`
`Priority Report
`
`A Model of Postsurgical Advanced Metastatic Breast Cancer
`
`More Accurately Replicates the Clinical Efficacy of
`
`Antiangiogenic Drugs
`
`Eric Guerin, Shah Man, Ping Xu, and Robert S. Kerbel
`
`Abstract
`
`The failure rate of randomized phase HI oncology clinical trials is extremely high, even when preceded by
`encouraging preclinical studies and phase II trial results of the same therapy. Thus, there is considerable effort
`being made to improve the predictive clinical potential of preclinical models, in addition to improving phase II
`trial design. With respect to the former, preclinical models have historically relied heavily on treatment ofprimary
`spontaneous or transplanted tumors rather than the more common and therapeutically challenging clinical trial
`circumstance ofadvanced metastatic disease. Here, we show that the oral antiangiogenic tyrosine kinase inhibitor
`(TKI), sunitinib, which failed to meet primary or secondary survival endpoints in 4 separate phase HI metastatic
`breast cancer (MBC) trials, either alone or with chemotherapy, similarly failed to show monotherapy or
`combination chemotherapy efficacy in a model of postsurgical advanced MBC using a metastatic variant of
`the MDA—MB—231 triple-negative human breast cancer. In contrast, the drug was effective when used to treat
`established orthotopic primary tumors. Similar results were obtained with pazopanib monotherapy, another
`antiangiogenic oral TKI. However, when an antibody targeting the VEGF pathway (DClOl) was tested, it showed a
`trend in modestly improving the efficacy of paclitaxel therapy, thus resembling to a degree prior phase HI clinical
`results of bevacizumab plus paclitaxel in MBC. Our results suggest the potential value of treating postsurgical
`advanced metastatic disease as a possible strategy to improve preclinical models for predicting outcomes in
`patients with metastatic disease. Cancer Res; 73(9); 2743—8. ©2013 AACR.
`
`Introduction
`
`An enduring problem in oncology experimental therapeu-
`tics has been the limited value ofmodels involving treatment of
`tumor-bearing mice to consistently predict outcomes later
`assessed in clinical trials, particularly at the randomized phase
`III level (1—4). A common scenario observed is positive and
`sometimes even remarkable preclinical activity, which is then
`followed by complete failure in the clinic (1—4). Such failures
`add substantially to the cost of approved agents as well as
`exposing patients with cancer enrolled in such trials to inef—
`fective therapies. As a result, there is considerable effort to
`identify potential causes for this discrepancy and develop
`significantly improved preclinical models (1—4) such as genet—
`ically engineered mouse models (GEMM) of cancer and
`patient-derived xenografts (PDX) as opposed to the historically
`
`more common use of transplantation of established cultured
`tumor cell lines grown as solid primary tumors.
`Although many factors have been proposed for the discrep-
`ant therapeutic outcomes observed between preclinical and
`clinical studies (1—4), one factor, which has received scant
`attention, is the failure to duplicate in mice treatment of
`advanced visceral metastatic disease (5, 6). Most phase I and
`II solid tumor clinical trials and the majority of phase III trials
`involve patients with such disease. In many or most cases the
`primary tumor has been surgically resected. The failure rate is
`extremely high in phase HI metastatic therapy trials (7) and
`when therapies succeed, the benefits in survival are frequently
`incremental (8). Therefore we have developed several models
`of postsurgical advanced metastatic disease using established
`human tumor cell lines grown in immunodeficient mice to
`mimic the more challenging circumstance of treating patients
`with metastatic disease (5). In most cases, the cell lines used are
`variants previously selected in viva for aggressive spontaneous
`metastatic spread after the primary orthotopic tumor has been
`surgically resected (5). One such variant, called LM2-4, was
`serially selected in Viva from the commonly used MDA-MB-231
`triple-negative human breast cancer cell line (9).
