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Abstract Background Since the Z0011 trial, the assessment of axillary lymph node status has been redirected from the previous assessment of the occurrence of lymph node metastasis alone to the assessment of the degree of lymph node loading. Our aim was to apply preoperative breast ultrasound and clinicopathological features to predict the diagnostic value of axillary lymph node load in early invasive breast cancer. Methods The 1247 lesions were divided into a high lymph node burden group and a limited lymph node burden group according to axillary lymph node status. Univariate and multifactorial analyses were used to predict the differences in clinicopathological characteristics and breast ultrasound characteristics between the two groups with high and limited lymph node burden. Pathological findings were used as the gold standard. Results Univariate analysis showed significant differences in ki-67, maximum diameter (MD), lesion distance from the nipple, lesion distance from the skin, MS, and some characteristic ultrasound features (P

Deregulation of cell cycles can result in a variety of cancers, including breast cancer (BC). In fact, abnormal regulation of cell cycle pathways is often observed in breast cancer, leading to malignant cell proliferation. CDK4/6 inhibitors (CDK4/6i) can block the G1 cell cycle through the cyclin D-cyclin dependent kinase 4/6-inhibitor of CDK4-retinoblastoma (cyclinD-CDK4/6-INK4-RB) pathway, thus blocking the proliferation of invasive cells, showing great therapeutic potential to inhibit the spread of BC. So far, three FDA-approved drugs have been shown to be effective in the management of advanced hormone receptor positive (HR+) BC: palbociclib, abemaciclib, and ribociclib. The combination strategy of CDK4/6i and endocrine therapy (ET) has become the standard therapeutic regimen and is increasingly applied to advanced BC patients. The present study aims to clarify whether CDK4/6i can also achieve a certain therapeutic effect on Human epidermal growth factor receptor 2 positive (HER2+) BC. Studies of CDK4/6i are not limited to patients with estrogen receptor positive/human epidermal growth factor receptor 2 negative (ER+/HER2-) advanced BC, but have also expanded to other types of BC. Several pre-clinical and clinical trials have demonstrated the potential of CDK4/6i in treating HER2+ BC. Therefore, this review summarizes the current knowledge and recent findings on the use of CDK4/6i in this type of BC, and provides ideas for the discovery of new treatment modalities.

Background and purpose: Colorectal cancer is one of the most common digestive system malignancies, and chemotherapy for colorectal cancer and drug resistance have always been concerns. 5-fluorouracil (5-FU) is the first-line chemotherapy drug for colorectal cancer, and its efficacy is often affected by drug resistance or adverse reactions. Dihydropyrimidine dehydrogenase (DPD) is a key rate-limiting enzyme in 5-FU metabolism, and its expression or degradation may be a factor affecting the efficacy of 5-FU. Autophagy is an important pathway of intracellular protein metabolism, and its role in chemotherapy-induced cell death or proliferation inhibition is still controversial. This study aimed to investigate the role of autophagy in colorectal cancer chemotherapy and the mechanism by which autophagy affects 5-FU drug resistance. Methods: Human colon cancer HCT-8, COLO205, LOVO and SW480 cell lines were cultured in vitro to observe the effect of DPD expression on 5-FU sensitivity. 5-FU sensitive cells were screened by drug sensitivity test. The expression of DPD, the level of autophagy and the expressions of microtubule-associated protein light chain 3 (LC3) and P62 in 5-FU-sensitive cell lines and their corresponding drug-resistant cell lines were detected. The effect of autophagy on the expression of DPD and the biological behavior and chemotherapy resistance of tumor cells were observed. UbiBrowser database screening and co-inmunoprecipitationn (Co-IP) experiment were used to verify the E3 ligase in the process of DPD degradation, and to investigate the molecular mechanism of reversing 5-FU resistance caused by autophagy degradation of DPD. Results: The COLO205 cell line had the highest DPD expression and was the most resistant to 5-FU, while the HCT-8 cell line had the lowest DPD expression and was the most sensitive to 5-FU. Compared with HCT-8/5-FU cells, HCT-8/FU-resistant cells expressed higher levels of DPD and lower levels of basal autophagy. Rapamycin (RAPA)-mediated autophagy activation enhanced the autophagy level, decreased the expression of DPD, and decreased cell proliferation, invasion and 5-FU resistance. The inhibition of autophagy mediated by 3-Methyladenine (3-MA) and hydroxychloroquine (HCQ) attenuated the autophagy level, increased the expression of DPD, and enhanced the drug resistance of 5-FU. The inhibitory effect of 5-FU combined with autophagy activator was much stronger compared with 5-FU alone or combined with autophagy inhibitor. The degradation of DPD required the participation of complete autophagic flow, among which the formation of autophagic lysosomes was one of the key steps of DPD degradation, however a single autophagosome could not degrade DPD. E3 ligase NEDD4 co-immunoprecipitated with DPD and P62 proteins in HCT-8/5-FU cells and played a role in autophagic degradation of DPD. Conclusion: The involvement of autophagy in DPD degradation affects the sensitivity of colorectal cancer cells to chemotherapy drugs. Activation of autophagy can promote the degradation of DPD and inhibit the catabolism of 5-FU, which may be a new way to reverse 5-FU resistance in colorectal cancer.

