Targeting Chaperone Dependence in Rhabdomyosarcoma

Amit J. Sabnis MD, Bivona LaboratoryUCSF Helen Diller Family Comprehensive Cancer Center

UCSF Benioff Children’s Hospital

Two Paradigms for Developing Cancer Therapies

Oncogene Dependence Non-Oncogene Dependence

Luo J et al., Cell 2009

HSP70 Chaperones are Cellular Buffers Against Proteotoxic Stress

Hypothesis: HSP70 inhibition will engage stress responses in rhabdomyosarcoma cells

and lead to cell death.

The HSP70 Inhibitor MAL3-101 Induces Apoptosis in Rhabdomyosarcoma (RMS) Cells

• Patient-derived rhabdomyosarcoma cell lines RMS13, Rh41, and Rh18 show micromolar sensitivity to MAL3-101.

• MAL3-101 induces apoptosis in sensitive cell lines.

RMS13 Cells: 24 Hr Treatment

The Integrated Stress Response (ISR) Links Chaperone Failure to Apoptosis

Hypothesis: Activation of the integrated stress response leads to cell death after MAL3-101 treatment.

Probing the Mechanism of Action of MAL3-101 with Derived Isogenic Resistant Cell Lines

Parental Line(MAL3-101 Sensitive)

Isogenic Resistant Lines(MAL3-101 Resistant)

2 Month MAL3-101

Dose Escalation

Treat with MAL3-101Measure Integrated Stress Response

RMS13 I.R. #1 - 4

ISR Activation is a Hallmark of MAL3-101 Sensitivity

ISR Activation is a Hallmark of MAL3-101 Sensitivity

Loss of CHOP Promotes Resistance to MAL3-101

CHOP Knockdown by shRNA Sensitivity to MAL3-101

Conclusions

• MAL3-101 is an HSP70 inhibitor that induces apoptosis in rhabdomyosarcoma cells.

• Apoptosis is a consequence of engaging the integrated stress response, and is CHOP dependent.

• Failure to upregulate CHOP after HSP70 inhibition is a biomarker of resistance to MAL3-101.

• Loss of CHOP causes partial resistance to MAL3-101.

Future Directions

• Test whether other stress sensors cooperate with the ISR in inducing apoptosis after HSP70 inhibition.

• Carry out an unbiased shRNA screen to comprehensively identify mechanisms underlying resistance in I.R. lines.

• Test MAL3-101 for activity in explant models of RMS as a prelude to clinical use.

Acknowledgements

Bivona LabTrever BivonaJonathan ShueAnin SayanaLuping LinCollin BlakelyEvangelos Pazarentzos

Support

Damon Runyon-Sohn Pediatric Cancer Fellow (A.J.S.)

St. Baldrick’s Foundation Fellow (A.J.S.)

HCIA Award (T.B.)

CollaboratorsJeff Brodsky (U. Pittsburgh)

Chris GuerrieroPeter Walter (UCSF)

Diego Acosta-AlvearCarmela Sidrauski

Jonathan Weissman (UCSF)Jason Gestwicki (UCSF)