myography, respectively. RESULTS: Tacrolimus induced a transient rise in systolic arterial pressure that was blocked by the RhoA/ROCK inhibitor Fasudil (12.0±0.9 versus 3.2±0.7; P<0.001). Moreover, tacrolimus reduced the glomerular filtration rate, which was also prevented by Fasudil (187±20 versus 281±8.5; P<0.001). Interestingly, tacrolimus enhanced the sensitivity of afferent arterioles and mesenteric arteries to Ang II (angiotensin II), likely due to increased intracellular Ca2+ mobilization and sensitization. Fasudil prevented increased Ang II-sensitivity and blocked Ca2+ mobilization and sensitization. Preincubation of mouse aortic vascular smooth muscle cells with tacrolimus activated the RhoA/ROCK/MYPT-1 (myosin phosphatase targeting subunit 1) pathway. Further, tacrolimus increased cytoplasmic reactive oxygen species generation in afferent arterioles (107±5.9 versus 163±6.4; P<0.001) and in cultured mouse aortic vascular smooth muscle cells (100±7.5 versus 160±23.2; P<0.01). Finally, the reactive oxygen species scavenger Tempol inhibited tacrolimus-induced Ang II hypersensitivity in afferent arterioles and mesenteric arteries. CONCLUSIONS: The RhoA/ROCK pathway may play an important role in tacrolimus-induced hypertension by enhancing Ang II-specific vasoconstriction, and reactive oxygen species may participate in this process by activating the RhoA/ROCK pathway.

Comparison of Immune Checkpoint Molecules PD-1 and PD-L1 in Paired Primary and Recurrent Glioma: Increasing Trend When Recurrence.

Purpose: This study aims to investigate PD-1/PD-L1 expression patterns in paired primary and recurrent gliomas. Methods: From January 2008 to December 2014, 42 patients who underwent surgical resections of primary and recurrent gliomas were retrospectively included. PD-1/PD-L1 protein expression in tumors was evaluated through immunohistochemistry. Results: In primary gliomas, PD-1 and PD-L1 expression was evident in 9 (22.0%) and 14 (33.3%) patients. In the paired recurrent glioma, PD-1 and PD-L1 expression was evident in 25 (61.0%) and 31 (74.0%) lesions. Both PD-1 and PD-L1 showed significantly enhanced expression after recurrence (p < 0.005; p < 0.005). For PD-L1 expression in recurrent gliomas, the adjuvant therapy group showed significantly increased expression compared to primary gliomas (p < 0.005). For PD-1- primary gliomas, if the matched recurrent gliomas showed PD-1+, the PFS became worse than the remaining recurrent gliomas PD-1- (12.7 vs. 25.9 months, p = 0.032). Interestingly, for PD-L1- primary gliomas, if the matched recurrent gliomas showed PD-L1+, the OS became better than the remaining recurrent gliomas PD-L1- (33.8 vs. 17.5 months, p < 0.001). Conclusions: In the study, we found the expression of PD-1/PD-L1 increased significantly in recurrent gliomas and the elevated level of PD-L1 was tightly associated with adjuvant treatment, suggesting the potential therapeutic and predictive value of PD-1 and PD-L1 in the treatment of recurrent gliomas.

Trimethylamine N-oxide promotes hyperoxaluria-induced calcium oxalate deposition and kidney injury by activating autophagy.

