1. Spectroscopic Window on Tumor Metabolism Michael Garwood, Ph.D. Univ. of Minnesota

2. Role of MRS in the Clinical Management of Cancer Diagnosis:  guide biopsy  avoid unnecessary/risky biopsies  ascertain aggressiveness/stage/prognosis Treatment:  guide choice of treatment  identify non-responders early → alter treatment regime  tool for follow up

3. Ackerstaff et al., J Cell Biochem 2003 High Res 1 H MRS of Cells Non-Malignant cells Malignant cells extract in vitro GPC → PCho switch Aboagye et al., Cancer Res 1999

4. R-CH 2 -CH 2 -N -C H H H C H H H - H H H - C + Choline-containing compounds

5. lipids suppressed water lipid Choline compounds (tCho) Frequency (ppm) 1 H MRS invasive ductal carcinoma MRI CMRR 4 Tesla In vivo 1 H MRS of breast cancer First reported studies: Roebuck et al, Radiol 1998; Gribbestad et al, JMRI 1998

6. Jacobs MA, Barker PB, et al. Proton magnetic resonance spectroscopic imaging of human breast cancer: a preliminary study. J Magn. Reson Imaging. 2004 Jan;19(1):68-75 Benign Focal Fibrosis Infiltrating ductal carcinoma P < 0.0008

7. Membrane Choline Phospholipid Metabolism Adapted from Aboagye EO, Bhujwalla ZM. Cancer Res 59:80-84 1999 Lysophosphatidic acid

8. Mechanisms of increased PC in cancer: Increased expression and activity of choline kinase [Ramirez de Molina et al., Oncogene 2002] Higher rate of choline transport [Katz-Brull & Degani, AntiCancer Res. 1996] Increased PLD activity [Noh et al., Cancer Lett. 2000] Increased PLA2 activity [Guthridge et al., Cancer Lett. 1994]

10. Glioblastoma Multiforme (High Grade Tumor) Cho NAALac FLAIR PPM4.03.02.01.0 T1T1 Cho Cr NAA Right Left slide courtesy of Peter Barker, Johns Hopkins U

11. Prostate Cancer Normal human prostate Tumor-bearing prostate Cheng LL, FEBS Lett. 2001

12. MRI/MRSI Targeted, TRUS-Guided Biopsies PSA - 12 ng/ml Two prior negative biopsies The sensitivity of TRUS guided biopsy is reduced in large prostates and when the cancer is located in difficult locations such as the apex or in the anterior or lateral aspects of the prostate. 55555 MR targeted TRUS guided biopsy positive Journal Urology 2000, 164(2) 400-404 The accuracy of cancer detection of MRI/MRSI targeted biopsy in men with prior negative biopsy ≈80%. (Yuen et al, J. Urol. 2004; Prando et al, Radiology 2005) courtesy of J.Kurhanewicz, UCSF

13. 13 Chemical Shift: Minimized with higher BW pulses X Y Z RF Gradients 90180 180 0 Standard pulses 90 0 Spectrum Center Frequency (-235 Hz) FF FF %CS :  F / BW RF FF Broadband pulses Courtesy of: G. Metzger

14. 14 OVS with over-prescription Courtesy of: G. Metzger

15. 15 Prostate Spectroscopy at 3T: Single Voxel Echo Time, Coupling and SNR TE = 260 ms TE = 100 ms Courtesy of: P. Choyke & G. Metzger

16. 16 Case Study: Slice 5 Cho Sp Cre Cit Courtesy of: P. Choyke & G. Metzger

17. Quantification Metabolite ratios (eg, tCho/NAA, (tCho+Cr)/Cit) External reference (eg, phantom of known conc) Reference to tissue water signal

18. a) 0123456 b) tCho 3 Tesla Normal breast MRI Devices 4-ch coil 3x3x3 cm voxel LASER Localization TE Averaging (60-300ms in 128 increments) NEX=2