`Here, we report the use of the aforementioned postsurgical
`model of LM2—4 to evaluate the impact of several antiangio—
`genic drugs, used alone or in combination with paclitaxel
`chemotherapy, and compare the results obtained with con-
`ventional treatment of established primary tumors. One of the
`Genentech 2081
`
`Authors' Affiliations: Department of Medical Biophysics, Biological
`Sciences Platform, Sunnybrook Research Institute, University of Toronto,
`Toronto, Ontario, Canada
`Note: R.S. Kerbel holds a Tier 1 Canada Research Chair.
`
`Corresponding Author: RS. Kerbel, Biological Sciences Platform, Sun-
`nybrook Research institute, Dept. of Medical Biophysics, University of
`Toronto, 8-217, 2075 Bayview Avenue, Toronto, ON M4N 3M5, Canada.
`Phone: 416-480-5711; Fax: 416—480-5884.
`
`doi: 10.1158/0008-5472.CAN-12-4183
`©2013 American Association for Cancer Research.
`
`www.aacrjourna|s.org
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`H
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`/44(Q AmericanAssociazimzfar CancerResearch
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`2743
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`Hospira v. Genentech
`IPR2017-00737
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`1
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`Guerin et a1.
`
`drugs we tested is sunitinib (Sutent), an oral tyrosine kinase
`inhibitor (TKI), which targets VEGF receptors (VEGFR) and
`platelet—derived growth factor receptors, among several others
`(10). Based partly on very encouraging preclinical results in 3
`different established primary breast cancer models (a trans-
`genic model, a chemically-induced rat model, and a human
`tumor xenograft model; ref. 10) and a bone colonization
`experiment (10), sunitinib was subsequently evaluated in
`patients with metastatic breast cancer (MBC); 4 independent
`phase III trials were undertaken (1 1- 15), 3 in combination with
`chemotherapy (paclitaxel, or docetaxel, or capecitabine). All 4
`trials failed to meet efficacy endpoints of survival (1 1-15). This
`stands in contrast to a phase HI trial involving the anti-VEGF
`antibody, bevacizumab (Avastin), when used with chemo-
`therapy, for example, paclitaxel, which provided a clinical
`benefit, at least in progression—free survival (PFS), though not
`in overall survival (OS; ref. 16). We also tested another anti-
`angiogenic TKI, pazopanib and a monoclonal antibody that
`targets the mouse VEGFR—2 (DC101). Sunitinib and DC101
`were also evaluated with concurrent paclitaxel chemotherapy.
`The purpose of these studies was to further validate the
`preclinical strategy of using postsurgical models of advanced
`metastatic disease to predict clinical outcomes involving
`treatment of patients with metastatic disease by addressing
`the following questions and comparing the results with prior
`phase III trial outcomes: (i) is it the case that antiangiogenic
`drug monotherapy has reduced or no therapeutic benefit when
`treating mice with advanced metastatic disease in contrast to
`established primary tumors? (ii) What is the impact on out-
`comes when chemotherapy is used in combination with the
`antiangiogenic agent? And, (iii) is there a difference in out-
`comes when using TKIs versus antibodies in combination with
`chemotherapy?
`
`Materials and Methods
`
`Female CB-17 severe combined immunodeficient (SCID)
`mice were purchased from Charles River, and female YFP
`SCID mice (17) were bred in house from breeding pairs
`generously provided by Dr. Janusz Rak (McGill University,
`Montreal). Mice at 6 to 8 weeks of age were used. MDA-MB
`231/LM2.4 is a variant cell line of MDA-MB 231 selected in
`
`Viva for aggressive spontaneous metastatic spread from
`established but resected primary tumors and was grown in
`cell culture as previously described (9). Cell line authenti-
`cation was carried out by genotyping using Illumina mouse
`linkage panel and confirmed to be human in origin. Routine
`mycoplasma screening is carried out in—house using com-
`mercial kits, which confirmed the cell line is mycoplasma
`free. Mammary fat pad injection (2 x 106 cells) was carried
`out as previously described (9). Weekly caliper measure-
`ments were carried out to determine tumor growth and
`tumor volume was calculated using the formula azb/Z where
`a is the width and b is the length. Treatment of primary
`tumors was initiated when average volume was approxi—
`mately 100 to 150 mms, that is, 12 to 15 days after cell
`injection. Surgical resection of the primary tumors was
`carried out when the average tumor size was 400 1111113.
