Identification of a key molecular regulator of liver metastasis in human pancreatic carcinoma using a novel quantitative model of metastasis in NOD/SCID/gammacnull (NOG) mice

We developed a reliable new model system for assaying liver metastasis using NOD/SCID/gamma(c)(null) (NOG) mice. Seven human pancreatic cancer cell lines were examined for their ability to form diverse metastatic foci in the livers of NOD/SCID and NOG mice. Capan-2 and PL45 showed no metastasis when seeded at up to 10(5) cells in both strains, and no BxPC-3 metastasis was observed in NOD/SCID mice. The NOD/SCID mouse model detected liver metastasis only in the AsPC-1 cell line when inoculated with >10(3) cells. In contrast, when inoculated with only 10(2) MIA PaCa-2, AsPC-1 and PANC-1 cells, liver metastasis was evident in 71.4% (5/7), 57.1% (4/7) and 37.5% (3/8) of the NOG mice, respectively. Capan-1 and BxPC-3 cells metastasized when seeded at 10(3) cells in 50% (5/10) and in 12.5% (1/8) of the mice, respectively. Using the NOG mouse model system, we established a highly metastatic cell line, liver metastasized-BxPC-3 (LM-BxPC-3), from liver metastatic foci formed by the relatively poorly metastatic parental BxPC-3 cell line. The gene expression profiles of parental and LM-BxPC-3 cells were compared, and we identified forty-five genes that were either upregulated or downregulated >4-fold in the LM-BxPC-3 cell line. We validated 9 candidate protein-coding sequences, and examined the correlation between their expression pattern and the in vivo liver metastatic potential of all 7 pancreatic cancer cell lines. Only S100A4 expression correlated with the ability to form liver metastases, as evaluated in our quantitative model of metastasis in NOG mice. These results suggested that S100A4 is a key regulator of liver metastasis in pancreatic cancer, and demonstrated the feasibility of using the quantitative metastasis model to search for and develop new anti-cancer therapies and novel drugs against this and other key molecules.     

Preclinical in vitro models from genetically engineered mice for breast and colon cancer (Review)

Genetically engineered mice with targeted alterations in clinically relevant oncogenes, tumor suppressor genes or DNA mismatch repair genes provide unique predictive animal models for human carcinogenesis, and cancer prevention/therapy. However, some of the genetically engineered mouse models lack target organ specificity for colon carcinogenesis. We have established, characterized and validated stable epithelial cell lines from 'normal' and 'genetically' predisposed target organs that offer innovative and mechanistic approaches, complementing in vivo studies on existing animal models for clinical breast and colon cancer. Epithelial cell lines with up- regulated Ras or myc oncogene, mutated Apc tumor suppressor gene and Mlh1 DNA mismatch repair gene provide facile experimental systems for organ site carcinogenesis and cancer prevention. Altered expression of cancer specific biomarkers and their modulation by several synthetic pharmacological agents such as retinoids, selective estrogen receptor modulators, non-steroidal anti-inflammatory drugs and specific enzyme inhibitors have been reported from our laboratory. Oncogene expressing MMEC-Ras and MMEC-myc mammary epithelial cells, Apc mutant 850Min COL and 1638N COL, and DNA mismatch repair/Apc mutant Mlh1/1638N COL colon epithelial cells exhibit aberrant cell cycle progression, down-regulated apoptosis and enhanced carcinogenic risk in vitro and tumor formation in vivo. We have reported that relative to the parental 'normal' non-neoplastic cells, genetically 'altered' pre-neoplastic cells exhibit enhanced sensitivity for growth arrest by multiple mechanistically distinct pharmacological agents. Comparative experiments on isogenic 'normal' and genetically 'altered' target cell lines facilitate cancer selective efficacy and identification of susceptible mechanistic pathways. Treatment of these genetically 'altered' pre-neoplastic cells with low dose combination of mechanistically distinct pharmacological agents as well as naturally occurring phytochemicals induce cytostatic growth arrest, alter cell cycle progression and reduce carcinogenic risk. The availability of validated technology for model development, and for mechanism based biomarker assays now establishes a novel platform to rapidly test carcinogenicity and preventive/therapeutic efficacy of novel pharmacological agents as well as naturally occurring phytochemicals. Thus, these data permit rational prioritization of efficacious lead compounds for preclinical testing and future clinical trials for prevention/therapy of breast and colon cancer.     