Abstract Background Papillary thyroid carcinoma (PTC) is the most common histological type of thyroid malignancy that tends to metastasize to cervical lymph nodes. In the present study, we aimed to investigate which clinicopathologic and ultrasound features of PTC are associated with clinical lymph node metastasis (LNM) and numbers of pathological LNM. Methods From January 2016 to December 2018, we identified a cohort of patients with PTC who underwent cervical ultrasonography and were diagnosed through operation and pathology. Clinical N1(cN1) and > 5 pathologic N1(pN1) were considered in the postoperative stratification to have an intermediate risk according to the 2015 ATA guidelines. Clinicopathological and ultrasound features in PTC patients were performed in accordance with the independent risk factors of cN1 and > 5pN1 respectively by using the univariate and multivariate analyses. Results We collected 748 PTC patients in the final inclusion criteria. There were 688 cN0 cases and 60 cN1 cases. From the analyses, primary tumor size > 2 cm, capsule contact, extrathyroidal extensions (ETE) and central LNM remained independent risk factors for cN1 in PTC patients. In the 748 PTC patients, 707 cases had ≤ 5 pN1, and 41 cases had > 5 pN1. Multifocality, primary tumor size > 2 cm, capsule contact and ETE are significant independent risk factors for > 5 pN1. Conclusions We concluded that multifocality, primary tumor size > 2 cm, capsule contact, ETE and central LNM were independent risk factors for the intermediate risk stratification in patients with PTC. Ultrasonography is a good technique for the preoperative lymph node staging of PTC and is helpful for detecting LNM.

Colorectal cancer is a common malignancy, and the incidence and mortality rates continue to rise. An important factor in the emergence of inflammation-induced colorectal carcinogenesis is elevated cyclooxygenase-2. Prostaglandin E2 (PGE2) over-production is frequently equated with cyclooxygenase-2 gene over-expression. PGE2 can be assessed by measuring the level of prostaglandin’s main metabolite, PGE-M, in urine. Colorectal adenoma is a precancerous lesion that can lead to colorectal cancer. We conducted research to evaluate the association between urinary levels of the PGE-M and the risk of colorectal adenomas. In a western Chinese population, we identified 152 cases of adenoma and 152 controls patients without polyps. Adenoma cases were categorized into control, low-risk and high-risk groups. There was no significant change in PGE-M levels, between the control group and the low-risk adenoma group. In the high-risk group, the PGE-M levels were 23% higher than the control group. When compared to people with the lowest urine PGE-M levels (first quartile), people with greater urinary PGE-M levels had a higher chance of developing high-risk colorectal adenomas, with an adjusted odds ratio (95% CI) of 1.65 (0.76-3.57) in the fourth quartile group, (p= 0.013). We conclude urinary PGE-M is associated with the risk of developing high-risk adenomas. Urinary PGE-M level may be used as a non-invasive indicator for estimating cancer risk.

