Induction Of Mammary Tumors Research Articles

Effects of non-contact electric fields on kidney and liver histology in tumour-induced rats

Background A novel modality of cancer treatment based on exposure to non-contact electric fields has been developed to reduce cancer incidence. However, the safety of the electric field exposure was not fully investigated. Therefore, This study aimed to observe the safety of electric field exposure on kidney and liver structures. Methods Female Sprague-Dawley rats were divided into one control group and three treatment groups. Animals were treated with 7,12-dimethylbenz[a]anthracene for mammary tumour induction and exposed to non-contact electric fields individually for 10 hours a day for three weeks. Fresh samples of the kidney and liver were collected for observation of structural damage in both organs. Both organs were prepared for histopathological cross-sectioning using the paraffin method and Hematoxylin and Eosin staining followed by histological scoring using the post-examination masking method. Results Damages found in the kidney were as follows: thickening of the Bowman capsule, karyolysis, karyorrhexis, pyknosis, cloudy swelling, epithelial sloughing, inflammation, haemorrhage, and congestion. In addition, the number of inflammation and haemorrhage in the kidney structure of healthy rats that were exposed to electric fields was significantly lower than that in the control group. All damages to the kidneys were also found in the liver, but each showed a different degree of damage. Exposure to this electric field can cause haemorrhagic damage to the livers of healthy rats, but not to rats with breast cancer. Conclusions Exposure to non-contact electric fields can cause haemorrhagic damage to the livers of healthy rats, however, in other liver tissues and the kidneys, exposure to this electric field was safe. It can even decrease the number of inflammations and haemorrhages in the kidneys.

Effects of non-contact electric fields on kidney and liver histology in tumour-induced rats

Background: There were an estimated 2.1 million breast cancer diagnoses in 2018 worldwide, which is about 11.6% of the total cancer incidence. A novel modality of cancer treatment based on exposure to non-contact electric fields has been developed to reduce cancer incidence. However, the safety of the electric field exposure was not fully investigated. Therefore, the purpose of this study is to observe the safety of the electric field exposure on renal and liver structure. Methods: Female Sprague-Dawley rats were divided into one control group and three treatment groups. Animals were treated with 7,12-dimethylbenz[a]anthracene for mammary tumour induction and exposed to non-contact electric fields individually for 10 hours a day for three weeks. Fresh samples of the kidney and liver were collected for observing structural damage in both organs. The two organs were prepared for histopathological cross-sectioning using the paraffin method and Hematoxylin & Eosin staining followed by histological scoring using the post-examination masking method. Results: The damages found in the kidney were the following: thickening of Bowman capsule, karyolysis, karyorrexhis, pyknosis, cloudy swelling, epithelial sloughing, inflammation, haemorrhage, and congestion. The number of inflammation and haemorrhage in the kidney structure of the placebo group was the lowest and significantly different from the three other groups. All damages in the kidney were also found in the liver, but each showed different levels of damage. The damages in the kidney and liver caused by the exposure were not significant. Conclusions: The non-contact electric fields were not harmful to renal and liver structure in tumour-induced rats. Instead, it may increase the renal function in normal rats.

Effects of non-contact electric fields on kidney and liver histology in tumour-induced rats

Background: A novel modality of cancer treatment based on exposure to non-contact electric fields has been developed to reduce the cancer incidence. However, the safety of this electric field exposure on vital organs has not been fully investigated. Therefore, the purpose of this study was to observe the safety of the electric field exposure on kidney and liver structures. Methods: Female Sprague-Dawley rats were divided into one control group and three treatment groups. Animals were treated with 7,12-dimethylbenz[a]anthracene for mammary tumour induction and exposed to non-contact electric fields individually for 10 hours a day for three weeks. Fresh samples of kidney and liver were collected for observation of structural damage in both organs. Both organs were prepared for histopathological cross-sectioning using the paraffin method and Hematoxylin & Eosin staining followed by histological scoring using the post-examination masking method. Results: Damages found in the kidney were as follows: thickening of Bowman capsule, karyolysis, karyorrexhis, pyknosis, cloudy swelling, epithelial sloughing, inflammation, haemorrhage, and congestion. The thickening of Bowman’s capsule was significant, but other damages to the rest of kidney tissue were not significant. In addition, the number of inflammations and haemorrhages in the kidney structure of the placebo group was the lowest and significantly different compared to the other three groups. All damages to the kidneys were also found in the liver, but each showed a different degree of damage. The damages to the liver caused by exposure to the electric fields were not significant. Conclusions: Together with the effects of DMBA administration, exposure to non-contact electric fields can cause thickening of Bowman's capsule in the glomerulus of the kidney, however, in other kidney tissues and in the liver, exposure to this electric field was safe. It can even decrease the number of inflammations and haemorrhages in the organs.