S100A4 expression with reduced E-cadherin expression predicts distant metastasis of human malignant melanoma cell lines in the NOD/SCID/gammaCnull (NOG) mouse model

Tumor xenografts in immune-deficient mice (athymic nude mice and SCID mice) are well-established animal models for the study of human cancer. Several human melanoma cell lines were reported to metastasize in the immune-deficient mice models. However, metastatic rates were extremely low in spite of large numbers of injections of cancer cells, more than 1 x 10(6) cells/mouse. The NOD/SCID/gamma(C)(null) (NOG) mouse shows multiple immunological dysfunctions, including cytokine production capability, in addition to the functional incompetence of T, B and natural killer (NK) cells. However, the immune-deficient mice, with preserved NK cell activity, might interfere with engraftment efficiency. We examined the distant metastasis of the human melanoma cell lines (A2058, A375, G361 and HMY-1, 1 x 10(4) cells/mouse) in the 6 weeks after intravenous inoculation. All four melanoma cell lines showed metastasis in the NOG mice, while no metastatic lesions were observed in the NOD/SCID mice. Metastatic lesions were noted in the liver and lung of 6/6 (100%) mice at A2058, 8/9 (89%) at A375, 2/6 (33%) at G361 and 2/8 (25%) at HMY-1. A2058 and A375 cell lines with high metastatic potentials show increased gene expression of S100A4. Western blot assay confirmed the increased protein levels of S100A4 in the A2058 and A375 cell lines. E-cadherin gene expression was conversely inhibited in these cell lines. The increased expression of S100A4 combined with inhibited E-cadherin expression resulted in high metastatic potentials of the human melanoma cell lines in vivo.     

大肠癌同时性肝转移癌应用植入式药泵25例治疗观察

目的探讨大肠癌同时性肝转移癌不能切除，术中同时应用植入式药泵化疗栓塞的作用。方法在切除原发病灶的同时对不能切除的肝转移癌在术中同时置入肝动脉植入式药泵并给予化疗栓塞。使用药物FUDR+DDP+EADM+碘化油，观察存活率。结果5年存活率8%、3年存活率56%、2年存活率24%、1年存活率12%。结论对于大肠癌同时性肝转移癌不能切除者采用植入式药泵化疗栓塞，具有明显提高存活率。使用植入式药泵操作简单、疗效显著、并发症少，具有较高的应用价值。     

Potential utility of eGFP-expressing NOG mice (NOG-EGFP) as a high purity cancer sampling system

Purpose

It is still technically difficult to collect high purity cancer cells from tumor tissues, which contain noncancerous cells. We hypothesized that xenograft models of NOG mice expressing enhanced green fluorescent protein (eGFP), referred to as NOG-EGFP mice, may be useful for obtaining such high purity cancer cells for detailed molecular and cellular analyses.

Methods

Pancreato-biliary cancer cell lines were implanted subcutaneously to compare the tumorigenicity between NOG-EGFP mice and nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice. To obtain high purity cancer cells, the subcutaneous tumors were harvested from the mice and enzymatically dissociated into single-cell suspensions. Then, the cells were sorted by fluorescence-activated cell sorting (FACS) for separation of the host cells and the cancer cells. Thereafter, the contamination rate of host cells in collected cancer cells was quantified by using FACS analysis. The viability of cancer cells after FACS sorting was evaluated by cell culture and subsequent subcutaneous reimplantation in NOG-EGFP mice.

Results

The tumorigenicity of NOG-EGFP mice was significantly better than that of NOD/SCID mice in all of the analyzed cell lines (p < 0.01). Sorting procedures enabled an almost pure collection of cancer cells with only slight contamination by host cells. Reimplantation of the sorted cancer cells formed tumors again, which demonstrated that cell viability after sorting was well maintained.

Conclusions

This method provides a novel cancer sampling system for molecular and cellular analysis with high accuracy and should contribute to the development of personalized medicine.     