19. Breast Anatomy Anatomy varies greatly Tissues are distributed heterogeneously  Intravoxel lipids are inevitable Lobules Fat Stroma Tavassoli, 1999 Adipose tissue Fibroglandular tissue Netter, 1997

20. Internal Referencing with Water [tCho] expressed in molal units (mmol tCho/kg water) No assumptions about volume or density NOT assuming constant water concentration Assuming a two-compartment model (water & fat) and all tCho is in the aqueous compartment Bolan et al., MRM 2003

21. Spectral Fitting Adapted TDFDFit (Slotboom et al., MRM 1998)  Time-Domain Model:  Minimize residuals in frequency-domain over narrow (0.4 ppm) band Fit 3 peaks independently: tCho, water, 1.3 ppm lipid Errors from Cramer-Rao Minimum Variance Bound; used for detection threshold model data residual 6 402 ppm Bolan et al., MRM 2003

22. Normal gland (Presumed) [tCho] = 0.75 ± 0.07 mmol/kg volume = 13.0 mL lipid fraction = 3.5% Invasive Ductal Carcinoma [tCho] = 6.8 ± 0.1 mmol/kg volume = 6.8 mL lipid fraction = 8% Atypical Hyperplasia [[tCho] = 1.5 ± 0.8 mmol/kg volume = 1.1 mL lipid fraction = 15% Bolan et al., MRM 2003

23. Frequency (ppm) invasive ductal carcinoma no Cho

24. Frequency (ppm) Reason for false negative? Spurious lipid sideband peaks! invasive ductal carcinoma

25. Sideband Artifacts -500 Hz -300 -100 100300 TE (ms) 45 57 sidebands water sidebands Sidebands have coherent, TE-dependent phase Averaging causes destructive interference Bolan et al., MRM 2002 Antisymmetric side peaks Amplitude >1% Caused by B 0 oscillation

26. Echo-time Averaging ppm -20468 Conventional single TE TE averaging No tCho tCho? NEX=64 TE=45ms TE=45-196ms 64 increments 2 Bolan et al., MRM 2002

27. In vivo 1 H spectrum of a voxel containing mainly adipose tissue

28. [Cho] = 0.642 mmol/kg Voxel of just the enhancing region [Cho] = 0.910 mmol/kg [Cho] = 0 mmol/kg Voxel of just the non enhancing region Day 127 (AC x 4 followed by Taxotere x 3) size = 3.0 x 2.7 x 3.0 cm 3

29. [tCho] = 0 ± 1.73 Lipid H2OH2O 71234560 ppm All 4 readers maintained their decision to biopsy Invasive Ductal Carcinoma SI time (sec) 4 2 3 0 0 1 013245 PrecontrastPostcontrastSubtraction tCho time (min) Meisamy et al, Radiology 2005

30. Conclusions about MRS for breast cancer diagnosis:  Adding quantitative 1 H MRS to breast MRI improves sensitivity, specificity, and accuracy, over MRI alone  Quantitative 1 H MRS is particularly useful in cases where lesion morphology and time-intensity curves are indeterminate Meisamy et al, Radiology 2005

31. Treatment Planning and Monitoring

32. MRSI for Radiation Treatment Planning of Brain Tumor Cho/CrGradeDose painting <10 ≥1-215040 ≥2-3≥2-325940 ≥3≥337020 MRSI-based radiation dose painting using the IMRT method Thakur, Chang, Huang, Koutcher, Narayana Memorial Sloan-Kettering Cancer Center

33. Models of tCho response Measured acute response to PX-478 (inhibits HIF1-alpha production) in mouse xenografts of HT-29 (colon) Methods: in vivo MRS at 4.7T, ex vivo validation Results: tCho dropped significantly at 12 and 24 hrs Jordan et al., NMR Biomed 2006Al-Safar et al., Cancer Res 2006 Measured acute response to MN58b (inhibits CK) in mouse xenografts of MDA- MB-231 (breast) and HT-29 (colon) Methods: in vivo MRS at 4.7T, ex vivo validation Results: tCho dropped significantly at 48hrs in both models PCho CK cell density