`
`All mice were randomized just before initiation oftreatment
`Antiangiogenic drugs were generously provided by the man-
`ufacturers, namely, sunitinib (Pfizer), pazopanib (GSK), and
`DC101, the monoclonal antibody targeting mouse VEGFR—Z
`(ImClone/Eli Lilly). All drugs were prepared according to
`manufacturer's specifications. Paclitaxel was dispensed by
`Sunnybrook Pharmacy Department, Odette Cancer Center
`(Toronto, Ontario, Canada) at 60 mg/mL and further diluted
`with normal saline to the appropriate concentration. Control
`mice received either vehicle and/or normal saline as appro-
`priate. Sunitinib was administered by gavage at 60 mg/kg dose
`daily for the first 14 days followed by 5 days daily with 2 days
`break thereafter to reduce toxicity as measured by weight loss.
`Pazopanib 150 mg/kg was administered by gavage daily with-
`out interruption. Paclitaxel was administered intraperitoneally
`(i.p.) at 50 mg/kg every 3 weeks in the studies, which included
`combination with DC101 but the dose and schedule was
`
`changed to 30 mg/kg once every 2 weeks in studies, which
`involved combination with sunitinib or pazopanib to minimize
`toxicity observed in SCID mice (18).
`
`Results and Discussion
`
`We first tested sunitinib on the grth of primary estab-
`lished orthotopic (mammary fat pad) tumors. Cells from the
`established variant of MDA-MB—231 called LM2.4, which was
`selected in viva for aggressive spontaneous metastatic spread
`after surgical resection of the primary tumor (9) were injected
`into the mammary fat pad of 6- to 8-week-old female SCID
`mice, as described previously (9) in Materials and Methods. We
`did not use a luciferase-tagged clone of LM2.4 (19) because we
`have found that these cells have a reduced ability for sponta-
`neous metastasis (unpublished observations), the basis of
`which is currently unknown. When the primary tumors
`reached a volume of approximately 100 to 150 m3, sunitinib
`was administered daily by gavage at a preclinically effective
`dose of 60 mg/kg, and the treatment continued until end point.
`As shown in Fig. 1A, a robust growth delay was observed,
`similar to Abrams and colleagues using another human breast
`cancer xenograft model (called MX—I; ref. 10). In the case of
`Abrams and colleagues, this was paralleled by an increase in
`OS, the extent of which was shown to be further enhanced by
`combination with chemotherapy, for example, docetaxel (10).
`In our case, the mice were sacrificed in this preliminary
`experiment at an earlier defined endpoint, namely, when the
`control group tumors reached an average size ofapproximately
`500 mm3 (Fig. 1A). However, in another experiment, shown
`in Fig. 1B where the primary tumor was surgically resected
`(at day 20) and the same therapy initiated 3 weeks later (i.e.,
`when the mice have established visceral metastatic disease
`
`based on numerous previous studies, e.g., ref. 9 in addition to
`reproducibility and lack of variability of the short median
`survival times), no impact of the same treatment on survival
`was observed. This pattern of discrepancy in outcomes is not
`specific for sunitinib as we observed a similar pattern using
`pazopanib (as shown in Fig. 1C and D) where primary tumors
`in control mice were allowed to grow in this case to endpoint of
`1,700 mm3 and the therapy maintained until endpoint.
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`
`
`
`
`
`Days
`
`Initiation
`
`of therapy
`
`Vehicle control
`
`Pazopanib
`
`_._ Vehicle control
`-I— Pazopanib 150 mg/kg
`
`
`
`D 110
`100
`90
`
`so
`70
`so
`so -----------
`40
`30
`20
`
`
`
`Percentsurvival
`
`10
`O
`
`
`
`50
`60
`70
`80
`
`Days after cell implantation
`
`
`
`
`
`Differential Drug Efficacy on Primary versus Metastatic Tumors
`
`700
`
`600
`
`500
`
`400
`300
`
`200
`100
`
`g:
`E
`v
`g
`§
`5
`E
`:3
`
`Vehicle control
`
`Sunitinib 60 mg/kg
`
`B 110
`
`100
`_ 9°
`E 80
`E
`70
`a 60
`0
`g
`50
`._
`40
`g:
`30
`20
`
`
`
`
`— Vehicle control
`— Sunitinib 60 mg/kg
`(therapy initiated at day41)
`
`
`
`
`
`
`
`40
`
`50
`
`60
`
`70
`
`80
`
`90
`
`Days after implantation
`
`.