Abstract Background Lung adenocarcinoma (LUAD) is the leading cause of cancer-related death. This study aimed to develop and validate reliable prognostic biomarkers and signature. Methods Differentially expressed genes were identified based on three Gene Expression Omnibus (GEO) datasets. Based on 1052 samples’ data from our cohort, GEO and The Cancer Genome Atlas, we explored the relationship of clinicopathological features and NEIL3 expression to determine clinical effect of NEIL3 in LUAD. Western blotting (22 pairs of tumor and normal tissues), Real-time quantitative PCR (19 pairs of tumor and normal tissues), and immunohistochemical analyses (406-tumor tissues subjected to microarray) were conducted. TIMER and ImmuCellAI analyzed relationship between NEIL3 expression and the abundance of tumor-infiltrating immune cells in LUAD. The co-expressed-gene prognostic signature was established based on the Cox regression analysis. Results This study identified 502 common differentially expressed genes and confirmed that NEIL3 was significantly overexpressed in LUAD samples (P

Abstract Background Increasing evidence has demonstrated that long noncoding RNAs (lncRNAs) have regulatory functions in hepatocellular carcinoma (HCC). The link between lincSCRG1 and HCC remains unclear. Methods To explore the lincSCRG1 regulation axis, bioinformatics, RIP and luciferase reporter assay were performed. The expressions of lincSCRG1-miR26a-SKP2 were detected in HCC tissues and cell lines through qPCR and western blot. The functions of HCC cells were investigated through in vitro assays (MTT, colony formation, transwell and flow cytometry) and the inner effect of lincSCRG1-miR26a in vivo was evaluated by xenografts and liver metatstatic nude mice models. Results LincSCRG1 was found to be strongly elevated in human HCC tissues and cell lines. MiR26a and S phase kinase-related protein 2 (SKP2) were predicted as the target miRNA for lincSCRG1 and the target gene for miR26a with direct binding sites, respectively. LincSCRG1 was verified as a competing endogenous RNA (ceRNA) via negative regulation of miR26a and derepression of SKP2 in HCC cells. Both overexpression of lincSCRG1 (ov-lincSCRG1) and inhibition of miR26a (in-miR26a) obviously stimulated cellular viability, colony formation, migration and proliferation of S phase cells and also significantly increased the protein levels of cyclinD1, CDK4, MMP2/3/9, Vimentin, and N-cadherin or inhibited the protein level of E-cadherin of HCC cells, while knockdown of lincSCRG1 (sh-lincSCRG1) and upregulation of miR26a (mi-miR26a) had the opposite effects on HCC cells. Cotransfection of in-miR26a or overexpression of SKP2 (ov-SKP2) with sh-lincSCRG1 could rescue the anticancer functions of sh-lincSCRG1, including suppressing proliferation and migration of HCC cells. Additionally, sh-lincSCRG1 could effectively inhibit the growth of subcutaneous xenograft tumours and lung metastasis, while the anticancer effect of sh-lincSCRG1 could be reversed by cotransfection of in-miR26a. Conclusions LincSCRG1 acts as a ceRNA of miR26a to restrict its ability to derepress SKP2, thereby inducing the proliferation and migration of HCC cells in vitro and in vivo. Depletion of lincSCRG1 could be used as a potential therapeutic approach in HCC.

Objective To evaluate the clinical efficacy and safety of apatinib in the treatment of advanced breast cancer(ABC) patients refractory to multiline treatments. Methods We retrospectively analyzed the clinical data of 29 ABC patients after the failure of multi-line treatments. Patients were treated with apatinib 500 mg/d orally. The clinical efficacy and adverse effects were evaluated. Symptomatic treatment and nursing care were given when the adverse effects were ≥grade Ⅲ and apatinib was suspended if not get relieved. When the adverse effects were ≤gradeⅠ, apatinib was administrated with reduced dosage of 250mg/d. Results Among 29 patients, there was no case with complete remission, 11(37.9%) patients with partial remission (PR), 13(44.8%) cases with stable disease (SD) and 5(17.2%) patients with progressive disease (PD). The disease control rate (PR+SD) was 82.8% (24/29). The median progression-free survival time (mPFS) was 126 days. The most common adverse effects were secondary hypertension(27.59%), hand-foot syndrome(20.69%), Secondary proteinuria(17.24%), nausea and fatigue(13.79%), Secondary oral mucositis(17.24%) and diarrhea(10.24%). Most of the side effects were degreeⅠ-Ⅱ. Log-rank univariate analysis showed a mPFS of 267 days for Luminal B(HER2-) ABC patients and 126 days for triple-negative group, compared with 33 days for HER2+ ABC patients (P=0.057). Patients with secondary hypertension or transient proteinuria had significantly longer mPFS than those without these adverse effects (P=0.025, P=0.058). Cox regression analysis identified molecular typing, secondary hypertension and proteinuria as the independent influence factors for mPFS of ABC patients treated with apatinib. Conclusion Apatinib has relatively high DCR and PFS in the treatment of Luminal B (HER2-) and triple-negative MBC patients, and adverse reactions could be controlled.