34. Neoadjuvant chemotherapy (primary systemic therapy, PST) is the preferred treatment for locally advanced breast cancer (Fisher et al. J Clin Oncol 1997, 1998) Advantages:  Tumor shrinkage; possible breast conserving procedures  In vivo monitoring of chemo-sensitivity (customize Tx  complete pathologic response) Treatment Monitoring in Breast Cancer

35. 4T Tx Monitoring in Breast Cancer: Results to Date 14/18 Responders had a decrease in [tCho] at Day 1 9/10 Non- responders had a increase in [tCho] at Day 1 Day 1 Rule: 82% accuracy in 28 subjects 0 1 2 3 4 5 6 7 8 9 [tCho] (mmol/kg) 0 1 2 3 4 5 6 7 8 9 BaselineDay 1BaselineDay 1 RespondersNon-Responders [tCho] (mmol/kg) Meisamy et al, Radiology 2004

36. Pre PST24 hrs AC X 1 AC X 4 [tCho] = 4.6 LD = 4.0 cm [tCho] = 3.7 LD = 4.0 cm [tCho] = 0.9 LD = 1.7 cm Responder to AC Meisamy et al, Radiology 2004

37. Pre PST24 hrs AC X 1 AC X 4 Taxol X 2 [tCho] = 4.1 LD = 1.7 cm [tCho] = 4.6 LD = 4.0 cm [tCho] = 3.7 LD = 4.0 cm [tCho] = 0.9 LD = 1.7 cm Responder to AC, but not Taxol Meisamy et al, Radiology 2004

38. choline citrate Therapeutic Selection and Monitoring Baseline Metabolic Atrophy 1 year Metabolic Atrophy 5 years courtesy of J.Kurhanewicz, UCSF

39. Is it possible to predict response from baseline MRS data?

40. Treatment Prediction / Phenotyping Inconsistent findings in brain MRS: Tzika, Neuroradiology 2001 – responders had lower tCho Preul, Neurosurgery 2000 – no difference Lazareff, J Neurooncol 1999 – no difference Baseline [tCho] was higher in responders than in non-responders (p=0.03) Higher [tCho] @ baseline associated with higher grade & positive nodes Can MRS identify responders before starting treatment?

41. Pretreatment 31 P spectrum from nodal disease of a HNSCC patient who experienced partial response Pretreatment PME/NTP ratio Pretreatment PME/NTP ratios from tumors; complete responders were different from incomplete response group P<0.001 Preliminary results with 31 P MRSI A. Shukla-Dave, et. al. Acad Radiol, 9:688-694, 2002

42. 31P MRS in Bone Sarcoma Zakian, et. al., Cancer Research 2003 Dec 15;63(24):9042-7 Baseline spectrum

43. Baseline Energetics Predicts Outcome in Bone Sarcoma Zakian, et. al., Cancer Research 2003 Dec 15;63(24):9042-7  NTP/Pi predicts longer survival

44. Future: More studies correlating with pathology, immunohistochemistry, and outcomes Further studies to assess reliability/reproducibility Results of multi-center trials Combine with other metrics (DCE-MRI, ADC,…) → multiparametric analyses 3T (and higher?)

45. Prostate spectroscopy at 1.5T with endorectal coil IMAPS (1.5T) The axial T2-weighted image (A) is used for matching voxel locations to histopathological specimens (D). One of the spectral maps (B), partially expanded in (E), reflects the quality of the MRSI data throughout the slice. Deviations in the (Cho + Cr)/Ci metabolite ratio map in (C) largely correspond to the tumor location indicated with the blue line in (D). Courtesy of T. Scheenen and Prof. A. Heerschap, Radboud University Nijmegen Medical Center, Dept. of Radiology The IMAPS community