Calcium oxalate (CaOx) is the most common component of kidney stones. Oxidative stress, inflammation and autophagy-induced cell death are the major causes of CaOx crystal deposition and CaOx crystal deposition can further lead to kidney injury. Trimethylamine N-oxide (TMAO), a gut microbiota-derived metabolite, plays an important role in the pathogenesis of many diseases, such as atherosclerosis, diabetes and chronic kidney disease, but the effect of TMAO on hyperoxaluria-induced CaOx crystal deposition and kidney injury remains unknown. We hypothesize that TMAO aggravates CaOx crystal deposition via promoting CaOx-mediated cell death. C57Bl/6 mice were given high-oxalate diet as a model of hyperoxaluria. TMAO was provided via drinking water. Serum TMAO levels increased 15 days after CaOx treatment (6.30 ± 0.17 µmol/L vs. 34.65 ± 8.95 µmol/L). High-oxalate diet induced inflammation, CaOx deposition and kidney injury, which TMAO aggravated. In accordance, TMAO intensified high-oxalate diet induced oxidative stress, autophagy and apoptosis. Moreover, TMAO enhanced CaOx crystal adhesion to HK-2 cells and reduced cell viability (from 88.9 ± 1.6% to 75.0 ± 2.7%). Protein kinase R-like endoplasmic reticulum kinase (PERK) may mediate these TMAO effects, as TMAO promoted PERK phosphorylation. Consistently, PERK knockdown alleviated TMAO-evoked CaOx-autophagy, apoptosis and oxidative stress in HK-2 cells. In conclusion, TMAO can aggravate hyperoxaluria-induced kidney injury by triggering the PERK/ROS pathway, which enhances autophagy, apoptosis and inflammation, and facilitates CaOx crystal deposition in renal tubular cells.

Gut microbiota dependent trimethylamine N-oxide aggravates angiotensin II-induced hypertension.

Gut microbiota produce Trimethylamine N-oxide (TMAO) by metabolizing dietary phosphatidylcholine, choline, l-carnitine and betaine. TMAO is implicated in the pathogenesis of chronic kidney disease (CKD), diabetes, obesity and atherosclerosis. We test, whether TMAO augments angiotensin II (Ang II)-induced vasoconstriction and hence promotes Ang II-induced hypertension. Plasma TMAO levels were indeed elevated in hypertensive patients, thus the potential pathways by which TMAO mediates these effects were explored. Ang II (400 ng/kg-1min-1) was chronically infused for 14 days via osmotic minipumps in C57Bl/6 mice. TMAO (1%) or antibiotics were given via drinking water. Vasoconstriction of renal afferent arterioles and mesenteric arteries were assessed by microperfusion and wire myograph, respectively. In Ang II-induced hypertensive mice, TMAO elevated systolic blood pressure and caused vasoconstriction, which was alleviated by antibiotics. TMAO enhanced the Ang II-induced acute pressor responses (12.2 ± 1.9 versus 20.6 ± 1.4 mmHg; P < 0.05) and vasoconstriction (32.3 ± 2.6 versus 55.9 ± 7.0%, P < 0.001). Ang II-induced intracellular Ca2+ release in afferent arterioles (147 ± 7 versus 234 ± 26%; P < 0.001) and mouse vascular smooth muscle cells (VSMC, 123 ± 3 versus 157 ± 9%; P < 0.001) increased by TMAO treatment. Preincubation of VSMC with TMAO activated the PERK/ROS/CaMKII/PLCß3 pathway. Pharmacological inhibition of PERK, ROS, CaMKII and PLCß3 impaired the effect of TMAO on Ca2+ release. Thus, TMAO facilitates Ang II-induced vasoconstriction, thereby promoting Ang II-induced hypertension, which involves the PERK/ROS/CaMKII/PLCß3 axis.

High phosphate impairs arterial endothelial function through AMPK-related pathways in mouse resistance arteries.

AIMS: In patients with renal disease, high serum phosphate shows a relationship with cardiovascular risk. We speculate that high phosphate (HP) impairs arterial vasodilation via the endothelium and explore potential underlying mechanisms. METHODS: Isolated vessel relaxation, endothelial function, glomerular filtration rate (GFR), oxidative stress status and protein expression were assessed in HP diet mice. Mitochondrial function and protein expression were assessed in HP-treated human umbilical vein endothelial cells (HUVECs). RESULTS: High phosphate (1.3%) diet for 12 weeks impaired endothelium-dependent relaxation in mesenteric arteries, kidney interlobar arteries and afferent arterioles; reduced GFR and the blood pressure responses to acute administration of acetylcholine. The PPARα/LKB1/AMPK/eNOS pathway was attenuated in the endothelium of mesenteric arteries from HP diet mice. The observed vasodilatory impairment of mesenteric arteries was ameliorated by PPARα agonist WY-14643. The phosphate transporter PiT-1 knockdown prevented HP-mediated suppression of eNOS activity by impeding phosphorus influx in HUVECs. Endothelium cytoplasmic and mitochondrial reactive oxygen species (ROS) were increased in HP diet mice. Moreover HP decreased the expression of mitochondrial-related antioxidant genes. Finally, mitochondrial membrane potential and PGC-1α expression were reduced by HP treatment in HUVECs, which was partly restored by AMPKα agonist. CONCLUSIONS: HP impairs endothelial function by reducing NO bioavailability via decreasing eNOS activity and increasing mitochondrial ROS, in which the AMPK-related signalling pathways may play a key role.