BackgroundImatinib (IM), a tyrosine kinase inhibitor (TKI), has markedly improved the survival and life quality of chronic myeloid leukemia (CML) patients. However, the lack of specific biomarkers for IM resistance remains a serious clinical challenge. Recently, growing evidence has suggested that exosome-harbored proteins were involved in tumor drug resistance and could be novel biomarkers for the diagnosis and drug sensitivity prediction of cancer. Therefore, we aimed to investigate the proteomic profile of plasma exosomes derived from CML patients to identify ideal biomarkers for IM resistance.MethodsWe extracted exosomes from pooled plasma samples of 9 imatinib-resistant CML patients and 9 imatinib-sensitive CML patients by ultracentrifugation. Then, we identified the expression levels of exosomal proteins by liquid chromatography-tandem mass spectrometry (LC-MS/MS) based label free quantification. Bioinformatics analyses were used to analyze the proteomic data. Finally, the western blot (WB) and parallel reaction monitoring (PRM) analyses were applied to validate the candidate proteins.ResultsA total of 2812 proteins were identified in plasma exosomes from imatinib-resistant and imatinib-sensitive CML patients, including 279 differentially expressed proteins (DEPs) with restricted criteria (fold change≥1.5 or ≤0.667, p

BackgroundThe previous investigations have identified that long non-coding RNA (lncRNAs) act as crucial regulators in gastric carcinoma. However, the function of lncRNA MIR4435-2HG in the modulation of gastric carcinoma remains elusive. Here, we aimed to explore the role of MIR4435-2HG in gastric carcinoma.MethodThe Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) were applied to select the differently expressed lncRNAs in gastric carcinoma. The qRT-PCR was applied to analyze MIR4435-2HG expression in carcinoma tissues and cell lines. The effect of MIR4435-2HG on proliferation, invasion, migration, and apoptosis of gastric carcinoma cells was detected by Cell Counting Kit-8 (CCK-8) assays, transwell assays, and flow cytometry in vitro. A subcutaneous tumor model was constructed to examine the tumor growth of gastric carcinoma cells after knocking out MIR4435-2HG. RNA immunoprecipitation and luciferase reporting assays were applied to evaluate the interaction of MIR4435-2HG, miR-138-5p, and Sox4.ResultsThe bioinformatics analysis based on TCGA and GEO databases indicated that MIR4435-2HG was obviously elevated in gastric carcinoma samples. The qRT-PCR analysis revealed that MIR4435-2HG was upregulated in clinical gastric carcinoma tissues and cells. The high expression of MIR4435-2HG is associated with the poor survival rate of patients. The knockout of MIR4435-2HG could repress the proliferation, invasion, migration, and epithelial–mesenchymal transition (EMT) and accelerate the apoptosis of gastric carcinoma cells. Moreover, the deletion of MIR4435-2HG was able to attenuate the tumor growth in vivo. Mechanically, we identified that MIR4435-2HG enhanced Sox4 expression by directly interacting with miR-138-5p as a competitive endogenous RNA (ceRNA) in gastric carcinoma cells, in which Sox4 was targeted by miR-138-5p.ConclusionMIR4435-2HG is elevated in gastric carcinoma cells and contributes to the growth, metastasis, and EMT of gastric carcinoma cells by targeting miR-138-5p/Sox4 axis. MIR4435-2HG may be applied as a potential therapeutic target in gastric carcinoma.