`
`
`
`C
`
`2000
`
`1,750
`
`isinmooo
`1.000
`750
`
`500
`
`250-
`
`
`
`
`
`Tumorvolume(mm3)
`
`5
`
`10
`
`35
`30
`25
`20
`15
`Days after cell implantation
`
`40
`
`45
`
`50
`
`Treatment
`initiated on day 14
`
`
`Figure 1. Differential impact of sunitinib or pazopanib monotherapy on primary breast tumor growth versus postsurgical advanced metastatic breast
`cancer. Sunitinib administered daily inhibits primary tumor growth (A), but has no survival benefit when treating advanced metastatic disease (B). The bottom
`shows similar results with pazopanib administered daily (C and D). For A and B, 2 x 106 MDA—MB 231/LM2-4 cells were implanted into the mammary fat
`pad of 6 08-17 SCID female mice; in the primary tumor study. treatment with sunitinib was initiated 12 days later when average tumor size was 100 mma;
`in the advanced metastasis therapy study, primary tumors were surgically resected 20 days after cell injection when average size was approximately 400 mm3
`and sunitinib treatment was initiated 21 days later. For C and D, 2 x 106 MDA-MB 231/LM2-4 cells were implanted in the primary tumor study (C),
`pazopanib treatment was initiated 14 days later when average tumorsize was 150 mm3; in the advanced metastasis therapy study (D), the primary tumors were
`surgically resected 20 days after cell injection when average size was approximately 400 mm3 and treatment was initiated 19 days later.
`
`to
`We next assessed the -impact of adding paclitaxel
`sunitinib. Shown in Fig. 2A and B is the impact on survival
`of mice with advanced metastatic disease when maximum
`
`tolerated dose (MTD) paclitaxel was combined with suniti-
`nib. Two independent experiments were carried out. If
`anything,
`there was a trend to reduced survival
`in the
`combination treatment group compared with the mice
`receiving paclitaxel alone, an observation that is consistent
`with a phase III breast cancer trial of sunitinib plus another
`taxane, docetaxel (12), where no added benefit in PPS was
`observed and OS was slightly reduced in the combination
`treatment group (12). We would note that in the 2 experi—
`ments shown in Fig. 2A and B, there was a difference in the
`therapeutic impact of paclitaxel alone. In one experiment
`(shown in Fig. 2A) the paclitaxel monotherapy treatment
`had no statistically significant benefit in median OS, whereas
`
`it did in the experiment shown in Fig. 2B. This difference
`may be due to the more aggressive tumor growth we noted
`in the experiment shown in Fig. 2A where all of the control
`mice died by day 60 compared with experiment 2B where all
`control mice died by day 70. Nevertheless in both experi-
`ments, sunitinib did not improve paclitaxel treatment out-
`comes. We also tested the effect of paclitaxel plus sunitinib
`in the established primary tumor model (Fig. 2C); sunitinib
`had a noticeable antitumor effect in contrast to paclitaxel;
`the 2 drug combination was not significantly different from
`the sunitinib-treated group.
`We next tested the antiangiogenic anti-VEGFR-2 antibody
`known as DC101 (20). The rationale for doing so was based on
`the results of the prior E2100 phase III trial evaluating bev-
`acizumab with paclitaxel in patients with MBC, which showed
`a gain in PPS of almost 6 months compared with paclitaxel
`
`
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`www.aacrjournals.org
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`Cancer Res; 73(9) May 1, 2013
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`Guerin et al.