ADAMTS13 inhibits oxidative stress and ameliorates progressive chronic kidney disease following ischaemia/reperfusion injury.

AIMS: Reduced A Disintegrin And Metalloproteinase with a ThromboSpondin type 1 motif member 13 (ADAMTS13) levels are observed in kidney disease. We test whether recombinant human ADAMTS13 (rhADAMTS13) mitigates renal injury in chronic kidney disease (CKD) and the potential mechanisms. METHODS: CKD was established 3 months after ischaemia/reperfusion (IR). ADAMTS13 and von Willebrand factor (vWF) levels, renal function and morphological changes were analysed. Afferent arteriolar responses to angiotensin II (Ang II) and acetylcholine (ACh) were measured. Oxidative stress-related molecules were detected. RESULTS: Higher vWF and lower ADAMTS13 levels were observed in CKD mice, which were markedly attenuated by rhADAMTS13. rhADAMTS13 alleviated renal dysfunction, as documented by decreased blood urea nitrogen (BUN), serum creatinine, kidney injury molecule-1 (KIM-1) and neutrophil gelatinase-associated lipocalin (NGAL) levels in CKD mice. Moreover, rhADAMTS13 attenuated transforming growth factor (TGF)-ß1/Smad3 activation. Plasma vWF: ADAMTS13 ratio showed positive correlations with malondialdehyde (MDA), hydrogen peroxide (H2 O2 ) and proteinuria, and correlated inversely with superoxide dismutase (SOD) and catalase (CAT). Finally, rhADAMTS13 inhibited reactive oxygen species (ROS) levels and improved microvascular functional disorders, accompanied by the inhibition of glycogen synthase kinase (GSK) 3ß hyperactivity and upregulation of nuclear factor erythroid 2-related factor 2 (Nrf2) expression. CONCLUSIONS: Acute kidney injury (AKI) reduces the expression of ADAMTS13 that contributes to progressive CKD, microvascular dysfunction, oxidative stress, inhibition of Nrf2 activity and renal histopathological damage. All of which can be alleviated by administration of rhADAMTS13.

Glioma SOX2 expression decreased after adjuvant therapy.

BACKGROUND: SOX2 is regarded as an important marker in stem cell. The change of SOX2 expression after adjuvant therapy in high grade glioma (HGG) remains unknown so far. Few patients with recurrent glioma have opportunity to undergo operation once again, so the recurrent glioma samples are scarce. This study tries to analyze SOX2 expression in paired primary and recurrent HGG, aims to better understand the transformation law of SOX2 after adjuvant therapy in HGG. METHODS: Twenty-four recurrent HGG patients who undergone a second resection were included. 16 patients received adjuvant therapy, the remaining 8 patients didn't receive any adjuvant therapy at all. The protein expression of SOX2 in paired primary and recurrent HGG was tested by immunohistochemistry. The statistical analysis was conducted by IBM SPSS Statistics 19.0. RESULTS: In primary HGG, SOX2 expression of 3 + , 2 + , 1+ and 0+ were seen in 20 (83.3%), 1 (4.2%), 1 (4.2%) and 2 cases (8.3%), respectively. The expression of SOX2 was decreased in recurrent HGG compared to the paired primary sample (p = 0.001). The decrease of SOX2 was often seen in patients received chemotherapy, radiotherapy or both (p = 0.003). Patients with SOX2 high expression in primary glioma had a longer median PFS than those with SOX2 low expression with marginal statistic significance (12.7 vs. 5.4 months, p = 0.083). For cases with SOX2 high expression in the primary glioma, those had SOX2 low expression after recurrence seemed to have worse prognosis as compared to patients with stable SOX2 high expression (PFS: 10.4 vs. 14.9 months, p = 0.036; OS: 27.0 vs 49.5 months, p = 0.005). CONCLUSIONS: This is the first study comparing the protein expression of SOX2 in recurrent HGG and its paired primary tumor. SOX2 high expression is common in brain HGG, a tendency of decreased SOX2 expression in recurrent gliomas was evidenced. Lower SOX2 expression was seen in those patients who received adjuvant chemotherapy and/or radiotherapy. Patients with low SOX2 expression in primary HGG usually have poorer prognosis, those with SOX2 expression decreased in recurrent HGG had worse outcome.