Kidney and liver histology in tumour-induced rats exposed to non-contact electric fields

Background: There were an estimated 2.1 million breast cancer diagnoses in 2018 worldwide, which is about 11.6% of the total cancer incidence. A novel modality of cancer treatment based on exposure to non-contact electric fields has been developed to reduce cancer incidence. However, the safety of the electric field exposure was not fully investigated. Therefore, the purpose of this study is to observe the safety of the electric field exposure on renal and liver structure. Methods: Female Sprague-Dawley rats were divided into one control group and three treatment groups. Animals were treated with 7,12-dimethylbenz[a]anthracene for mammary tumour induction and exposed to non-contact electric fields individually for 10 hours a day for three weeks. Fresh samples of the kidney and liver were collected for observing structural damage in both organs. The two organs were prepared for histopathological cross-sectioning using the paraffin method and Hematoxylin & Eosin staining followed by histological scoring using the post-examination masking method. Results: The damages found in the kidney were the following: thickening of Bowman capsule, karyolysis, karyorrexhis, pyknosis, cloudy swelling, epithelial sloughing, inflammation, haemorrhage, and congestion. The number of inflammation and haemorrhage in the kidney structure of the placebo group was the lowest and significantly different from the three other groups. All damages in the kidney were also found in the liver, but each showed different levels of damage. The damages in the kidney and liver caused by the exposure were not significant. Conclusions: The non-contact electric fields were not harmful to renal and liver structure in tumour-induced rats. Instead, it may increase the renal function in normal rats.

Tumor-associated macrophage heterogeneity is driven by tissue territories in breast cancer.

Tissue-resident macrophages adapt to local signals within tissues to acquire specific functions. Neoplasia transforms the tissue, raising the question as to how the environmental perturbations contribute to tumor-associated macrophage (TAM) identity and functions. Combining single-cell RNA sequencing (scRNA-seq) with spatial localization of distinct TAM subsets by imaging, we discover that TAM transcriptomic programs follow two main differentiation paths according to their localization in the stroma or in the neoplastic epithelium of the mammary duct. Furthermore, this diversity is exclusively detected in a spontaneous tumor model and tracks the different tissue territories as well as the type of tumor lesion. These TAM subsets harbor distinct capacity to activate CD8+ Tcells and phagocyte tumor cells, supporting that specific tumor regions, rather than defined activation states, are the major drivers of TAM plasticity and heterogeneity. The distinctions created here provide a framework to design cancer treatment targeting specific TAM niches.

Cytotoxic T cells response with decreased CD4/CD8 ratio during mammary tumors inhibition in rats induced by non-contact electric fields.

Background: Breast cancer is the most common cancer in women worldwide and is the leading cause of death amongst women with cancer. One novel therapy used for breast cancer treatmentconstitutes non-contact electric fields and is called electro-capacitive cancer therapy (ECCT) with intermediate frequency and low intensity. The objective of this study was to examine the effect of ECCT on mammary tumors growth in rats and observing the immune responses that play a role in fighting the tumor. Methods: Female SD rats were used and divided into four groups, namely control (NINT), placebo (NIT), non- therapy (INT), and therapy (IT) groups with 6 biological replicates in each group. Rats in INT and IT groups were treated with 7,12-dimethylbenz[a]anthracene for mammary tumor induction. Only rats in NIT and IT groups were exposed to ECCT individually for 10 hours per day for 21 days. The size of all tumors was measured with a digital caliper. The distributions of PCNA, ErbB2, caspase-3, CD68, CD4, and CD8-positive cells were observed with immunohistochemistry and scoring with ImageJ. Results: The growth rate of mammary tumors in IT group was significantly lower (p<0.05) than that in INT group. The number of mitotic figures and the percentage of PCNA, caspase-3, and CD68-positive cells in IT group were significantly lower (p<0.05) than those in INT group. Conversely, the percentage of CD8-positive T cells in IT group was significantly higher (p<0.05) than that in INT group. Moreover, the CD4/CD8 ratio in IT group was found to have decreased. Some tumor tissues were blackened and detached from the surrounding tissue, resulting in an open wound which then healed upon exposure. Conclusions: Non-contact electric fields exposure showed inhibition on mammary tumor growth in rats while inducing CD8+ T cells, leading to tumor cell death and potentially helping wounds heal.