非小细胞肺癌病人源性NOD/SCID小鼠和BALB/c裸鼠移植瘤模型的建立及比较

目的：构建人非小细胞肺癌的NOD/SCID小鼠和BALB/c裸鼠移植瘤模型,并探讨移植瘤成瘤性与小鼠品系间的关系,为后续建立优良的动物模型奠定基础。方法：取手术切除获得的23例非小细胞肺癌组织,1 h内移植于NOD/SCID小鼠或BALB/c裸鼠皮下,观察移植瘤成瘤情况,测量移植瘤体积,绘制生长曲线图,计算成瘤率、成瘤潜伏时间和成瘤时间,分析并比较移植成瘤组和未成瘤组所对应患者的相关临床病理指标,并取移植成瘤组和所对应患者的组织标本进行病理学分析。结果：NOD/SCID小鼠皮下移植瘤模型成瘤率（55.6%）,BALB/c裸鼠皮下移植瘤模型（20%）,两者间差异无统计学意义（P > 0.05）,但前者的成瘤潜伏时间和成瘤时间均短于后者（P < 0.05）。患者相关临床病理指标中鳞癌、TNM分期和淋巴结转移情况与移植瘤成瘤建模无明显相关（P=0.109、0.153、0.077）,NOD/SCID小鼠和BALB/c裸鼠皮下移植瘤模型均能保持病人肺癌肿瘤组织的形态学特征。结论：成功构建了NOD/SCID小鼠和BALB/c裸鼠皮下移植瘤模型,NOD/SCID小鼠更适宜用于肺癌病人源性皮下移植瘤模型的建立,非小细胞肺癌移植瘤模型的建立为进行体外抗肿瘤药物的筛选提供了良好的研究工具。     

A suitable model for experimental liver metastasis of human colon cancer xenografts using mice with severe combined immunodeficiency

Studies on liver metastasis of human colon cancer are limited because of a lack of suitable animal models. In this study, the usefulness of mice with severe combined immunodeficiency (SCID), which congenitally lack functional T and B lymphocytes, was evaluated in comparison with currently available nude mice. Three human colon cancer xenografts transplantable into nude mice were disaggregated enzymatically to obtain tumor cell suspensions, and implanted intrasplenically into SCID and nude mice. The incidence of splenic tumorigenesis and of liver metastases were significantly greater in SCID mice for all xenografts, in comparison with nude mice. In total, 33 of 36 SCID mice and 17 of 43 nude mice developed liver metastases. On the basis of this result, we conclude that SCID mice would be a more suitable model than nude mice for studying liver metastasis of human colon cancer. © 1993 Wiley-Liss, Inc.     

A surgical (right lobectomy) case of liver metastasis of colon cancer after chemotherapy using mFOLFIRI and cetuximab

A man in his early seventies underwent low anterior resection and partial resection of the liver for the rectal cancer and liver metastasis. However, 4 months after the surgery, he was found to have a liver tumor at S5 and S7 by abdominal CT scan. Then, he underwent chemotherapy (mFOLFOX6), but the metastatic tumor was progressive. We selected FOLFIRI + cetuximab regimen for second-line therapy to resect the metastatic tumor. As the metastatic lesion was become smaller after 4-course of the regimen including cetuximab, we decided to perform a radical resection. We conducted a right lobectomy of the liver, and the tumor was completely resected.     

Orthotopic models of cancer for preclinical drug evaluation: advantages and disadvantages

Considering the enormous effort that has taken place over the years to discover new chemotherapeutic drugs for treating the common cancers, the conventional murine and xenograft test systems used to test efficacy for drug development have identified only a limited number of useful agents that are active clinically at well tolerated doses. In recent years, considerable effort has been made to develop more clinically relevant models by the use of orthotopic transplantation of tumour material in rodents. It has been shown that it is now possible to transplant tumour material from a variety of tumour types into the appropriate anatomical site and often these tumours will metastasise in a similar manner and to similar locations as the same tumour type will in human cancer. As yet, although a body of literature has amassed on the technique itself and its implications for metastasis, there are relatively few laboratories using these test systems in drug development programmes. Nevertheless, given the expertise now being developed and some interesting observations being made on the role of the tumour site on response to therapeutic agents, it is likely that the use of orthotopic systems will strengthen our ability to select the most appropriate molecules for recommended use in clinical studies.     

Prevention of early-onset familial/hereditary colon cancer: new models and mechanistic biomarkers (review)