Stereotactic body radiotherapy based treatment for hepatocellular carcinoma with extensive portal vein tumor thrombosis.

BACKGROUND: There is currently no worldwide consensus for the management of hepatocellular carcinoma (HCC) with portal vein tumor thrombus (PVTT). We evaluated the efficacy of stereotactic body radiotherapy (SBRT) as the initial treatment for HCC with extensive PVTT based on a relatively large number of patients. METHODS: In our multidisciplinary approach for patients with hepatobiliary tumors, SBRT is recommended for unresectable HCC with PVTT or those with contraindication for transarterial chemoembolization (TACE). The aim is to shrink the tumor thrombus and preserve adequate portal venous flow, thus facilitating subsequent treatments such as TACE and tumor resection. In the present study, 70 continuous cases of HCC patients with extensive PVTT initially treated with SBRT were studied. The median follow-up period was 9.5 months (range, 1.0-21.0 months). The dynamic changes of tumor thrombosis with time after SBRT were also analyzed. RESULTS: The median survival time for the whole group was 10.0 months (95% CI, 7.7-12.3 months), with a 6- and 12-month overall survival (OS) rate of 67.3%, and 40.0% respectively. Patients who received combined SBRT and TACE showed significantly longer OS than those without indication for TACE after SBRT (12.0 ± 1.6 vs. 3.0 ± 1.0 months). Patients with good response to radiation usually had better survival. SBRT was well tolerated in our patient series. CONCLUSIONS: In conclusion, SBRT used as the initial treatment for HCC patients with extensive PVTT originally unsuitable for resection or TACE can achieve adequate thrombus shrinkage and portal vein flow restoration in the majority of cases. It could thus offer the patients an opportunity to undergo further treatment such as resection or TACE procedure. Such therapeutic strategy may result in survival advantage, especially for those who do receive combined modality with SBRT.

Dynamic MRI follow-up of radiation encephalopathy in the temporal lobe following nasopharyngeal carcinoma radiotherapy.

The natural course of radiation encephalopathy following nasopharyngeal carcinoma (NPC) radiotherapy remains poorly understood. The present study aimed to investigate the magnetic resonance imaging (MRI) characteristics and evolution of radiation encephalopathy. A series of 162 follow-up MRI examinations from 68 NPC patients with radiation encephalopathy in the temporal lobes were analyzed retrospectively. Each component of radiation encephalopathy was defined as follows: i) contrast enhanced lesions were enhanced lesions on contrast enhanced T1-weighted images (T1WI); ii) white matter lesions were lesions of homogeneous hyperintensity on T2-weighted images (T2WI) and hypointensity on T1WI; iii) cysts were round or oval well-defined lesions of hyperintensity on T2WI; iv) hemosiderin deposition were nodular or annular hypointense lesions with lower hypointense than normal white matter on both T1WI and T2WI; v) gray matter lesions were defined as disruption or erosion of hyperintensity in the cortex on T2WI. Contrast enhanced lesions, white matter lesions, gray matter lesions, cysts and hemosiderin deposition were detected in 105 (100.0%), 98 (93.3%), 94 (89.5%), 2 (1.7%) and 2 (1.7%) cases of the 105 initial diagnosed temporal lobe lesions. Contrast enhanced lesions were the most commonly observed, followed by white matter lesions, gray matter lesions, temporal lobe atrophy, cysts and hemosiderin deposition. In addition, 12 new lesions were identified during the follow-up, 4 of which presented as solid enhanced nodular lesions. Importantly, in 11 of the 117 (9.4%) affected temporal lobes, solid enhanced nodular lesions were observed to be the only initial abnormalities to occur. For those enhanced nodular lesions measuring <0.8 cm, no necrosis could be detected. On the contrary, all the contrast enhanced lesions measuring >2.0 cm exhibited a necrotic core. To the best of our knowledge, the present study revealed for the first time solid enhanced nodular lesions as the earliest MRI abnormalities of radiation encephalopathy following NPC radiotherapy.