`
`—-— Vehicle control
`
`—-— Sunitinib 60 mg/kg
`
`-- Paclitaxel so mg/kg*#
`-‘- Sunitinib + paclitaxel
`
`
`
`Percentsurvival
`
`-— Vehicle control
`—I— Sunitinib 60 mg/kg
`_._ Paclitaxel 30 mg/kg
`-I- Sunitinib + paclitaxel
`
`
`
`Percentsurvival
`
`75
`65
`55
`Da safterim lantation
`y
`p
`
`.
`.
`Median survnval:
`Vehicle control: 54 days
`Paclitaxel: 61 days
`Sunitinib: 65 days
`Sunitinib + paclitaxel: 50 days
`
`Therapy initiated on day 14
`
`so
`Days after implantation
`
`511
`
`4'0
`Median survival:
`Vehicle control: 65 days
`Paclitaxel: 88.5 days
`Sunitinib: 66 days
`Sunitinib + paclitaxel: 76 days
`
`Vehicle control
`Paclitaxel
`
`Sunitinib
`Sunitinib + paclitaxel
`
`10
`
`35
`3D
`25
`20
`15
`Days after cell implantation
`
`40
`
`45
`
`Figure 2. Differential impact of sunitinib plus paclitaxel chemotherapy when treating established primary tumors versus postsurgical advanced
`metastatic disease. Two independent metastatic therapy experiments are shown in A and B. Paclitaxel alone administered intraperitoneally at an MTD of 30
`mg/kg once every 2 weeks shows extensions of median survival in A and B, which was not statistically significant in one case (A). Combination of sunitinib with
`paclitaxel does not improve the survival advantages over paclitaxel alone (8) and may even worsen outcome (A). In the advanced metastasis studies,
`median survival for (i) control vehicle treatment was 54 to 65 days, (ii) paclitaxel treatment was 61 to 88.5 days, (iii) sunitinib treatment was 65 to 66 days, and (iv)
`sunitinib plus paclitaxel was 50 to 76 days; P values were not significant for A; B, ‘, paclitaxel versus control P z: 0.03; #, sunitinib + paclitaxel versus
`control P = 0.003. In established primary tumors, MTD paclitaxel alone shows no activity, whereas sunitinib alone or in combination with MTD
`paclitaxel inhibited tumor growth (C). In the advanced metastasis study (B), primary tumors were surgically resected 20 days after injection of 2 x 106 cells
`when average size was approximately 400 mm3 and treatment was initiated 20 days later; in the primary tumor study (0), treatment was initiated 14 days
`after cell injection when average tumor size was 150 mma.
`
`alone (16). As shown in Fig. 3A, DC101 had a robust antitumor
`effect when used to treat established LM2.4 primary tumors.
`The paclitaxel treatment, once again, did not have an antitu—
`mor effect when used to treat primary tumors. The 2 drug
`combination was the most effective. However, as shown in Fig.
`3B the pattern of response as assessed by survival analysis was
`somewhat different in the postsurgical advanced metastatic
`setting. Similar to sunitinib or pazopariib, DC101 was seem-
`ingly devoid ofactivity, at least when used as monotherapy and
`assessed by its impact on survival, because median survival was
`64 to 65 days in both the control and DC101 treatment groups.