BackgroundChronic myeloid leukemia (CML) is an acquired hematopoietic stem malignant disease originating from the myeloid system. Long non-coding RNAs (lncRNAs) have been widely explored in cancer tumorigenesis. However, their roles in CML remain largely unclear.MethodsThe peripheral blood mononuclear cells (PBMCs) and CML cell lines (K562, KCL22, MEG01, BV173) were collected for in vitro research. Real-time quantitative polymerase chain reaction was used to determine the mRNA expression levels. Cell viability and apoptosis were analyzed by cell counting kit 8 and flow cytometry assays. The targeting relationships were predicted using Starbase and TargetScan and ulteriorly verified by RNA pull-down and luciferase reporter assays. Western blotting assay was performed to assess the protein expressions. N6-methyladenosine (m6A) modification sites were predicted by SRAMP and confirmed by Methylated RNA immunoprecipitation (MeRIP) assay.ResultsLncRNA nuclear-enriched abundant transcript 1 (NEAT1) expression levels were decreased in the CML cell lines and PBMCs of CML patients. Moreover, METTL3-mediated m6A modification induced the aberrant expression of NEAT1 in CML. Overexpression of NEAT1 inhibited cell viability and promoted the apoptosis of CML cells. Additionally, miR-766-5p was upregulated in CML PBMCs and abrogated the effects of NEAT1 on cell viability and apoptosis of the CML cells. Further, CDKN1A was proved to be the target gene of miR-766-5p and was downregulated in the CML PBMCs. Knockdown of CDKN1A reversed the effects of NEAT1.ConclusionThe current research elucidates a novel METTL3/NEAT1/miR-766-5p/CDKN1A axis which plays a critical role in the progression of CML.

Abstract Background A key factor underlying the control of the cellular growth, size and proliferation involves the regulation of the total protein synthesis. Most often, the initial stages of mRNA translation are rate limiting, which involves a group of eukaryotic translation initiation factors (EIFs). Research advances focused on the inhibition of their expression and activity hold the key to the initiation and progression of tumor and tumor prognosis. Method We performed RNA interference (RNAi) with the lentivirus vector system to silence the EIF3B gene using the colon cancer cell strain SW1116. The negative control included the normal target cells infected with the negative control virus whereas the knockdown cells included the normal target cells transfected with the RNAi target virus. We tested the inhibition resulting from the decreased expression of EIF3B gene on the proliferation rate of SW1116 cells, including the cell cycle, apoptosis and clonability. Results Compared with the negative control, the impact of EIF3B gene expression in SW1116 cells on the levels of mRNA and protein in the knockdown group, was significantly inhibited (P P P P P P Conclusions The silencing of EIF3B gene expression inhibits the proliferation of colon cancer cells.

Abstract Background X-ray repair cross-complementing group 1 (XRCC1) protein plays an important role in the repair of DNA damage and adducts. Single nucleotide polymorphisms (SNPs) of XRCC1 are suspected to have some relationship with response to chemotherapy and overall survival of lung cancer. This meta-analysis aimed to summarize published data on the association between the commonest SNPs of XRCC1 (Arg194Trp, C > T, rs1799782 and Arg399Gln, G > A, rs25487) and clinical outcome of lung cancer patients. Methods We retrieved the relevant articles from PubMed, EMBASE and the China National Knowledge Infrastructure (CNKI) databases. Studies were selected using specific inclusion and exclusion criteria. Primary outcomes included objective response (i.e., complete response + partial response vs. progressive disease + stable disease) and overall survival (OS). Odds ratio (OR) or hazard ratio (HR) with 95% confidence interval (CI) were estimated. All analyses were performed using the Stata software. Results Twenty-two articles were included in the present analysis. XRCC1 Arg194Trp and Arg399Gln polymorphisms were significantly associated with response to treatment in lung cancer patients. Patients with C/T genotype, T/T genotype and minor variant T allele at Arg194Trp were more likely to respond to platinum-based chemotherapy compared with those with C/C genotype (C/T vs. C/C: OR, 2.54; 95%CI, 1.95-3.31; T/T vs. C/C: OR, 2.06; 95%CI, 1.39-3.06; C/T+T/T vs. C/C: OR, 2.42; 95% CI, 1.88-3.10). For XRCC1 Arg399Gln, G/A genotype, A/A genotype and minor variant A allele were associated with objective response in all patients (G/A vs. G/G: OR, 0.67; 95%CI, 0.50-0.90; A/A vs. G/G: OR, 0.43; 95%CI, 0.25-0.73; A/A+G/A vs. G/G: OR, 0.63; 95%CI, 0.49-0.83). Both G/A and A/A genotypes of XRCC1 Arg399Gln could influence overall survival of lung cancer patients (G/A vs. G/G: HR, 1.23; 95%CI, 1.06-1.44; A/A vs. G/G: HR, 2.03; 95%CI, 1.20-3.45). Interaction analysis suggested that compared with the patients carrying C/T+T/T genotype at XRCC1 194 and G/G genotype at XRCC1 399, the patients carrying 194 C/C and 399 G/A+A/A or 194 C/C and 399 G/G genotype showed much worse objective response. Conclusions Genetic polymorphisms in XRCC1 gene might be associated with overall survival and response to platinum-based chemotherapy in lung cancer patients.