Human colon cancer is a multi-factorial, multi-step disease wherein genetic and dietary factors represent important regulators of initiation, promotion and progression. While the etiology of sporadic colon cancer remains largely unidentified, familial adenomatous polyposis (FAP) and hereditary non-polyposis colon cancer (HNPCC) represent predisposing genetic syndromes for early-onset familial/hereditary colon cancer. These syndromes are characterized by germ-line mutations in the adenomatous polyposis coli (APC) and/or DNA mismatch repair genes, respectively. Currently available preclinical animal models for human FAP and HNPCC syndromes, expressing clinically relevant germ-line mutations, exhibit adenomas in the small intestine rather than in the colorectum. These models are, therefore, subject to extrapolation for direct clinical translatability of the data for colon carcinogenesis and chemoprevention. Experimental models expressing clinically relevant genetic defects (APC and/or DNA mismatch repair gene mutations) in an appropriate target site (colon) may represent novel approaches that reduce extrapolation of the data for their clinical relevance. This report provides an overview on carcinogenesis and chemoprevention in preclinical models of FAP and HNPCC syndromes, and summarizes recent data on i) development of new cell culture models for FAP and HNPCC syndromes; and ii) validation of developed models for rapid, mechanism-based screening of new pharmacological or naturally occurring chemopreventive agents.     

Inhibition of haematogenous metastasis of colon cancer in mice by a selective COX-2 inhibitor, JTE-522

It is proposed that non-steroidal anti-inflammatory drugs (NSAIDs) reduce colorectal tumorigenesis by inhibition of cyclooxygenase (COX). COX is a key enzyme in the conversion of arachidonic acid to prostaglandins and two isoforms of COX have been characterized, COX-1 and COX-2. Multiple studies have shown that COX-2 is expressed at high levels in colorectal tumours and play a role in colorectal tumorigenesis. Recently it has been reported that selective inhibition of COX-2 inhibits colon cancer cell growth. In this study we investigated the effect of a selective COX-2 inhibitor (JTE-522) on haematogenous metastasis of colon cancer. For this purpose, we selected a murine colon cancer cell line, colon-26, that constitutively expresses the COX-2 protein. The subclone P expressed a high level of COX-2 and the subclone 5 expressed a low level. The colon-26 subclones were injected into the tail vein of BALB/c mice. JTE-522 was given intraperitoneally every day from the day prior to cancer cell injection, and the mice were sacrificed 16 days after cell injection. Lung metastases were compared between groups with and without JTE-522. In the mice injected with subclone P, the number of lung metastatic nodules was significantly reduced in the treated group. However, in the mice injected with subclone 5, there was little difference between the control and the treated groups. These results indicate that there may be a direct link between inhibition of haematogenous metastasis of colon cancer and selective inhibition of COX-2, and that selective COX-2 inhibitors may be a novel class of therapeutic agents not only for colorectal tumorigenesis but also for haematogenous metastasis of colon cancer.     

A case of pathological complete response after treatment with uracil/tegafur (UFT) and folinate (Leucovorin) for liver metastasis of colon cancer

The patient is a 62-year-old female who underwent a right hemicolectomy for type-2 ascending colon cancer (moderately-differentiated adenocarcinoma, ss, n0, H0, P0, M0, stage II). Six months after the surgery, a solitary metastatic focus was expressed in the liver S3. Because schizophrenia was present concurrently, tegafur and uracil/folinate (UFT/Leucovorin) treatment was selected and performed for 3 months. Because the tumor shrank afterward, a partial hepatectomy was performed to obtain a curative resection. In a pathological examination of the resected focus, cicatricial/necrotic findings were observed, but no cancer cells were observed; hence, it was determined to be a pathological complete response (CR). In regard to chemotherapy for distant metastasis of colorectal cancer, many molecular-targeted agents are being introduced, thus resulting in more treatment options; however, depending on the patient's background, UFT/LV treatment can be an effective treatment option.     

1,2-Dimethylhydrazine-induced colon carcinoma and lymphoma in msh2(-/-) mice

BACKGROUND: Defective mismatch repair (MMR) in humans is particularly associated with familial colorectal cancer, but defective repair in mice is generally associated with lymphoma in the absence of experimental exposure to carcinogens. Loss of MMR also confers resistance to the toxic effects of methylating agents. We investigated whether resistance to methylation contributes to increased susceptibility to colorectal cancer in mice by exposing mice with defects in the MMR gene msh2 to a methylating agent. METHODS: Tumor incidence and time of death in msh2(+/+), msh2(+/-), and msh2(-/-) mice were analyzed after weekly exposure (until tumor appearance) to the methylating agent 1,2-dimethylhydrazine (DMH). Chemically induced and spontaneous tumors were characterized by frequency, type, and location. The tumor incidence in untreated and treated mice of each genotype was compared by a Mann-Whitney U test. Carcinogen-induced apoptosis in histologic sections of small and large intestines was also determined. All statistical tests were two-sided. RESULTS: Homozygous inactivation of the msh2 gene statistically significantly accelerated (P<.0001) death due to the development of DMH-induced colorectal tumors and lymphomas. Rates of death from DMH-induced colorectal adenocarcinoma were similar in msh2 heterozygous and wild-type mice, but only msh2 heterozygotes (msh(+/-)) developed additional, noncolorectal malignancies (notably trichofolliculoma [two of 21], angiosarcoma of the kidney capsule [two of 21], and lymphoma [one of 21]), suggesting that heterozygosity for msh2 slightly increases DMH susceptibility. DMH induced apoptosis in small intestinal and colonic epithelial crypts that was dependent on active msh2. CONCLUSIONS: Inactivation of msh2 allows the proliferation of gastrointestinal tract cells damaged by methylating agents. Furthermore, MMR constitutes a powerful defense against colorectal cancer induced by DNA methylation.     