Cytotoxic T cells response with decreased CD4/CD8 ratio during mammary tumors inhibition in rats induced by non-contact electric fields

Background: Breast cancer is the most common cancer in women worldwide and is the leading cause of death amongst women with cancer. One novel therapy used for breast cancer treatment constitutes non-contact electric fields and is called electro-capacitive cancer therapy (ECCT) with intermediate frequency and low intensity. The objective of this study was to examine the effect of ECCT on mammary tumors growth in rats and observing the immune responses that play a role in fighting the tumor. Methods: Female SD rats were used and divided into four groups, namely control (NINT), placebo (NIT), non- therapy (INT), and therapy (IT) groups with 6 biological replicates in each group. Rats in INT and IT groups were treated with 7,12-dimethylbenz[a]anthracene for mammary tumor induction. Only rats in NIT and IT groups were exposed to ECCT individually for 10 hours per day for 21 days. The size of all tumors was measured with a digital caliper. The distributions of PCNA, ErbB2, caspase-3, CD68, CD4, and CD8-positive cells were observed with immunohistochemistry and scoring with ImageJ. Results: The growth rate of mammary tumors in IT group was significantly lower (p<0.05) than that in INT group. The number of mitotic figures and the percentage of PCNA, caspase-3, and CD68-positive cells in IT group were significantly lower (p<0.05) than those in INT group. Conversely, the percentage of CD8-positive T cells in IT group was significantly higher (p<0.05) than that in INT group. Moreover, the CD4/CD8 ratio in IT group was found to have decreased. Some tumor tissues were blackened and detached from the surrounding tissue, resulting in an open wound which then healed upon exposure. Conclusions: Non-contact electric fields exposure showed inhibition on mammary tumor growth in rats while inducing CD8+ T cells, leading to tumor cell death and potentially helping wounds heal.

Cytotoxic T cells response with decreased CD4/CD8 ratio during mammary tumors inhibition in rats induced by non-contact electric fields.

Background: Breast cancer is the most common cancer in women worldwide and is the leading cause of death in women with cancer. One novel therapy used for breast cancer treatment is non-contact electric fields called electro-capacitive cancer therapy (ECCT) with intermediate frequency (100 kHz) and low intensity (18 Vpp). The objective of this study was to examine the effect of ECCT on mammary tumors growth in rats and observing the immune responses that play a role in fighting the tumor. Methods: Female SD rats were used and divided into four groups, namely control (NINT), placebo (NIT), non- therapy (INT), and therapy (IT) groups with 6 biological replicates in each group. Rats in INT and IT groups were treated with 7,12-dimethylbenz[a]anthracene for mammary tumor induction. Only rats in NIT and IT groups were exposed to ECCT individually for 10 hours per day for 21 days. The size of all tumors was measured with a digital caliper. The distributions of PCNA, ErbB2, caspase-3, CD68, CD4 and CD8-positive cells were observed with immunohistochemistry and scoring with ImageJ. Results: The growth rate of mammary tumors in IT group was significantly lower (p<0.05) than that in the INT group. The number of mitotic figures and the percentage of PCNA, caspase-3, and CD68- positive cells in IT group were significantly lower (p<0.05) than those in INT group. Conversely, the percentage of CD8-positive T cells in IT group was significantly higher (p<0.05) than that in INT group. Moreover, the CD4/CD8 ratio in IT group was decreased. Some tumor tissues were blackened and detached from the surrounding tissue, resulting in an open wound which then healed up upon exposure. Conclusions: Non-contact electric fields exposure showed inhibition on mammary tumor growth in rats while inducing CD8+ T cells that lead to tumor cells death and potentially helps wound healing.

Liver cell membrane antibody, kidney injury molecule and lungs tumour antigen expression on the MCF-7 cell induced breast cancer administered with Citrus limon juice and tamoxifen in Sprawgue dawley rats