Pancreatic cancer adjuvant radiotherapy target volume design: based on the postoperative local recurrence spatial location.

OBJECTIVES: To explore the areas at highest risk for postoperative pancreatic cancer local recurrence according to the spatial location of local failures, with the aim to provide a precise target volume for pancreatic cancer adjuvant radiotherapy. METHODS: Patients with pancreatic cancer who had undergone surgery for the primary tumor in pancreas at our institution from January 2010 to August 2015 were retrospectively analyzed. All local recurrences were plotted on the computed tomography (CT) image of a representative patient according to their relative coordinates to superior mesenteric artery (SMA) or celiac axis (CA). Adjuvant radiation clinical target volume (CTV)-90 and CTV-80 were created to cover 90 % and 80 % plotted recurrences. This planning approach was applied in four simulated cases with comparison to the plan according to RTOG 0848 contouring consensus guidelines. Raystation v4.5.1.14 was used for analyzing high throughput physics data. RESULTS: Eighty-three patients with local recurrence were included from 305 postoperative pancreatic cancer patients who did not receive adjuvant radiotherapy. Thirty-one (37 %) patients did not have adjuvant therapy at all, 52 (63 %) patients undergone adjuvant chemotherapy alone. Spatial location of local failure was created. Most recurrences occurred near CA or SMA. CTV-90 was generated through expanding the combined SMA and CA contours by 30 mm right-lateral, 21 mm left-lateral, 20 mm anterior, 13 mm posterior, 10 mm superior, and 20 mm inferior. CTV-80, smaller in volume, was also created for simultaneous integrated boost. Through comparison and analysis of the simulated cases, the radiation volumes proposed were much smaller than those with RTOG 0848 contouring consensus guidelines (average volume: PTV-80 = 120 ml, PTV-90 = 220 ml, RTOG PTV = 490 ml). Accordingly, the organs at risk received less irradiation dose with the proposed CTV-90 and CTV-80. CONCLUSIONS: Smaller adjuvant radiotherapy CTVs targeting the high-risk local failure areas of postoperative pancreatic cancer were proposed, according to the three-dimensional spatial location of local recurrences. This may help to minimize radiation-related toxicities, achieve dose escalation, and finally reduce local recurrence.

Changes in c-Kit expression levels during the course of radiation therapy for nasopharyngeal carcinoma.

In the era of intensity-modulated radiotherapy, distant metastasis is currently the main cause of treatment failure for nasopharyngeal carcinoma (NPC). Additional therapeutic strategies are required to control the metastasis and improve survival. One strategy is targeted therapy, for example against c-Kit. In the current study, the frequency of c-Kit expression was determined immunohistochemically in 106 NPC patients. c-Kit expression changes during the course of radiation therapy were detected in 41 cases via weekly biopsy. Twelve cases (11.3%) had c-Kit expression scores of 3+ and 16 (15.1%) had scores of 2+. Thus, c-Kit overexpression (2+ or 3+) was observed in 28 (26.4%) patients. There were 35 (33.0%) and 43 (40.6%) patients with c-Kit expression scores of 1+ and 0, respectively. Furthermore, a trend of decreased c-Kit expression was observed after commencing radiotherapy according to the 41 NPC patients who were biopsied weekly. Therefore, c-Kit overexpression was identified to be common in NPC, and evaluating c-Kit as a therapeutic target for metastatic NPC via c-Kit overexpression subsequent to first line treatment may be of interest. To the best of our knowledge, the present study is the first to demonstrate a trend of decreased c-Kit expression during the course of radiotherapy.