Abstract Background In this study, we pilot tested an in vitro assay of cancer killing activity (CKA) in circulating leukocytes of 22 cancer cases and 25 healthy controls. Methods Using a human cervical cancer cell line, HeLa, as target cells, we compared the CKA in circulating leukocytes, as effector cells, of cancer cases and controls. The CKA was normalized as percentages of total target cells during selected periods of incubation time and at selected effector/target cell ratios in comparison to no-effector-cell controls. Results Our results showed that CKA similar to that of our previous study of SR/CR mice was present in human circulating leukocytes but at profoundly different levels in individuals. Overall, males have a significantly higher CKA than females. The CKA levels in cancer cases were lower than that in healthy controls (mean ± SD: 36.97 ± 21.39 vs. 46.28 ± 27.22). Below-median CKA was significantly associated with case status (odds ratio = 4.36; 95% Confidence Interval = 1.06, 17.88) after adjustment of gender and race. Conclusions In freshly isolated human leukocytes, we were able to detect an apparent CKA in a similar manner to that of cancer-resistant SR/CR mice. The finding of CKA at lower levels in cancer patients suggests the possibility that it may be of a consequence of genetic, physiological, or pathological conditions, pending future studies with larger sample size.

Abstract Background Double stranded RNA (dsRNA) has multiple anti-tumor mechanisms. Over the past several decades, there have been numerous attempts to utilize synthetic dsRNA to control tumor growth in animal models and clinical trials. Recently, it became clear that intracellular dsRNA is more effective than extracellular dsRNA on promoting apoptosis and orchestrating adaptive immune responses. To overcome the difficulty in delivering a large dose of synthetic dsRNA into tumors, we propose to deliver a RNA replicase-based plasmid DNA, hypothesizing that the dsRNA generated by the replicase-based plasmid in tumor cells will inhibit tumor growth. Methods The anti-tumor activity of a plasmid (pSIN-β) that encodes the sindbis RNA replicase genes (nsp1-4) was evaluated in mice with model tumors (TC-1 lung cancer cells or B16 melanoma cells) and compared to a traditional pCMV-β plasmid. Results In cell culture, transfection of tumor cells with pSIN-β generated dsRNA. In mice with model tumors, pSIN-β more effectively delayed tumor growth than pCMV-β, and in some cases, eradicated the tumors. Conclusion RNA replicase-based plasmid may be exploited to generate intracellular dsRNA to control tumor growth.

Abstract Background Spontaneous regression/complete resistance (SR/CR) mice are a unique colony of mice that possess an inheritable, natural cancer resistance mediated primarily by innate cellular immunity. This resistance is effective against sarcoma 180 (S180) at exceptionally high doses and these mice remain healthy. Methods In this study, we challenged SR/CR mice with additional lethal transplantable mouse cancer cell lines to determine their resistance spectrum. The ability of these transplantable cancer cell lines to induce leukocyte infiltration was quantified and the percentage of different populations of responding immune cells was determined using flow cytometry. Results In comparison to wild type (WT) mice, SR/CR mice showed significantly higher resistance to all cancer cell lines tested. However, SR/CR mice were more sensitive to MethA sarcoma (MethA), B16 melanoma (B16), LL/2 lung carcinoma (LL/2) and J774 lymphoma (J774) than to sarcoma 180 (S180) and EL-4 lymphoma (EL-4). Further mechanistic studies revealed that this lower resistance to MethA and LL/2 was due to the inability of these cancer cells to attract SR/CR leukocytes, leading to tumor cell escape from resistance mechanism. This escape mechanism was overcome by co-injection with S180, which could attract SR/CR leukocytes allowing the mice to resist higher doses of MethA and LL/2. S180-induced cell-free ascites fluid (CFAF) co-injection recapitulated the results obtained with live S180 cells, suggesting that this chemoattraction by cancer cells is mediated by diffusible molecules. We also tested for the first time whether SR/CR mice were able to resist additional cancer cell lines prior to S180 exposure. We found that SR/CR mice had an innate resistance against EL-4 and J774. Conclusions Our results suggest that the cancer resistance in SR/CR mice is based on at least two separate processes: leukocyte migration/infiltration to the site of cancer cells and recognition of common surface properties on cancer cells. The infiltration of SR/CR leukocytes was based on both the innate ability of leukocytes to respond to chemotactic signals produced by cancer cells and on whether cancer cells produced these chemotactic signals. We found that some cancer cells could escape from SR/CR resistance because they did not induce infiltration of SR/CR leukocytes. However, if infiltration of leukocytes was induced by co-injection with chemotactic factors, these same cancer cells could be effectively recognized and killed by SR/CR leukocytes.