Monitoring and evaluation have been one of the major measures to determine possible progression of response of disorder to the alternative treatments, however Anti-Liver Cell membrane Antibody, Kidney Injury Molecule and Lungs Tumour Antigen Expression on the MCF-7 Cell Induced Breast Cancer Rats administered with Citrus Limon juice and Tamoxifen were carried out, Over one hundred and twenty Sprague dawley rats of 40 days old average body weight 180-220g were divided into ten (10) containing of 12 animals per group, group 1 was control, fed only with rat chow and water, group 2 was MCF-7 cell line induced rats alone (BCIR only), group 3 Citrus Limon juice (CLJ) at 8.88%, Group 4 Citrus Limon juice (CLJ) at 17.32%, group 5 Citrus Limon juice (CLJ) at 25.98%, group 6 was given 0.2mg/kg of Tamoxifen alone, group 7 (BCIR+CLJ at 8.88%), group 8 (BCIR+CLJ at 17.32%), group 9 (BCIR+CLJ at 25.98%) and group (BCIR+ 0.2mg/kg of Tamoxifen). Acute and sub – acute toxicity were carried out after the establishment of safety dose following determination of LD50, on the fresh Citrus Limon juice Breast cancer induction and Tumor sizes were analysed. At the end of the administration, animals were sacrificed, fresh blood was taken, centrifuged at 3000 rpm, serum was collected and stored at 8oc Breast Cancer Tumor Associated Assays, Kidney Injury, Liver cell Membrane Antibody and Lung Tumor Antigens were analysed. Result, Mammary tumor induction was succeeded in all the MCF-7 cell line induced groups with minimum number of morbidity. Tumor size was significantly increase in tumor mass BCIR only at P&gt;0.001 when compared to BCIR + CLJ and BCIR + Tamoxifen groups. Immunoogical expression of Breast Cancer Associated Tumor revealed significant increase suppressor gene/protein, Immunocytochemical in-vivo of Breast Cancer Associated Organs showed significant increase in KIM , LCMA and LTA at P&gt;0.001 in BCIR when compared to all the BCIR + CLJ, BCIR + Tamoxifen and other non-BCIR groups. In conclusion, this study showed that KIM , LCMA and LTA exhibited strong Immunochemical expression of Citrus Limon juice potency possessing possible anticancer activities without posing pathological conditions on the body during and after usage, hence KIM, LCMA and LTA are demonstrated to be valuable in the monitoring and evaluation progression of tumor as Citrus Limon Juice could be used as alternative therapy in the treatment of Breast cancer that are hormonal dependant in similar manner to Tamoxifen.

Acrylamide levels in coffee powder, potato chips and French fries in Addis Ababa city of Ethiopia

Acrylamide is a processing contaminant with genotoxic and carcinogenic properties. It is formed in food when free asparagine reacts with reducing sugars at high temperatures. This study evaluated the levels of acrylamide in three commercial important heat-processed foods: roasted coffee powder (n = 30), potato chips (n = 30), and French fries (n = 30) marketed in Addis Ababa, Ethiopia. Acrylamide levels obtained ranged from 135 μg/kg to 1139 μg/kg in roasted coffee powder, from 211 μg/kg to 3515 μg/kg in potato chips, and from 36 μg/kg to 1411 μg/kg in French fries. Compared to French fries, potato chips samples showed a higher mean acrylamide content. 43% of the coffee powder (n = 13), 57% of the potato chips (n = 17), and 40% of the French fries samples (n = 12) had acrylamide higher than the levels recommended by the European Commission. Acrylamide levels in brewed coffee will be much lower than coffee powder as coffee powder undergoes further processing before consumption. Exposure to acrylamide is based on a food's acrylamide content and frequency of consumption of that food. Considering Ethiopia is largest coffee consuming country in Africa, assessing the risk of exposure to acrylamide from coffee consumption could be something worth investigating. The dietary exposure of children (n = 30) to acrylamide from consumption of French fries was estimated to be 0.66 μg/kg body weight/day. The margins of exposure (MOE) based on Harderian gland and mammary tumor induction endpoints were estimated to be 272 and 470. These low MOE values may indicate that the risk of exposure to acrylamide from French fries consumption could be of public concern and requires attention.

Control of Initiation and Aggressiveness of DMBA‐Induced Breast Cancer in Rats by Dietary Consumption of Hinute‐AM.