`The paclitaxel monotherapy treatment showed a trend for a
`survival benefit (from 65 to 78 days) but this did not reach
`
`statistical significance. However, median survival was signif-
`icantly prolonged to 88 days in the combination treatment
`group compared with control, untreated mice. The difference
`in median survival between the DC101 + paclitaxel group
`(88 days) and the paclitaxel group (78 days) was not statistically
`significant, but would likely have led to a benefit in PPS had we
`been able to undertake such an assessment, and if so, would
`mirror the results of the E2100 phase III trial. Taken together,
`there does seem to be a modest benefit in improving the
`therapeutic impact of paclitaxel by combination with the
`VEGF pathway targeting antibody, but not with sunitinib,
`observations that seem to reflect previous phase III clinical
`trial results (13, 16), including a head-to—head comparison of
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`Cancer Res; 73(9) May 1, 2013
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`4
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`Differential Drug Efficacy on Primary versus Metastatic Tumors
`
`2,500
`
`2,250
`2,000
`1,750
`
`1,500
`1,250
`1,000
`750
`500
`
`250
`
`
`
`
`
`Tumorvolume(mm3)
`
`Control
`
`
`
`
`
`Paclitaxel
`
`1 1 0
`1 00
`90
`80
`7O
`60
`50
`40
`30
`20
`1 0
`
`
`
`Percentsurvival
`
`
`
`
`-'- Control
`
`-'- Paclitaxel 50 mg/kg
`"" DC101
`
`i D0101 + paclitaxel""+
`
`
`
`40
`
`so
`
`100110120
`so
`so
`7'0
`so
`Days after cell implantation
`
`D0101+paclitaxel
`
`70
`
`80
`
`
`
`
`10
`
`20
`
`30
`
`40
`
`50
`
`60
`
`Days after cell implantation
`
`
`
`Median survival:
`Control: 65 days
`Paclitaxel: 78 days
`D0101: 64 days
`DC101+ paclitaxelz 88 days.
`_ ____——l
`
`Figure 3. Differential impact of an antibody to VEGFFt-2 (D0101) plus MTD paclitaxel on established orthotopic primary LM2-4 tumors versus postsurgical
`advanced metastatic disease. D0101 800 rig/mouse administered intraperitoneally twice per week showed marked antitumor activity when treating
`established primary tumors (A) but no survival activity in the setting of postsurgical advanced metastasis therapy (B); MTD paclitaxel 50 mg/kg i.p. once every 3
`weeks showed no activity in the primary tumor therapy model but showed an increase, although not statistically significant, in median survival in the
`postsurgical advanced metastasis model; combination of D0101 with MTD paclitaxel PTX increased the extent of inhibition of primary tumor growth (A)
`and had a significant survival benefit in advanced metastatic model (B). in the advanced metastasis studies (B), median survival for (i) control treatment was
`65 days, (ii) paclitaxel treatment was 78 days, (iin DC101 treatment was 64 days, and (iv) 00101 + paclitaxel was 88 days; ‘, D0101 + paclitaxel
`versus control P .— 0.0006; + D0101 + paclitaxel versus PTX P = 0.42. In the primary tumor study (A), treatment with D0101 was initiated 15 days later when
`average tumor size was 150 mm"; in the advanced metastasis study (B), primary tumors were surgically resected 21 days after cell injection when
`average size was approximately 400 mm3 and treatment was initiated 25 days later.
`
`sunitinib plus paclitaxel versus bevacizumab plus paclitaxel
`(13) as well as the known modest to minimal PFS activity of
`single-agent paclitaxel in metastatic triple-negative breast
`cancer (16).
`Several questions are raised by our results. First, what is the
`basis for the widely divergent effects we have observed when
`treating primary tumor-bearing mice versus mice with postsur-
`gical advanced metastatic disease with antiangiogenic drugs?
`Some possibilities include reduced expression of VEGF, or
`VEGFR—2 in the tumor or tumor vasculature of established
`
`metastases compared with the primary tumors. The qualitative
`characteristics of the vasculature in the slightly "older" metasta-
`ses may be substantially different from the primary tumors such
`that there is a greater proportion of late/mature vessels that are
`known to be less responsive to VEGF pathway targeting drugs
`(21). Metastases in vasculature rich organs such as the lung and
`liver may be more adept at co-opting the existing vasculature
`than tumors growing in the mammary fat pad (22). Second.
`would a similar pattern of results be observed if using GEMMs or
`PDXs? With respect to GEMMs, surgical resection ofthe multiple
`asynchronously arising primary tumors and the well-known
`observed lack of distant metastases in most such models make
`
`this difficult to answer. Nevertheless, some GEMM primary
`tumor therapy studies have shown a remarkable retrospective
`correlation with prior phase 111 clinical trial PFS or OS results of
`
`the respective tumor types (lung and pancreatic cancer) in part
`by using clinically relevant endpoints of tumor response (23).