46. Slide courtesy of Michael Jacobs, JHU

47. Current Multiparametric (MRI/DTI/MRSI) Prostate Imaging Exam Polyamines Creatine 3.0PPM2.02.5 Citrate Choline Creatine 3.02.02.5 Lipid HealthyCancer T 2 weighted MRI Diffusion weighted MRI ADC Map MRSI (0.3 cc) Elevated choline Reduced citrate Reduced polyamines Decreased Signal Intensity on T2 weighted Imaging Reduced water diffusion Slide courtesy J. Kurhanewicz UCSF

48. 3T MRSI vs 1.5T MRSI: Improved Detection of Residual Cancer 3T 1.5T Cho 0.16 cc 0.34 cc Choline Creatine

49. U of Minn Researchers Patrick Bolan Greg Metzger Sina Meisamy Adeka McIntosh Curt Corum Angela Styczynski Nate Powell Djaudat Idiyatullin Jang-Yeon Park Carl Snyder James Boyum Doug Yee Michael Nelson Tim Emory Lenore Everson Todd Tuttle Evin Gulbahce Tommy Vaughan Thanks for Sending Slides Arend Heerschap Jason Koutcher John Kurhanewicz Michael Jacobs Peter Barker Wei Huang Funding Sources National Institutes of Health (CA92004, RR08079) Acknowledgements

50. Biomarker of Effective Therapy - Metabolic Atrophy Pickett, Int J Radiat Oncol Biol Phys 2004; 60:1047 Pickett, Int J Radiat Oncol Biol Phys 2004; 59:665 water On average increasing metabolic atrophy paralleled decreasing serum PSA with time after Brachytherapy and EBRT, but occurred significantly earlier. The presence of residual abnormal metabolism did not correlate with PSA in individual patients after radiation.

51. Biomarker of Therapeutic Failure - Elevated Choline The presence of three or more abnormal voxels (choline/creatine >1.5) demonstrated a sensitivity and specificity of 87% and 72%, respectively, and an overall accuracy of 81% for the diagnosis of local recurrent/residual disease. Choline Creatine Coakley, Radiology 2004; 233:441-448. Cancer Radiation atypia

52. MRI/MRSI based Radiation Planning MRI/MRSI was used to identify prostate anatomy and up to 2 DIL’s in patients undergoing HDR Brachytherapy treatment planning. The DIL dose was successfully escalated to a minimum of 120%, while simultaneously treating the entire prostate and without increasing the dose to surrounding normal tissues. Pouliot J, Int J Radiat Oncol Biol Phys 2004; 59:1196-1207.

53. 3T MRSI: Increased Spatial and Spectral Resolution 5 5 5 5 1 3 4 5 1 1 2 1 1 1 1 1 1 1 1 1 1 0.15 cc Spatial Resolution Spectral Data using a MLEV-PRESS sequence PSA - 7.5 Gleason 3+3 cancer - right midgland and apex Cunningham CH, Magn Reson Med 2005; 53:1033- 1039 135 Cr + polyamine Cho polyamine Reduced partial voluming and increased resolution of choline from polyamines Early ECE

54. Future: High Field (3T) Multi-parametric Approach - MRI/MRSI/DTI Cancer ADC Map T2-Weighted FSE Choline Patient 1: Biopsy- confirmed prostate cancer in apex prior to therapy Polyamine Ceatine ADC map T2-Weighted FSE Choline Patient 2: Recurrent/residual prostate cancer 3 years after hormone therapy. Creatine

55. Diagnosing Suspicious Breast Lesions at 4T 0 2 4 6 8 10 Malignant (n=58)Benign (n=54)Normal (n=5) [tCho] (mmol/kg)

56. Diagnosing Suspicious Breast Lesions at 4T 0 2 4 6 8 10 Malignant (n=58)Benign (n=54)Normal (n=5) [tCho] (mmol/kg) ROC cutoff = 1.0 mmol/kg sensitivity 72% specificity 83% ROC cutoff = 1.0 mmol/kg sensitivity 72% specificity 83%