We reported earlier that the consumption of soy protein controlled not only the proliferation of dimethylbenz [a] anthracene (DMBA)‐induced breast cancer in rats but also its aggressiveness. Soy protein is hydrolyzed in the gut to small peptides which are easily absorbed and are involved in various cellular functions. Fuji Oil Company, Osaka, Japan, has commercially marketed a soy protein hydrolysate, termed Hinute‐AM, composed of primarily di‐ and tripeptides, to be used as a health food, and it has anti‐inflammatory property. The purpose of the present study was to investigate whether dietary intake of Hinute‐AM can delay the development of DMBA‐induced breast cancer and its aggressiveness in rats. For this study, we obtained 22‐days old female Sprague Dawley rats from Charles River (Wilmington, MA). Animals were divided into 5 groups, each containing 5 animals. Group 1 received standard AIN‐76A diet (Envigo – Teklad Diets, Madison, WI) containing 20% casein and 0% Hinute‐AM; Group 2 received the same diet containing 15% casein + 5% Hinute‐AM; Group 3 received the same diet, but containing 10% casein + 10% Hinute‐AM; Group 4 received the same diet but containing 5% casein + 15% Hinute‐AM; and Group 5 received the same diet but containing 0% casein + 20% Hinute‐AM. For induction of mammary tumors, all groups received a single intragastric administration of DMBA (Sigma Chemical Co., St. Louis) dissolved in sesame oil (80 mg/kg b.wt.) at 50 days of age. At 120 days post DMBA injection, the animals were euthanized by CO2 asphyxiation. All tumors were weighed and measured for volume, and a section of tumor was subjected to histological grading by a certified pathologist. Our preliminary data revealed that replacement of casein by Hinute‐AM as a source of dietary protein caused a dose‐dependent delay in the development of breast tumor, decrease in breast cancer incidence, a decrease in tumor volume, and tumor grade.Support or Funding InformationThis work was supported by grants from Fuji Oil Company, Osaka, Japan and by the Meharry Translational Research Center grant NIH 5U54MD007593

Potency ofMurraya koenigiiLeaves as Anti-Cancer Mammary in 7,12 dimethylbenz(α) anthracene (DMBA) induced-Sprague Dawley Rats

The purpose of the research was to study the potency ofMurraya koenigiileaves extract to overcome the mammary tumor in Sprague Dawley rat. Thirty-five female rats were divided into seven groups: control (P1), tumor without therapy (P2), methotrexate group (P3), curative groups (P4 and P5) were given extract after the tumor was formed, and preventive groups (P6 and P7) were given extract before the tumor was formed with dose of 300 and 400 mg/kg, respectively. The induction of mammary tumor in rats were carried out using 7,12 dimethylbenz(α) anthracene (DMBA) subcutaneously. Bodyweight and tumor size were measured every week for 4 weeks. At the end of treatment, rats were euthanized and mammary glands were collected for histopathological examination. The result showed tumor size in P2 was significantly higher (p&lt;0.05) than in other groups. On the other hand, tumor size in P4 and P6 were significantly smaller (p&lt;0.05) compared to P5 and P7. Histopathological changes showed PMN cells, 1-3 layers of cuboid epithelial and solid collagen fibers proliferation in P2, while in P3 to P7 showed moderate collagen fibers proliferation. In conclusion, the administration of the extract at a dose of 300 mg/kg can decelerate tumor development in Sprague Dawley rat mammary gland.

Rasam (South Indian Spice Soup) - Attenuates the Mammary Tumor Induction Magnitude of 7,12-Dimethylbenz[a] anthracene in Sprague–Dawley Rats

Background: Recently, we have reported rasam (South Indian spice soup) for antiproliferative activity against Michigan Cancer Foundation-7 cell lines. Objectives: Breast cancer seems to be more common in the younger age group. Considering the deviation of the younger group from the traditional food habits as one of the many reasons of breast cancer incidence, rasam was investigated for its chemopreventive effect on mammary carcinoma. Materials and Methods: Rasam at 3 and 4 mL/kg dose was administered to female Spraque–Dawley rats for 30 days before 7,12-dimethylbenz[a] anthracene (DMBA) induction of mammary carcinoma. Tamoxifen was used as the standard drug. Body weight, thiobarbituric acid reactive substances (TBARSs), phase I enzymes, hexokinase (HK), aldolase (AD), glucose 6-phosphatase (G6P), fructose 1,6-bisphosphatase (F16P), citric acid cycle (CAC) enzymes, histopathology, tumor weight, tumor latency, and tumor incidence were studied. Results: In all the studied parameters such as body weight, TBARS, phase I enzymes, HK, AD, G6P, F16P, and CAC enzymes, rasam-pretreated groups showed better prevention than tamoxifen-treated groups. Histochemical findings also clearly confirm the chemopreventive effect of rasam. Particularly at the dose of 4 mL/kg, rasam was efficient in reducing the percentage of incidence of tumor, number of tumor, and tumor weight. Conclusion: The present study reveals that rasam attenuates the mammary tumor induction magnitude of DMBA in female rats.

Therapeutic and Preventive Effects of Osteoclastogenesis Inhibitory Factor on Osteolysis, Proliferation of Mammary Tumor Cell and Induction of Cancer Stem Cells in the Bone Microenvironment.