Efficient engraftment of primary adult T-cell leukemia cells in newborn NOD/SCID/beta2-microglobulin(null) mice.

Adult T-cell leukemia (ATL) develops via multiple oncogenic steps in human T-cell leukemia virus type I (HTLV-I) carriers. To better understand pathogenesis of ATL, we developed a novel xenogeneic engraftment model in which primary ATL cells are intravenously transplanted into neonatal nonobese diabetic (NOD)/severe-combined immunodeficiency (SCID)/beta2-microglobulin(null) (NOD/SCID/beta2m(null)) mice. Acute-type ATL cells engrafted in the peripheral blood and in the lymph nodes of recipients at a high efficiency. Engrafted ATL cells were dually positive for human CD4 and CD25, and displayed patterns of HTLV-I integration identical to those of donors by Southern blot analysis. These cells infiltrated into recipients' liver, and formed nodular lesions, recapitulating the clinical feature of each patient. In contrast, in smoldering-type ATL cases, multiple clones of ATL cells engrafted efficiently in NOD/SCID/beta2m(null) mice. When smoldering-type ATL cells were retransplanted into secondary NOD/SCID/beta2m(null) recipients, single HTLV-I-infected clones became predominant, suggesting that clones with dominant proliferative activity can be competitively selected in this xenogeneic system. Taken together, the NOD/SCID/beta2m(null) newborn system is useful to understand kinetics, metastasis, and disease progression of ATL in vivo.     

Radioimmunotherapy of human colon cancer xenografts by using 131I labeled-CAb1 F(ab')2

PURPOSE: Therapeutic efficacy, suitable dose, and administration times of 131I-CAb1 F(ab')2, a new monoclonal antibody therapeutics specifically directed against a cell surface-associated glycoprotein of colon cancer, were investigated in this article. METHODS AND MATERIALS: In human colon cancer xenografts, 131I-CAb1 F(ab')2 at the dose of 125 muCi, 375 muCi, and 1125 muCi were administrated intraperitoneally on Days 6 and 18 after implantation of HR8348 cells with CAb1 high reactivity. Survival time and tumor growth inhibition rate were used to evaluate the efficacy and safety of 131I-CAb1 F(ab')2 in treatment of colon cancer xenografts. RESULTS: Treatment of 125, 375, and 1125 muCi 131I-CAb1 F(ab')2 did not significantly decrease the mean survival time of nude mice when compared with nontreated groups (p = 0.276, 0.865, 0.582, respectively). Moreover, the mean survival times of nude mice receiving 375 muCi and 1125 muCi 131I-CAb1 F(ab')2 were significantly longer than that of 5-FU-treated groups (p = 0.018 and 0.042). Tumor growth inhibition rates of the first therapy were 35.67% and 41.37%, with corresponding 131I-labeled antibody dosage of 375 muCi and 1125 muCi. After single attack dosage, second reinforcement therapy may rise efficacy significantly. Tumor growth inhibition rates of 125 muCi, 375 muCi, and 1125 muCi 131I-labeled antibody on Day 20 posttherapy were 42.65%, 56.56%, and 84.41%, respectively. Histopathology examination revealed that tissue necrosis of various degrees was found in 131I-CAb1 F(ab')2-treated groups. CONCLUSION: 131I-CAb1 F(ab')2 is safe and effective for colon cancer. It may be a novel and potentially adjuvant therapeutics for colon cancer.     

