Patient-derived xenografts faithfully replicated clinical outcome in a phase II co-clinical trial of arsenic trioxide in relapsed small cell lung cancer

Abstract

Background: SCLC has limited treatment options and inadequate preclinical models. Promising activity of arsenic trioxide (ASO) recorded in conventional preclinical models of SCLC supported the clinical evaluation of ASO in patients. We assessed the efficacy of ASO in relapsed SCLC patients and in corresponding patient-derived xenografts (PDX).

Methods: Single arm, Simon 2-stage, phase II trial to enroll patients with relapsed SCLC who have failed at least one line of therapy. ASO was administered as an intravenous infusion over 1-2 h daily for 4 days in week 1 and for 2 days in weeks 2-6 of an 8-week cycle. Treatment continued until disease progression. Pretreatment tumor biopsy was employed for PDX generation through direct implantation into subcutaneous pockets of SCID mice without in vitro manipulation and serially propagated for five generations. Ex vivo efficacy of cisplatin (3 mg/kg i.p. weekly) and ASO (3.75 mg/kg i.p. every other day) was tested in PDX representative of platinum sensitive and platinum refractory SCLC.

Results: The best response in 17 evaluable patients was stable disease in 2 (12 %), progressive disease in 15 (88 %) patients and median time-to-progression of seven (range 1-7) weeks. PDX was successfully grown in 5 of 9 (56 %) transplanted biopsy samples. Serially-propagated PDXs preserved characteristic small cell histology and genomic stability confirmed by immunohistochemistry, short tandem repeat (STR) profiling and targeted sequencing. ASO showed in vitro cytotoxicity but lacked in vivo efficacy against SCLC PDX tumor growth.

Conclusions: Cisplatin inhibited growth of PDX derived from platinum-sensitive SCLC but was ineffective against PDX from platinum-refractory SCLC. Strong concordance between clinical and ex vivo effects of ASO and cisplatin in SCLC supports the use of PDX models to prescreen promising anticancer agents prior to clinical testing in SCLC patients. Trial Registration The study was registered at http://www.clinicaltrials.gov (NCT01470248).

Keywords: Arsenic trioxide; Clinical trial; Efficacy; Ex vivo; Patient-derived xenograft; Small cell lung cancer; Survival.

Fig. 1

Subcutaneous growth of patient-derived xenograft in a SCID mouse host just prior to euthanasia. Harvested tumor from bilateral subcutaneous pockets; 3 × 3 mm sized sections were immediately propagated to the next generation of mice through implantation into subcutaneous pockets over the hind legs of the mice without in vitro manipulation (top panel). Histopathologic confirmation of small cell lung carcinoma histology by hematoxylin and eosin stain (X400) and immunohistochemistry for neuroendocrine differentiation showing intense diffusely positive staining for CD56 (middle panel), moderately intense staining for synaptophysin and focal areas of weakly positive chromogranin A staining (bottom panel)

Fig. 2

Electropherogram showing unique profile of each of five different PDX models generated from SCLC. Note the identical STR profile of tumor samples harvested from animals bearing first generation (passage 1) and second generation (passage 2) PDX

Fig. 3

SCLC cell line derived directly from a tumor biopsy specimen employed for the generation of TKO-002 PDX was employed for in vitro cytotoxicity assessment. TKO-002 cells were seeded in 96-well plates and allowed to grow overnight. Exponentially growing cells were treated the next day with vehicle or serially increasing concentrations of cisplatin (2–216 μM) and ASO (0.2–24 μM). After 72 h of continuous drug exposure, cell numbers were estimated using MTS assay. IC₅₀ concentration was estimated from the growth inhibition using GraphPad prism software. The IC₅₀ concentration for cisplatin (top panel) and ASO (middle panel) was estimated at 11.25 and 3.08 μM, respectively. There was no demonstrable additive or synergistic effect of the combination of ASO (6 μM) and cisplatin (2.5 or 10 μM) over each agent alone against TKO-002 cells (bottom panel)

Fig. 4

Efficacy of ASO (3.75 mg/kg i.p. every other day) and cisplatin (3 mg/kg i.p. weekly) was tested in TKO-002, a PDX model of platinum refractory SCLC. Tumor volume (mm³) and body weight of animals were measured at least twice weekly while on treatment. There was no significant tumor growth inhibition by ASO (*p = 0.48) or cisplatin (**p = 0.42) in comparison to vehicle-treated control animals at the end of the treatment period (top). There was also no significant difference in the harvested tumor weights from animals treated with ASO compared to control animals treated with vehicle (*p = 0.33) (middle). There was no significant increase in toxicity (measured by body weight of the animals) with active therapy in comparison to controls (bottom)

Fig. 5

To assess the efficacy of ASO and cisplatin (CDDP) in TKO-005, a PDX model of platinum sensitive SCLC, animals were treated and monitored for tumor growth and body weight as described in Fig. 4. In addition, a matching group of tumor-bearing mice was treated with rigosertib (250 mg/kg i.p. daily). At the end of the treatment period, there was no significant reduction in tumor volume in animals treated with ASO (*p = 0.40) but a significant reduction was achieved with cisplatin (**p = 0.048) and a strong trend toward reduced tumor volume was noted with rigosertib (p = 0.058) in comparison to vehicle-treated control animals. Similarly, harvested tumor weights were significantly lower from animals treated with cisplatin (**p = 0.04) and rigosertib (p = 0.038) but not from animals treated with ASO (*p = 0.46) in comparison to control animals. There was no significant increase in toxicity as measured by body weight of the animals on active therapy in comparison to controls. Furthermore, rigosertib (ON-01910.Na) efficacy was comparable to cisplatin both in terms of growth inhibition (p = 0.24) and harvested tumor weights (p = 0.32) at the end of treatment

Fig. 6

Efficacy of ASO and cisplatin (CDDP) singly and in combination was tested in TKO-002, a PDX model of platinum refractory SCLC. Kaplan–Meier survival curves for animal groups treated with vehicle, ASO (7.5 mg/kg i.p. daily), cisplatin (3 mg/kg i.p. weekly) and the combination of ASO plus cisplatin. There was significant toxicity with rapid death of mice treated with ASO alone or in combination with cisplatin (top). A reduced dose of ASO (3.75 mg/kg every other day) was better tolerated but showed negligible efficacy and failed to potentiate the minimal growth inhibition achieved by cisplatin (3 mg/kg i.p. weekly) in this PDX model derived from a patient with platinum resistant SCLC (middle and bottom panel)