Background: We examined the effects of recombinant human osteoclastogenesis inhibitory factor (hOCIF) on osteolysis, proliferation of mammary tumor cells, and induction of cancer stem cells (CSCs) in the tumor-bone and tumor-subcutaneous microenvironments (TB- and TS-microE). Methods: Mouse mammary tumor cells were transplanted onto the calvaria or into a subcutaneous lesion of female mice, creating a TB-microE and a TS-microE, and the mice were then treated with hOCIF. To investigate the preventive effects of hOCIF, mice were treated with hOCIF before tumor cell implantation onto the calvaria (Pre), after (Post), and both before and after (Whole). The number of CSCs and cytokine levels were evaluated by IHC and ELISA assay, respectively. Results: hOCIF suppressed osteolysis, and growth of mammary tumors in the TB-microE, but not in the TS-microE. In the Pre, Post, and Whole groups, hOCIF suppressed osteolysis, and cell proliferation. hOCIF increased mouse osteoprotegrin (mOPG) levels in vivo, which suppressed mammary tumor cell proliferation in vitro. These preventive effects were observed in the dose-dependent. hOCIF did not affect the induction of CSCs in either microenvironment. Conclusion: While receptor activator of NF-κB ligand (RANKL) targeting therapy may not affect the induction of CSCs, RANKL is a potential target for prevention as well as treatment of breast cancer bone metastasis.

Effect of Age at Exposure on the Incidence of Lung and Mammary Cancer after Thoracic X-Ray Irradiation in Wistar Rats.

Epidemiology studies have shown that children are at greater overall risk of radiation-induced cancer, but the modifying effect of age at exposure in different tissues is heterogeneous. Early epidemiology findings of increased lung cancer risk with increasing age at the time of exposure have been dismissed, with suggestions that the trend is an artefact from a failure to adequately correct for the effects of tobacco smoking. Yet, differing models used in subsequent analyses have shown that the increased susceptibility with age, counter to the overall solid tumor trend, can either be confirmed or discounted depending on the model parameters used. In this study, we analyzed the induction of tumors in female Wistar rats exposed to increasing thoracic doses of X-ray as neonates, juveniles or young adults, to allow the effect of age at exposure in this early period to be observed in the absence of any interactions with smoking. Histology was used to compare tumor subtypes among groups, and genomic DNA copy number alterations in a number of tumors arising after irradiation at different ages were examined. Induction of lung cancers increased with radiation dose, with the frequency of early occurring lung adenomas greater in rats irradiated at older ages. At the highest dose, the rats irradiated at 5 or 15 weeks of age showed increased age-specific rates of lung adenocarcinomas in later life compared to those irradiated at 1 week of age. However, thoracic mammary gland tumors induced by the highest dose at the later ages significantly decreased the lifespan in these groups, reducing the number of rats at risk of radiation-induced lung adenocarcinoma. There was no induction of mammary tumors outside of the irradiated field. Lung adenocarcinomas showed widespread DNA copy number aberrations at the chromosome level, but the only recurrent lesions were intragenic Fhit deletions and losses on chromosome 4. The results presented here suggest that the risk of radiation-induced lung cancer after irradiation may not monotonically decrease with age, and demonstrate that increasing lung cancer risk with exposure age can be observed independent of corrections for smoking, and that mammary tumors may show a similar relationship with age.

Differences in the Rate of in Situ Mammary Gland Development and Other Developmental Endpoints in Three Strains of Female Rat Commonly Used in Mammary Carcinogenesis Studies: Implications for Timing of Carcinogen Exposure.

The potential of chemicals to alter susceptibility to mammary tumor formation is often assessed using a carcinogen-induced study design in various rat strains. The rate of mammary gland (MG) development must be considered so that the timing of carcinogen administration is impactful. In this study, in situ MG development was assessed in females of the Harlan Sprague-Dawley (Hsd:SD), Charles River Sprague-Dawley (Crl:SD), and Charles River Long-Evans (Crl:LE) rat strains at postnatal days 25, 33, and 45. Development was evaluated by physical assessment of growth parameters, developmental scoring, and quantitative morphometric analysis. Although body weight (BW) was consistently lower and day of vaginal opening (VO) occurred latest in female Hsd:SD rats, they exhibited accelerated pre- and peripubertal MG development compared to other strains. Glands of Crl:SD and Crl:LE rats exhibited significantly more terminal end buds (TEBs) and TEB/mm than Hsd:SD rats around the time of VO. These data suggest a considerable difference in the rate of MG development across commonly used strains, which is independent of BW and timing of VO. In mammary tumor induction studies employing these strains, administration of the carcinogen should be timed appropriately, based on strain, to specifically target the peak of TEB occurrence.