`As for PDXs, some recent studies have shown the presence of
`metastases in clinically relevant patterns in nonresected primary
`tIHnor-bearing patient-derived breast cancer xenografis obtain-
`ed from patients with different breast cancer major subtypes
`(24), thus making it possible to use these models for preclinical
`adjuvant and metastatic therapy investigations.
`Finally, is there any evidence that an investigational therapy
`previously shown to be highly effective in the postsurgical
`preclinical metastatic setting shows prospective clinical
`activity? In this regard, we have reported that doublet oral
`low-dose metronomic chemotherapy using cyclophosphamide
`and a 5-fluorouracil prodrug (UFT), that is, tegafur + uracil,
`had potent activity in the postsurgical metastatic setting using
`the LM2-4 breast cancer model (9), whereas the activity was
`much less impressive when treating primary tumors in control
`unresected mice (9). A similar version of this metronomic
`chemotherapy was tested in phase II MBC trial, in combination
`with bevacizumab, with very encouraging results (25). How-
`ever, as discussed in the Introduction, such phase H trial results
`have to be confirmed in a larger randomized phase III trial,
`of which one is underway evaluating metronomic doublet
`cyclophosphamide and capecitabine with bevacizumab
`(NCT01131195; www.clinicaltrials.gov). Nevertheless, for now,
`
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`Cancer Res; 73(9) May 1, 2013
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`Guerin et al.
`
`our results suggest the value of preclinical modeling postsur-
`gical advanced metastatic disease as a potential strategy to
`improve how they might predict clinical outcomes. The same
`may be the case for postsurgical adjuvant therapy models of
`early stage microscopic (minimal residual) metastatic disease,
`which can also respond to therapy in a manner different from
`primary tumors in control experiments (19, 26). Although not
`practical for routine drug screening, use of such models may be
`useful to confirm prior preclinical studies using conventional
`primary tumor therapy models, before embarking on expensive
`phase II or III metastatic therapy clinical trials.
`
`Disclosure of Potential Conflicts of Interest
`No potential conflicts of interest were disclosed
`
`Authors' Contributions
`Conception and design: E. Guerin, S. Man, R.S. Kerbel
`Development of methodology: S. Man, R.S. Kerbel
`Acquisition of data (provided animals, acquired and managed patients,
`provided facilities, etc.): E. Guerin, S. Man
`
`References
`
`Analysis and interpretation ofdata (e.g., statistical analysis, hiostatistics,
`computational analysis): E. Guerin, S. Man
`Writing, review, and/or revision ofthe manuscript: E. Guerin, S. Man, P. Xu,
`R.S. Kerbel
`Administrative, technical, or material support (i.e., reporting or orga—
`nizing data, constructing databases): S. Man, P. Xu
`Study supervision: R.S. Kerbel
`
`Acknowledgments
`The authors thank GSK through Dr. Rakesh Kumar for pazopanib, Pfizer
`through Dr.J. Christensen for sunitinib, and ImClone System through Dr. Bronek
`Pytowski for the DC101 antibody.
`
`Grant Support
`This work was supported by grants from the Ontario Institute for Cancer
`Research, the Canadian Breast Cancer Foundation, and the NIH (CA—41233;
`R.S. Kerbel).
`The costs ofpublication ofthis article were defrayed in part by the payment of
`page charges. This article must therefore be hereby marked advertisement in
`accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
`
`Received November 14, 2012; revised February 1, 2013; accepted March 5, 2013;
`published OnlineFirst April 22, 2013.
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`Cancer Research
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`Cancer Res; 73(9) May 1, 2013
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`Cancer Research
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`The journal of Cancer Research (1916—1930)
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`| The American Journal of Cancer (1 931 ~1940)
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`for Cancer Research
`AAC-R American Association
`
`A Model of Postsurgical Advanced Metastatic Breast Cancer More
`Accurately Replicates the Clinical Efficacy of Antiangiogenic Drug:
`
`Eric Guerin, Shari Man, Ping Xu, et al.
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`Cancer Res 2013;713:2743-2748. Published OnlineFirst April 22, 2013.
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