RAD001 (everolimus) improves the efficacy of replicating adenoviruses that target colon cancer

Selectively replicating adenoviruses have the potential to cure cancer but have shown little efficacy in clinical trials. We have tested the ability of the mTOR kinase inhibitor RAD001 (everolimus) to enhance the response of xenografts to an oncolytic adenovirus. The virus has Tcf sites inserted in the early viral promoters and replicates selectively in cells with activation of the Wnt signaling pathway. To enhance tumor cell infection, an integrin targeting peptide (CDCRGDCFC) was inserted into the fiber gene of the virus. RAD001 combines three useful properties: it inhibits tumor cell growth directly, blocks angiogenesis, and suppresses the immune response. RAD001 does not block viral protein expression, DNA replication, or cytopathic effect in tumor cells in vitro. After 6 weeks of daily RAD001 treatment, ongoing viral DNA replication could be detected in tumor xenografts, showing that RAD001 does not inhibit virus replication in vivo. I.v. injection of virus alone produced a small delay in xenograft growth, whereas combination therapy substantially prolonged the survival of the mice. We suggest that collapsing the tumor vasculature after the initial infection traps the virus and facilitates local spread within the tumor. Unlike conventional drugs, which require continued access to the tumor through the vascular system, oncolytic viruses are in principle less sensitive to late reductions in perfusion because they are produced locally within the tumor.     

口服DNA疫苗pcDNA3.1+/flk-1(n1-7)抗小鼠结肠癌肝转移

背景与目的：血管内皮生长因子（vascular endothelial growth factor，VEGF）及其主要受体血管内皮生长因子受体-2（vascular endothelial growth factor receptor-2，VEGFR-2）在肿瘤血管生成过程中起着重要作用。VEG—FR-2在鼠中为flk-1，本研究以减毒沙门氏菌SL3261为载体制备了抗肿瘤血管生成口服DNA疫苗pcDNA3．1＋／flk-1（n1-7），研究该疫苗抗小鼠结肠癌肝转移的作用，并探讨其可能的作用机制。方法：RT—PCR法扩增鼠VEGFR-2胞外cDNA全长序列flk-1（n1-7），构建重组质粒pcDNA3．1＋／flk-1（n1-7）。将重组质粒转化减毒沙门氏菌SL3261，制备成以SL3261为载体的口服DNA疫苗。将18只小鼠随机分组，口服接种疫苗两周后，用CT-26细胞建立结肠癌肝转移动物模型，15d后处死小鼠，体视学方法计数肝脏的肿瘤结节数目。流式细胞仪检测小鼠外周血CD4^＋细胞和CD8^＋细胞变化。结果：构建成重组质粒pcDNA3．1（＋）／flk-1（n1-7），制备成以SL3261为载体的口服DNA疫苗pcD—NA3．1（＋）／flk-1（n1-7）。疫苗组小鼠肝脏内的肿瘤转移病灶明显低于对照组，P〈0．05。结论：成功构建了重组质粒pcDNA3．1（＋）／flk-1（n1-7），制备成以SL3261为载体的口服DNA疫苗pcDNA3．1（＋）／flk-1（n1-7）。这种疫苗能抑制小鼠结肠癌的肝脏转移。     

Improved engraftment of human acute myeloid leukemia progenitor cells in beta 2-microglobulin-deficient NOD/SCID mice and in NOD/SCID mice transgenic for human growth factors.

Primitive malignant progenitors defined as nonobese diabetic/severe combined immunodeficient (NOD/SCID) leukemia-initiating cells or NOD/SL-IC from patients with acute myeloid leukemia (AML) can be detected and quantitated in sublethally irradiated NOD/SCID mice. However, there is variability in the levels of bone marrow (BM) engraftment obtained after intravenous injection of cells from different AML samples. In the current study, AML cell engraftment in standard NOD/SCID mice was compared to that obtained with NOD/SCID mice transgenic for the human growth factor genes Steel factor (SF), interleukin-3 (IL-3) and granulocyte macrophage-colony-stimulating factor (GM-CSF) (N/S-S/GM/3) as well as beta 2 microglobulin-null NOD/SCID (N/S-beta 2m(-/-)) mice. Three of the eight AML samples that failed to engraft in standard NOD/SCID animals showed easily detectable and up to 70-fold increased in the number of leukemic cells in BM 8-12 weeks post-transplantation in each of the N/S-beta 2m(-/-) and N/S-S/GM/3 mouse strains. In two of the four AML samples studied at limiting dilution, the frequency of NOD/SL-IC detected was increased six- and seven-fold. Thus, in these novel mouse strains a broader spectrum of AML patient samples can be evaluated for their progenitor content and potentially studied for their response to innovative therapeutics in vivo.