Abstract 3219: Prior vaccination is critical to PD1/PDL-1 treatment efficacy in a mouse breast cancer allograft

Abstract We previously generated the MuPrime® mBR6004 model, by engrafting the breast adenocarcinoma derived from MMTV-PyVT transgenic mice (GEMM)1 into the syngeneic FVB/N mice2,3. The allograft grows robustly, maintains the histopathology features of the original GEMM, and metastasizes to the lungs in all examined cases when implanted orthotopically2. Additionally, histopathology shows that the models is HER2++, but ER-/PR-. We profiled drug responses to standard of care (SoC)2 and checkpoint inhibitors3. Interestingly, among all checkpoints inhibitors tested, our model is insensitive to PD1/PDL-1 inhibitors, but responds to CTLA-4 inhibitors3. To further explore the underlying mechanism of response, we performed extensive tumor immuno-profiling for the presence of infiltrating immune cells, e.g. TIL, CTL, Treg, immune-suppressive macrophages, NK, etc3. We observed a good pharmacodynamic (PD) correlation between the presence of tumor infiltrating T-cells, particularly CD8+ TIL, and NK and a positive response to therapy, regardless the treatment type. Given this preliminary observation, we attempted animal vaccination with tumor lysates prior to tumor engraftment or treatments. Interestingly, while vaccination had minimal effects on the engraftment take rate (100% take for all animals) and on the growth kinetics of the engraftments, it had profound effects on the tumor response to anti-PD1/anti-PDL-1 antibody treatments, significantly enhancing tumor response to these agents in a model that was otherwise unresponsive. Our results suggest that the immunological status of the animal, at least with regard to the specific anti-tumor immunity, is critically important to determine the response to checkpoint inhibitors. Our observations suggest that vaccination is critical for the overall success of immunotherapy (I/O) treatments utilizing checkpoint inhibitors as well other treatment strategies.

Abstract P3-03-03: A mouse model of sporadic breast tumor with a conditional P53 mutation

Abstract Missense mutations in the tumor suppressor gene p53 are present in more than 50% of human tumors. In particular, the arginine-to-histidine mutation at codon 175 (p53R175H) has been found in more than 4% of human breast cancers. p53 mutations fall into two general categories: germline mutations that are associated with hereditary tumors, and somatic mutations that cause sporadic tumors. Mouse models for cancers induced by p53 germline mutations have been established and characterized. However, hitherto there are no accurate animal models for sporadic tumors induced by p53 somatic missense mutations, although this mechanism of inactivating p53 occurs in a large fraction of human cancers. We have generated a mouse allele, p53WM, that carries coding sequences for both wild type (WT) and R172H mutant p53 (corresponding to R175H in humans) at the p53 endogenous locus. The coding sequence for WT p53 is flanked by loxP sites and therefore can be deleted by Cre recombinase. In the absence of Cre, following radiation, the p53WM/+ mice have the similar ability as the p53 WT mice to activate p53, as demonstrated by transcriptional activation of p53 targets and induction of apoptosis. In addition, like WT mouse embryonic fibroblasts (MEFs), the p53WM/+ MEFs also underwent cell cycle arrest following radiation. After crossing p53WM/+ mice with Zp3-Cre mice which express the Cre recombinase in oocytes, the coding sequence for WT p53 was removed, and the mutant p53R172H allele was regenerated as determined by sequencing, western blot analysis and real time reverse transcription PCR. To investigate whether a somatic p53R172H mutation in mammary epithelium can induce breast tumors, adenoviruses expressing Cre (Ad-Cre) were injected into the mammary duct of p53WM/+ mice. Limiting the adenovirus dose allowed us to induce the p53R172H mutation in 1 of 20, 100 or 1000 cells, generating a mutant p53 surrounded by normal cells which more accurately mimics the clinical situation of sporadic tumors. Meanwhile, p53WM/+; K14-Cre and p53WM/+; WAP-Cre mice have been generated, in which p53R172H mutation are induced through Cre transgene specifically in the mammary epithelial cells. Lastly, to examine how somatic p53 mutation cooperates with an oncogene in mammary tumor induction, MMTV-neu; p53WM/+ mice were also subjected to the mammary intraductal Ad-Cre injection. We are currently monitoring mice and will further investigate the molecular mechanisms of tumorigenesis by studying the loss of heterozygosity, and the co-evolution of tumor epithelial cells and their microenvironment. This mouse model is more likely to share the underlying molecular pathology with human sporadic tumors. Therefore, it will be more predictive of human responses to drugs, and thus a more valuable tool in preclinical testing. Citation Format: Zhang Y, Xiong S, Pant V, El-Naggar A, Lozano G. A mouse model of sporadic breast tumor with a conditional P53 mutation. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P3-03-03.