Abstract Background Spontaneous Regression/Complete Resistant (SR/CR) mice are a colony of cancer-resistant mice that can detect and rapidly destroy malignant cells with innate cellular immunity, predominately mediated by granulocytes. Our previous studies suggest that several effector mechanisms, such as perforin, granzymes, or complements, may be involved in the killing of cancer cells. However, none of these effector mechanisms is known as critical for granulocytes. Additionally, it is unclear which effector mechanisms are required for the cancer killing activity of specific leukocyte populations and the survival of SR/CR mice against the challenges of lethal cancer cells. We hypothesized that if any of these effector mechanisms was required for the resistance to cancer cells, its functional knockout in SR/CR mice should render them sensitive to cancer challenges. This was tested by cross breeding SR/CR mice into the individual genetic knockout backgrounds of perforin (Prf-/-), superoxide (Cybb-/), or inducible nitric oxide (Nos2-/). Methods SR/CR mice were bred into individual Prf-/-, Cybb-/-, or Nos2-/- genetic backgrounds and then challenged with sarcoma 180 (S180). Their overall survival was compared to controls. The cancer killing efficiency of purified populations of macrophages and neutrophils from these immunodeficient mice was also examined. Results When these genetically engineered mice were challenged with cancer cells, the knockout backgrounds of Prf-/-, Cybb-/-, or Nos2-/- did not completely abolish the SR/CR cancer resistant phenotype. However, the Nos2-/- background did appear to weaken the resistance. Incidentally, it was also observed that the male mice in these immunocompromised backgrounds tended to be less cancer-resistant than SR/CR controls. Conclusion Despite the previously known roles of perforin, superoxide or nitric oxide in the effector mechanisms of innate immune responses, these effector mechanisms were not required for cancer-resistance in SR/CR mice. The resistance was functional when any one of these effector mechanisms was completely absent, except some noticeably reduced penetrance, but not abolishment, of the phenotype in the male background in comparison to female background. These results also indicate that some other effector mechanism(s) of granulocytes may be involved in the killing of cancer cells in SR/CR mice.

Abstract Background Spontaneous Regression/Complete Resistant (SR/CR) mice are resistant to cancer through a mechanism that is mediated entirely by leukocytes of innate immunity. Transfer of leukocytes from SR/CR mice can confer cancer resistance in wild-type (WT) recipients in both preventative and therapeutic settings. In the current studies, we investigated factors that may impact the efficacy and functionality of SR/CR donor leukocytes in recipients. Results In sex-mismatched transfers, functionality of female donor leukocytes was not affected in male recipients. In contrast, male donor leukocytes were greatly affected in the female recipients. In MHC-mismatches, recipients of different MHC backgrounds, or mice of different strains, showed a greater negative impact on donor leukocytes than sex-mismatches. The negative effects of sex-mismatch and MHC-mismatch on donor leukocytes were additive. Old donor leukocytes performed worse than young donor leukocytes in all settings including in young recipients. Young recipients were not able to revive the declining function of old donor leukocytes. However, the function of young donor leukocytes declined gradually in old recipients, suggesting that an aged environment may contain factors that are deleterious to cellular functions. The irradiation of donor leukocytes prior to transfers had a profound suppressive effect on donor leukocyte functions, possibly as a result of impaired transcription. The cryopreserving of donor leukocytes in liquid nitrogen had no apparent effect on donor leukocyte functions, except for a small loss of cell number after revival from freezing. Conclusion Despite the functional suppression of donor leukocytes in sex- and MHC-mismatched recipients, as well as old recipients, there was a therapeutic time period during the initial few weeks during which donor leukocytes were functional before their eventual rejection or functional decline. The eventual rejection of donor leukocytes will likely prevent donor leukocyte engraftment which would help minimize the risk of transfusion-associated graft-versus-host disease. Therefore, using leukocytes from healthy donors with high anti-cancer activity may be a feasible therapeutic concept for treating malignant diseases.