Studies on the biochemical and molecular effects of some natural herbs on experimental-induced breast cancer in Wistar rats

Background: Breast cancer is the most frequent malignant tumor in women, and its metastatic state represents the second cause of mortality. To treat cancer, we must prevent tumor cell proliferation and angiogenesis. The usage of herbal medication for treatment of some diseases as diabetes is increased nowadays for their wide safety margin. Aims and Objectives: Therefore, the current study was designed to examine the protective effect of ginger extract (GE) and cinnamon extract (CE) in controlling the mammary gland incidence in female Wistar rats. Materials and Methods: A total of 75 female virgin Wistar rats were allocated into 5 groups. Control group without any treatment; mammary gland group administered 7, 12-dimethyl-benz[a]anthracene at a dose of 20 mg/kg orally in corn oil. Rats in 3-5 served as tumor groups and received GE for group 3 (0.125% in water), CE for group 4 (100 mg/kg), and a mixture of GE plus CE for group 5. GE and CE were administered 2 weeks before cancer induction and continued for 4 months. Serum levels of breast cancer biomarker (CA125) and oxidative stress biomarkers were measured. Tissues of tumors were examined at molecular and histopathological levels. Results: There was a significant increase in the serum levels of CA125 in untreated mammary gland tumor group when compared with untreated healthy rats. There was a significant decrease in the serum levels of glutathione (GSH) peroxidase, catalase, superoxide dismutase, and GSH reductase in the untreated mammary gland tumor groups compared to control healthy rats. Administration of ginger and CE normalized the decrease in antioxidants levels. Induction of mammary gland tumor upregulated the genes associated with tumor incidences such as GST-P, CYP1A1, CYP1B1, and vascular endothelial growth factor-receptor 1 and administration of both herbal plants normalized these changes. Coadministration of GE and CE-induced additive inhibitory effect on genes upregulated due to tumor incidence. Bax expression was downregulated in tumor group and increased after GE and CE expression either alone or in combination. At histopathological levels, breast cancer group showed adenocarcinoma that is decreased with plants extracts administration when compared with breast cancer and control group. Conclusion: Results suggest that extracts of ginger and cinnamon have anticarcinogenic protective activities and support the hypothesis that these plants help in the prevention of mammary cancer by controlling the expression of carcinogenesis-associated genes.

Abstract B97: Response to checkpoint inhibition by GEMM breast cancer allograft

Abstract Background. Cancer patient's response to immunotherapy, e.g. checkpoint inhibitors, differ greatly, likely due to their genetic composition (e.g. neo-mutation) or immunological profile of tumors. Experimental models may facilitate the identification of responders or of the approach to enhance response. Syngeneic mouse tumor models have been widely used as experimental model for testing surrogate immunotherapy, but limited by the fact that only a few are available and respond to the current checkpoint inhibitors. Allografts of mouse spontaneous tumors (MuPrimeTM) may be used as new type of I/O model with advantages: 1) its primary natures with “stem cell diseases” and relevant tumor microenvironment as seen in PDX1-5; 2) diverse cancer types and deriving from wider range of available spontaneous mouse tumors. Method. We created a MuPrime, MBR6004, by allografting the breast adenocarcinoma derived from GEMM-MMTV-PyVT in FVB/N mice1. We then characterized the model by examine it subcutaneous and orthotopic growth, and its ability to metastasis. We also transcriptome-sequenced and immune-profiled the tumor, and assessed its response to different treatments, including I/O therapy. Results. MBR6004 was found to over-express HER2 but negative for ER and PR. It maintains original primary tumour histopathology, grows robustly, and is confirmed to express its original transgene. It is also resistant to docetaxel. The orthotopic implantation resulted in lung metastasis. We can detected and quantitatively confirmed tumor-infiltrating immune cells, e.g. TIL, CTL, Treg, immune-suppression macrophage, NK, etc. We confirmed that the tumor cell expressed relatively low PD-L1, but inducible, per flow analysis. Our preliminary data indicated that it partially respond to anti-mouse PD1 and anti-mouse CTLA-4 antibodies when mice are preconditioned. Currently, we are investigating whether the responses are associated with the reduced Treg and increased CD8+ TIL in tumors, and also whether the combination therapy can enhance its tumor response synergistically. Conclusion. Our data seem to suggest that MuPrime could be another type of I/O model alternative to commonly used syngeneic cell line derived models at present.